The olfactory amygdala in amniotes: an evo-devo approach.
In tetrapods, the medial amygdala is a forebrain center that integrates olfactory and/or vomeronasal signals with the endocrine and autonomic systems, playing a key role in different social behaviors. The vomeronasal system has undergone important changes during evolution, which may be behind some interspecies differences in chemosensory-mediated social behavior. These evolutionary changes are associated with variations in vomeronasal-recipient brain structures, including the medial amygdala. Herein, we employed an evolutionary developmental biology approach for trying to understand the function and evolution of the medial amygdala. For that purpose, we reviewed published data on fate mapping in mouse, and the expression of orthologous developmental regulatory genes (Nkx2.1, Lhx6, Shh, Tbr1, Lhx9, Lhx5, Otp, and Pax6) in embryos of mouse, chicken, emydid turtles, and a pipid frog. We also analyzed novel data on Lhx9 and Otp in a lacertid lizard. Based on distinct embryonic origin and genetic profile, at least five neuronal subpopulations exist in the medial amygdala of rodents, expressing either Nkx2.1/Lhx6, Shh, Lhx9, Otp/Lhx5, or Pax6. Each neuronal subpopulation appears involved in different functional pathways. For example, Lhx6 cells are specifically activated by sex pheromones and project to preoptic and hypothalamic centers involved in reproduction. Based on data in nonmammals, at least three of these neuronal subtypes might have been present in the medial amygdala of the amniote common ancestor. During mammalian evolution, the downregulation of Nkx2.1 in the alar hypothalamus may have been a driving force for an increment of the Otp/Lhx5 subpopulation.


Exercise induced muscle weakness in a young adult: McArdle's disease unusual presentation.
McArdle's disease (Glycogen storage disease type V) is a rare inherited autosomal recessive disease involving defect in enzyme, glycogen phosphorylase (PYGM) which results in accumulation of glycogen mainly affecting skeletal muscles. It commonly presents in childhood and rarely in adults with symptoms like exercise intolerance, muscle weakness, cramps and fatigue. Herein, we report an unusual case of a 22 years old male in Pakistan with probable McArdle's Disease presenting with repeated episodes of generalized cramping muscle pain, exercise intolerance and haematuria. The diagnostic approach to identifying this disease as well as the differentials of other rare types of skeletal muscle disorders that should be kept in mind while dealing with a similar clinical picture, irrespective of the age of presentation, have been discussed.


Psychological intervention to prevent ICU-related PTSD: who, when and for how long?
Experiencing treatment on a modern intensive care unit (ICU) is a potentially traumatic event. People who experience traumatic events have an increased risk of depression, anxiety disorders and post-traumatic stress disorder (PTSD). Extended follow-up has confirmed that many patients suffer physical and psychological consequences of the ICU treatment up to 12 months after hospital discharge. PTSD in particular has become increasingly relevant in both the immediate and longer-term follow-up care of these patients. The extent to which the consequences of critical illness and the treatments received in the ICU contribute to the development of PTSD is poorly understood and more rigorous studies are needed. Understanding the factors associated with a poor psychological recovery after critical illness is essential to generate models of causality and prognosis, and to guide the delivery of effective, timely interventions.


Human pluripotent stem cells differentiated in fully defined medium generate hematopoietic CD34- and CD34+ progenitors with distinct characteristics.
Differentiation of pluripotent stem cells in vitro provides a powerful means to investigate early developmental fates, including hematopoiesis. In particular, the use of a fully defined medium (FDM) would avoid biases induced by unidentified factors contained in serum, and would also allow key molecular mediators involved in such a process to be identified. Our goal was to induce in vitro, the differentiation of human embryonic stem cells (ESC) and induced pluripotent stem cells (iPSC) into morphologically and phenotypically mature leukocytes and erythrocytes, in the complete absence of serum and feeder cells. ESC and iPSC were sequentially induced in liquid cultures for 4 days with bone morphogenic protein-4, and for 4 days with FLT3-ligand, stem cell factor, thrombopoietin and vascular endothelium growth factor. Cell differentiation status was investigated by both mRNA expression and FACS expression profiles. Cells were further sorted and assayed for their hematopoietic properties in colony-forming unit (CFU) assays. In liquid cultures, cells progressively down-modulated Oct-4 expression while a sizeable cell fraction expressed CD34 de novo. SCL/Tal1 and Runx1 transcripts were exclusively detected in CD34(+) cells. In clonal assays, both ESC and iPSC-derived cells generated CFU, albeit with a 150-fold lower efficacy than cord blood (CB) CD34(+) cells. ESC-derived CD34(+) cells generated myeloid and fully hemoglobinized erythroid cells whereas CD34(-) cells almost exclusively generated small erythroid colonies. Both ESC and iPSC-derived erythroid cells expressed embryonic and fetal globins but were unable to synthesize adult β-globin in contrast with CB cells, suggesting that they had differentiated from primitive rather than from definitive hematopoietic progenitors. Short-term, animal protein-free culture conditions are sufficient to sustain the differentiation of human ESC and iPSC into primitive hematopoietic progenitors, which, in turn, produce more mature blood cell types. However, additional factors have yet to be identified to allow their differentiation into definitive erythroid cultures.


Inhibitory mechanism of inteferon-gamma on human fibroblasts from Tenon's capsule.
The inhibitory mechanism of interferon-gamma (IFN-gamma) on the fibroblasts from Tenon's capsule was studied. By using immunohistochemical SP method and pathological image system, the inhibitory effects of IFN-gamma on the expression of transforming growth factor beta receptor I in the in vitro cultured fibroblasts from Tenon's capsule were quantitatively analyzed. The results showed that IFN-gamma could reduce the expression of transforming growth factor beta receptor I in the fibroblasts with the following dose-effect relationship: Y = 1937.5-134.2 Igx (r=-0.971, P<0.01). It was concluded that IFN-gamma could inhibit the expression of transforming growth factor beta receptor I in the fibroblasts from Tenon's capsule. The modulation of the transforming growth factor beta receptor I expression by IFN-gamma may be beneficial to the alleviation of the hyperplasia of scar after trabeculectomy.


beta-carotene: friend or foe?
This symposium focused on the research which documents benefit and toxicity in beta-carotene supplementation. Reflecting on past and current studies, the panel of experts discussed: (1) the potential harm of a high intake of beta-carotene on selected populations, (2) biochemical antioxidant/prooxidant mechanisms of beta-carotene at the cellular level, (3) potential benefits of other carotenoids and antioxidants, and (4) future directions for research in beta-carotene and other antioxidants.


Renal medullary carcinoma: molecular, immunohistochemistry, and morphologic correlation.
Renal medullary carcinoma, a highly aggressive tumor mainly occurring in patients with sickle cell hemoglobinopathy, is characterized by advanced stage at the time of presentation and poor response to treatment. Currently, the pathogenesis of this tumor is not well understood. In this study, the clinicopathologic features and molecular changes of 15 renal medullary carcinoma cases were evaluated. These cases demonstrated male predominance (M:F=2:1) with a median age of 26 years. The tumors occurred predominantly in the right kidney with an average size of 5.9 cm. Immunohistochemistry analysis showed that the neoplastic cells were positive for CEA (7/8), AE1/3 (8/8), CAM5.2 (7/7), CK7 (5/5), CK20 (4/6), and vimentin (6/6). Absence of SMARCB1 protein expression in tumor cells was demonstrated in all of the 7 cases analyzed. By polymerase chain reaction-based microsatellite analysis, loss of heterozygosity of SMARCB1 was identified in 9 of 10 cases. These data suggest that inactivation of SMARCB1 may play a role in the pathogenesis of renal medullary carcinoma.


Effect of N-bromoacetamide on single sodium channel currents in excised membrane patches.
We have studied the effect of N-bromoacetamide (NBA) on the behavior of single sodium channel currents in excised patches of rat myotube membrane at 10 degree C. Inward sodium currents were activated by voltage steps from holding potentials of about -100 mV to test potentials of -40 mV. The cytoplasmic-face solution was isotonic CsF. Application of NBA or pronase to the cytoplasmic face of the membrane irreversibly removed sodium channel inactivation, as determined by averaged single-channel records. Teh lifetime of the open channel at -40 mV was increased about 10-fold by NBA treatment without affecting the amplitude of single-channel currents. A binomial analysis was used both before and after treatment to determine the number of channels within the excised patch. NBA was shown to have little effect on activation kinetics, as determined by an examination of both the rising phase of averaged currents and measurements f the delay between the start of the pulse and the first channel opening. Our data support a kinetic model of sodium channel activation in which the rate constant leading back from the open state to the last closed state is slower than expected from a strict Hodgkin-Huxley model. The data also suggest that the normal open-channel lifetime is primarily determined by the inactivation process in the voltage range we have examined.


A sphenodontine (Rhynchocephalia) from the Miocene of New Zealand and palaeobiogeography of the tuatara (Sphenodon).
Jaws and dentition closely resembling those of the extant tuatara (Sphenodon) are described from the Manuherikia Group (Early Miocene; 19-16 million years ago, Mya) of Central Otago, New Zealand. This material is significant in bridging a gap of nearly 70 million years in the rhynchocephalian fossil record between the Late Pleistocene of New Zealand and the Late Cretaceous of Argentina. It provides the first pre-Pleistocene record of Rhynchocephalia in New Zealand, a finding consistent with the view that the ancestors of Sphenodon have been on the landmass since it separated from the rest of Gondwana 82-60 Mya. However, if New Zealand was completely submerged near the Oligo-Miocene boundary (25-22 Mya), as recently suggested, an ancestral sphenodontine would need to have colonized the re-emergent landmass via ocean rafting from a currently unrecorded and now extinct Miocene population. Although an Early Miocene record does not preclude that possibility, it substantially reduces the temporal window of opportunity. Irrespective of pre-Miocene biogeographic history, this material also provides the first direct evidence that the ancestors of the tuatara, an animal often perceived as unsophisticated, survived in New Zealand despite substantial local climatic and environmental changes.


