Relative and absolute lung function change in a general population aged 60-102 years.
Data on longitudinal lung function change in the elderly are scarce. Uncertainty remains about whether to use absolute or relative change and how it relates to subject demographics.We studied absolute and relative forced expiratory volume in 1 s (FEV1) and forced vital capacity (FVC) change in a population-based geriatric sample using a repeated measurements model adjusted for age, sex, smoking habits, heart failure, hypertension, diabetes, coronary heart disease, educational level, occupation, alcohol consumption, C-reactive protein (CRP) and body mass index. 3736 participants aged 60-102 years completed between one and five spirometries during 13.5 years of follow-up. Lung volumes, FEV1 quotient (Q) and Global Lung Initiative (GLI)-2012 and National Health and Nutrition Examination Survey (NHANES) III z-scores were presented from 6932 spirometries.Adjusted absolute change per year (95% CI) was -51.7 (-63.7--39.9) mL for FEV1 and -56.2 (-73.6--38.8) mL for FVC. Adjusted relative change per year was -2.97 (-3.53--2.40)% for FEV1 and -2.46 (-3.07--1.85)% for FVC. Risk factors for increased relative FVC and FEV1 decline were female sex, higher age, current smoking habits, elevated CRP (nonsignificant for FEV1, p=0.057) and low educational level. For increased absolute decline the risk factors were male sex and being a current smoker for FEV1 and low education for FVC.Relative but not absolute change correlated significantly with clinically relevant markers of functional status and may be superior to absolute change in risk factor analysis. Cross-sectional reduction in terms of FEV1Q was ∼1 unit per 10 years for both sexes. Proportions of subjects with results below lower limit of normal using NHANES III were close to anticipated, but were two to four times higher than expected using GLI-2012.


Obesity and increased contractile activity influence the protein content of UCP2 in human skeletal muscle.
The newly discovered uncoupling protein-2 (UCP2) has been proposed to play a critical role in thermoregulatory and substrate oxidation processes. Skeletal muscle mRNA expression and, more recently, the protein content of UCP2 were investigated in humans. These studies have shown that the content of this protein varies quite substantially and that several factors could be responsible for its variation in human skeletal muscle. The aim of this review is to determine whether obesity and low-intensity increased contractile activity contribute to variation in muscle UCP2 content. A recent study from our laboratories revealed that, in obesity, UCP2 content in skeletal muscle is over-expressed by about 1.5-fold compared to lean. Body weight loss in obese subjects did not cause any change in skeletal muscle UCP2 content. On the other hand, when increased muscular contractile activity of knee extensor muscles is induced by several weeks of low-frequency electrical stimulation, UCP2 content increased by about 15%. Obesity and increased contractile activity do not appear sufficient, however, to explain the magnitude of the human skeletal muscle variation in UCP2 content. Since intensive efforts are being devoted to this area of research, it is expected that our understanding of the causes contributing to its variation in humans will soon be substantially improved.


Flufenamic acid as an ion channel modulator.
Flufenamic acid has been known since the 1960s to have anti-inflammatory properties attributable to the reduction of prostaglandin synthesis. Thirty years later, flufenamic acid appeared to be an ion channel modulator. Thus, while its use in medicine diminished, its use in ionic channel research expanded. Flufenamic acid commonly not only affects non-selective cation channels and chloride channels, but also modulates potassium, calcium and sodium channels with effective concentrations ranging from 10(-6)M in TRPM4 channel inhibition to 10(-3)M in two-pore outwardly rectifying potassium channel activation. Because flufenamic acid effects develop and reverse rapidly, it is a convenient and widely used tool. However, given the broad spectrum of its targets, experimental results have to be interpreted cautiously. Here we provide an overview of ion channels targeted by flufenamic acid to aid in interpreting its effects at the molecular, cellular, and system levels. If it is used with good practices, flufenamic acid remains a useful tool for ion channel research. Understanding the targets of FFA may help reevaluate its physiological impacts and revive interest in its therapeutic potential.


