Prediction of hormone sensitivity for breast cancers.
The classic action that leads to transcriptional activation of estrogen response genes mediated through estrogen receptors (ER) and the estrogen complex plays a pivotal role in the development of ER-positive breast cancers. In addition to this pathway, non-classic action and non-genomic action, both estrogen-dependent and estrogen-independent genomic actions have also been found to contribute to ER-positive tumor growth. Although the details of these mechanisms are not well known, participation of the growth factor signaling pathway is likely to be the most significant factor for acquisition of resistance to hormonal therapy. This resistance is mediated not only directly through cell growth promotion by growth factor signaling, but also through enhancement of alternative ER signaling pathways in addition to classic action. The reason why tamoxifen-insensitive ER-positive breast cancers respond to aromatase inhibitors may be explained, at least in part, by the different estrogen-related signaling pathways in which aromatase inhibitors may block estrogen signaling. In this paper we discuss the molecular mechanisms for resistance to hormonal therapy based on an understanding of estrogen signaling pathways.


Oxidative stress and ischemic injuries in heat stroke.
When rats were exposed to high environmental temperature (e.g., 42 or 43 degrees C), hyperthermia, hypotension, and cerebral ischemia and damage occurred during heat stroke were associated with increased production of free radicals (specifically hydroxyl radicals and superoxide anions), higher lipid peroxidation, lower enzymatic antioxidant defenses, and higher enzymatic pro-oxidants in the brain of heat stroke-affected rats. Pretreatment with conventional hydroxyl radical scavengers (e.g., mannitol or alpha-tocopherol) prevented increased production of hydroxyl radicals, increased levels of lipid peroxidation, and ischemic neuronal damage in different brain structures attenuated with heat stroke and increased subsequent survival time. Heat shock preconditioning (a mild sublethal heat exposure for 15min) or regular, daily exercise for at least 3 weeks, in addition to inducing overproduction of heat shock protein 72 in multiple organs including brain, significantly attenuated the heat stroke-induced hyperthermia, hypotension, cerebral ischemia and damage, and overproduction of hydroxyl radicals and lipid peroxidation. The precise function of heat shock protein 72 are unknown, but there is considerable evidence that these proteins are essential for survival at both normal and elevated temperatures. They also play a critical role in the development of thermotolerance and protection from oxidative damage associated with cerebral ischemia and energy depletion during heat stroke. In addition, Shengmai San or magnolol (Chinese herbal medicines) or hypervolemic hemodilution (produced by intravenous infusion of 10% human albumin) is effective for prevention and repair of ischemic and oxidative damage in the brain during heat stroke. Thus, it appears that heat shock protein 72 preconditioning induced by prior heat shock or regular exercise training, as well as pretreatment with Shengmai San or magnolol is able to prevent the oxidative damage during heat stroke. On the other hand, hypervolemic hemodilution, Shengmai San, or magnolol is able to treat the oxidative damage after heat stroke onset.


Uterine epithelial changes during placentation in the viviparous skink Eulamprus tympanum.
We used scanning electron microscopy (SEM) and transmission electron microscopy (TEM) to describe the complete ontogeny of simple placentation and the development of both the yolk sac placentae and chorioallantoic placentae from nonreproductive through postparturition phases in the maternal uterine epithelium of the Australian skink, Eulamprus tympanum. We chose E. tympanum, a species with a simple, noninvasive placenta, and which we know, has little net nutrient uptake during gestation to develop hypotheses about placental function and to identify any difference between the oviparous and viviparous conditions. Placental differentiation into the chorioallantoic placenta and yolk sac placenta occurs from embryonic Stage 29; both placentae are simple structures without specialized features for materno/fetal connection. The uterine epithelial cells are not squamous as previously described by Claire Weekes, but are columnar, becoming increasingly attenuated because of the pressure of the impinging underlying capillaries as gestation progresses. When the females are nonreproductive, the luminal uterine surface is flat and the microvillous cells that contain electron-dense vesicles partly obscure the ciliated cells. As vitellogenesis progresses, the microvillous cells are less hypertrophied than in nonreproductive females. After ovulation and fertilization, there is no regional differentiation of the uterine epithelium around the circumference of the egg. The first differentiation, associated with the chorioallantoic placentae and yolk sac placentae, occurs at embryonic Stage 29 and continues through to Stage 39. As gestation proceeds, the uterine chorioallantoic placenta forms ridges, the microvillous cells become less hypertrophied, ciliated cells are less abundant, the underlying blood vessels increase in size, and the gland openings at the uterine surface are more apparent. In contrast, the yolk sac placenta has no particular folding with cells having a random orientation and where the microvillous cells remain hypertrophied throughout gestation. However, the ciliated cells become less abundant as gestation proceeds, as also seen in the chorioallantoic placenta. Secretory vesicles are visible in the uterine lumen. All placental differentiation and cell detail is lost at Stage 40, and the uterine structure has returned to the nonreproductive condition within 2 weeks. Circulating progesterone concentrations begin to rise during late vitellogenesis, peak at embryonic Stages 28-30, and decline after Stage 35 in the later stages of gestation. The coincidence between the time of oviposition and placental differentiation demonstrates a similarity during gestation in the uterus between oviparous and simple placental viviparous squamates.


Altered reward processing underpins emotional apathy in dementia.
While apathy is broadly defined as a loss of motivation, it is increasingly recognised as a multidimensional syndrome spanning executive, emotional, and initiation domains. Emotional apathy is purportedly driven by deficits in using socioemotional rewards to guide behaviour, yet the link between these symptoms and reward processing, and their common neural correlates, has not been directly examined. Sixty-four patients (33 behavioural-variant frontotemporal dementia, 14 Alzheimer's disease, 8 semantic dementia, 6 progressive nonfluent aphasia, 3 logopenic progressive aphasia) were classified into high (HEA; n = 36) and low (LEA; n = 28) emotional apathy groups based on emotional apathy subscale scores on the Dimensional Apathy Scale. Patients and age-matched healthy controls (n = 27) performed an instrumental reward learning task where they learned to associate cues with either social or monetary outcomes. HEA patients showed impaired learning on both the social and monetary reward conditions, relative to LEA patients (p = 0.016) and controls (p = 0.005). Conversely, the LEA group did not differ from controls (p = 0.925). Importantly, multiple regression analyses indicated that social reward learning significantly predicted emotional apathy. Voxel-based morphometry analyses revealed that emotional apathy and social reward learning were both associated with orbitofrontal cortex, ventral striatum, and insula atrophy. Our results demonstrate a unique link between impaired social reward learning and emotional apathy in dementia and reveal a shared neurobiological basis. Greater understanding of these neurocognitive mechanisms of reward processing will help improve the identification of emotional apathy in dementia and inform the development of novel interventions to address these symptoms.


Poly(I:C) induces anti-inflammatory response against secondary LPS challenge in zebrafish larvae.
Poly(I:C) is known as an agonist of the TLR3 receptor which could prime inflammation and elicit the host immune response, which is widely applied as adjuvant or antivirus treatment. However, the negative effects of poly(I:C) on regulating immune response to protect the host from inflammatory diseases remain largely unknown. Here, we established an in vivo model to pre-treat zebrafish larvae with poly(I:C) at 2 dpf, then challenge them with LPS at 6 dpf, and found that poly(I:C) training could significantly alleviate the LPS challenge-induced septic shock and inflammatory phenotype. Moreover, the poly(I:C)-trained larvae exhibit decreased number of macrophages, but not neutrophils, after secondary LPS challenge. Furthermore, training the larvae with poly(I:C) could elevate the transcripts of mTOR signaling and heighten the H3K4me3-mediated epigenetic modifications. And interestingly, we found that inhibiting the H3K4me3 modification, rather than mTOR signaling, could recover the number of macrophages in poly(I:C)-trained larvae, which is consistent with the observations of inflammatory phenotypes. Taken together, these results suggest that poly(I:C) training could induce epigenetic rewiring to mediate the anti-inflammatory response against secondary LPS challenge-induced septic shock through decreasing macrophages' number in vivo, which might expand our understanding of poly(I:C) in regulating fish immune response.


Change in head posture and character of nystagmus in a patient with neurological upbeat nystagmus.
To report a case of a patient with chin-up head posture and presumed congenital toxoplasmosis chorioretinal scars, who had a change in the character of the nystagmus and therefore the head posture following treatment for a neurological upbeat nystagmus. A 5 month old female presented with a chin up head posture and upbeat nystagmus. Magnetic resonance imaging of the brain revealed an arachnoid cyst in the area of the pineal gland. Nine months after cyst-peritoneal shunt surgery, the upbeat nystagmus was dampened but change in character to a rotary nystagmus worse on the left gaze. The patient had assumed a left face turn, shifting the null point from the vertical to the horizontal plane. The left face turn was successfully corrected at age eight years with a Kestenbaum procedure. This case emphasizes the possibility of having two distinct types of nystagmus associated with two etiologies. In this case, an acquired upbeat nystagmus secondary to an arachnoid cyst, and a congenital left rotary nystagmus from the chorioretinal scars. Furthermore, there can be a change in head position and character of nystagmus after treating the cause of the central motility disorder, thereby affecting the choice and timing of surgical intervention to correct the head positioning.


Pneumothorax in AIDS: case reviews and proposed clinical management.
Pneumothorax is a not uncommon complication of advanced HIV infection, and may prove difficult to manage in view of its recalcitrant and recurrent nature. In this group where immunosuppression and reduced life expectancy are a feature, standard protocols are often abandoned in favour of a more conservative approach. This is often unsuccessful. Patients attending the Department of Genitourinary Medicine, Chelsea and Westminster Hospital who sustained pneumothorax between 1988 and 1992 were identified retrospectively and their notes reviewed. Fifteen patients were identified of whom three had post-procedural pneumothoraces. In the remaining 12 patients, 10 had previously had Pneumocystic carinii pneumonia (PCP), whilst all 12 had some evidence to suggest current PCP (seven proven, five presumptive). In those six patients with a single, unilateral pneumothorax, four were managed successfully with intercostal drainage alone (one patient died early, one required pleurectomy). In those with recurrent pneumothoraces or pneumothoraces that did not respond to prolonged intercostal drainage, failure of medical treatment was judged to have occurred and surgery was performed. Overall, conservative management failed in 7/11 patients. Conversely surgery resulted in resolution in 7/7 with recurrence seen in one individual. Median survival was similar in the two groups. Pneumothorax in patients with AIDS is associated with a high rate of intercurrent PCP; a low threshold for treating this infection presumptively is indicated. Intercostal drainage was successful in patients with a single, unilateral pneumothorax. However, in patients with recurrent or bilateral pneumothorax extended periods on intercostal drainage were uniformly unsuccessful. Early surgical referral should be considered in this group.