Fractures of the Nasal Bones: Is External Splinting Really Warranted?
It has been advocated that reduction of nasal bone fractures should be followed by internal packing and/or external splinting. Despite the ample literature concerning the advantages and limitations of various splint types, the necessity and effectiveness of external splinting has not been well documented. To present the authors' experience and review the literature on treatment of nasal bone fractures, focusing on the indications and effectiveness of external splinting following closed reduction. Retrospective analysis and literature review. Medical records of all patients, treated at the Department of Oral and Maxillofacial Surgery of the "KAT" General Hospital of Attica between January 2010 and December 2016 for facial trauma including nasal bone fractures, were retrospectively reviewed. Patient demographic data, fracture type, applied treatment, complications, and final outcome were registered. A total of 77 patients (58 males; 19 females) were included in the study. The age range was 18 to 65 years (mean, 37.8). Closed reduction without external splinting was performed in 63 patients and open reduction with internal fixation in 6; 8 severely comminuted fractures were treated with closed reduction and external splinting. The mean follow-up was 4.8 months. All severely comminuted fractures presented complications. External splinting following closed reduction of nasal bone fractures should not be used routinely but only in selected patients with severe comminution. Since the pertinent literature is inconclusive on the indications and effectiveness of external splinting, randomized controlled studies are warranted to fully elucidate the issue.


Effects of fishing technique on assessing species composition in aquatic systems in semi-arid Brazil.
In most ecological field research, appropriate sampling is critical for the understanding of processes underlying fish populations and communities, and is even more important in heterogeneous environments such as the aquatic systems of the semi-arid region of Brazil. This study intends to make a contribution to the development of sampling programs and gear selection in aquatic systems of semi-arid Brazil by evaluating the effects of different fishing techniques on the assessment of richness and composition of the fish fauna in selected aquatic environments. Six sites were selected to represent typical artificial (reservoirs) and natural (intermittent streams) environments and four different types of sampling gear were applied to each site during four occasions. The present study shows that when selecting sampling techniques to be used in aquatic systems in semi-arid Brazil, one must consider the objectives of the study, e.g. ecological or taxonomic, in order to decide on inclusion of rare species in the sampling population. Also, the effect of the sampling gear on natural abundances of fish must be considered given that some sampling techniques are highly detrimental to fish population numbers.


Visceral White Adipose Tissue after Chronic Intermittent and Sustained Hypoxia in Mice.
Angiogenesis, a process induced by hypoxia in visceral white adipose tissues (vWAT) in the context of obesity, mediates obesity-induced metabolic dysfunction and insulin resistance. Chronic intermittent hypoxia (IH) and sustained hypoxia (SH) induce body weight reductions and insulin resistance of different magnitudes, suggesting different hypoxia inducible factor (HIF)-1α-related activity. Eight-week-old male C57BL/6J mice (n = 10-12/group) were exposed to either IH, SH, or room air (RA). vWAT were analyzed for insulin sensitivity (phosphorylated (pAKT)/AKT), HIF-1α transcription using chromatin immunoprecipitation (ChIP)-sequencing, angiogenesis using immunohistochemistry, and gene expression of different fat cell markers and HIF-1α gene targets using quantitative polymerase chain reaction or microarrays. Body and vWAT weights were reduced in hypoxia (SH > IH > RA; P < 0.001), with vWAT in IH manifesting vascular rarefaction and increased proinflammatory macrophages. HIF-1α ChIP-sequencing showed markedly increased binding sites in SH-exposed vWAT both at 6 hours and at 6 weeks compared with IH, the latter also showing decreased vascular endothelial growth factor, endothelial nitric oxide synthase, P2RX5, and PAT2 expression, and insulin resistance (IH > > > SH = RA; P < 0.001). IH induces preferential whitening of vWAT, as opposed to prominent browning in SH. Unlike SH, IH elicits early HIF-1α activity that is unsustained over time and is accompanied by concurrent vascular rarefaction, inflammation, and insulin resistance. Thus, the dichotomous changes in HIF-1α transcriptional activity and brown/beige/white fat balance in IH and SH should enable exploration of mechanisms by which altered sympathetic outflow, such as that which occurs in apneic patients, results in whitening, rather than the anticipated browning of adipose tissues that occurs in SH.


Stereotactic navigation with the O-arm for placement of S-2 alar iliac screws in pelvic lumbar fixation.
Pelvic fixation is a crucial adjunct to many lumbar fusions to avoid L5-S1 pseudarthrosis. It is useful for treatment of kyphoscoliosis, high-grade spondylolisthesis, L5-S1 pseudarthrosis, sacral tumors, lumbosacral dislocations, and osteomyelitis. The most popular method, iliac fixation, has drawbacks including hardware prominence, extensive muscle dissection, and the need for connection devices. S-2 alar iliac fixation provides a useful primary or salvage alternative. The authors describe their techniques for using stereotactic navigation for screw placement. The O-arm Surgical Imaging System allowed for CT-quality multiplanar reconstructions of the pelvis, and registration to a StealthStation Treon provided intraoperative guidance. The authors describe their technique for performing computer-assisted S-2 alar iliac fixation for various indications in 18 patients during an 18-month period. All patients underwent successful bilateral placement of screws 80-100 mm in length. All placements were confirmed with a second multiplanar reconstruction. One screw was moved because of apparent anterior breach of the ilium. There were no immediate neurological or vascular complications due to screw placement. The screw length required additional instruments including a longer pedicle finder and tap. Stereotactic guidance to navigate the placement of distal pelvic fixation with bilateral S-2 alar iliac fixation can be safely performed in patients with a variety of pathological conditions. Crossing the sacroiliac joint, choosing trajectory, and ensuring adequate screw length can all be enhanced with 3D image guidance. Long-term outcome studies are underway, specifically evaluating the sacroiliac joint.


[Esophageal and gastric fistulas following surgery of the esophagogastric junction].
Seven cases of oesophageal and gastric fistulae after hiatal hernia surgery are reported: 4 fistulae occurred after an abdominal anti-reflux repair; 3 after an intrathoracic Nissen repair. The clinical symptomatology of these complications highly influences their therapeutical strategy. The fistulae with only an abdominal manifestation (subphrenic abscess or peritonitis) are opposed to those with mediastinal or pleural symptoms. The different modalities of treatment and their results are analysed. The authors prefer in case of a fistula with abdominal manifestation a directed fistulization associated with enteral alimentation; in case of a fistulae with thoracic manifestation an oesophageal exclusion with delayed restoration of G.I. tract continuity is indicated.


Antiretroviral drug transporters and metabolic enzymes in human testicular tissue: potential contribution to HIV-1 sanctuary site.
The testes are a potential viral sanctuary site for HIV-1 infection. Our study aims to provide insight into the expression and localization of key drug transporters and metabolic enzymes relevant to ART in this tissue compartment. We characterized gene and protein expression of 12 representative drug transporters and two metabolic enzymes in testicular tissue samples obtained from uninfected (n = 8) and virally suppressed HIV-1-infected subjects on ART (n = 5) and quantified antiretroviral drug concentrations in plasma and testicular tissues using LC/MS/MS from HIV-1-infected subjects. Our data demonstrate that key ABC drug transporters (permeability glycoprotein, multidrug-resistance protein 1, 2 and 4, and breast cancer resistance protein), solute carrier transporters (organic anion transporting polypeptides 1B1 and 2B1, organic anion transporter 1, concentrative nucleoside transporter 1, equilibrative nucleoside transporter 2) and cytochrome P450 metabolic enzymes (CYP3A4 and CYP2D6) previously shown to interact with many commonly used antiretroviral drugs are expressed at the mRNA and protein level in the testes of both subject groups and localize primarily at the blood-testis barrier, with no significant differences between the two groups. Furthermore, we observed that PIs known to be substrates for ATP-binding cassette membrane transporters, displayed variable testicular tissue penetration, with darunavir concentrations falling below therapeutic values. In contrast, the NRTIs emtricitabine, lamivudine and tenofovir displayed favourable tissue penetration, reaching concentrations comparable to plasma levels. We also demonstrated that nuclear receptors, peroxisome proliferator-activated receptors α and γ exhibited higher gene expression in the testicular tissue compared with pregnane X receptor and constitutive androstane receptor, suggesting a potential regulatory pathway governing drug transporter and metabolic enzyme expression in this tissue compartment. Our data suggest the testes are a complex pharmacological compartment that can restrict the distribution of certain antiretroviral drugs and potentially contribute to HIV-1 persistence.


Affinity of mismatch-binding protein MutS for heteroduplexes containing different mismatches.
We have used bandshift analysis to measure the interaction between the Escherichia coli mismatch-binding protein MutS and synthetic DNA fragments containing all possible DNA mismatches as well as an unpaired T (DeltaT). The order of affinity is found to be DeltaT>GT>GG>AA approximately TT approximately TC>CA>GA>CC>GC. We find that the affinity for GT mismatches is affected by the flanking sequence and decreases in the order G(n)C(n)>(GC)(n)>A(n)T(n)>(AT)(n). Studies with base analogues show good binding to phiT (where phi represents 1',2'-dideoxyribose), but much weaker binding to Gphi.


Longitudinal and shear mode ultrasound propagation in human skull bone.
Recent studies have attempted to dispel the idea of the longitudinal mode being the only significant mode of ultrasound energy transport through the skull bone. The inclusion of shear waves in propagation models has been largely ignored because of an assumption that shear mode conversions from the skull interfaces to the surrounding media rendered the resulting acoustic field insignificant in amplitude and overly distorted. Experimental investigations with isotropic phantom materials and ex vivo human skulls demonstrated that, in certain cases, a shear mode propagation scenario not only can be less distorted, but at times allowed for a substantial (as much as 36% of the longitudinal pressure amplitude) transmission of energy. The phase speed of 1.0-MHz shear mode propagation through ex vivo human skull specimens has been measured to be nearly half of that of the longitudinal mode (shear sound speed = 1500 +/- 140 m/s, longitudinal sound speed = 2820 +/- 40 m/s), demonstrating that a closer match in impedance can be achieved between the skull and surrounding soft tissues with shear mode transmission. By comparing propagation model results with measurements of transcranial ultrasound transmission obtained by a radiation force method, the attenuation coefficient for the longitudinal mode of propagation was determined to between 14 Np/m and 70 Np/m for the frequency range studied, while the same for shear waves was found to be between 94 Np/m and 213 Np/m. This study was performed within the frequency range of 0.2 to 0.9 MHz.