A multi-level parcellation approach for brain functional connectivity analysis.
Most brain functional connectivity methods in fMRI require a brain parcellation into functionally homogeneous regions. In this work we propose a novel parcellation approach based on a spatial hierarchical clustering, that provides clusters within a multi-level framework. The method has the advantage of producing several brain parcellations rather than a single one from a fixed size-homogeneity criterion. Results obtained on real data demonstrate the relevance of the approach. Finally, a connectivity study shows the benefit of a prior multi-level parcellation of the brain.


Reduced sinoatrial cAMP content plays a role in postnatal heart rate slowing in the rabbit.
1. Decreasing heart rate during development is known to be the result of parasympathetic nervous system maturation that depresses the pacemaker current (If) by acetylcholine (ACh). However, a direct effect of ACh on If has been ruled out and the involvement of other secondary messengers, such as cAMP, was verified in previous studies. Therefore, we hypothesized that reduced basal cAMP production in sinoatrial (SA) nodal cells may contribute to the slowing of heart rate after birth. 2. The electrocardiogram and heart rate variability (HRV) were documented and measured in vivo and in vitro (in isolated perfused Langendorff preparations) for rabbits aged 2, 4, 6, 8 and 12 weeks. Sinoatrial node action potential (AP) recording and perforated patch-clamp analyses were used to investigate the spontaneous depolarization rate and pacemaker If currents. Concentrations of cAMP in SA nodal tissues were determined by radioimmunoassay. Relative expression of adenylate cyclases (ADCY1, 5) and phosphodiesterases (PDE1A, 4A and 8A) were quantified by real-time reverse transcription-polymerase chain reaction. 3. Significantly reduced heart rate, but unchanged HRV, was observed in perfused hearts in the older age groups, accompanied with a slowed phase 4 spontaneous depolarization rate (90.5 +/- 4.7 vs 49.6 +/- 2.6 mV/s for 2 week vs 4 week hearts, respectively; n = 5; P < 0.05), a negative shift of the If threshold potential (-45.5 +/- 3.0 vs -51.1 +/- 6.0 mV for 2 week vs 4 week hearts, respectively; n = 9; P < 0.05) and decreasing basal levels of SA nodal cAMP (0.31 +/- 0.05 vs 0.025 +/- 0.002 micromol/L for 2 week vs 4 week hearts, respectively; n = 6; P < 0.05). Gene expression levels of PDE1A, 4A and 8A were increased in the 12 week group compared with the 2 week group 1.5-, 2- and 1.8-fold, respectively (P < 0.05), with little change in ADCY1 and 5. 4. These data suggest that, in addition to autonomic innervation, slowing of heart rate during postnatal maturation can be attributed to a negative shift of the If activation caused by diminished baseline cAMP content in SA nodal cells.


Renal phosphate wasting in the absence of adenylyl cyclase 6.
Parathyroid hormone (PTH) and fibroblast growth factor 23 (FGF-23) enhance phosphate excretion by the proximal tubule of the kidney by retrieval of the sodium-dependent phosphate transporters (Npt2a and Npt2c) from the apical plasma membrane. PTH activates adenylyl cyclase (AC) through PTH 1 receptors and stimulates the cAMP/PKA signaling pathway. However, the precise role and isoform(s) of AC in phosphate homeostasis are not known. We report here that mice lacking AC6 (AC6(-/-)) have increased plasma PTH and FGF-23 levels compared with wild-type (WT) mice but comparable plasma phosphate concentrations. Acute activation of the calcium-sensing receptor or feeding a zero phosphate diet almost completely suppressed plasma PTH levels in both AC6(-/-) and WT mice, indicating a secondary cause for hyperparathyroidism. Pharmacologic blockade of FGF receptors resulted in a comparable increase in plasma phosphate between genotypes, whereas urinary phosphate remained significantly higher in AC6(-/-) mice. Compared with WT mice, AC6(-/-) mice had reduced renal Npt2a and Npt2c protein abundance, with approximately 80% of Npt2a residing in lysosomes. WT mice responded to exogenous PTH with redistribution of Npt2a from proximal tubule microvilli to intracellular compartments and lysosomes alongside a PTH-induced dose-response relationship for fractional phosphate excretion and urinary cAMP excretion. These responses were absent in AC6(-/-) mice. In conclusion, AC6 in the proximal tubule modulates cAMP formation, Npt2a trafficking, and urinary phosphate excretion, which are highlighted by renal phosphate wasting in AC6(-/-) mice.