Cervical cancer screening in Bulgaria--past and present experience.
In Bulgaria the previously (1970-1985) existing population based cervical cancer screening was replaced in the early 1990s with an opportunistic model due to political and socioeconomic reasons. As a result, in the last 20 years, cervical cancer incidence and mortality rates steady increased. The objective of the EUROCHIP project in Bulgaria was to evaluate the readiness of the health system as well as health providers' attitudes to implementation in the country of a population based screening program for cervical cancer. Using a structured questionnaire, a convenience sample of medical specialists representing different actors involved in cervical cancer prevention, treatment, financing and policy were interviewed. The majority of interviewed practitioners worried that organization and implementation of an effective population-based cervical cancer screening program is not possible in the current unstable health system. A nostalgic attitude to the cervical cancer screening, performed in the past and pessimistic view on the capability of the current health system to cope are strong. As main barriers to implementation of an effective program were pointed financial and organizational ones. Motivation for gynecologists to perform smear test should include better information, organization and payment. Medical specialists in Bulgaria are aware of the alarming rates of cervical cancer incidence and mortality in the country. However, due to the insufficient communication and interaction between policy makers and front-line health care staff, they do not have enough information on the ongoing programs. Absence of health policy regarding screening is considered as main barrier for implementation of an effective screening program.


Kidney-resident macrophages promote a proangiogenic environment in the normal and chronically ischemic mouse kidney.
Renal artery stenosis (RAS) caused by narrowing of arteries is characterized by microvascular damage. Macrophages are implicated in repair and injury, but the specific populations responsible for these divergent roles have not been identified. Here, we characterized murine kidney F4/80+CD64+ macrophages in three transcriptionally unique populations. Using fate-mapping and parabiosis studies, we demonstrate that CD11b/cint are long-lived kidney-resident (KRM) while CD11chiMϕ, CD11cloMϕ are monocyte-derived macrophages. In a murine model of RAS, KRM self-renewed, while CD11chiMϕ and CD11cloMϕ increased significantly, which was associated with loss of peritubular capillaries. Replacing the native KRM with monocyte-derived KRM using liposomal clodronate and bone marrow transplantation followed by RAS, amplified loss of peritubular capillaries. To further elucidate the nature of interactions between KRM and peritubular endothelial cells, we performed RNA-sequencing on flow-sorted macrophages from Sham and RAS kidneys. KRM showed a prominent activation pattern in RAS with significant enrichment in reparative pathways, like angiogenesis and wound healing. In culture, KRM increased proliferation of renal peritubular endothelial cells implying direct pro-angiogenic properties. Human homologs of KRM identified as CD11bintCD11cintCD68+ increased in post-stenotic kidney biopsies from RAS patients compared to healthy human kidneys, and inversely correlated to kidney function. Thus, KRM may play protective roles in stenotic kidney injury through expansion and upregulation of pro-angiogenic pathways.


Characterization of the kinase activity of a WNK4 protein complex.
Mutations in WNK4 protein kinase cause pseudohypoaldosteronism type II (PHAII), a genetic disorder that is characterized by renal NaCl and K(+) retention leading to hypertension and hyperkalemia. Consistent with this, WNK4 is known to regulate several renal tubule transporters, including the NaCl cotransporter, NCC, and the K(+) channel, ROMK, but the mechanisms are incompletely understood, and the role of the kinase activity in its actions is highly controversial. To assay WNK4 kinase activity, we have now succeeded in expressing and purifying full-length, enzymatically active WNK4 protein from HEK293 cells. We show that full-length wild-type WNK4 phosphorylates oxidative stress response kinase 1 (OSR1) and Ste20/SPS1-related proline/alanine-rich kinase (SPAK) in vitro. Introducing the PHAII-associated mutations, E559K, D561A, and Q562E, into our protein had no significant effect on this phosphorylation. We conclude that PHAII is unlikely to be caused by abnormal WNK4 kinase activity. We also made the intriguing observation that inactivating mutations of the WNK4 kinase domain did not completely abolish in vitro phosphorylation of OSR1/SPAK. Led by this, we identified a novel 40-kDa kinase that associates specifically with the COOH-terminal half of WNK4 and is able to phosphorylate both WNK4 and SPAK/OSR1. We suggest that this 40-kDa kinase functions in the WNK4 signal transduction pathway and may mediate some of the physiological actions attributed to WNK4.


Intermittent availability of ethanol does not always lead to elevated drinking in mice.
Intermittent access (IA) to an alcohol (ethanol) solution can lead rats to higher ethanol intakes than continuous access, and a recent report showed increased drinking in C57BL/6J mice offered 20% ethanol vs. water 3X/week (Prior studies have offered ethanol during 24 h periods, either continuously or intermittently.). We tested the high-preference C57BL/6J inbred mice: we also studied High Drinking in the Dark (HDID) mice, a line we have selectively bred to reach intoxicating blood ethanol levels after a short period of access to a single bottle of 20% ethanol. Neither HDID or C57BL/6J male mice offered ethanol every other day during only a 4-h access period showed greater daily intake than mice offered ethanol daily for 4 h. There was a small increase in drinking with 24 h IA in C57BL/6J mice. An experiment with HDID mice and their control heterogeneous stock stock modeled closely after a published study with C57BL/6J mice (Hwa, Chu, Levinson SA et al. Persistent escalation of alcohol drinking in C57BL/6J mice with intermittent access to 20% ethanol. Alcohol Clin Exp Res 2011;35:1938-1947) showed no significant elevation with 24 h IA exposure in either sex of any genotype. Finally, a near replication of the Hwa et al. study showed modestly greater intake in C57BL/6J mice, confirming the efficacy of 24 h IA. We conclude that 4 h of IA is likely insufficient to elevate drinking in mice. The lack of effect in HDID mice and their controls further suggests that not all genotypes respond to intermittency.


Modelling predicts that heat stress, not drought, will increase vulnerability of wheat in Europe.
New crop cultivars will be required for a changing climate characterised by increased summer drought and heat stress in Europe. However, the uncertainty in climate predictions poses a challenge to crop scientists and breeders who have limited time and resources and must select the most appropriate traits for improvement. Modelling is a powerful tool to quantify future threats to crops and hence identify targets for improvement. We have used a wheat simulation model combined with local-scale climate scenarios to predict impacts of heat stress and drought on winter wheat in Europe. Despite the lower summer precipitation projected for 2050s across Europe, relative yield losses from drought is predicted to be smaller in the future, because wheat will mature earlier avoiding severe drought. By contrast, the risk of heat stress around flowering will increase, potentially resulting in substantial yield losses for heat sensitive cultivars commonly grown in northern Europe.


Rhabdomyosarcomatous differentiation in gastrointestinal stromal tumors after tyrosine kinase inhibitor therapy: a novel form of tumor progression.
Approximately 80% of advanced metastatic gastrointestinal stromal tumors (GISTs) respond to treatment with the tyrosine kinase inhibitor (TKI) imatinib mesylate. However, the majority of patients suffer disease progression at a median of 2 years due to drug resistance. In general, progressing GISTs retain their typical morphology. Herein, we report 5 cases of progressing metastatic GIST with heterologous rhabdomyoblastic differentiation after TKI treatment. Histologic, immunohistochemical, and mutational analyses were performed on histologically classic GISTs and components with rhabdomyoblastic differentiation. There were 3 men and 2 women (ranging from 35 to 66 y of age). Three tumors were localized at presentation (2 stomach and 1 small bowel) and 2 presented with metastases. All localized primary tumors were high risk. Two tumors showed spindle cell morphology and 3 were epithelioid, including 1 with marked pleomorphism. After resection of the 3 localized primary tumors, intra-abdominal (2 patients) and liver (1 patient) metastases developed. All patients were treated with imatinib and showed partial clinical responses (4 patient) or stable disease (1 patient). Four patients subsequently progressed; 2 patients were treated with sunitinib after progression with minor responses. Four patients underwent surgical debulking. At last follow-up (range: 20 to 87 mo), 2 patients died of disease, 2 were alive with metastatic disease resistant to TKIs, and 1 was alive without evidence of disease. In all cases, rhabdomyoblastic differentiation was identified adjacent to areas with classic GIST morphology in at least 1 metastatic site; in 1 case, the primary tumor (after treatment with TKIs) showed heterologous differentiation. The rhabdomyoblastic components showed strong and diffuse positivity for desmin and expressed myogenin, whereas KIT was negative in the rhabdomyoblastic component in all cases. Primary KIT mutations were detected in both the conventional GIST and rhabdomyoblastic components from all patients: KIT exon 11 mutations in 4 cases and a platelet-derived growth factor receptor alpha gene exon 18 deletion in 1 case. No secondary mutations of the type associated with TKI resistance were identified in the rhabdomyoblastic areas. This is the first report of rhabdomyoblastic differentiation occurring in GISTs that progressed on TKI therapy. It is associated with loss of KIT expression, but retention of the receptor tyrosine kinase mutation of the precursor GIST. The rhabdomyoblastic differentiation can represent a diagnostic pitfall. The molecular mechanisms for this form of TKI-resistant clonal evolution remain to be determined.


Morality and management: an oxymoron? fNIRS and neuromanagement perspective explain us why things are not like this.
The neuroscience interest for moral decision-making has recently increased. To investigate the processes underlying moral behavior, this research aimed to investigate neurophysiological and behavioral correlates of decision-making in moral contexts. Specifically, functional Near-infrared spectroscopy (fNIRS) allowed to record oxygenated (O2Hb) and deoxygenated (HHb) cerebral hemoglobin concentrations during different moral conditions (professional fit, company fit, social fit) and offers types (fair, unfair, neutral). Moreover, individuals' responses to offers types and reaction time (RTs) were considered. Specifically, from hemodynamic results emerged a difference in O2Hb and HHb activity according to moral conditions and offers types in different brain regions. In particular, O2Hb increase and a HHb decrease were observed in ventromedial and dorsolateral prefrontal cortex (VMPFC, DLPFC) for fair offers in professional fit condition and in superior temporal sulcus (STS) for unfair offers in social fit condition. Moreover, an increase of left O2Hb activity in professional fit condition and in right VMPFC for unfair offers in company fit condition was observed. In addition, from behavioral results, an RTs increase in company and social fit condition for fair and unfair offers emerged. This study, therefore, shows the behavioral and neurophysiological correlates of moral decision-making that guide moral behavior in different context, such as company one.