Effects of immigration on the incidence of congenital hypothyroidism.
The incidence of congenital hypothyroidism (CH) has been shown to vary among different parts of the world. This could result from environmental or hereditary factors. Studies of other congenital diseases have shown that immigrants tend to retain the incidence of their country of origin while their children acquire the incidence of their new homeland, suggesting an environmental influence. This study aimed to assess the differences in the incidence of CH among immigrants from different parts of the world and to study the effects of immigration on its occurrence. During the 9-year period between 1979 and 1987, 196 Jewish infants with primary CH were born in Israel; this constitutes an incidence of 1:3354 live births. We collected data from hospitals, endocrine pediatric clinics and the children's parents regarding the birth place of the parents and grandparents of those infants. These data were compared with the birth place of the parents and grandparents of all infants born in Israel during that period in order to learn about the incidence of CH among infants of different origins and to compare the incidence between children of parents born in Israel and those of immigrants of the same grandparental origin. CH incidence was lower among offspring of mothers and fathers of Israeli origin (1:4717 and 1:4255 live births respectively) and higher among those of African mothers (1:2950) and Asian fathers (1:2941). Parents of Asian or African origin, born in Israel have a lower incidence of CH-affected children compared with parents of the same origin born in their own continent. This trend is reversed for European and American parents, for whom being born in Israel is related to an increase in the CH incidence in their children. The difference in CH incidence between offspring of parents born in Israel and those of parents born in their original country was statistically significant (P < 0.05). In the different origin groups the gender of the parent did not influence significantly the incidence of CH. Environmental changes resulting from immigration can influence the incidence of congenital hypothyroidism.


Barriers and Facilitators of Conducting Medication Reviews in Nursing Home Residents: A Qualitative Study.
Objectives: Inappropriate medication prescribing is a recognized clinical problem in nursing home residents of whom many have polypharmacy. However, results about the effectiveness of medication reviews targeted at improving prescribing and deprescribing have been equivocal. We therefore examined barriers and facilitators of conducting medication reviews. Method: We purposively sampled medication reviews to capture salient barriers and facilitators of conducting medication reviews both in nursing home care units for dementia and disabling conditions. We held semi-structured interviews about consecutive steps of medication reviews. Interviews were transcribed verbatim and analyzed with the "method of constant comparison." Results: Six nursing home residents/relatives of nursing home residents, 8 elder care physicians, 5 pharmacists, and 10 nurses took part in the semi-structured interviews. We observed four overarching themes of barriers and facilitators: "realizing fidelity of the patient perspective (theme 1)," "level of comprehensiveness of medication reviews (theme 2)," "inclinations of healthcare providers (theme 3)," and "inter-professional collaboration and alliances (theme 4)." Theme 1 "realizing fidelity of the patient perspective" referred to the observation that assessing the patient perspective was a delicate balance between the value and the impediments of a proper assessment of the patient perspective. Theme 2 "level of comprehensiveness of medication reviews" reflected the struggle of practitioners to find an optimum between medication reviews being both comprehensive and feasible. Theme 3 "inclinations of healthcare providers" concerned setting intervention targets that were complementary to the practices of physicians and keeping the pharmacist blind to the patient perspective as a countermeasure to physicians' inclinations. Finally, theme 4 "inter-professional collaboration and alliances" highlighted mutual support and inter-professional collaboration to strengthen healthcare practitioners' contributions. Discussion: These themes of barriers and facilitators emphasize the need to improve meta-communication during the medication review process. This pertains to the need for healthcare providers to appraise the fidelity of the patient perspective in a dialogue with residents/relatives. Furthermore, discourse between healthcare practitioners is needed beforehand about the level of comprehensiveness intervention targets, and inter-professional collaboration.


[Behavior patterns of the population in relation to health].
In order to identify the basic patterns of behavior that reflect the attitude of Russians to their health, a large corpus of relevant scientific literature was analyzed, as well as actual statistical data and the results of representative sociological studies concerning various aspects of both self-preserving and self-destructive behavior of Russians. As a result, four basic models of behavior were formed and described, which differ in a set of positive and negative attitudes and characteristic lines of behavior in relation to one's health («a convinced healthy lifestyle person», «a beginner healthy lifestyle person», «irresponsible» and «self-destructive»). Based on the analysis of statistics and sociological information, it was concluded that even the behavioral model of a «beginner healthy lifestyle person» is implemented by a very minority of Russians. At best, the majority of the population adheres to the «irresponsible» pattern of behavior that involves refusing or reducing the consumption of alcohol and tobacco, but not assuming a rational diet, adequate sleep, physical and medical activity. Hence the task of everyone who has the opportunity to influence citizens, including medical workers, to convey to them the idea that health and longevity are achieved only through systematic and comprehensive efforts. In addition, a number of common misconceptions and unrealistic expectations related to healthy lifestyles were highlighted. Attention is drawn to the fact that subjects capable of exerting an educational impact on citizens, including medical specialists, should not only provide patients with useful knowledge, but also try to protect them from misconceptions and unfounded expectations.


NMDA receptor-mediated control of presynaptic calcium and neurotransmitter release.
Before action potential-evoked Ca2+ transients, basal presynaptic Ca2+ concentration may profoundly affect the amplitude of subsequent neurotransmitter release. Reticulospinal axons of the lamprey spinal cord receive glutamatergic synaptic input. We have investigated the effect of this input on presynaptic Ca2+ concentrations and evoked release of neurotransmitter. Paired recordings were made between reticulospinal axons and the neurons that make axo-axonic synapses onto those axons. Both excitatory and inhibitory paired-cell responses were recorded in the axons. Excitatory synaptic inputs were blocked by the AMPA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 10 microM) and by the NMDA receptor antagonist 2-amino-5-phosphonopentanoate (AP-5; 50 microM). Application of NMDA evoked an increase in presynaptic Ca2+ in reticulospinal axons. Extracellular stimulation evoked Ca2+ transients in axons when applied either directly over the axon or lateral to the axons. Transients evoked by the two types of stimulation differed in magnitude and sensitivity to AP-5. Simultaneous microelectrode recordings from the axons during Ca2+ imaging revealed that stimulation of synaptic inputs directed to the axons evoked Ca2+ entry. By the use of paired-cell recordings between reticulospinal axons and their postsynaptic targets, NMDA receptor activation was shown to enhance evoked release of transmitter from the axons that received axoaxonic inputs. When the synaptic input to the axon was stimulated before eliciting an action potential in the axon, transmitter release from the axon was enhanced. We conclude that NMDA receptor-mediated input to reticulospinal axons increases basal Ca2+ within the axons and that this Ca2+ is sufficient to enhance release from the axons.


Effects of Mindfulness-Based Interventions in High School and College Athletes for Reducing Stress and Injury, and Improving Quality of Life.
Clinical Scenario: Student athletes experience a variety of stressors from school and social activities, as well as the additional demands of sport participation. Mindfulness-based interventions can help increase mental awareness and acceptance, as well as mitigate negative thoughts and emotions. The use of mindfulness-based interventions may be beneficial for reducing thoughts of stress, injury reduction, and improving overall wellbeing. Does the use of mindfulness-based interventions for student-athletes aged 13-24 years reduce stress and injury as well as improve overall quality of life? The literature was searched for studies that investigated the use of mindfulness-based strategies for student-athletes specifically for reducing stress and injury and/or improving quality of life. The literature search returned 8 possible studies related to the clinical question and 3 studies met the inclusion criteria (1 randomized control trial, 2 nonrandomized control cohort studies). All 3 included studies demonstrated overall improved levels of mindfulness among student-athletes after the use of a mindfulness-based intervention. Mindfulness-based interventions had positive effects for reducing negative thoughts and levels of perceived stress. The number of injury occurrences were found to decrease following the integration of a mindfulness-based intervention within an athletic population. Clinical Bottom Line: There is sufficient evidence supporting the use of mindfulness-based interventions with student-athletes for increasing mindfulness, managing negative emotions and perceived stress, as well as improving overall well-being. There is also current literature that advocates the use of mindfulness-based interventions for reducing injury, but further research is needed for support. Strength of Recommendation: Grade B evidence exists to support that the use of mindfulness-based interventions for student-athletes will reduce stress and improve overall well-being as well as support the possibility that if a student-athlete is more mindful, it may help decrease risk of injury incurred if the student-athlete is under mental stress.


[The role of the adenylate cyclase system in the inhibition of gastric secretion by products of limited hydrolysis of kappa-casein].
The influence of the peptide inhibiting gastric acid secretion and derived from kappa-casein on the levels of cAMP, cGMP and adenylate cyclase activity of rat gastric mucosa was studied. Intravenous peptide preparation administration failed to alter cyclic nucleotide levels. The revealed decrease of adenylate cyclase activity was due to the presence of calcium in the investigated peptide preparation. It was assumed that cyclase system of the rat gastric mucosa cells were not involved in the inhibition of gastric acid secretion by the peptide preparations under study.


Ayahuasca, dimethyltryptamine, and psychosis: a systematic review of human studies.
Ayahuasca is a hallucinogen brew traditionally used for ritual and therapeutic purposes in Northwestern Amazon. It is rich in the tryptamine hallucinogens dimethyltryptamine (DMT), which acts as a serotonin 5-HT2A agonist. This mechanism of action is similar to other compounds such as lysergic acid diethylamide (LSD) and psilocybin. The controlled use of LSD and psilocybin in experimental settings is associated with a low incidence of psychotic episodes, and population studies corroborate these findings. Both the controlled use of DMT in experimental settings and the use of ayahuasca in experimental and ritual settings are not usually associated with psychotic episodes, but little is known regarding ayahuasca or DMT use outside these controlled contexts. Thus, we performed a systematic review of the published case reports describing psychotic episodes associated with ayahuasca and DMT intake. We found three case series and two case reports describing psychotic episodes associated with ayahuasca intake, and three case reports describing psychotic episodes associated with DMT. Several reports describe subjects with a personal and possibly a family history of psychosis (including schizophrenia, schizophreniform disorders, psychotic mania, psychotic depression), nonpsychotic mania, or concomitant use of other drugs. However, some cases also described psychotic episodes in subjects without these previous characteristics. Overall, the incidence of such episodes appears to be rare in both the ritual and the recreational/noncontrolled settings. Performance of a psychiatric screening before administration of these drugs, and other hallucinogens, in controlled settings seems to significantly reduce the possibility of adverse reactions with psychotic symptomatology. Individuals with a personal or family history of any psychotic illness or nonpsychotic mania should avoid hallucinogen intake.