Trends in Utilization of Total Hip Arthroplasty for Femoral Neck Fractures in the United States.
The ideal mode of fixation for patients with femoral neck fractures is not well defined in the current literature. This study describes the recent trends in surgical management of femoral neck fractures with an analysis on perioperative outcomes. The National Hospital Discharge Survey was used to identify femoral neck fractures in the United States between 1990 and 2007 (n = 1,155,960) treated with open reduction and internal fixation (ORIF), total hip arthroplasty (THA), or hemiarthroplasty (HA). Trends were examined over the following 3 time periods: 1990 to 1995 (group 1), 1996 to 2001 (group 2), and 2002 to 2007 (group 3). Elixhauser Comorbidity Index and perioperative complications were calculated. Use of HA increased (74.4% to 84.6%), whereas that of THA (7.3% to 4.9%) and ORIF (18.3% to 10.6%) decreased, from group 1 to group 3 in the age group of >80 years. The use of ORIF increased (63.9% to 81.4%), whereas the use of both HA and THA decreased, from group 1 to group 3 in the age group of <50 years. The rate of adverse events increased across all fixation types but was greatest among THA (32.2% to 48.3%). The femoral neck patient population is now older and has more medical comorbidities. We observed a trend toward performing HA in older patients and ORIF in younger patients. Despite superior functional outcomes reported in THA, this study found a decreased utilization of THA in all age groups along with an increase in adverse events and nonroutine discharges for patients with femoral neck fractures treated with THA.


Misperceptions and management of LDL-cholesterol in secondary prevention of patients with familial hypercholesterolemia in cardiology practice: Real-life evidence from the EPHESUS registry.
Familial hypercholesterolemia (FH) is a common inherited disease, leading to premature atherosclerotic cardiovascular disease (ASCVD) due to elevated low-density lipoprotein cholesterol (LDL-C) levels. Achieving LDL-C goals is extremely important for preventing the complications of this fatal disease. We evaluated the management of FH patients with ASCVD in cardiology practice. We analyzed patients with ASCVD from the nationwide EPHESUS registry, which was conducted in 40 cardiology outpatient clinics, and compared those with and without FH. Of the 1482 consecutively enrolled patients with ASCVD, 618 (41.7%) had FH, among which 455 were categorized as 'Possible FH' and 163 as 'Probable or Definite FH'. Proposed LDL-C goals were not attained in more than 90% of the patients with FH. The proportion of those on statin therapy was 77% for possible and 91% for probable or definite FH, whereas 34.2 % and 59.4% were in use of high-intensity statins, respectively. None of the patients were on PCSK-9 inhibitors, and only 2 used ezetimibe. Adverse media coverage was the most common cause of statin discontinuation (32.5% in 'possible FH' and 45.7% in 'probable/definite FH'). The negative impact of media in the decision to stop lipid lowering therapy (LLT) was increasing with education level. In real life most of the FH patients with ASCVD are undertreated in cardiology practice regarding statin dosing and combined LLT. Drug discontinuation rates are notably high and are mostly media-related, and side effects very rarely cause cessation of LLT. Urgent measures are needed to increase the awareness of FH among healthcare providers and patients and to develop improved treatment strategies aimed at preventing the complications of FH.