Neuroprotective effects of glycine for therapy of acute ischaemic stroke.
The aim of this randomized, double-blind, placebo-controlled trial was to assess the safety and the efficacy of the pharmaceutic drug glycine in 200 patients with acute (<6 h) ischaemic stroke in the carotid artery territory. Fifty patients received placebo, 49 glycine 0.5 g/day, 51 glycine 1.0 g/day and 50 glycine 2.0 g/day for 5 days in each group. The efficacy of glycine was assessed by clinical analysis, by an enzyme-linked immunosorbent assay of levels of blood serum autoantibodies to NMDA-binding proteines, by detection of excitatory (glutamate, aspartate) and inhibitory (glycine, GABA) amino acid concentrations and lipid peroxidation products (TBARS) in CSF. The trial confirmed the safety profile of the glycine treatment. Slight sedation was observed in 9 patients (4. 5%) as a side-effect. Other marked side-effects or adverse events were absent. The glycine treatment at the dose of 1.0-2.0 g/day was accompanied by a tendency to a decreased 30-day mortality (5.9% in 1. 0 g/day glycine and 10% in 2.0 g/day glycine groups vs. 14% in the placebo and 14.3% in 0.5 g/day glycine groups), to an improved clinical outcome on the Orgogozo Stroke Scale (p < 0.01) and the Scandinavian Stroke Scale (p < 0.01) and to a favourable functional outcome on the Barthel index (p < 0.01 in 1.0 g/day glycine vs. placebo group in patients with no or mild disability). An early normalization of autoantibody titres to NMDA-binding proteins in serum was found (p < 0.01 vs. placebo), a reduction of glutamate and aspartate levels (p < 0.05 vs. placebo), an increase in GABA concentrations (p < 0.01 vs. placebo in severe stroke patients) and also a reduction of TBARS levels (p < 0.05 vs. placebo) in CSF by day 3. Thus, the trial suggests that sublingual application of 1.0-2. 0 g/day glycine started within 6 h after the onset of acute ischaemic stroke in the carotid artery territory is safe and can exert favourable clinical effects. These results will be verified in further trials with a larger number of patients.


Shoulder joint hemiarthroplasty for treatment of a severe osteochondritis dissecans lesion in a dog.
Partial resurfacing of the humeral head has been reported in humans to treat humeral osteochondritis dissecans. The aim is to describe a custom-made humeral resurfacing prosthesis for treatment of severe humeral head osteochondritis disse-cans in a dog. A seven-month-old female entire St. Bernard dog was presented with a 10 week history of severe left thoracic limb lameness. Radiography, arthroscopy and magnetic resonance imaging confirmed an extensive osteochondritis dissecans lesion affecting the caudal, medial and central regions of the humeral head. A prosthesis designed from computed tomography (CT) images was manufactured in polished stainless steel alloy with a hydroxyapatite coated base and central finned humeral stem for cementless insertion. A standard caudal approach to the shoulder was used to place the prosthesis following reaming of the caudal humeral head. Radiography and CT imaging revealed appropriate topographical placement on the humerus. Force plate analysis demonstrated initial reduction in ground reaction force at six weeks, followed by gradual improvement at three months. There was no radiographic evidence of implant loosening and the range of shoulder joint motion was comparable to the contralateral joint at three months; these findings were maintained to final follow-up at 24 months. Placement of a humeral head resurfacing prosthesis was an effective technique for the management of severe osteochondritis dissecans-related shoulder lameness. To the authors' knowledge, this is the first report of the clinical use of shoulder hemiarthroplasty in a dog.


An official American Thoracic Society/European Respiratory Society statement: update on limb muscle dysfunction in chronic obstructive pulmonary disease.
Limb muscle dysfunction is prevalent in chronic obstructive pulmonary disease (COPD) and it has important clinical implications, such as reduced exercise tolerance, quality of life, and even survival. Since the previous American Thoracic Society/European Respiratory Society (ATS/ERS) statement on limb muscle dysfunction, important progress has been made on the characterization of this problem and on our understanding of its pathophysiology and clinical implications. The purpose of this document is to update the 1999 ATS/ERS statement on limb muscle dysfunction in COPD. An interdisciplinary committee of experts from the ATS and ERS Pulmonary Rehabilitation and Clinical Problems assemblies determined that the scope of this document should be limited to limb muscles. Committee members conducted focused reviews of the literature on several topics. A librarian also performed a literature search. An ATS methodologist provided advice to the committee, ensuring that the methodological approach was consistent with ATS standards. We identified important advances in our understanding of the extent and nature of the structural alterations in limb muscles in patients with COPD. Since the last update, landmark studies were published on the mechanisms of development of limb muscle dysfunction in COPD and on the treatment of this condition. We now have a better understanding of the clinical implications of limb muscle dysfunction. Although exercise training is the most potent intervention to address this condition, other therapies, such as neuromuscular electrical stimulation, are emerging. Assessment of limb muscle function can identify patients who are at increased risk of poor clinical outcomes, such as exercise intolerance and premature mortality. Limb muscle dysfunction is a key systemic consequence of COPD. However, there are still important gaps in our knowledge about the mechanisms of development of this problem. Strategies for early detection and specific treatments for this condition are also needed.


Toxicity of cationic lipid-ribozyme complexes in human prostate tumor cells can mimic ribozyme activity.
Prostate tumor cell lines have been shown to both produce interleukin-6 (IL-6) and express the IL-6 receptor, suggesting a potential autocrine growth regulatory role for IL-6. We explored the role of IL-6 in the proliferation of the human prostatic carcinoma cell line, DU145, using ribozymes to inhibit IL-6 expression. Hammerhead-type ribozymes targeted against IL-6 mRNA sequences were prepared, and in vitro analyses were used to demonstrate that these molecules catalyzed the cleavage of IL-6 mRNA poly- nucleotide fragments. To test in situ activity, these ribozymes were transfected into DU145 cells using cationic transfection lipids, cytofectins. Treatment of cultured cells with ribozyme/cationic lipid complexes resulted in a reduction of IL-6 protein levels in the supernatant and reduced numbers of DU145 cells 48 h after treatment. However, similar results were also seen following treatment with control RNA/lipid complexes. This reduction in IL-6 levels and cell numbers was a function of the RNA/lipid complexes and was not seen with either lipid or RNA alone. Therefore, the reductions in IL-6 levels and cell numbers observed were not due to ribozyme-mediated cleavage of IL-6 mRNA, but rather reflected a dose-dependent, nonspecific toxic effect of the treatment with ribozyme/cytofectin complexes. This effect can resemble functional ribozyme activity, complicating analysis of the activity of synthetic ribozymes after transfection into cultured cells.


Arterial spin-labeling in routine clinical practice, part 1: technique and artifacts.
The routine use of arterial spin-labeling (ASL) in a clinical population has led to the depiction of diverse brain pathologic features. Unique challenges in the acquisition, postprocessing, and analysis of cerebral blood flow (CBF) maps are encountered in such a population, and high-quality ASL CBF maps can be generated consistently with attention to quality control and with the use of a dedicated postprocessing pipeline. Familiarity with commonly encountered artifacts can help avoid pitfalls in the interpretation of CBF maps. The purpose of this review was to describe our experience with a heterogeneous collection of ASL perfusion cases with an emphasis on methodology and common artifacts encountered with the technique. In a period of 1 year, more than 3000 pulsed ASL cases were performed as a component of routine clinical brain MR evaluation at both 1.5 and 3T. These ASL studies were analyzed with respect to overall image quality and patterns of perfusion on final gray-scale DICOM images and color Joint Photographic Experts Group (JPEG) CBF maps, and common artifacts and their impact on final image quality were categorized.


The influence of heart rate on augmentation index and central arterial pressure in humans.
Arterial stiffness is an important determinant of cardiovascular risk. Augmentation index (AIx) is a measure of systemic arterial stiffness derived from the ascending aortic pressure waveform. The aim of the present study was to assess the effect of heart rate on AIx. We elected to use cardiac pacing rather than chronotropic drugs to minimize confounding effects on the systemic circulation and myocardial contractility. Twenty-two subjects (13 male) with a mean age of 63 years and permanent cardiac pacemakers in situ were studied. Pulse wave analysis was used to determine central arterial pressure waveforms, non-invasively, during incremental pacing (from 60 to 110 beats min-1), from which AIx and central blood pressure were calculated. Peripheral blood pressure was recorded non-invasively from the brachial artery. There was a significant, inverse, linear relationship between AIx and heart rate (r = -0.76; P < 0.001). For a 10 beats min-1 increment, AIx fell by around 4 %. Ejection duration and heart rate were also inversely related (r = -0. 51; P < 0.001). Peripheral systolic, diastolic and mean arterial pressure increased significantly during incremental pacing. Although central diastolic pressure increased significantly with pacing, central systolic pressure did not. There was a significant increase in the ratio of peripheral to central pulse pressure (P < 0.001), which was accounted for by the observed change in central pressure augmentation. These results demonstrate an inverse, linear relationship between AIx and heart rate. This is likely to be due to alterations in the timing of the reflected pressure wave, produced by changes in the absolute duration of systole. Consideration of wave reflection and aortic pressure augmentation may explain the lack of rise in central systolic pressure during incremental pacing despite an increase in peripheral pressure.


Robust Blur Kernel Estimation for License Plate Images From Fast Moving Vehicles.
As the unique identification of a vehicle, license plate is a key clue to uncover over-speed vehicles or the ones involved in hit-and-run accidents. However, the snapshot of over-speed vehicle captured by surveillance camera is frequently blurred due to fast motion, which is even unrecognizable by human. Those observed plate images are usually in low resolution and suffer severe loss of edge information, which cast great challenge to existing blind deblurring methods. For license plate image blurring caused by fast motion, the blur kernel can be viewed as linear uniform convolution and parametrically modeled with angle and length. In this paper, we propose a novel scheme based on sparse representation to identify the blur kernel. By analyzing the sparse representation coefficients of the recovered image, we determine the angle of the kernel based on the observation that the recovered image has the most sparse representation when the kernel angle corresponds to the genuine motion angle. Then, we estimate the length of the motion kernel with Radon transform in Fourier domain. Our scheme can well handle large motion blur even when the license plate is unrecognizable by human. We evaluate our approach on real-world images and compare with several popular state-of-the-art blind image deblurring algorithms. Experimental results demonstrate the superiority of our proposed approach in terms of effectiveness and robustness.