Experimental orthotopic heart and bilateral lung transplantation completed without cardiopulmonary bypass.
Most experimental studies of orthotopic heart and lung graft failure are complicated by an inability to eliminate the rejection-specific inflammatory mediator from the cardiopulmonary bypass. The following model was developed in our laboratory to investigate the feasibility of performing an orthotopic heart and bilateral lung transplantation without performing a cardiopulmonary bypass. Nineteen transplants were attempted using 19 pairs of mongrel dogs. The recipient dog (mean weight, 23 kg) was anesthetized, and the ascending aorta, the superior vena cava (SVC), the inferior vena cava (IVC), and the main bronchus were dissected. Then, the donor dog (mean weight, 20 kg) was anesthetized, and the heart and lung block was prepared and explanted from the chest under cardioplegic arrest. A Gore-tex shunt (W. L. Gore; Flagstaff, AZ) was placed side-to-side between the recipient IVC and SVC, and then the donor right atrium was anastomosed to the Gore-tex shunt. The donor ascending aorta was anastomosed to the recipient ascending aorta with a partial clamp. On completion of these anastomoses, the donor heart was reperfused by the recipient heart and allowed to beat. When hemodynamic conditions were stable with double hearts, the recipient SVC and IVC were ligated just proximal to the venous anastomosis and the recipient aorta was ligated proximal to the anastomotic site. The recipient trachea was anastomosed to the donor trachea with an end-to-end anastomosis. Finally, the recipient heart and lungs were removed from the chest and the sternum was closed. Four of the 19 transplants failed. Three died due to left ventricular dysfunction, and one died due to bleeding. Mean (+/- SD) ischemic time was 67 +/- 11 min with a mean (+/- SD) anastomotic time of 54 +/- 12 min. The 15 survivors were hemodynamically stable with or without the minimal use of inotropic support (dopamine, 2 to 3 microg/kg/min) 6 h after grafting, with normal cardiac output, satisfactory oxygenation, and normal wall motion. The sternotomy was repaired without loss of cardiopulmonary function. On the basis of our experiences, the experimental model of orthotopic heart and bilateral lung transplantation completed "off pump" can be technically feasible without the loss of cardiac and pulmonary functions.


Disease severity in ankylosing spondylitis: variation by region and local area deprivation.
To investigate whether patient disease severity in ankylosing spondylitis (AS) varies among regions or by local area social deprivation. Eight hundred patients with AS from 8 specialist rheumatology centers across England were invited to participate in a cross-sectional survey. Sociodemographic and disease-related variables were collected [pain (numerical rating scale), disease activity (Bath AS Disease Activity Index), and physical function (Bath AS Functional Index)]. Deprivation was measured using the Index of Multiple Deprivation 2004. Of the 800 patients invited, 468 responded (adjusted response rate 62.8%). Most were male (72.9%), with a mean age of 50.2 years (SD 12.1), and a mean diagnosed disease duration of 17 years (SD 11.4). Across all centers, those living in more deprived areas demonstrated significantly greater disease severity and poorer psychological health. After controlling for age, gender, disease duration, and region, greater deprivation was significantly associated with greater disease activity (OR 3.39; 95% CI 1.65, 6.98) and poorer function (OR 4.46; 95% CI 2.11, 9.44). There was a nonsignificant trend toward more pain (OR 1.98; 95% CI 0.97, 4.07). There was also a significant independent association between region and disease severity. The need for healthcare is greatest for patients with AS who are living in more socially deprived areas. With the growing use of interventional therapies, these findings have important implications if health service resources are to be allocated equitably; particularly as deprived patients are known to access healthcare less frequently.


Spatially resolving the secretome within the mycelium of the cell factory Aspergillus niger.
Aspergillus niger is an important cell factory for the industrial production of enzymes. These enzymes are released into the culture medium, from which they can be easily isolated. Here, we determined with stable isotope dimethyl labeling the secretome of five concentric zones of 7-day-old xylose-grown colonies of A. niger that had either or not been treated with cycloheximide. As expected, cycloheximide blocked secretion of proteins at the periphery of the colony. Unexpectedly, protein release was increased by cycloheximide in the intermediate and central zones of the mycelium when compared to nontreated colonies. Electron microscopy indicated that this is due to partial degradation of the cell wall. In total, 124 proteins were identified in cycloheximide-treated colonies, of which 19 secreted proteins had not been identified before. Within the pool of 124 proteins, 53 secreted proteins were absent in nontreated colonies, and additionally, 35 proteins were released ≥4-fold in the central and subperipheral zones of cycloheximide-treated colonies when compared to nontreated colonies. The composition of the secretome in each of the five concentric zones differed. This study thus describes spatial release of proteins in A. niger, which is instrumental in understanding how fungi degrade complex substrates in nature.


Methylation of the CpG sites only on the sense strand of the APC gene is specific for hepatocellular carcinoma.
Hypermethylation of the promoter of the tumor suppressor gene, adenomatous polyposis coli (APC), occurs in various malignancies, including hepatocellular carcinoma (HCC). However, reports on the specificity of the methylation of the APC gene for HCC have varied. To gain insight into how these variations occur, bisulfite PCR sequencing was performed to analyze the methylation status of both sense and antisense strands of the APC gene in samples of HCC tissue, matched adjacent non-HCC liver tissue, hepatitis, cirrhosis, and normal liver tissues. DNA derived from fetal liver and 12 nonhepatic normal tissue was also examined. These experiments revealed liver-specific, antisense strand-biased CpG methylation of the APC gene and suggested that, although methylation of the antisense strand of the APC gene exists in normal liver and other non-HCC disease liver tissue, methylation of the sense strand of the APC gene occurs predominantly in HCC. To determine the effect of the DNA strand on the specificity of the methylated APC gene as a biomarker for HCC detection, quantitative methylation-specific PCR assays for sense and antisense strand DNA were developed and performed on DNA isolated from HCC (n = 58), matched adjacent non-HCC (n = 58), cirrhosis (n = 41), and hepatitis (n = 39). Receiver operating characteristic curves were constructed. With the cutoff value set at the limit of detection, the specificity of sense and antisense strand methylation was 84% and 43%, respectively, and sensitivity was 67.2% and 72.4%, respectively. This result demonstrated that the identity of the methylated DNA strand impacted the specificity of APC for HCC detection. Interestingly, methylation of the sense strand of APC occurred in 40% of HCCs from patients with serum AFP levels less than 20 ng/mL, suggesting a potential role for APC as a biomarker to complement AFP in HCC screening.


Reimagining WHO: leadership and action for a new Director-General.
Three candidates to be the next WHO Director-General remain: Tedros Adhanom Ghebreyesus, David Nabarro, and Sania Nishtar. The World Health Assembly's ultimate choice will lead an organisation facing daunting internal and external challenges, from its own funding shortfalls to antimicrobial resistance and immense health inequities. The new Director-General must transform WHO into a 21st century institution guided by the right to health. Topping the incoming Director-General's agenda will be a host of growing threats-risks to global health security, antimicrobial resistance, non-communicable diseases, and climate change-but also the transformative potential of the Sustainable Development Goals, including their universal health coverage target. Throughout, the next Director-General should emphasise equality, including through national health equity strategies and, more boldly still, advancing the Framework Convention on Global Health. Success in these areas will require a reinvigorated WHO, with sustainable financing, greater multisector engagement, enhanced accountability and transparency, and strengthened normative leadership. WHO must also evolve its governance to become far more welcoming of civil society and communities. To create the foundation for these transformative changes, the Director-General will need to focus first on gaining political support. This entails improving accountability and transparency to gain member state trust, and enabling meaningful civil society participation in WHO's governance and standing up for the right to health to gain civil society support. Ultimately, in the face of a global environment marked by heightened nationalism and xenophobia, member states must empower the next Director-General to enable WHO to be a bulwark for health and human rights, serving as an inspiring contra-example to today's destructive politics, demonstrating that the community of nations are indeed stronger together.


Association of paternal age with perinatal outcomes between 2007 and 2016 in the United States: population based cohort study.
To evaluate the impact of advanced paternal age on maternal and perinatal outcomes in the United States. Retrospective, population based cohort study. US. 40 529 905 documented live births between 2007 and 2016. Primary perinatal outcomes were gestational age, birth weight, Apgar score at five minutes, admission to a neonatal intensive care unit, need for postpartum antibiotics, and seizures. Primary maternal outcomes were gestational diabetes and pre-eclampsia. Secondary outcome was the number of preventable perinatal events. Higher paternal age was associated with an increased risk of premature birth, low birth weight, and low Apgar score. After adjustment for maternal age, infants born to fathers aged 45 years or older had 14% higher odds of premature birth (odds ratio 1.14, 95% confidence interval 1.13 to 1.15), independent of gestational age, and 18% higher odds of seizures (1.18, 0.97 to 1.44) compared with infants of fathers aged 25 to 34 years. The odds of gestational diabetes was 34% higher (1.34, 1.29 to 1.38) in mothers with the oldest partners. 13.2% (95% confidence interval 12.5% to 13.9%) of premature births and 18.2% (17.5% to 18.9%) of gestational diabetes in births associated with older fathers were estimated to be attributable to advanced paternal age. Advanced paternal age is associated with negative effects on both mothers and offspring. Given the relatively low prevalence of advanced paternal age in the US, population level impacts are currently modest. Nevertheless, as advanced paternal age has doubled in the US over the past generation, further investigation is warranted of the impact on birth outcomes and public health.


Are Brain-Computer Interfaces Feasible With Integrated Photonic Chips?
The present paper examines the viability of a radically novel idea for brain-computer interface (BCI), which could lead to novel technological, experimental, and clinical applications. BCIs are computer-based systems that enable either one-way or two-way communication between a living brain and an external machine. BCIs read-out brain signals and transduce them into task commands, which are performed by a machine. In closed loop, the machine can stimulate the brain with appropriate signals. In recent years, it has been shown that there is some ultraweak light emission from neurons within or close to the visible and near-infrared parts of the optical spectrum. Such ultraweak photon emission (UPE) reflects the cellular (and body) oxidative status, and compelling pieces of evidence are beginning to emerge that UPE may well play an informational role in neuronal functions. In fact, several experiments point to a direct correlation between UPE intensity and neural activity, oxidative reactions, EEG activity, cerebral blood flow, cerebral energy metabolism, and release of glutamate. Therefore, we propose a novel skull implant BCI that uses UPE. We suggest that a photonic integrated chip installed on the interior surface of the skull may enable a new form of extraction of the relevant features from the UPE signals. In the current technology landscape, photonic technologies are advancing rapidly and poised to overtake many electrical technologies, due to their unique advantages, such as miniaturization, high speed, low thermal effects, and large integration capacity that allow for high yield, volume manufacturing, and lower cost. For our proposed BCI, we are making some very major conjectures, which need to be experimentally verified, and therefore we discuss the controversial parts, feasibility of technology and limitations, and potential impact of this envisaged technology if successfully implemented in the future.