Induction of in vitro and in vivo NK cell cytotoxicity using high-avidity immunoligands targeting prostate-specific membrane antigen in prostate carcinoma.
Cancer that might develop as host natural killer (NK) cells fail to detect ligands for their activating NK receptors. Immunoligands represent promising immunotherapeutic tools to overcome this deficit. These are fusion proteins containing a single-chain antibody fragment (scFv) to target an available tumor antigen and ULBP2 to activate host NK cells by targeting the activatory receptor NKG2D. Prostate-specific membrane antigen (PSMA) is an integral non-shed type 2 membrane protein that is highly and specifically expressed on prostate epithelial cells and strongly upregulated in prostate cancer. Here, we compare the impact of various anti-PSMA immunoligand formats on the therapeutic efficacy against prostate carcinoma cells by activating NK cells via NKG2D. Shortening of the linker separating the heavy and light chain antibody domain leads to the formation of dimers, trimers, and higher molecular mass oligomers. NK cells are most efficiently activated by multimeric immunoligands, thus showing an altered cytokine release pattern. The high avidity format is also superior in in vitro NK-mediated tumor cell targeting as shown in cytotoxicity assays. Finally, the efficacy of a multimeric immunoligand is shown in a prostate carcinoma mouse xenograft model showing a strong activity against advanced established tumors.


Neurophysiologic studies and cognitive function in congenital hypothyroid children.
Minor neurologic and intellectual impairments have been described in some congenital hypothyroid (CH) children in spite of early detection by neonatal screening. The aim of our study was to assess cognitive functions as well as neurophysiologic parameters in hypothyroid children and to compare children detected by neonatal screening (group A) versus hypothyroid patients clinically diagnosed before the beginning of the screening program (group B). Group A consisted of 15 children (13 girls, mean age at the beginning of treatment 33 d). Group B consisted of 11 patients (7 girls, mean age at the start of treatment 10.1 mo). Twenty age-matched healthy children were studied as a control group for neurophysiologic tests. Neurophysiologic tests (Auditory P 300, long latency somatosensory evoked potentials (LL-SEP) were performed along with IQ evaluation. Abnormalities of neurophysiologic tests were detected in 82% of clinically diagnosed hypothyroid children. Surprisingly, 47% of the children detected by neonatal screening, having normal mental development index, showed at least one abnormal neurophysiologic test. LL-SEP latencies were found significantly increased in both groups of CH patients compared with controls. Our data are suggestive for a prenatal or perinatal CNS damage in some children with congenital hypothyroidism, despite early treatment.


Thyroid hormone is required for the phenotype transitions induced by the pharmacological inhibition of calcineurin in adult soleus muscle of rats.
The present experiment was designed to examine the effects of hypothyroidism and calcineurin inhibition induced by cyclosporin A (CsA) administration on both contractile and metabolic soleus muscle phenotypes, with a novel approach to the signaling pathway controlling mitochondrial biogenesis. Twenty-eight rats were randomly assigned to four groups, normothyroid, hypothyroid, and orally treated with either CsA (25 mg/kg, N-CsA and H-CsA) or vehicle (N-Vh and H-Vh), for 3 wk. Muscle phenotype was estimated by the MHC profile and activities of oxidative and glycolytic enzymes. We measured mRNA levels of the peroxisome proliferator-activated receptor-gamma coactivator-1 alpha (PGC-1 alpha), the major regulator of mitochondrial content. We also studied the expression of the catalytic A-subunit of calcineurin (CnA) both at protein and transcript levels and mRNA levels of modulatory calcineurin inhibitor proteins (MCIP)-1 and -2, which are differentially regulated by calcineurin activity and thyroid hormone, respectively. CsA-administration induced a slow-to-fast MHC transition limited to the type IIA isoform, which is associated with increased oxidative capacities. Hypothyroidism strongly decreased both the expression of fast MHC isoforms and oxidative capacities. Effects of CsA administration on muscle phenotype were blocked in conditions of thyroid hormone deficiency. Changes in the oxidative profile were strongly related to PGC-1 alpha changes and associated with phosphorylation of p38 MAPK. Calcineurin and MCIPs mRNA levels were decreased by both hypothyroidism and CsA without additive effects. Taken together, these results suggest that adult muscle phenotype is primarily under the control of thyroid state. Physiological levels of thyroid hormone are required for the effects of calcineurin inhibition on slow oxidative muscle phenotype.