Dynamic organization of DNA replication in mammalian cell nuclei: spatially and temporally defined replication of chromosome-specific alpha-satellite DNA sequences.
Five distinct patterns of DNA replication have been identified during S-phase in asynchronous and synchronous cultures of mammalian cells by conventional fluorescence microscopy, confocal laser scanning microscopy, and immunoelectron microscopy. During early S-phase, replicating DNA (as identified by 5-bromodeoxyuridine incorporation) appears to be distributed at sites throughout the nucleoplasm, excluding the nucleolus. In CHO cells, this pattern of replication peaks at 30 min into S-phase and is consistent with the localization of euchromatin. As S-phase continues, replication of euchromatin decreases and the peripheral regions of heterochromatin begin to replicate. This pattern of replication peaks at 2 h into S-phase. At 5 h, perinucleolar chromatin as well as peripheral areas of heterochromatin peak in replication. 7 h into S-phase interconnecting patches of electron-dense chromatin replicate. At the end of S-phase (9 h), replication occurs at a few large regions of electron-dense chromatin. Similar or identical patterns have been identified in a variety of mammalian cell types. The replication of specific chromosomal regions within the context of the BrdU-labeling patterns has been examined on an hourly basis in synchronized HeLa cells. Double labeling of DNA replication sites and chromosome-specific alpha-satellite DNA sequences indicates that the alpha-satellite DNA replicates during mid S-phase (characterized by the third pattern of replication) in a variety of human cell types. Our data demonstrates that specific DNA sequences replicate at spatially and temporally defined points during the cell cycle and supports a spatially dynamic model of DNA replication.


Effects of dexmedetomidine on heart arrhythmia prevention in off-pump coronary artery bypass surgery: A randomized clinical trial.
Arrhythmia occurring during and after surgery is one of the major complications in open-heart surgery. Dexmedetomidine is an intravenous alpha-2 agonist and very specific short-acting drug to protect the various organs against ischemic injuries and blood reflow. However, the effect of dexmedetomidine for preventing intraoperative heart arrhythmias has not been recognized. This study aimed to determine the effect of dexmedetomidine on the incidence rate of heart arrhythmias and anesthetic required in off-pump coronary artery bypass surgery. This randomized clinical trial was conducted on patients who were candidates for off-pump coronary artery bypass referring to Imam Reza Hospital of Mashhad, Iran, from July 2016 through January 2017. The patients were randomly assigned to two groups of intervention (infusion of 0.5 mcg/kg/h dexmedetomidine together with induction followed by infusion of 0.5 mcg/kg/h by the end of the surgery) or control (saline infusion). Mean arterial pressure (MAP) and heart rate (HR) were measured before induction, during surgery operation and ICU admission. Data were analyzed by SPSS version 18 using Chi Square and independent-samples t-test. A total of 76 patients with a mean age of 59.8 ± 8.2 years (in two groups of 38) were studied. The two groups had no statistically significant difference in terms of background variables. The MAP and HR values before induction, during surgery and ICU admission were significantly higher in the control group than in the intervention group (p=0.001). Out of the studied arrhythmias, the values of PAC (55.2% vs. 15.7%), PVC (81.5% vs. 21.0%), AF (26.3% vs. 7.8%), VTAC (21.0% vs. 2.6%) were significantly lower in dexmedetomidine group (p=0.001). It seems that dexmedetomidine administration during induction and surgery can cause significant reduction in most of the common arrhythmias in off-pump coronary bypass surgery. The use of dexmedetomidine maintains MAP and HR at significantly lower values, and changes compared to the control group as well as reduces the need for anesthetic compounds. The present study has been registered at the Iranian Registry of Clinical Trials (www.IRCT.IR) with a code of IRCT2016072413159N9 before starting the study. This study was fully sponsored by the Research Deputy at Mashhad University of Medical Sciences, Iran (grant number 941413).


Near-Field Ultrasound Imaging During Radiofrequency Catheter Ablation: Tissue Thickness and Epicardial Wall Visualization and Assessment of Radiofrequency Ablation Lesion Formation and Depth.
Safe and successful radiofrequency catheter ablation depends on creation of transmural lesions without collateral injury to contiguous structures. Near-field ultrasound (NFUS) imaging through transducers in the tip of an ablation catheter may provide important information about catheter contact, wall thickness, and ablation lesion formation. NFUS imaging was performed using a specially designed open-irrigated radiofrequency ablation catheter incorporating 4 ultrasound transducers. Tissue/phantom thickness was measured in vitro with varying contact angles. In vivo testing was performed in 19 dogs with NFUS catheters positioned in 4 chambers. Wall thickness measurements were made at 222 sites (excluding the left ventricle) and compared with measurements from intracardiac echocardiography. Imaging was used to identify the epicardium with saline infusion into the pericardial space at 39 sites. In vitro, the measured exceeded actual tissue/phantom thickness by 13% to 20%. In vivo, NFUS reliably visualized electrode-tissue contact, but sensitivity of epicardial imaging was 92%. The chamber wall thickness measured by NFUS correlated well with intracardiac echocardiography (r=0.86; P<0.0001). Sensitivity of lesion identification by NFUS was 94% for atrial and 95% for ventricular ablations. NFUS was the best parameter to predict lesion depth in right and left ventricle (r=0.47; P<0.0001; multiple regression P=0.0025). Lesion transmurality was correctly identified in 87% of atrial lesions. NFUS catheter imaging reliably assesses electrode-tissue contact and wall thickness. Its use during radiofrequency catheter ablation may allow the operator to assess the depth of ablation required for transmural lesion formation to optimize power delivery.


Recall by expert medical practitioners and novices as a record of processing attention.
Using recall of clinical protocols as a measure of expertise in medicine has yielded disappointingly small effects. Experiments using recall of clinical laboratory data are presented to provide an explanation. In one experiment, subjects either deliberately memorized or first diagnosed and then were incidentally asked for memory. With incidental instructions, experts recalled over twice as much data as did students, but with memorization instructions, student performance approximated that of experts. Experts also showed a large advantage over students in incidental recall of data that were not relevant to the problem solution. These results suggest that expert processing in this "discrete, independent inputs" domain requires effortful analysis with minimal reliance on default values, rather than relatively effortless pattern perception reported in highly visual areas of expertise. For this area, intentional memory is a misleading measure of expertise. However, incidental memory is a valuable measure of processing during diagnosis.


Hemichorea associated with nonketotic hyperglycemia in a female.
A case of a 61-year-old woman with hemichorea associated with nonketotic hyperglycemia is reported. The typical presentation of this disease is nonketotic hyperglycemia, hemichorea, hyper-intense signal on T1-weighted magnetic resonance imaging (MRI) and high-density on computed tomography (CT) in the contralateral striatum. With good glycemic control, the clinical symptoms disappeared.


Empirical correction of thermal responses in the Solar Occultation for Ice Experiment nitric oxide measurements and initial data validation results.
The Solar Occultation for Ice Experiment (SOFIE) makes broadband transmission measurements centered at 5.32 μm to determine the concentration profile of nitric oxide (NO). These measurements show a signal oscillation due to detector temperature variations that severely limit the accuracy of NO retrievals if corrections are not applied. An empirical correction was developed to remove this instrumental error. This paper describes the correction, its impact on the retrieval, and presents a comparison from 87 to 105 km versus coincident atmospheric chemistry experiment-Fourier transform spectrometer (ACE-FTS) measurements. The southern hemisphere (SH) shows excellent agreement between the datasets, with statistically insignificant differences. The northern hemisphere (NH) SOFIE measurements exhibit a low bias of -18.5% compared to ACE-FTS. NH measurements (sunrise observations) are still under study, and only SH NO data (sunset observations) are currently publicly available as of SOFIE data version 1.2.


Role of tumour necrosis factor receptor-1 and nuclear factor-κB in production of TNF-α-induced pro-inflammatory microparticles in endothelial cells.
Tumour necrosis factor-α (TNF-α) is upregulated in many inflammatory diseases and is also a potent agent for microparticle (MP) generation. Here, we describe an essential role of TNF-α in the production of endothelial cell-derived microparticles (EMPs) in vivo and the function of TNF-α-induced EMPs in endothelial cells. We found that TNF-α rapidly increased blood levels of EMPs in mice. Treatment of human umbilical vein endothelial cells (HUVECs) with TNF-α also induced EMP formation in a time-dependent manner. Silencing of TNF receptor (TNFR)-1 or inhibition of the nuclear factor-κB (NF-κB) in HUVECs impaired the production of TNF-α-induced EMP. Incubation of HUVECs with PKH-67-stained EMPs showed that endothelial cells readily engulfed EMPs, and the engulfed TNF-α-induced EMPs promoted the expression of pro-apoptotic molecules and upregulated intercellular adhesion molecule-1 level on the cell surface, which led to monocyte adhesion. Collectively, our findings indicate that the generation of TNF-α-induced EMPs was mediated by TNFR1 or NF-κB and that EMPs can contribute to apoptosis and inflammation of endothelial cells.


Cloning, expression, and localization of a mouse retinal gamma-aminobutyric acid transporter.
To isolate a cDNA clone encoding a high-affinity gamma-aminobutyric acid (GABA) transporter from mouse retina, to examine its biochemical and pharmacologic properties, and to determine the sites of its mRNA expression in retinal cells. A mouse retinal cDNA library was screened using a fragment of a rat brain GABA transporter (GAT-1) cDNA as a probe. One homologous clone, mouse retinal GAT-1, was chosen for further characterization. RNA transcribed from mouse retinal GAT-1 was microinjected into Xenopus oocytes, and pharmacologic properties of the expressed transporter were determined. Sites of mouse retinal GAT-1 mRNA expression were examined by in situ hybridization. The protein sequence deduced from the DNA sequence of mouse retinal GAT-1 cDNA was virtually identical to that of the rat and the mouse brain GAT-1. RNA transcribed from this clone induced a [3H]-GABA uptake activity in microinjected Xenopus oocytes that was both sodium and chloride dependent. The apparent Km and Vmax for the GABA uptake were 8.3 microM and 40.0 pmol/egg per hour, respectively. The mouse retinal GAT-1 induced GABA uptake was inhibited by L-diaminobutyric acid, guvacine, cis-4-hydroxynipecotic acid, nipecotic acid, and 4,5,6,7-tetrahydroisoxazolo [4,5c]-pyridin-3-ol with IC50 values of 320, 79, 71, 7.1, and 200 microM, respectively. However, beta-alanine was unable to inhibit the induced GABA uptake significantly (IC50 approximately 2,500 microM). In situ hybridization studies showed that mouse retinal GAT-1 mRNA was present in a subpopulation of amacrine, interplexiform, and displaced amacrine cells. Hybridization signal in the Müller cells was significantly lower, and GAT-1 transcripts were not detected in the bipolar, horizontal, or photoreceptor cells of mouse retina. The mouse retinal GAT-1 cDNA encodes a Na(+)-dependent, high-affinity GABA transporter that is mainly expressed in a subset of mouse retinal inter neurons.