Limited diagnostic usefulness of antibodies to cytoplasmic proteins of Brucella in early-treated human brucellosis.
Antibodies to cytoplasmic proteins (CP) of Brucella have been shown to be useful for the diagnosis of human brucellosis; however, some early-diagnosed patients lack such an antibody response while having high titers of antibodies to lipopolysaccharide (LPS). To address which factors determine this serological discrepancy in the early stages of brucellosis we examined the antibody response to CP and LPS of 21 patients involved in an outbreak of B. melitensis infection who had a short duration of clinical illness at diagnosis (3-40 d). At diagnosis, antibodies to LPS (IgM and/or IgG) were found in all patients, while anti-CP antibodies were detected in 16 subjects (76%). At 6 weeks post-diagnosis IgG to CP (with or without IgM) had been detected in 13 patients and IgM alone had been found in 4; however, 4 other patients (19%) had no response to CP. No significant differences were found between these 3 groups in terms of age, gender, antimicrobial agents or factors that could hamper the immune response. Notably, however, the 4 non-responders and 3 of the 4 patients having only IgM to CP had started antibiotic therapy within 14 d post-symptoms, while treatment was started later in 9 of 13 patients who developed anti-CP IgG. In addition, maximum titers of IgG to CP tended to be lower in early-treated patients. These results suggest that very early antibiotic therapy hampers the antibody response to Brucella CP but has little impact on the anti-LPS response. Given the higher specificity of the former and the higher sensitivity of the latter, both reactivities should be measured in order to diagnose human brucellosis.


Combined treatment with alendronate and Drynaria rhizome extracts : Effect on fracture healing in osteoporotic rats.
The effects of both alendronate (ALN) and Drynaria rhizome extracts (DRE) alone could promote bone healing in osteoporotic fractures but there are no reports about the combined use of ALN and DRE for promotion of bone healing of fractures in osteoporotic settings. This study investigated the effects of ALN plus DRE on fractures in osteopenic rats. Osteopenic rats underwent unilateral transverse osteotomy on the femur fixed by a sterilized Kirschner wire 2 weeks after intragastric administration of retinoic acid (80 mg/kg body weight/day). Subsequently, the animals were randomly divided into four groups: control, ALN, DRE and ALN + DRE. All rats from groups ALN, DRE and ALN + DRE received ALN (40 mg/kg, weekly), DRE (90 mg/kg/day), or both for 2, 4 and 6 weeks. The results of our study indicated that all treatment promoted fracture healing and callus formation compared to controls but ALN + DRE treatment showed significantly stronger effects than ALN or DRE alone in histological, X‑ray and biomechanical tests. These results seem to indicate that combined treatment with ALN and DRE has an additive effect on fracture healing and callus formation in osteoporotic rats.


[Cyclosporin A for treatment in nephrotic syndrome of childhood--a preliminary report].
We treated five, three steroid resistant and 2 steroid dependent nephrotic syndrome with oral cyclosporin A and prednisolone. All children received more than eight weeks course of prednisolone, and were in a critically ill status from their nephrotic syndrome and by steroid-toxic side effects. Cyclosporin A was started at 7 mg/kg/day and titrated to maintain serum level of 150-250 ng/ml. Prednisolone was given initially at a dose of 1 mg/kg/day x 1 month, subsequently at a dose of 0.4 mg/kg/day x 1 month, then 0.2 mg/kg/day x 4 months. After a total course of six months, cyclosporin A was tapered to 3.5 mg/kg/day. Prednisolone was maintained at a dose of 0.2 mg/kg/day. Renal biopsies of the 3 steroid resistant children showed membranous glomerulonephritis with focal segmental glomerulosclerosis, focal segmental glomerulosclerosis and IgM nephropathy respectively. These three cases all went into complete remission within four weeks. Among them, one case relapsed twice within one year. Renal biopsies of the two steroid dependent children showed minimal change and focal segmental glomerulosclerosis respectively. These two cases went into complete remission within three weeks sustained up to a year. Side effects of cyclosporin A were observed in one patient with gum hypertrophy and two patients with hirsutism. Other side effects were not found. After cyclosporin A treatment for one year, the renal biopsy of the relapsing patient showed no interstitial fibrosis or tubular atrophy. In conclusion, cyclosporin A was found to be effective in steroid resistant and dependent patients. The duration of cyclosporin A treatment is so far unknown. It needs further evaluation in the future.


Evidence for somatic rearrangement of immunoglobulin genes coding for variable and constant regions.
A high-molecular-weight DNA from Balb/c mouse early embryo or from MOPC 321 plasmacytoma (a k-chain producer) was digested to completion with Bacillus amyloliquefaciens strain H restriction enzyme (BamH I). The resulting DNA fragments were fractionated according to size in preparative agarose gel electrophoresis. DNA fragments carrying gene sequences coding for the variable or constant region of k chains were detected by hybridization with purified, 125I-labeled, whole MOPC 321 K MRNA and with its 3'-end half. The pattern of hybridization was completely different in the genomes of embryo cells and of the plasmacytoma. The pattern of embryo DNA showed two components, one of which (molecular weight=6.0 million) hybridized with C-gene sequences and the other (molecular weight=3.9 million) with V-gene sequences. The pattern of the tumor DNA showed a single component that hybridized with both V-gene and C-gene sequences and that is smaller (molecular weight=2.4 million) than either of the components in embryo DNA. The results were interpreted to mean that the Vk and Ck genes, which are some distance away from each other in the embryo cells, are joined to form a contiguous polynucleotide stretch during differentiation of lymphocytes. Such joining occurs in both of the homologous chromosomes. Relevance of these findings with respect to models for V-C gene joining, activation of a specific V k gene, and allelic exclusion in immunoglobulin gene loci is discussed.


Effects of discrimination by sex and race on the early careers of British medical graduates during 1981-7.
To examine the possible effects of discrimination by sex and race on the career patterns of doctors up to six years after qualifying. Postal questionnaire follow up survey. 1572 Doctors who graduated from five British medical schools in 1981, 1983, and 1985, including 587 women and 131 doctors from ethnic minorities. Reported success rates of applications for training posts. Comparison of the career patterns of women and men yielded no evidence of discrimination against women in competition for posts. In contrast, there were striking differences in career patterns between graduates of native European origin and those of ethnic minority origin. Graduates from ethnic minorities reported lower success rates and more difficulty in obtaining house officer posts, registrar posts, and places in vocational training schemes in general practice. Most of this discrimination seemed to occur at the stage of shortlisting for interview. Graduates from ethnic minorities were more likely than graduates of native European origin to have experienced spells of unemployment while seeking work. They were also more likely to have changed their original choice of career because of difficulty in obtaining suitable training posts or unfavourable career prospects. Discrimination against ethnic minorities occurs in the competition for training posts among graduates from British medical schools. There was no evidence of discrimination against women graduates. Shortlisting procedures based on objective scoring systems may help to ensure equality of opportunity in future.


Endogenous syntaxins 2, 3 and 4 exhibit distinct but overlapping patterns of expression at the hepatocyte plasma membrane.
To investigate the mechanisms regulating polarized vesicle delivery to the cell surface in hepatocytes, we have characterized the endogenous plasma membrane (PM)-associated syntaxins. These integral membrane proteins are components of the membrane docking/fusion apparatus and are thought to function as vesicle receptors at the PM. In hepatocytes, the PM is divided into two domains, the apical and basolateral. If syntaxins are mediating the specific recognition of vesicles delivered to either membrane surface, the simple prediction is that each domain expresses one syntaxin isoform. However, we report that rat hepatocytes express three endogenous PM-associated syntaxin isoforms, syntaxins 2, 3 and 4. By biochemical subfractionation, we determined that the syntaxins exhibit distinct, but overlapping patterns of expression among the PM domains. Syntaxin 4 is primarily expressed at the basolateral surface while syntaxins 2 and 3 are enriched at the apical PM. The immunolocalization of syntaxins 2 and 4 in rat hepatocytes and PM sheets revealed similarly complex patterns of PM expression with enhanced apical staining for both. A significant proportion of syntaxin 3 (25%) was detected in subcellular fractions containing transport vesicles. We have used quantitative immunoblotting to determine that the syntaxins are relatively abundant PM molecules (11-260 nM) in rat liver, spleen and kidney. Also, we determined that the syntaxin binding protein, Munc-18, is present at concentrations from 1.5-20 nM in the same tissues. Although this fundamental quantitative and morphological information is lacking in other systems, it is critical not only for defining syntaxin function, but also for predicting the specific mechanisms that regulate vesicle targeting in hepatocytes and other tissues.


Tumor cell cycle arrest induced by shear stress: Roles of integrins and Smad.
Interstitial flow in and around tumor tissue affects the mechanical microenvironment to modulate tumor cell growth and metastasis. We investigated the roles of flow-induced shear stress in modulating cell cycle distribution in four tumor cell lines and the underlying mechanisms. In all four cell lines, incubation under static conditions for 24 or 48 h led to G(0)/G(1) arrest; in contrast, shear stress (12 dynes/cm(2)) induced G(2)/M arrest. The molecular basis of the shear effect was analyzed, and the presentation on molecular mechanism is focused on human MG63 osteosarcoma cells. Shear stress induced increased expressions of cyclin B1 and p21(CIP1) and decreased expressions of cyclins A, D1, and E, cyclin-dependent protein kinases (Cdk)-1, -2, -4, and -6, and p27(KIP1) as well as a decrease in Cdk1 activity. Using specific antibodies and small interfering RNA, we found that the shear-induced G(2)/M arrest and corresponding changes in G(2)/M regulatory protein expression and activity were mediated by alpha(v)beta(3) and beta(1) integrins through bone morphogenetic protein receptor type IA-specific Smad1 and Smad5. Shear stress also down-regulated runt-related transcription factor 2 (Runx2) binding activity and osteocalcin and alkaline phosphatase expressions in MG63 cells; these responses were mediated by alpha(v)beta(3) and beta(1) integrins through Smad5. Our findings provide insights into the mechanism by which shear stress induces G(2)/M arrest in tumor cells and inhibits cell differentiation and demonstrate the importance of mechanical microenvironment in modulating molecular signaling, gene expression, cell cycle, and functions in tumor cells.