Hospices and rituals after death: a survey of British hospice chaplains.
This article presents the results of a survey asking British hospice chaplains to describe hospice involvement in post-mortem, funeral and memorial rituals. The findings are followed by discussion and comment from the author, who takes a sociologist's perspective on the issues raised. It was found that many hospices provided rites for the family, both immediately after death and in the year to come, although there were differing views regarding the extent to which other patients should be involved in such rites. The only hospices that became involved with the funeral itself were those who care for children and patients with AIDS. This may be related to the longer time over which a patient can build up relationships with other patients and staff. In some other hospices, funeral and memorial services were perceived by managers as morbid, fitting uncomfortably with the hospice emphasis on life. The article suggests that hospices should choose whether or not this is an area they wish to be involved in.


Family physicians' attitude and practice of infertility management at primary care--Suez Canal University, Egypt.
The very particular natures of infertility problem and infertility care make them different from other medical problems and services in developing countries. Even after the referral to specialists, the family physicians are expected to provide continuous support for these couples. This place the primary care service at the heart of all issues related to infertility. to improve family physicians' attitude and practice about the approach to infertility management within primary care setting. This study was conducted in the between June and December 2010. The study sample comprised 100 family physician trainees in the family medicine department and working in family practice centers or primary care units. They were asked to fill a questionnaire about their personal characteristics, attitude, and practice towards support, investigations, and treatment of infertile couples. Hundred family physicians were included in the study. They were previously received training in infertility management. Favorable attitude scores were detected among (68%) of physicians and primary care was considered a suitable place for infertility management among (77%) of participants. There was statistically significant difference regarding each of age groups, gender and years of experience with the physicians' attitude. There was statistically significant difference regarding gender, perceiving PHC as an appropriate place to manage infertility and attitude towards processes of infertility management with the physicians' practice. Favorable attitude and practice were determined among the study sample. Supporting the structure of primary care and evidence-based training regarding infertility management are required to improve family physicians' attitude and practice towards infertility management.


Alterations of Hepatic Metabolism in Chronic Kidney Disease via D-box-binding Protein Aggravate the Renal Dysfunction.
Chronic kidney disease (CKD) is associated with an increase in serum retinol; however, the underlying mechanisms of this disorder are poorly characterized. Here, we found that the alteration of hepatic metabolism induced the accumulation of serum retinol in 5/6 nephrectomy (5/6Nx) mice. The liver is the major organ responsible for retinol metabolism; accordingly, microarray analysis revealed that the hepatic expression of most CYP genes was changed in 5/6Nx mice. In addition, D-box-binding protein (DBP), which controls the expression of several CYP genes, was significantly decreased in these mice. Cyp3a11 and Cyp26a1, encoding key proteins in retinol metabolism, showed the greatest decrease in expression in 5/6Nx mice, a process mediated by the decreased expression of DBP. Furthermore, an increase of plasma transforming growth factor-β1 (TGF-β1) in 5/6Nx mice led to the decreased expression of the Dbp gene. Consistent with these findings, the alterations of retinol metabolism and renal dysfunction in 5/6Nx mice were ameliorated by administration of an anti-TGF-β1 antibody. We also show that the accumulation of serum retinol induced renal apoptosis in 5/6Nx mice fed a normal diet, whereas renal dysfunction was reduced in mice fed a retinol-free diet. These findings indicate that constitutive Dbp expression plays an important role in mediating hepatic dysfunction under CKD. Thus, the aggravation of renal dysfunction in patients with CKD might be prevented by a recovery of hepatic function, potentially through therapies targeting DBP and retinol.