Monoclonal antibodies for the treatment of severe asthma.
Patients with severe asthma have a significant unmet need with persistent symptoms and/or frequent exacerbations despite treatment with high-dose steroid and other currently available therapies. These patients are also at risk of developing steroid-related side effects, and their severe, unrelenting symptoms have a huge impact on health care resources due to frequent hospital admissions and requirement for intensive medication use. Consequently, a compelling need exists for more effective and safer pharmacotherapies to help them achieve adequate disease control. Recent novel therapies for severe asthma are now emerging, some of the most promising of which are monoclonal antibodies. Monoclonal antibodies represent a form of immunotherapy used in a wide variety of therapeutic roles. The spectrum of disease states in which monoclonal antibodies have been approved for therapeutic use now includes respiratory and allergic diseases. At present, only one drug is licensed for allergic asthmatics with severe disease, omalizumab. We review some of the currently available biologics that are approved or under investigation for use in severe asthma. Some have shown to be useful in specifically targeted subpopulations of patients with severe asthma, whereas other have proven to be unsafe and/or unsuccessful. Despite these developments, more effort should be devoted to identifying new molecular targets, testing innovative approaches, and establishing the best use of what is available. Regarding this latter point, identifying individual characteristics that predict successful responses to these treatments is highly desirable.


Rodent malaria models: insights into human disease and parasite biology.
The use of rodents as model organisms to study human disease is based on the genetic and physiological similarities between the species. Successful molecular methods to generate transgenic reporter or humanized rodents has rendered rodents as powerful tools for understanding biological processes and host-pathogen interactions relevant to humans. In malaria research, rodent models have been pivotal for the study of liver stages, syndromes arising from blood stages of infection, and malaria transmission to and from the mammalian host. Importantly, many in vivo findings are comparable to pathology observed in humans only when adequate combinations of rodent strains and Plasmodium parasites are used.


IGF-I inhibits apoptosis induced by serum withdrawal, but potentiates TNF-alpha-induced apoptosis, in 3T3-L1 preadipocytes.
We have previously shown that human preadipocytes in primary culture undergo apoptosis in response to serum deprivation and addition of tumour necrosis factor alpha (TNF-alpha), and have proposed that regulation of preadipocyte apoptosis in vivo may contribute to the overall control of adipose mass. In the present study we have investigated both pro- and anti-apoptotic factors, and the signalling pathways by which they act, in murine 3T3-L1 preadipocytes. Apoptotic indices (fraction of cells undergoing apoptosis) were determined by microscopic examination of acridine orange-stained cells, fluorescence-activated cell sorting of propidium iodide-stained cells, or phase-contrast video microscopy. Murine 3T3-L1 cells were more susceptible to apoptosis than human preadipocytes. In medium containing 10% newborn calf serum, the basal apoptotic index was very low (<2%), but the number of apoptotic cells increased significantly following serum withdrawal (10% after 24 h). Addition of TNF-alpha (6 nM) stimulated apoptosis in both serum-containing and serum-free media (apoptotic indices of 12% and 20% respectively after 24 h). IGF-I inhibited by approximately 50% the apoptosis induced by serum withdrawal, but increased by 25% the apoptosis induced by TNF-alpha in serum-free medium. It was shown by using specific inhibitors of lipid and protein kinases (LY294002, rapamycin, PD98059, SB203580) that both phosphoinositide 3-kinase and MAP kinase pathways contribute to the anti-apoptotic action of IGF-I on serum-starved cells, while phosphoinositide 3-kinase but not MAP kinase activity is required for the paradoxical pro-apoptotic action of IGF-I in the presence of TNF-alpha. We conclude that, in addition to its previously described anti-apoptotic action, IGF-I can also be pro-apoptotic in 3T3-L1 cells in the presence of TNF-alpha, and that both the anti- and pro-apoptotic effects of IGF-I require the activation of phosphoinositide 3-kinase.


Non-catalytic tyrosine-phosphorylated receptors.
Leukocytes play a critical role in recognizing and responding to infection and cancer. Central to this function is an array of cell-surface receptors that lack sequence homology. Many of these receptors have in common the fact that their signaling involves phosphorylation of cytoplasmic domains by extrinsic tyrosine kinases. These non-catalytic tyrosine-phosphorylated receptors (NTRs) share a number of other features, including small size and optimal stimulation by surface-associated ligands. We argue here that NTRs are also likely to share the same kinetic-segregation triggering mechanism, which involves segregation of the engaged NTR from receptor tyrosine phosphatases with large ectodomains such as CD45 and CD148. NTRs signal through tyrosine-containing cytoplasmic motifs, which recruit distinct cytoplasmic signaling proteins when phosphorylated, transducing activatory or inhibitory signals. They have two features that make them uniquely well suited to their role in immune recognition of infection and cancer. Their modular structure enables the coupling of many rapidly evolving receptors with diverse ligand specificities to the same conserved signaling machinery. Their similarity in size and shared signaling machinery enables them to colocalize at cell-cell interfaces when they engage ligands, facilitating the integration of activatory and inhibitory signals from multiple receptors at the cell surface.


Epitope spreading to citrullinated antigens in mouse models of autoimmune arthritis and demyelination.
Anti-citrullinated protein antibodies have a diagnostic role in rheumatoid arthritis (RA); however, little is known about their origins and contribution to pathogenesis. Citrullination is the post-translational conversion of arginine to citrulline by peptidyl arginine deiminase, and increased citrullination of proteins is observed in the joint tissue in RA and in brain tissue in multiple sclerosis (MS). We applied synovial and myelin protein arrays to examine epitope spreading of B cell responses to citrullinated epitopes in both the collagen-induced arthritis (CIA) model for RA and the experimental autoimmune encephalomyelitis (EAE) model for MS. Synovial and myelin protein arrays contain a spectrum of proteins and peptides, including native and citrullinated forms, representing candidate autoantigens in RA and MS, respectively. We applied these arrays to characterise the specificity of autoantibodies in serial serum samples derived from mice with acute and chronic stages of CIA and EAE. In samples from pre-disease CIA and acute-disease EAE, we observed autoantibody targeting of the immunising antigen and responses to a limited set of citrullinated epitopes. Over the course of diseases, the autoantibody responses expanded to target multiple citrullinated epitopes in both CIA and EAE. Using immunoblotting and mass spectrometry analysis, we identified citrullination of multiple polypeptides in CIA joint and EAE brain tissue that have not previously been described as citrullinated. Our results suggest that anti-citrulline antibody responses develop in the early stages of CIA and EAE, and that autoimmune inflammation results in citrullination of joint proteins in CIA and brain proteins in EAE, thereby creating neoantigens that become additional targets in epitope spreading of autoimmune responses.


Gene expression profile of trisomy 21 placentas: a potential approach for designing noninvasive techniques of prenatal diagnosis.
The purpose of this study was to evaluate the feasibility of the DNA microarray technology for the study of the differences in gene expression between placentas of pregnancies with trisomy 21 fetuses and placentas of fetuses with a normal karyotype. Complementary DNA samples from 7 second-trimester placentas of fetuses with trisomy 21 was compared with 7 matched and 7 nonmatched complementary DNA samples from normal karyotype placentas as control samples. For gene expression comparison, a microarray that contained 8976 expressed sequence tags was used. Four experiments were performed: (1) the study samples were compared with matched control samples; (2) the study samples were compared with nonmatched control samples; (3) the study samples were compared with themselves; and (4) the matched and nonmatched control samples were compared. Gene overexpression and underexpression were defined as sample signal differences of >1.7 over each other. Gene overexpression and underexpression were confirmed by Northern blotting studies. Seven expressed sequence tags of 8976 tags that were examined have met the selection criteria. Overexpression of 2 expressed sequence tags in placentas with trisomy 21 was confirmed by Northern blotting studies. Our study demonstrates the feasibility of the microarray technique in the study of the differences of gene expression in trisomic placentas compared with placentas with a normal karyotype. This technique may contribute to the identification of additional maternal serum biochemical markers in aneuploid pregnancies.


ICU Admission, Discharge, and Triage Guidelines: A Framework to Enhance Clinical Operations, Development of Institutional Policies, and Further Research.
To update the Society of Critical Care Medicine's guidelines for ICU admission, discharge, and triage, providing a framework for clinical practice, the development of institutional policies, and further research. An appointed Task Force followed a standard, systematic, and evidence-based approach in reviewing the literature to develop these guidelines. The assessment of the evidence and recommendations was based on the principles of the Grading of Recommendations Assessment, Development and Evaluation system. The general subject was addressed in sections: admission criteria and benefits of different levels of care, triage, discharge timing and strategies, use of outreach programs to supplement ICU care, quality assurance/improvement and metrics, nonbeneficial treatment in the ICU, and rationing considerations. The literature searches yielded 2,404 articles published from January 1998 to October 2013 for review. Following the appraisal of the literature, discussion, and consensus, recommendations were written. Although these are administrative guidelines, the subjects addressed encompass complex ethical and medico-legal aspects of patient care that affect daily clinical practice. A limited amount of high-quality evidence made it difficult to answer all the questions asked related to ICU admission, discharge, and triage. Despite these limitations, the members of the Task Force believe that these recommendations provide a comprehensive framework to guide practitioners in making informed decisions during the admission, discharge, and triage process as well as in resolving issues of nonbeneficial treatment and rationing. We need to further develop preventive strategies to reduce the burden of critical illness, educate our noncritical care colleagues about these interventions, and improve our outreach, developing early identification and intervention systems.


Practical Aspects of microRNA Target Prediction.
microRNAs (miRNAs) are endogenous non-coding RNAs that control gene expression at the posttranscriptional level. These small regulatory molecules play a key role in the majority of biological processes and their expression is also tightly regulated. Both the deregulation of genes controlled by miRNAs and the altered miRNA expression have been linked to many disorders, including cancer, cardiovascular, metabolic and neurodegenerative diseases. Therefore, it is of particular interest to reliably predict potential miRNA targets which might be involved in these diseases. However, interactions between miRNAs and their targets are complex and very often there are numerous putative miRNA recognition sites in mRNAs. Many miRNA targets have been computationally predicted but only a limited number of these were experimentally validated. Although a variety of miRNA target prediction algorithms are available, results of their application are often inconsistent. Hence, finding a functional miRNA target is still a challenging task. In this review, currently available and frequently used computational tools for miRNA target prediction, i.e., PicTar, TargetScan, DIANA-microT, miRanda, rna22 and PITA are outlined and various practical aspects of miRNA target analysis are extensively discussed. Moreover, the performance of three algorithms (PicTar, TargetScan and DIANA-microT) is both demonstrated and evaluated by performing an in-depth analysis of miRNA interactions with mRNAs derived from genes triggering hereditary neurological disorders known as trinucleotide repeat expansion diseases (TREDs), such as Huntington's disease (HD), a number of spinocerebellar ataxias (SCAs), and myotonic dystrophy type 1 (DM1).