Chatbots for HIV Prevention and Care: a Narrative Review.
To explore the intersection of chatbots and HIV prevention and care. Current applications of chatbots in HIV services, the challenges faced, recent advancements, and future research directions are presented and discussed. Chatbots facilitate sensitive discussions about HIV thereby promoting prevention and care strategies. Trustworthiness and accuracy of information were identified as primary factors influencing user engagement with chatbots. Additionally, the integration of AI-driven models that process and generate human-like text into chatbots poses both breakthroughs and challenges in terms of privacy, bias, resources, and ethical issues. Chatbots in HIV prevention and care show potential; however, significant work remains in addressing associated ethical and practical concerns. The integration of large language models into chatbots is a promising future direction for their effective deployment in HIV services. Encouraging future research, collaboration among stakeholders, and bold innovative thinking will be pivotal in harnessing the full potential of chatbot interventions.


Signal-amplifying nanoparticle/hydrogel hybrid microarray biosensor for metal-enhanced fluorescence detection of organophosphorus compounds.
In this study, we developed an enzyme-based miniaturized fluorescence biosensor to detect paraoxon, one of the most well-known neurotoxic organophosphorus compounds. The biosensor was fabricated with poly(ethylene glycol) (PEG) hydrogel microarrays that entrapped acetylcholinesterase (AChE) and quantum dots (QDs) as fluorescence reporters. Metal-enhanced fluorescence (MEF) was utilized to amplify the fluorescence signal, which was achieved by decorating QDs on the surface of silica-coated silver nanoparticles (Ag@Silica). The MEF effects of Ag@Silica were optimized by tuning the thickness of the silica shells, and under the optimized conditions, the fluorescence intensity was shown to be increased 5 fold, compared with the system without MEF. PEG hydrogel microarray entrapping QD-decorated Ag@Silica and AChE was prepared via photopatterning process. The entrapped AChE hydrolyzed paraoxon to produce p-nitrophenol within the hydrogel microstructure, which subsequently quenched the fluorescence of the QDs on the surface of Ag@Silica. The MEF-assisted fluorescence detection resulted in a significant enhancement of paraoxon detection. The detection limit was approximately 1.0 × 10-10 M and 2.0 × 10-7 M for sensing with and without MEF, respectively. The successful integration of a hydrogel microarray system with a microfluidic system was demonstrated to be a potential application for the MEF-based micro-total-analysis-system.


Use of isoelectric focusing and a chromophoric organomercurial to monitor urea-induced conformational changes of yeast phosphoglycerate kinase.
The effects of urea in concentrations from 0 to 6M on the following properties of yeast phosphoglycerate kinase were studied: the kinetics of inactivation of the enzyme, the spectrum of 2-chloromercuri-4-nitrophenol bound to the single thiol group of the enzyme, the rate of reaction between the mercurial and enzyme, and the isoelectric point. The enzyme was inactivated by as much as 30% in 1M-urea, and the other data were interpreted as a possible 'tightening' of enzyme structure. The catalytic behaviour of the enzyme in 2M-urea was time-dependent, the initial effects being similar to those in 1M-urea. Polyacrylamide-gel isoelectric focusing of the enzyme in the presence of 2M-urea showed a single species of enzyme with an isoelectric point intermediate between those in 1M- and 3M-urea; a species with an identical isoelectric point was obtained after an 11-day exposure at 4 degrees C to the denaturant at 2M. The enzyme was rapidly inactivated in 3M-urea, with the thiol group fully exposed and the isoelectric point 0.9pH unit higher than in the absence of urea. No further conformational changes could be demonstrated with urea concentrations of 4M or greater. It is suggested that the equilibrium species that exists in 2M-urea has one of two buried lysine residues exposed. The second lysine residue is exposed in 3M or greater concentrations of the denaturant.


Relapses in recurrent depression 1 year after psychoeducational treatment: the role of therapist adherence and competence, and the therapeutic alliance.
Psychoeducation has proved to be an effective treatment method for the prevention of relapse in recurrent depression. However, little is known about the processes which could account for the effects of psychoeducational treatment. In this study, patients with recurrent depression (currently remitted) received, over a period of 8 months, 16 sessions of psychoeducational treatment, in order to prevent relapse. Therapist adherence and competence, and the therapeutic alliance, were investigated as predictors of reducing the recurrence risk in depression. Videotapes of 43 participants in a psychoeducational treatment for depression were analyzed, in order to evaluate therapist adherence and competence. Additionally, the therapeutic alliance was assessed by means of a questionnaire. One year after treatment, no associations were found between therapist adherence or competence and the risk of relapse. The patients' view of the therapeutic alliance was moderately associated with the time to relapse. However, the correlation disappeared when controlled for the number of previous depressive episodes. The latter was the most important predictor of time to relapse, explaining 15% of variance.


Antimalarial aminothiazoles and aminopyridines from phenotypic whole-cell screening of a SoftFocus(®) library.
The current state of antimalarial drug resistance emphasizes the need for new therapies with novel modes of action that will add a significant benefit compared with current standards. In this regard, high throughput phenotypic whole-cell screening aids the discovery of novel antiplasmodial scaffolds that are inherently suited to hit-to-lead and lead-optimization efforts. The aminothiazoles and aminopyridines exemplify two such compound classes stemming from whole-cell screening. Respective structure-activity relationship determinations and subsequent optimization around these scaffolds led to frontrunner compounds in each series, which possess the desired antimalarial efficacy, bioavailability and metabolic stability to further progress medicinal chemistry programs.


Genomic distribution of simple sequence repeats in Brassica rapa.
Simple Sequence Repeats (SSRs) represent short tandem duplications found within all eukaryotic organisms. To examine the distribution of SSRs in the genome of Brassica rapa ssp. pekinensis, SSRs from different genomic regions representing 17.7 Mb of genomic sequence were surveyed. SSRs appear more abundant in non-coding regions (86.6%) than in coding regions (13.4%). Comparison of SSR densities in different genomic regions demonstrated that SSR density was greatest within the 5'-flanking regions of the predicted genes. The proportion of different repeat motifs varied between genomic regions, with trinucleotide SSRs more prevalent in predicted coding regions, reflecting the codon structure in these regions. SSRs were also preferentially associated with gene-rich regions, with peri-centromeric heterochromatin SSRs mostly associated with retrotransposons. These results indicate that the distribution of SSRs in the genome is non-random. Comparison of SSR abundance between B. rapa and the closely related species Arabidopsis thaliana suggests a greater abundance of SSRs in B. rapa, which may be due to the proposed genome triplication. Our results provide a comprehensive view of SSR genomic distribution and evolution in Brassica for comparison with the sequenced genomes of A. thaliana and Oryza sativa.


Nephrogenic metaplasia of urinary tract in children: report of three cases and review of the literature.
Nephrogenic metaplasia of the urinary tract, originally thought to be a benign tumor with possible malignant potential, is commonly called nephrogenic adenoma. It predominantly affects male adults and is rarely seen in children. In this report 18 pediatric cases are reviewed and some clinical and pathologic parameters are compared with the condition in adults. Male to female ratio is reversed (3.7:1 in adults and 1:3.5 in children). Recurrences are more common and are more frequently multiple in the pediatric age group. Although no malignant transformation has been reported, prolonged follow-up is recommended since the natural history of this lesion is still uncertain.


Inflammatory markers in skeletal muscle of older adults.
Older adults have an increase in circulating markers of inflammation. The current study examined whether there is an increase in the expression of inflammatory markers within the vastus lateralis, a major locomotive muscle, of older adults, and if so, whether the reduction in muscle strength and aerobic capacity in older adults is related to increased muscle inflammation. Skeletal muscle biopsies were taken from older adults (n = 17, 67 ± 1.6 years) and young individuals (n = 16, 24 ± 0.6 years) under resting and fasting conditions. Muscle was analyzed for mRNA levels of intracellular inflammatory molecules (MCP1, TNFα and IL-1β) and total cellular protein abundance of cytokines, chemokines and kinases (IL-6, IL-8, MCP1, TNFα, p65 (NF-κB), JNK1/2 and STAT3). MCP1 expression was significantly higher (p < 0.05; 50 %, mRNA and 40 %, protein) in elderly than younger participants, as was IL-8 (4 %). No detectable difference in kinase protein expression was observed for STAT3, JNK or p65 (NF-κB), TNFα or IL-6. Muscle strength was lower in the elderly compared to the young group (1.55 ± 0.17 vs. 2.56 ± 0.13 Nm/kg, p < 0.001). The elderly group also had a significantly lower VO(2peak) compared to the young group (24.9 ± 1.9 vs. 39.3 ± 1.9, p < 0.001), but muscle strength and VO(2peak) were not correlated with the examined inflammatory markers. Older adults have increased MCP1 (mRNA and protein abundance) and IL-8 (protein abundance) and also reduced muscle strength and VO(2peak). However, the reduction in muscle strength and VO(2peak) was not related to the increase in muscle inflammatory markers in this cohort.


Leveraging the Biology of Adversity and Resilience to Transform Pediatric Practice.
Advances in science are fundamentally changing the way we understand how inextricable interactions among genetic predispositions, physical and social environments, and developmental timing influence early childhood development and the foundations of health and how significant early adversity can lead to a lifetime of chronic health impairments. This article and companion article illustrate the extent to which differential outcomes are shaped by ongoing interactive adaptations to context that begin at or even before conception and continue throughout life, with increasing evidence pointing to the importance of the prenatal period and early infancy for the developing brain, the immune system, and metabolic regulation. Although new discoveries in the basic sciences are transforming tertiary medical care and producing breakthrough outcomes in treating disease, this knowledge is not being leveraged effectively to inform new approaches to promoting whole-child development and preventing illness. The opportunity for pediatrics to serve as the leading edge of science-based innovation across the early childhood ecosystem has never been more compelling. In this article, we present a framework for leveraging the frontiers of scientific discovery to inform new strategies in pediatric practice and advocacy to protect all developing biological systems from the disruptive effects of excessive early adversity beyond providing information on child development for parents and enriched learning experiences for young children.