Update on lupus epidemiology: advancing health disparities research through the study of minority populations.
The current review focuses on recent population-based studies that have examined the burden of lupus, disease outcomes, and gaps in quality of care, with an emphasis in research addressing health disparities. The Centers for Disease Control and Prevention National Lupus Registries underscored higher susceptibility of both systemic lupus erythematosus (SLE) and primary cutaneous lupus among people of color, compared with whites. Not only does SLE disproportionately strike people from racial and ethnic minorities, those individuals are also at increased risk of developing severe manifestations following SLE diagnosis. Mortality is higher and death occurs at a younger age among blacks, compared with whites. Furthermore, ongoing Centers for Disease Control and Prevention-supported population-based lupus cohorts, along with research by other groups, have provided new insight into the role of social determinants on outcomes and opportunities to improve care in diverse lupus populations. While descriptive epidemiological efforts have been critical to providing more accurate estimates of the burden and mortality of lupus across diverse demographic groups, emerging research suggests a significant influence of psychosocial and healthcare system factors on disease outcomes. These current efforts represent important steps toward the development of clinical and public health interventions aimed at eliminating health disparities in lupus populations.


Treatment of bleeding in patients with liver disease.
Patients with cirrhosis frequently have complex alterations in their hemostatic system. Although routine diagnostic tests of hemostasis in cirrhosis (platelet count, prothrombin time, fibrinogen level) are suggestive of a bleeding tendency, it is now widely accepted that these tests do not reflect hemostatic competence in this population. Rather, patients with cirrhosis appear to have a rebalanced hemostatic system with hypercoagulable elements. Therefore, routine correction of hemostasis laboratory values, for example by fresh frozen plasma or platelet concentrates, with the aim to avoid spontaneous or procedure-related bleeding is not indicated as is outlined in recent clinical guidance documents. However, little guidance on how to manage patients with cirrhosis that are actively bleeding is available. Here we present three common bleeding scenarios, variceal bleeding, post-procedural bleeding and bleeding in a critically ill cirrhosis patient, with specific management suggestions. As patients with cirrhosis generally have adequate hemostatic competence and as bleeding complications may be unrelated to hemostatic failure, prohemostatic therapy is not the first line of management in bleeding patients with cirrhosis, even in the presence of markedly abnormal platelet counts and/or prothrombin times. We provide a rationale for the restrictive approach to prohemostatic therapy in bleeding patients with cirrhosis.


DNA plasmid coding for Phlebotomus sergenti salivary protein PsSP9, a member of the SP15 family of proteins, protects against Leishmania tropica.
The vector-borne disease leishmaniasis is transmitted to humans by infected female sand flies, which transmits Leishmania parasites together with saliva during blood feeding. In Iran, cutaneous leishmaniasis (CL) is caused by Leishmania (L.) major and L. tropica, and their main vectors are Phlebotomus (Ph.) papatasi and Ph. sergenti, respectively. Previous studies have demonstrated that mice immunized with the salivary gland homogenate (SGH) of Ph. papatasi or subjected to bites from uninfected sand flies are protected against L. major infection. In this work we tested the immune response in BALB/c mice to 14 different plasmids coding for the most abundant salivary proteins of Ph. sergenti. The plasmid coding for the salivary protein PsSP9 induced a DTH response in the presence of a significant increase of IFN-γ expression in draining lymph nodes (dLN) as compared to control plasmid and no detectable PsSP9 antibody response. Animals immunized with whole Ph. sergenti SGH developed only a saliva-specific antibody response and no DTH response. Mice immunized with whole Ph. sergenti saliva and challenged intradermally with L. tropica plus Ph. sergenti SGH in their ears, exhibited no protective effect. In contrast, PsSP9-immunized mice showed protection against L. tropica infection resulting in a reduction in nodule size, disease burden and parasite burden compared to controls. Two months post infection, protection was associated with a significant increase in the ratio of IFN-γ to IL-5 expression in the dLN compared to controls. This study demonstrates that while immunity to the whole Ph. sergenti saliva does not induce a protective response against cutaneous leishmaniasis in BALB/c mice, PsSP9, a member of the PpSP15 family of Ph. sergenti salivary proteins, provides protection against L. tropica infection. These results suggest that this family of proteins in Ph. sergenti, Ph. duboscqi and Ph. papatasi may have similar immunogenic and protective properties against different Leishmania species. Indeed, this anti-saliva immunity may act as an adjuvant to accelerate the cell-mediated immune response to co-administered Leishmania antigens, or even cause the activation of infected macrophages to remove parasites more efficiently. These findings highlight the idea of applying arthropod saliva components in vaccination approaches for diseases caused by vector-borne pathogens.