One-dimensional rainbow thermometry system by using slit apertures.
A new rainbow thermometry system by using slit apertures and a laser light sheet, called a one-dimensional rainbow thermometry (ORT) system, has been developed as an extension of global rainbow thermometry (GRT). This system is capable of one-dimensional or line measurements of the size and refractive index of droplets in the spray space, while the conventional GRT system is normally considered a typical "single-point" or "small volume" measurement method. The performance of this new system was tested and verified with both water and ethanol spray. The results show the feasibility and potential of ORT in simultaneous and one-dimensional measurement of the size and refractive index of liquid droplets, especially in the research field of spray evaporation and combustion.


Serum concentration of integrin-linked kinase in malignant pleural mesothelioma and after asbestos exposure.
Integrin-linked kinase (ILK) is an intracellular protein implicated in chronic inflammation and neoplastic transformation. In a recently accomplished pilot study, we showed that ILK can be detected in the serum of patients with benign and malignant chest diseases, including malignant pleural mesothelioma (MPM). Interestingly, average serum ILK concentrations were 10 times higher in MPM patients when compared with the rest of the study population, and a diagnostic test solely based on serum ILK concentration could discriminate between MPM and non-MPM with considerable accuracy. This study aimed to investigate whether serum ILK concentration could also be used to discriminate between MPM and asbestos exposure only. Using a self-developed sandwich enzyme-linked immunosorbent assay, we measured serum ILK concentrations in 101 MPM patients, and 96 asbestos-exposed, but healthy insulation workers. Seventy-three MPM patients had an epitheloid subtype (72.3%), and 42 had a Stage I or II disease (41.6%). When compared with asbestos-exposed individuals, MPM patients of all clinical stages had significantly higher (mean ± standard deviation, median) serum ILK concentrations (10.7 ± 13.6, median 7 ng/ml vs 3.1 ± 4.6, median 1.4 ng/ml; P < 0.001). Among MPM patients, the serum ILK concentration was significantly higher at advanced disease stages III + IV than at early stages I + II (13.7 ± 15.9, median 8.5 ng/ml vs 6.7 ± 7.8, median 3.5 ng/ml; P = 0.02). Using serum ILK to discriminate between MPM patients and asbestos-exposed individuals yielded an area under the curve of 0.69 (95% confidence interval 0.63-0.76). The corresponding sensitivity and specificity for a cut-off of 4.49 ng/ml ILK are 61.4 and 80.2%, respectively. These data show significant differences between MPM patients and asbestos-exposed but healthy individuals concerning their serum ILK concentration. Furthermore, since ILK levels are increased in advanced MPM stages in comparison with early MPM stages, we suggest evaluating its potential use as a marker of disease progression in MPM.


Long term prognostic importance of late gadolinium enhancement in first-presentation non-ischaemic dilated cardiomyopathy.
Presence of myocardial fibrosis in well-established non-ischaemic dilated cardiomyopathy (NIDCM) is associated with adverse clinical outcomes. However, the impact of myocardial fibrosis at first presentation in NIDCM, and its long-term association with left ventricular (LV) dysfunction, heart failure (HF) and ventricular arrhythmia (VA) remains unclear. We investigated whether the presence of myocardial fibrosis quantified by late gadolinium enhancement cardiovascular magnetic resonance (LGE-CMR) at presentation, is independently associated with long-term major adverse cardiovascular events (MACE) in patients with first presentation NIDCM. Consecutive patients with a first diagnosis of NIDCM were recruited. Patients underwent LGE-CMR at baseline. Replacement myocardial fibrosis by LGE-CMR was quantified by experienced observers blinded to patient outcome. MACE was defined as a composite end-point including cardiac death, HF rehospitalisation and the occurrence of sustained VA. Fifty-one patients with first presentation NIDCM were included, of which 49 (96%) had follow up and outcome data. Median follow up was 8.2 years. Both the LGE positive and LGE negative groups had similar clinical characteristics at follow up. In univariate Cox regression analysis, positive LGE was associated with MACE (HR:3.44; 95% CI:1.89 to 6.24, p-value < 0.001) and HF rehospitalisation (HR:2.89; 95% CI:1.42 to 5.85, p-value = 0.003). In multivariate Cox regression, positive LGE-CMR was independently associated with MACE (HR:3.53; 95% CI:1.51 to 8.27, p-value = 0.004) and HF rehospitalisation (HR:3.07; 95% CI:1.24 to 7.59, p-value = 0.015). The presence of myocardial fibrosis in first presentation NIDCM is independently associated with an increased risk of HF rehospitalisation, at long term follow-up.


Paclitaxel (Taxol)/doxorubicin combinations in advanced breast cancer: the Eastern Cooperative Oncology Group experience.
Paclitaxel (Taxol; Bristol-Myers Squibb Company, Princeton, NJ), the first taxane to enter routine clinical practice, has aroused considerable interest due to its novel mechanism of action and its significant activity in metastatic breast cancer. Given this activity, it seemed logical to attempt to combine paclitaxel with doxorubicin, the other most active single agent in metastatic breast cancer. The Eastern Cooperative Oncology Group performed two trials investigating paclitaxel/doxorubicin combinations in patients with advanced breast cancer in an attempt to identify a tolerable dose and schedule of the combination. In the first trial, paclitaxel and doxorubicin were alternated every 3 weeks in doses of 200 mg/m2 and 75 mg/m2, respectively, for patients who had received no more than one prior chemotherapeutic regimen. Therapy was well tolerated in this setting. At these doses, paclitaxel induced more granulocytopenia and less thrombocytopenia than did doxorubicin. Objective responses (complete and partial responses) were seen in seven of 12 patients; two other patients had improved disease (relief of pain in bony metastases). A second limited-institution Eastern Cooperative Oncology Group trial evaluated paclitaxel and doxorubicin given in combination. In this phase I trial, doxorubicin was given as an intravenous push and paclitaxel as a 24-hour continuous infusion. The sequence of drug administration (D-->P or P-->D) was alternated both between and within patients, so that we might evaluate the effect of administration schedule on toxicity. Therapy was begun at an initial paclitaxel dose of 150 mg/m2 and an initial doxorubicin dose of 50 mg/m2 in six patients, with a subsequent six patients receiving 175 and 60 mg/m2, respectively, of paclitaxel and doxorubicin. In addition, patients received granulocyte colony-stimulating factor 5 micrograms/kg/d. While therapy at the initial dose level was well tolerated, dose-limiting mucositis was seen at the second dose level, although only when paclitaxel preceded doxorubicin. This suggests that sequence of drug administration in paclitaxel-based regimens may play an important role as a determinant of toxicity and (perhaps) efficacy, a finding similar to that seen when paclitaxel and cisplatin were combined in patients with ovarian cancer. Based on this study, we identified the sequence of doxorubicin (50 mg/m2) followed by paclitaxel (150 mg/m2) to be the maximum tolerated dose. This combination is currently being compared with paclitaxel alone and doxorubicin alone in patients with advanced breast cancer in an intergroup trial led by the Eastern Cooperative Oncology Group.


Uterine compression sutures for the management of severe postpartum hemorrhage.
To assess maternal outcomes after uterine compression suture use and to characterize the risk factors for failure, defined as cases that proceeded to hysterectomy. A prospective population-based study of 1.2 million women delivering was conducted using the U.K. Obstetric Surveillance System to identify all women in the United Kingdom delivering between September 2007 and March 2009 and treated with uterine compression sutures. Two hundred eleven women were treated with a uterine compression suture to control postpartum hemorrhage. The overall rate of failure, leading to hysterectomy, was 25% (95% confidence interval, 19–31%); there were no significant differences in failure rates among B-Lynch sutures, modified B-Lynch sutures, and other suture techniques. Women were more likely to have a hysterectomy if they were aged 35 years or older (33% compared with 20% aged younger than 35 years), multiparous (33% compared with 14% in nulliparous), in unemployed and routine or manual occupational groups (28% compared with 17% in managerial or professional groups), had a vaginal delivery (47% compared with 22% in the cesarean delivery group), or a delay of between 2 and 6 hours from delivery to uterine suture compression (42% compared with 16% with delay less than 1 hour). A prolonged delay of 2–6 hours between delivery and uterine compression suture was independently associated with a fourfold increase in the odds of hysterectomy. These data emphasize the need for a careful evaluation of blood loss after delivery to avoid any prolonged delay in recognition of hemorrhage. III


Clinical, Histochemical, and Molecular Study of Three Turkish Siblings Diagnosed with H Syndrome, and Literature Review.
The term "H syndrome" was coined to denote the major clinical findings, which include hyperpigmentation, hypertrichosis, hearing loss, hepatosplenomegaly, hyperglycaemia, hypogonadism, hallux flexion contractures, and short height. To report the clinical, endocrinological, histochemical, and genetic findings of three siblings. Skin and liver biopsies were taken to investigate the histochemical characteristics of hyperpigmented hypertrichotic skin lesions and massive hepatomegaly. The levels of basal serum thyroid hormones, oestradiol, total testosterone, follicle-stimulating hormone, luteinising hormone, and stimulated growth hormone (GH) were measured to investigate the endocrine aspects of the syndrome. Mutation analysis was carried out in all six exons and exon-intron boundaries of SLC29A3 by direct sequencing. Physical examination of the patients revealed common charac-teristic findings of H syndrome. Additional clinical findings were sectorial iris atrophy in the younger sister. Laboratory evaluation revealed microcytic anaemia, markedly increased erythrocyte sedimentation rate and C-reactive protein levels, and humoral immune deficiency in the younger siblings, who presented with recurrent fever and sinopulmonary infection. Two different GH stimulation tests revealed GH deficiency in the younger sister with short stature. Liver and skin biopsies revealed polyclonal lymphohistiocytic and plasma cell infiltration. Sequencing of SLC29A3 in the three siblings revealed a novel homozygous mutation in exon 6, which caused the transition of arginine to tryptophan. This study not only extended the clinical and mutation spectrum of SLC29A3 in H syndrome, but also showed that short children should be assessed according to the guidelines for short stature in children.