Protein vaccine NVX-CoV2373 elicits functional T cell immunity.
The SARS-CoV-2 vaccine NVX-CoV2373 is a protein-based vaccine that might circumvent the difficulties in distributing mRNA vaccines to regions with limited access to cold-chain and refrigeration. However, the NVX-CoV2373-induced T cell and antibody responses remain poorly understood. In this issue of the JCI, Moderbacher et al. characterized SARS-CoV-2-specific CD4+ and CD8+ T cell responses elicited by one or two doses of NVX-CoV2373 in individuals enrolled in a phase I/IIa trial. Substantially increased spike-specific CD4+ and T follicular helper cells were found after the first or second vaccine dose, with some individuals developing a modest spike-specific CD8+ T cell response. Correlation analysis revealed an association between spike-specific CD4+ T cells and neutralizing antibody titers. Notably, preexisting T cell immunity showed negligible effects on NVX-CoV2373-induced T cell responses. These findings indicate that the protein-based vaccine NVX-CoV2373 induces robust T cell immunity capable of recognizing SARS-CoV-2 antigens and supporting humoral immune responses.


Kinetic and thermodynamic studies on removal of Cu(II) from aqueous solutions using soil nanoclays.
Soil clays (< 2,000 nm) (SC) and soil nanoclays (< 100 nm) (SNC) were used as adsorbents for removal of Cu(II) from aqueous solution. The experiments were conducted with variables including pH, interaction time, concentration of Cu(II) and temperature. Four kinetic models have been employed to investigate adsorption mechanisms, and the experimental data more closely resemble a second-order process of the kinetic model. Adsorption studies on soil nanoclays have been shown to be highly effective in removing of Cu(II) from aqueous solution. This adsorbent is widely available as a natural material, is mechanically stable and, most importantly, it is environmentally appealing. The maximum Cu(II) adsorption capacity of soil nanoclays (31.7 mg/g) is more than three times higher than natural soil clays (10.2 mg/g). Our study demonstrates that soil nanoclays can be used effectively for removal of Cu(II) from aqueous systems to achieve environmental cleaning purposes.


Prolactin levels in paternal striped mouse (Rhabdomys pumilio) fathers.
Paternal behavior is associated with an increase in prolactin levels in fish, birds and mammals, including rodents. The striped mouse (Rhabdomys pumilio) from southern Africa shows highly developed paternal care. We investigated whether striped mouse fathers have higher prolactin levels than nonfathers, and whether there is a relationship between tactile stimulation with pups and prolactin secretion in fathers. We measured serum prolactin in 42 male striped mice assigned to one of four different experimental groups (single males, paired males, fathers housed with mother and pups, and fathers separated from their family by a wire-mesh partition). Our results revealed no increases in prolactin levels in fathers, and fathers with tactile contact with pups did not have higher prolactin levels than the fathers that were prevented from making tactile contact with pups. In contrast, experienced males had higher prolactin levels than inexperienced males. Male striped mice are polygynous in nature, living in groups, with three breeding females, and are permanently associated with pups during the breeding season. In a field study, males had higher prolactin levels during the breeding season than during the nonbreeding season. Thus, prolactin secretion in the polygynous striped mouse might be regulated by environmental stimuli, whereas social stimuli might be important for monogamous species. This is the first study to demonstrate seasonal changes in prolactin levels in a free-living male mammal.


Barriers to providing dual energy x-ray absorptiometry services to individuals with spinal cord injury.
To assess barriers to bone mineral density testing in individuals with chronic spinal cord injury, a cross-sectional study of 20 individuals with spinal cord injury was conducted with assessment of physical and logistical barriers to dual energy x-ray absorptiometry scanning. We identified several barriers, including scanner design and configuration in the scanning room that limited accessibility, increased typical scanning time, and made additional staff necessary. For dual energy x-ray absorptiometry to become a routine component of ongoing care in spinal cord injury medicine, we recommend the following changes: (1) install ceiling-mounted hydraulic lifts and grab bars to facilitate transfers in the scanning room; (2) increase staffing during scans; (3) increase time allotment for scans; (4) install the scanner in an adequately-sized room; and (5) partner with administrators and staff to raise awareness of access issues faced by individuals with spinal cord injury.


Phytochrome regulation of pea phototropin.
Type 1 phototropin, one of the blue light receptors responsible for phototropism, is encoded in peas by at least two genes, PsPHOT1A and PsPHOT1B (formerly PsPK4 and PsPK5), both of which are more similar to Arabidopsis PHOT1 than to Arabidopsis PHOT2. We show here that PsPHOT1B encodes a full-length phototropin, whose expression pattern suggests that Psphot1b is the predominant phot1-type phototropin in etiolated seedlings. The gene encoding the other type 1 phototropin, PsPHOT1A, is expressed at low levels, with its highest levels in the leaves and stems of more mature, light-grown plants. Studies with phyA, phyB and the phyAphyB double mutants show that phyA and phyB have partially redundant roles in the reduction of PsPHOT1B expression under red light.


Least Possible Fixation Techniques of 4-Part Valgus Impacted Fractures of the Proximal Humerus: A Systematic Review.
The valgus-impacted (VI) 4-part fractures are a subset of fractures of the proximal humerus with a unique anatomic configuration characterized by a relatively lower incidence of avascular necrosis after operative intervention. We systematically reviewed clinical studies assessing the benefits and harms of least possible fixation techniques (LPFT) for this unique fracture type. Such information would be potentially helpful in developing an evidence-based approach in the management of these complex injuries. We performed analytic searches of PubMed, Embase, Web of Science, Google Scholar and the Cochrane Library, restricting it to the years 1991-2014. Included studies had to describe outcomes and complications after primary osteosynthesis with any type of LPFT apart from plate-screws and intramedullary nailing. Eligibility criteria were also included English language, more than 5 cases, minimum follow up of one year and report of clinical outcome using at least one relevant score (Constant, Neer or ASES). Based on 292 database hits we identified 12 eligible studies including 190 four-part valgus impacted fractures in 188 patients. All eligible studies were case series composed of min 8 to max 45 patients per study. The gender distribution was 60% (112) female and 40% (76) male. The average age of the patients at the time of injury was 54.5 years. In 8/12 studies an open reduction was used for fracture fixation using different surgical techniques including KW, cerclage wires, cannulated screws and osteosutures. Closed reduction and percutaneous fixation was used in 4 studies. Mean follow-up time ranged from 24 to 69 months. A good functional outcome (constant score >80) was reported in 9/12 studies. The most common complication was avascular necrosis of the humeral head with an overall incidence of 11% (range, 0-26.3%). Total avascular necrosis (AVN) was found in 15/188 patients (7.9%) and was more common in percutaneous techniques and partial AVN in 6/188 (3.1%) being more common in open techniques. The overall re-operation rate was very low (3.7%). Insufficient study designs cannot provide definite treatment recommendations and quantitative data synthesis of outcome. In general, LPFT for 4-part VI fractures leads to satisfactory outcomes with low incidence of complications. A considerable risk of biases can be attributed to fracture classification, proper radiological control, mean age of patients, mixed types of fixation methods, nonage adjusted clinical scoring and small follow up periods. These factors are discussed in detail. systematic review of literature (level IV) as most of the studies were level IV.


Interactive Isogeometric Volume Visualization with Pixel-Accurate Geometry.
A recent development, called isogeometric analysis, provides a unified approach for design, analysis and optimization of functional products in industry. Traditional volume rendering methods for inspecting the results from the numerical simulations cannot be applied directly to isogeometric models. We present a novel approach for interactive visualization of isogeometric analysis results, ensuring correct, i.e., pixel-accurate geometry of the volume including its bounding surfaces. The entire OpenGL pipeline is used in a multi-stage algorithm leveraging techniques from surface rendering, order-independent transparency, as well as theory and numerical methods for ordinary differential equations. We showcase the efficiency of our approach on different models relevant to industry, ranging from quality inspection of the parametrization of the geometry, to stress analysis in linear elasticity, to visualization of computational fluid dynamics results.


Oxidative stress, mammospheres and Nrf2-new implication for breast cancer therapy?
Mammosphere culture of breast cancer cell lines is an important approach used for enrichment of cancer stem cells (CSCs), which exhibit high tumorigenicity and chemoresistance features. Evidence shows that CSCs maintain lower ROS levels due to elevated expression of ROS-scavenging molecules and antioxidative enzymes, which favors the survival of the CSCs and their chemoresistance. The transcription factor NF-E2-related factor 2 (Nrf2) has emerged as the master regulator of cellular redox homeostasis, by up-regulating antioxidant response element (ARE)-bearing genes products. Although Nrf2 has long-term been regarded as a beneficial defense mechanism, accumulating studies have revealed the "dark side" of Nrf2. High constitutive levels of Nrf2 was observed in many types of tumors and cancer cell lines promoting their resistance to chemotherapeutics. In this study, we report a high expression of Nrf2 and its target genes in mammospheres compared to corresponding adherent cells. In MCF-7 and MDA-MB-231 mammmosphere cells, the Nrf2-mediated cellular protective response is significantly elevated which is associated with increased resistance to taxol and anchorage-independent growth. Brusatol, an inhibitor of the Nrf2 pathway, suppressed the protein level of Nrf2 and its target genes, enhanced intracellular ROS and sensitized mammospheres to taxol, and reduced the anchorage-independent growth. These results suggest that mammospheres rely on abnormal up-regulation of Nrf2 to maintain low intracellular ROS levels. Nrf2 inhibitors, such as brusatol, have the potential to be developed into novel adjuvant chemotherapeutic drug combinations in order to combat refractory tumor initiating CSCs.


Predictive factors of mortality from nonvariceal upper gastrointestinal hemorrhage: a multicenter study.
From an Italian Registry of patients with upper gastrointestinal hemorrhage (UGIH), we assessed the clinical outcomes and explored the roles of clinical, endoscopic, and therapeutic factors on 30-day mortality in a real life setting. Prospective analysis of consecutive patients endoscoped for UGIH at 23 community and tertiary care institutions from 2003 to 2004. Covariates and outcomes were defined a priori and 30-day follow-up obtained. Logistic regression analysis identified predictors of mortality. One thousand and twenty patients were included. A total of 46 patients died for an overall 4.5% mortality rate. In all, 85% of deaths were associated with one or more major comorbidity. Sixteen of 46 patients (35%) died within the first 24 h of the onset of bleeding. Of these, eight had been categorized as ASA class 1 or 2 and none of them was operated upon, despite a failure of endoscopic intention to treatment in four. Regression analysis showed advanced age, presence of severe comorbidity, low hemoglobin levels at presentation, and worsening health status as the only independent predictors of 30-day mortality (P < 0.001). The acute use of a PPI exerted a protective effect (OR 0.23, 95% CI 0.09-0.73). Recurrent bleeding was low (3.2%). Rebleeders accounted for only 11% of the total patients deceased (OR 3.27, 95% CI 1.5-11.2). These results indicate that 30-day mortality for nonvariceal bleeding is low. Deaths occurred predominantly in elderly patients with severe comorbidities or those with failure of endoscopic intention to treatment.