Impact of community pharmacists advocating immunization on influenza vaccination rates among the elderly.
This study was carried out to investigate whether the personal advocacy of influenza vaccination by community pharmacists to people aged 65 years and above affected the vaccination rate and number of influenza patients. A cluster randomized controlled trial was conducted with the cooperation of 84 community pharmacies in the wards of Suginami and Nerima, Tokyo. Participants were aged 65 years and above living in Suginami and Nerima wards, Tokyo, receiving dispensing services in their community pharmacies. The intervention was that pharmacists in the intervention pharmacy group provided information on the risk of influenza and benefits of influenza vaccination. Main outcome measures were the self-reported influenza vaccination rate in January 2004, and the number of participants with influenza, as confirmed by inspection of their prescriptions from January to May 2004. The vaccination rate in the intervention pharmacy group (81.6%) was significantly higher than that in the control pharmacy group (64.9%). The number of participants with influenza among the intervention group (2/881) was significantly lower than that among the control group (11/895). The personal advocacy of influenza vaccination by community pharmacists among people aged 65 years and above increases the vaccination rate and decreases the number of influenza patients.


A Prospective Study Evaluating the Clinical Utility of the Tag-Less Patency Capsule with Extended Time for Confirming Functional Patency.
Patency confirmation is useful for avoiding the retention of capsule endoscopy (CE). We first evaluated the clinical utility of the tag-less PillCamTM Patency Capsule (PPC) for up to 72 h after ingestion prospectively. Excretion of an intact capsule or intact body within 72 h, or positive PPC image in the colon or negative PPC image by abdominal X-ray at 30 h was defined as confirmed functional patency. In patients with unconfirmed functional patency, balloon-assisted enteroscopy was performed to evaluate the reason. Functional patency was confirmed in 44 of 57 patients. Patency was confirmed in 38 patients (66.7%) at 30 h. In 6 of 19 patients (31.6%) in whom patency was not confirmed at 30 h, patency was confirmed within 72 h and no capsule retention (CR) occurred. The rate of pan-enteroscopy did not differ between patients whose patency was confirmed at 30 h and those whose patency was confirmed within 72 h. Excretion time of the PPC significantly correlated with that of the capsule (r = 0.650). Severe adhesions (>5 cm) interfered with PPC passage in 2 patients, despite the absence of critical stricture. The rate of no CR in patients with functional patency confirmed by prior PPC was 97.7%. Misjudgment of the location of the PPC is a risk for CR. Extending the time to confirm functional patency to 72 h may be acceptable and increase the possibility to perform CE safely in whom CE can perform intrinsically. University Hospital Medical Information Network registration No: 000002564.


Monoclonal antibodies specific for murine p55 and p75 tumor necrosis factor receptors: identification of a novel in vivo role for p75.
Monoclonal antibodies (mAbs) specific for the murine p55 and p75 tumor necrosis factor (TNF) receptors were produced after immunization of Armenian hamsters with the purified soluble extracellular domains of each receptor protein. Four p55- (55R) and five p75 (TR75)-reactive mAbs immunoprecipitated the appropriate receptor from the surface of L929 cells. None of the mAbs cross-reacted with the other TNF receptor form. The mAbs were functionally characterized by their ability to inhibit ligand binding and influence TNF-dependent L cell cytolytic activity or proliferation of the murine cytolytic T cell clone CT6. One p55-specific mAb, 55R-593, displayed agonist activity, while two other p55-specific mAbs (55R-170 and -176) were found to be TNF antagonists. The fourth mAb (55R-286) had no functional effects on cells. Several antibodies specific for the p75 TNF receptor partially inhibited recombinant murine TNF-alpha-dependent cytolytic activity (60%). Blocking mAbs specific for p75 but not anti-p55 inhibited TNF-mediated proliferation of CT6 T cells. When used in vivo, p55- but not p75-specific mAbs protected mice from lethal endotoxin shock and blocked development of a protective response against Listeria monocytogenes infection. In contrast, both p55 and p75 mAbs individually blocked development of skin necrosis in mice treated with murine TNF-alpha. These data thus demonstrate the utility of the two families of murine TNF receptor-specific mAbs and identify a novel function of the p75 TNF receptor in vivo.


Combining QOF data with the care bundle approach may provide a more meaningful measure of quality in general practice.
A significant minority of patients do not receive all the evidence-based care recommended for their conditions. Health care quality may be improved by reducing this observed variation. Composite measures offer a different patient-centred perspective on quality and are utilized in acute hospitals via the 'care bundle' concept as indicators of the reliability of specific (evidence-based) care delivery tasks and improved outcomes. A care bundle consists of a number of time-specific interventions that should be delivered to every patient every time. We aimed to apply the care bundle concept to selected QOF data to measure the quality of evidence-based care provision. Care bundles and components were selected from QOF indicators according to defined criteria. Five clinical conditions were suitable for care bundles: Secondary Prevention of Coronary Heart Disease (CHD), Stroke & Transient Ischaemic Attack (TIA), Chronic Kidney Disease (CKD), Chronic Obstructive Pulmonary Disease (COPD) and Diabetes Mellitus (DM). Each bundle has 3-8 components. A retrospective audit was undertaken in a convenience sample of nine general medical practices in the West of Scotland. Collected data included delivery (or not) of individual bundle components to all patients included on specific disease registers. Practice level and overall compliance with bundles and components were calculated in SPSS and expressed as a percentage. Nine practices (64.3%) with a combined patient population of 56,948 were able to provide data in the format requested. Overall compliance with developed QOF-based care bundles (composite measures) was as follows: CHD 64.0%, range 35.0-71.9%; Stroke/TIA 74.1%, range 51.6-82.8%; CKD 69.0%, range 64.0-81.4%; and COPD 82.0%, range 47.9-95.8%; and DM 58.4%, range 50.3-65.2%. In this small study compliance with individual QOF-based care bundle components was high, but overall ('all or nothing') compliance was substantially lower. Care bundles may provide a more informed measure of care quality than existing methods. However, the acceptability, feasibility and potential impact on clinical outcomes are unknown.


Inflammatory pseudotumor of the urinary bladder with an aberrant expression of cytokeratin.
A case of inflammatory pseudotumor of the urinary bladder in a 47 year old Japanese male patient is presented. Inflammatory pseudotumor of the urinary bladder is a benign but rare proliferative lesion of the submucosal stroma, easily mistaken for a malignant neoplasm. Based on the clinical diagnosis of bladder cancer by cystoscopy and magnetic resonance imaging (MRI), urologists started chemotherapy before results of the histological report were available which described inflammatory pseudotumor on the biopsy. Biopsied materials showed marked proliferation of irregularly bundled spindle cells, varied in size and shape and separated in severe loose myxoid stroma with moderate infiltration of the inflammatory cells and capillary proliferations. At a glance, these findings resemble the sarcomatous pattern. However neither severe nuclear atypism nor atypical mitoses were present. Immunohistochemically, these spindle cells, which were positive for vimentin and alpha-smooth muscle actin, showed a diffuse aberrant expression of cytokeratin. Some of them were positive for phosphotungstic acid hematoxylin. Electron microscopy revealed only the fibroblasts. No recurrence has been observed for 10 months. These findings indicate that inflammatory pseudotumor is a benign mesenchymal lesion that must be discriminated from true sarcoma to avoid subjecting the patient to unnecessary therapy. Only careful histological examination can enable a successful diagnosis.


Analyzing Policies Through a DOHaD Lens: What Can We Learn?
Social, health, and environmental policies are critical tools for providing the conditions needed for healthy populations. However, current policy analyses fall short of capturing their full potential impacts across the life course and from generation to generation. We argue that the field of Developmental Origins of Health and Disease (DOHaD), a conceptual and research framework positing that early life experiences significantly affect health trajectories across the lifespan and into future generations, provides an important lens through which to analyze social policies. To illustrate this point, we synthesized evidence related to policies from three domains-family leave, nutrition, and housing-to examine the health implications for multiple generations. We selected these policy domains because they represent increasing distance from a reproductive health focus, each with a growing evidence base to support a potential impact on pregnant women and their offspring. Each of these examples represents an opportunity to extend our understanding of policy impact using a DOHaD lens, taking into account the potential life course and intergenerational effects that have previously been overlooked.


Effects of hyper- and hypoprolactinemia on gonadotropin secretion, rat testicular luteinizing hormone/human chorionic gonadotropin receptors and testosterone production by isolated Leydig cells.
The effect of prolactin (Prl) on gonadotropin secretion, testicular luteinizing hormone (LH)/human chorionic gonadotropin (hCG) receptors, and testosterone (T) production by isolated Leydig cells has been studied in 60-day-old rats treated for 4 days, 4 and 8 weeks with sulpiride (SLP), a dopaminergic antagonist, or for 4 days and 4 weeks with bromocriptine (CB), a dopaminergic agonist. Plasma Prl concentrations were significantly greater in the SLP groups (204 +/- 6 ng/ml) and lower in the CB groups (3.0 +/- 0.2 ng/ml) than those measured in the control groups (54 +/- 6 ng/ml). The plasma concentrations of gonadotropin were not affected by a 4-day treatment with SLP or CB, nor were they after a 4-week treatment with CB. However, the hyperprolactinemia induced by an 8-week treatment with SLP was associated with a reduced secretion of gonadotropin (LH, 16 +/- 4 vs. 35 +/- 6 ng/ml; FSH, 166 +/- 12 vs. 307 +/- 14 ng/ml). In SLP-induced hyperprolactinemia, a 30% increase in the density of the LH/hCG testicular binding sites was observed (178 +/- 12 fmol/mg protein), whereas a 60% decrease was measured in hypoprolactinemia (55 +/- 5 vs. control 133 +/- 5 fmol/mg protein). Plasma T levels were increased in 4-day and 4-week hyperprolactinemic animals (4.3 +/- 0.4 and 3.9 +/- 0.4 ng/ml, respectively), but returned to normal levels in the 8-week group (3.0 +/- 0.5 vs. C: 2.3 +/- 0.2 ng/ml). No T modifications were observed in hypoprolactinemic animals. Two distinct populations of Leydig cells (I and II) were obtained by centrifugation of dispersed testicular cells on a 0-45% continuous Metrizamide gradient. Both possess LH/hCG binding sites. However, the T production from Leydig cells of population II increased in the presence of hCG, whereas that of cell population I which also contain immature germinal cells did not respond. The basal and stimulated T secretions from cell populations I and II obtained from CB-treated animals were similar to controls, whereas from 4 days to 8 weeks of hyperprolactinemia, basal and hCG induced T productions from cell population II decreased progressively. These data show that hyperprolactinemia causes, in a time-dependent manner, a trophic effect on the density of LH/hCG testicular receptors; reduces basal and hCG-stimulated T production from isolated Leydig cells type II; and results in an elevated plasma T concentration which decreases with time. The latter suggests a slower T catabolism and/or an impaired peripheral conversion of T into 5 alpha-dihydrotestosterone (DHT). Although hypoprolactinemia is associated with a marked reduction in testicular LH receptors, it does not affect T production.


Angular amplification by a diffraction grating for chiro-optical measurements.
The angles at which a light beam gets diffracted by a grating depend strongly on the direction of incidence for diffraction angles close to a right angle. Accordingly, it is possible to amplify small beam deflections by placing a grating at an optimal orientation to the light path. We use this principle to amplify small beam deviations arising out of a light beam refracting at the interface of an optically active medium, and demonstrate a new technique of enhancing the limit of detection of chiro-optical measurements.


Competency based medical education in gastrointestinal motility.
Traditional apprenticeship-based medical education methods focusing on subjective evaluations and case-volume requirements do not reliably produce clinicians that provide high-quality care in unsupervised practice. Consequently, training approaches are shifting towards competency based medical education, which incorporates robust assessment methods and credible standards of physician proficiency. However, current gastroenterology and hepatology training in the US continues to utilize procedural volume and global impressions without standardized criteria as markers of competence. In particular, efforts to optimize competency based training in gastrointestinal (GI) motility are not underway, even though GI motility disorders account for nearly half of outpatient gastroenterology visits. These deficiencies compromise the quality of patient care. Thus, there is a great need and opportunity to shift our focus in GI motility training towards a competency based approach. First, we need to clarify the variable rates of learning for individual diagnostic tests. We must develop integrated systems that standardize training and monitor physician competency for GI motility diagnostics. Finally, as a profession and society, we must create certification processes to credential competent physicians. These advances are critical to optimizing the quality of GI motility diagnostics in practice.


A Dynamic Core in Human NQO1 Controls the Functional and Stability Effects of Ligand Binding and Their Communication across the Enzyme Dimer.
Human NAD(P)H:quinone oxidoreductase 1 (NQO1) is a multi-functional protein whose alteration is associated with cancer, Parkinson's and Alzheimer´s diseases. NQO1 displays a remarkable functional chemistry, capable of binding different functional ligands that modulate its activity, stability and interaction with proteins and nucleic acids. Our understanding of this functional chemistry is limited by the difficulty of obtaining structural and dynamic information on many of these states. Herein, we have used hydrogen/deuterium exchange monitored by mass spectrometry (HDXMS) to investigate the structural dynamics of NQO1 in three ligation states: without ligands (NQO1apo), with FAD (NQO1holo) and with FAD and the inhibitor dicoumarol (NQO1dic). We show that NQO1apo has a minimally stable folded core holding the protein dimer, with FAD and dicoumarol binding sites populating binding non-competent conformations. Binding of FAD significantly decreases protein dynamics and stabilizes the FAD and dicoumarol binding sites as well as the monomer:monomer interface. Dicoumarol binding further stabilizes all three functional sites, a result not previously anticipated by available crystallographic models. Our work provides an experimental perspective into the communication of stability effects through the NQO1 dimer, which is valuable for understanding at the molecular level the effects of disease-associated variants, post-translational modifications and ligand binding cooperativity in NQO1.


Novel small molecule drugs inhibit tumor cell metabolism and show potent anti-tumorigenic potential.
Rapidly dividing tumor cells have an increased demand for nutrients to support their characteristic unabated growth; this demand is met by an increased availability of nutrients such as amino acids through vasculogenesis and by the enhanced cellular entry of nutrients through the upregulation of specific transporters. Deprivation of intracellular amino acids or block of amino acid uptake has been shown to be cytotoxic to many established human cancer cell lines in vitro and in human cancer xenograft models. In this paper, we provide evidence that the two small molecule oxyphenisatine analogs TOP001 and TOP216 exert their anti-cancer effect by affecting tumor cell metabolism and inducing intracellular amino acid deprivation, leading to a block of cell proliferation. GCN2-mediated phosphorylation of eIF2α as well as mTOR pathway inhibition supports the above notion. In addition, these novel anti-cancer compounds inhibit DNA and protein synthesis and induce apoptosis in a broad spectrum of cancer cell lines. In vivo, the compounds induce tumor stasis and regression in mouse xenograft models of human breast, prostate, ovarian and pancreatic cancer, both when administered intravenously and orally. In conclusion, these small molecules, built on a 1,3-dihydroindole-2-one scaffold, elicit strong anti-proliferative and cytotoxic activity, and importantly, a strong anti-tumorigenicity is observed in in vivo xenograft models of human breast, ovary, prostate and pancreatic cancers encouraging the translation of this class of compounds into the clinic.


Multiple-mouse magnetic resonance imaging with cryogenic radiofrequency probes for evaluation of brain development.
Multiple-mouse magnetic resonance imaging (MRI) increases scan throughput by imaging several mice simultaneously in the same magnet bore, enabling multiple images to be obtained in the same time as a single scan. This increase in throughput enables larger studies than otherwise feasible and is particularly advantageous in longitudinal study designs where frequent imaging time points result in high demand for MRI resources. Cryogenically-cooled radiofrequency probes (CryoProbes) have been demonstrated to have significant signal-to-noise ratio benefits over comparable room temperature coils for in vivo mouse imaging. In this work, we demonstrate implementation of a multiple-mouse MRI system using CryoProbes, achieved by mounting four such coils in a 30-cm, 7-Tesla magnet bore. The approach is demonstrated for longitudinal quantification of brain structure from infancy to early adulthood in a mouse model of Sanfilippo syndrome (mucopolysaccharidosis type III), generated by knockout of the Hgsnat gene. We find that Hgsnat-/- mice have regionally increased growth rates compared to Hgsnat+/+ mice in a number of brain regions, notably including the ventricles, amygdala and superior colliculus. A strong sex dependence was also noted, with the lateral ventricle volume growing at an accelerated rate in males, but several structures in the brain parenchyma growing faster in females. This approach is broadly applicable to other mouse models of human disease and the increased throughput may be particularly beneficial in studying mouse models of neurodevelopmental disorders.


Infant milk-feeding practices and cardiovascular disease outcomes in offspring: a systematic review.
During the Pregnancy and Birth to 24 Months Project, the US Departments of Agriculture and Health and Human Services initiated a review of evidence on diet and health in these populations. The aim of these systematic reviews was to examine the relation of 1) never versus ever feeding human milk, 2) shorter versus longer durations of any human milk feeding, 3) shorter versus longer durations of exclusive human milk feeding, and 4) lower versus higher intensities of human milk fed to mixed-fed infants with intermediate and endpoint cardiovascular disease (CVD) outcomes in offspring. The Nutrition Evidence Systematic Review team conducted systematic reviews with external experts. We searched CINAHL, Cochrane, Embase, and PubMed for articles published January 1980-March 2016, dual-screened the results using predetermined criteria, extracted data from and assessed the risk of bias for each included study, qualitatively synthesized the evidence, developed conclusion statements, and graded the strength of the evidence. The 4 systematic reviews included 13, 24, 6, and 0 articles, respectively. The evidence was insufficient to draw conclusions about endpoint CVD outcomes across all 4 systematic reviews. Limited evidence suggests that never versus ever being fed human milk is associated with higher blood pressure within a normal range at 6-7 y of age. Moderate evidence suggests there is no association between the duration of any human milk feeding and childhood blood pressure. Limited evidence suggests there is no association between the duration of exclusive human milk feeding and blood pressure or metabolic syndrome in childhood. Additional evidence about intermediate outcomes for the 4 systematic reviews was scant or inconclusive. There is insufficient evidence to draw conclusions about the relationships between infant milk-feeding practices and endpoint CVD outcomes; however, some evidence suggests that feeding less or no human milk is not associated with childhood hypertension.


What Box: A task for assessing language lateralization in young children.
The assessment of active language lateralization in infants and toddlers is challenging. It requires an imaging tool that is unintimidating, quick to setup, and robust to movement, in addition to an engaging and cognitively simple language processing task. Functional Transcranial Doppler Ultrasound (fTCD) offers a suitable technique and here we report on a suitable method to elicit active language production in young children. The 34-second "What Box" trial presents an animated face "searching" for an object. The face "finds" a box that opens to reveal a to-be-labelled object. In a sample of 95 children (1 to 5 years of age), 81% completed the task-32% with ≥10 trials. The task was validated (ρ = 0.4) against the gold standard Word Generation task in a group of older adults (n = 65, 60-85 years of age), though was less likely to categorize lateralization as left or right, indicative of greater measurement variability. Existing methods for active language production have been used with 2-year-old children while passive listening has been conducted with sleeping 6-month-olds. This is the first active method to be successfully employed with infants through to pre-schoolers, forming a useful tool for populations in which complex instructions are problematic.


Feeding on grains containing pesticide residues is detrimental to offspring development through parental effects in grey partridge.
Numerous toxicological studies have shown that ingestion of pesticides can induce physiological stress in breeding birds, with adverse consequences on egg laying parameters and offspring quality through parental effects. However, previous studies do not mimic current levels of pesticide residues in typical landscapes, and they do not consider potential cocktail effects of pesticides as they occur in the wild. Herein, we explored whether realistic pesticide exposure affected reproduction parameters and offspring condition through parental effects in Grey partridge. We fed 24 breeding pairs with either seeds from conventional agriculture crops treated with various pesticides during cropping, or organic grains without pesticide residues as controls. The conventional and organic grain diets mimicked food options potentially encountered by wild birds in the field. The results showed that ingesting low pesticide doses over a long period had consequences on reproduction and offspring quality without altering mortality in parents or chicks. Compared with organic pairs, conventional pairs yielded smaller chicks at hatching that had a lower body mass index at 24 days old. Additionally, these chicks displayed lower haematocrit when body mass index was higher. Therefore, ingestion of conventional grains by parents resulted in chronic exposure to pesticide residues, even at low doses, and this had detrimental consequences on offspring. These results demonstrate a sublethal effect of pesticide residues through parental effects. The consequences of parental exposure on chicks might partly explain the decline in wild Grey partridge populations, which raises questions for avian conservation and demography if current agrosystem approaches are continued.