Mammalian muscle fiber types: comparison of excitation-contraction coupling mechanisms.
The mechanism of fatigue of single skeletal muscle fibers appears to involve failure or inactivation of excitation-contraction (EC) coupling. The communication between transverse tubule (TT) and sarcoplasmic reticulum (SR) membranes, whereby plasmalemma voltage controls SR Ca2+ release, may involve inositol trisphosphate (IP3) as a chemical signal. IP3 has already been shown to cause SR Ca2+ release in skeletal fast-twitch fibers; the purpose of this paper is to show that IP3 has similar effects on rabbit slow twitch fibers from soleus muscle. IP3 appears to trigger SR Ca2+ release via a mechanism which is independent of the Ca2+ induced one, since it elicits regenerative SR Ca2+ release in the presence of 20 mM procaine. IP3 also acts at a step in EC coupling beyond TT depolarization, making it a likely candidate for a TT-SR chemical signal. The possible relationship of an IP3 chemical signaling mechanism to skeletal muscle fiber fatigue and mammalian fiber type differences are discussed.


Revision anterior cruciate ligament reconstruction using tibial or hamstring tendon allografts.
To report outcomes of revision anterior cruciate ligament (ACL) reconstruction using tibial or hamstring tendon allografts and to compare with another study using non-irradiated fresh-frozen bone-patellar tendon-bone allografts. Records of 12 men and 7 women aged 18 to 53 (mean, 33) years who underwent revision ACL reconstructions using tibial tendon (n=17) or hamstring tendon (n=2) allografts were retrospectively reviewed. At the time of primary ACL reconstruction, hamstring autografts (n=8) and bonepatellar tendon-bone allografts (n=11) were used. The mean time interval between surgeries was 93 (range, 11-225) months. The causes of failure were traumatic injury (n=7) and technical or biological reasons (n=12). The physical activity level was high in 2 patients, medium in 10, and low in 7. For clinical assessment, the Lysholm test, International Knee Documentation Committee (IKDC) scale, and visual analogue scale (VAS) for pain were used. Patient satisfaction was also assessed. Four of the patients had laxity and were dissatisfied or very dissatisfied with the outcome; the failure rate was 21%. The mean IKDC score was 63% (range, 25-100%), and the mean Lysholm score was 74% (range, 30-100%). Comparing our patients with those in another study using bone-patellar-bone allografts, there was no significant difference in terms of the VAS for pain, IKDC score, and Lysholm score. Comparing our patients with and without chondral and/or meniscal lesions, there was significant difference in terms of the Lysholm score only (86±11 vs. 57±28, p=0.043). Comparing patients who had used hamstring tendon autografts at the primary ACL reconstruction with those who had used bonepatellar tendon-bone autografts, there was significant difference in terms of the VAS for pain only (4.4±3.1 vs. 1.6±1.0, p=0.020). Revision ACL reconstruction using tibial or hamstring tendon allografts provided acceptable results, similar to those using the bone-patellar tendon-bone allografts.


Utility and cutoff value of hair nicotine as a biomarker of long-term tobacco smoke exposure, compared to salivary cotinine.
While hair samples are easier to collect and less expensive to store and transport than biological fluids, and hair nicotine characterizes tobacco exposure over a longer time period than blood or urine cotinine, information on its utility, compared with salivary cotinine, is still limited. We conducted a cross-sectional study with 289 participants (107 active smokers, 105 passive smokers with self-reported secondhand smoke (SHS) exposure, and 77 non-smokers with no SHS exposure) in Baltimore (Maryland, USA). A subset of the study participants (n = 52) were followed longitudinally over a two-month interval. Median baseline hair nicotine concentrations for active, passive and non-smokers were 16.2, 0.36, and 0.23 ng/mg, respectively, while those for salivary cotinine were 181.0, 0.27, and 0.27 ng/mL, respectively. Hair nicotine concentrations for 10% of passive or non-smokers were higher than the 25th percentile value for active smokers while all corresponding salivary cotinine concentrations for them were lower than the value for active smokers. This study showed that hair nicotine concentration values could be used to distinguish active or heavy passive adult smokers from non-SHS exposed non-smokers. Our results indicate that hair nicotine is a useful biomarker for the assessment of long-term exposure to tobacco smoke.


Activity-dependent synaptic competition in vitro: heterosynaptic suppression of developing synapses.
The development and stability of synaptic connections in the nervous system are influenced by the pattern of electrical activity and the competitive interaction between the adjacent nerve terminals. To investigate this influence, a culture system of nerve and muscle cells has been developed in which a single embryonic muscle cell is coinnervated by two spinal neurons. The effect of electrical activity on the synaptic efficacy was examined after repetitive electrical stimulation was applied to one or both neurons. Brief tetanic stimulation of one neuron resulted in immediate functional suppression of the synapse made by the unstimulated neuron innervating the same muscle cell. This heterosynaptic suppression was largely absent when the tetanic stimulation was applied concurrently to both neurons. This result demonstrates that activity-dependent synaptic competition can be studied in vitro at a cellular level.


Novel target genes of RUNX2 transcription factor and 1,25-dihydroxyvitamin D3.
The RUNX2 transcription factor is indispensable for skeletal development and controls bone formation by acting as a signaling hub and transcriptional regulator to coordinate target gene expression. A signaling partner of RUNX2 is the nuclear vitamin D receptor (VDR) that becomes active when bound by its ligand 1,25-dihydroxyvitamin D3 (VD3). RUNX2 and VDR unite to cooperatively regulate the expression of numerous genes. In this study, we overexpressed RUNX2 in NIH3T3 fibroblasts concomitantly treated with VD3 and show that RUNX2 alone, or in combination with VD3, failed to promote an osteoblastic phenotype in NIH3T3 cells. However, the expression of numerous osteoblast-related genes was up-regulated by RUNX2 and large-scale gene expression profiling using microarrays identified over 800 transcripts that displayed a twofold of greater change in expression in response to RUNX2 overexpression or VD3 treatment. Functional analysis using gene ontology (GO) revealed GO terms for ossification, cellular motility, biological adhesion, and chromosome organization were enriched in the pool of genes regulated by RUNX2. For the set of genes whose expression was modulated by VD3, the GO terms response to hormone stimulus, chemotaxis, and metalloendopeptidase activity where overrepresented. Our study provides a functional insight into the consequences of RUNX2 overexpression and VD3 treatment in NIH3T3 cells in addition to identifying candidate genes whose expression is controlled by either factor individually or through their functional cooperation.


From Communication Skills to Skillful Communication: A Longitudinal Integrated Curriculum for Critical Care Medicine Fellows.
Communication with patients and families in critical care medicine (CCM) can be complex and challenging. A longitudinal curricular model integrating multiple techniques within classroom and clinical milieus may facilitate skillful communication across diverse settings. In 2014-2015, the authors developed and implemented a curriculum for CCM fellows at the Cumming School of Medicine, University of Calgary, to promote the longitudinal development of skillful communication. A departmental needs assessment informed curriculum development. Five 4-hour classroom sessions were developed: basic communication principles, family meetings about goals and transitions of care, discussing patient safety incidents, addressing conflict, and offering organ donation. Teaching methods-including instructor-led presentations incorporating a consistent framework for approaching challenging conversations, simulation and clinical practice, and feedback from peers, trained facilitators, family members, and clinicians-supported integration of skills into the clinical setting and longitudinal development of skillful communication. Seven fellows participated during the first year of the curriculum. CCM fellows engaged enthusiastically in the program, commented that the framework provided was helpful, and highly valued the opportunity to practice challenging communication scenarios, learn from observing their peers, and receive immediate feedback. More detailed accounts of fellows', patients', and family members' experiences will be obtained to guide curricular development. The curriculum will be expanded to involve other members of the multidisciplinary intensive care unit team, and faculty education initiatives will be offered to enhance the quality of the feedback provided. The impact of the curriculum on initial skill development, retention, and progression will be assessed.


Targeting the epitope spreader Pep19 by naïve human CD45RA+ regulatory T cells dictates a distinct suppressive T cell fate in a novel form of immunotherapy.
Beyond the limited scope of non-specific polyclonal regulatory T cell (Treg)-based immunotherapy, which depends largely on serendipity, the present study explored a target Treg subset appropriate for the delivery of a novel epitope spreader Pep19 antigen as part of a sophisticated form of immunotherapy with defined antigen specificity that induces immune tolerance. Human polyclonal CD4+CD25+CD127lo- Tregs (127-Tregs) and naïve CD4+CD25+CD45RA+ Tregs (45RA-Tregs) were isolated and were stimulated with target peptide 19 (Pep19)-pulsed dendritic cells in a tolerogenic milieu followed by ex vivo expansion. Low-dose interleukin-2 (IL-2) and rapamycin were added to selectively exclude the outgrowth of contaminating effector T cells (Teffs). The following parameters were investigated in the expanded antigen-specific Tregs: the distinct expression of the immunosuppressive Treg marker Foxp3, epigenetic stability (demethylation in the Treg-specific demethylated region), the suppression of Teffs, expression of the homing receptors CD62L/CCR7, and CD95L-mediated apoptosis. The expanded Tregs were adoptively transferred into an NOD/scid/IL-2Rγ-/- mouse model of collagen-induced arthritis. Epitope-spreader Pep19 targeting by 45RA-Tregs led to an outstanding in vitro suppressive T cell fate characterized by robust ex vivo expansion, the salient expression of Foxp3, high epigenetic stability, enhanced T cell suppression, modest expression of CD62L/CCR7, and higher resistance to CD95L-mediated apoptosis. After adoptive transfer, the distinct fate of these T cells demonstrated a potent in vivo immunotherapeutic capability, as indicated by the complete elimination of footpad swelling, prolonged survival, minimal histopathological changes, and preferential localization of CD4+CD25+ Tregs at the articular joints in a mechanistic and orchestrated way. We propose human naïve CD4+CD25+CD45RA+ Tregs and the epitope spreader Pep19 as cellular and molecular targets for a novel antigen-specific Treg-based vaccination against collagen-induced arthritis.