Connexin 39.9 protein is necessary for coordinated activation of slow-twitch muscle and normal behavior in zebrafish.
In many tissues and organs, connexin proteins assemble between neighboring cells to form gap junctions. These gap junctions facilitate direct intercellular communication between adjoining cells, allowing for the transmission of both chemical and electrical signals. In rodents, gap junctions are found in differentiating myoblasts and are important for myogenesis. Although gap junctions were once believed to be absent from differentiated skeletal muscle in mammals, recent studies in teleosts revealed that differentiated muscle does express connexins and is electrically coupled, at least at the larval stage. These findings raised questions regarding the functional significance of gap junctions in differentiated muscle. Our analysis of gap junctions in muscle began with the isolation of a zebrafish motor mutant that displayed weak coiling at day 1 of development, a behavior known to be driven by slow-twitch muscle (slow muscle). We identified a missense mutation in the gene encoding Connexin 39.9. In situ hybridization found connexin 39.9 to be expressed by slow muscle. Paired muscle recordings uncovered that wild-type slow muscles are electrically coupled, whereas mutant slow muscles are not. The further examination of cellular activity revealed aberrant, arrhythmic touch-evoked Ca(2+) transients in mutant slow muscle and a reduction in the number of muscle fibers contracting in response to touch in mutants. These results indicate that Connexin 39.9 facilitates the spreading of neuronal inputs, which is irregular during motor development, beyond the muscle cells and that gap junctions play an essential role in the efficient recruitment of slow muscle fibers.


Rotavirus strain types circulating in Africa: Review of studies published during 1997-2006.
Rotavirus is responsible for more than half a million deaths among infants and young children worldwide each year; many of these deaths could be prevented by widespread use of an effective rotavirus vaccine. The diversity of rotavirus strains in many developing countries, where most rotavirus deaths occur, could represent a significant challenge to the efficacy of current vaccines. In anticipation of rotavirus vaccine introduction, we examined studies published over a 10-year period (1997-2006) from countries in Africa that examined the distribution of VP7 (G) and VP4 (P) rotavirus strain types in symptomatic children and in neonates, together with studies that undertook a more detailed characterization of unusual rotavirus strains. Compared with recently published global reviews of rotavirus strain types and a previous review of the African literature published before 1997, the current data indicate a substantially increased diversity of rotavirus strains across the continent. Notable findings included a reduction in the proportion of globally common serotypes; a high proportion of unusual P/G combinations, suggesting viral reassortment; evidence for zoonotic rotavirus transmission; the emergence and spread across Africa of serotype G9; and a high prevalence of the P[6] VP4 genotype. These data imply that rotavirus vaccines will need to confer protection against a wide variety of strain types in Africa and emphasize the importance of continued strain surveillance before and after the introduction of routine rotavirus vaccination.


Single human B cell-derived monoclonal anti-Candida antibodies enhance phagocytosis and protect against disseminated candidiasis.
The high global burden of over one million annual lethal fungal infections reflects a lack of protective vaccines, late diagnosis and inadequate chemotherapy. Here, we have generated a unique set of fully human anti-Candida monoclonal antibodies (mAbs) with diagnostic and therapeutic potential by expressing recombinant antibodies from genes cloned from the B cells of patients suffering from candidiasis. Single class switched memory B cells isolated from donors serum-positive for anti-Candida IgG were differentiated in vitro and screened against recombinant Candida albicans Hyr1 cell wall protein and whole fungal cell wall preparations. Antibody genes from Candida-reactive B cell cultures were cloned and expressed in Expi293F human embryonic kidney cells to generate a panel of human recombinant anti-Candida mAbs that demonstrate morphology-specific, high avidity binding to the cell wall. The species-specific and pan-Candida mAbs generated through this technology display favourable properties for diagnostics, strong opsono-phagocytic activity of macrophages in vitro, and protection in a murine model of disseminated candidiasis.


The birthing experiences of rural Aboriginal women in context: implications for nursing.
It has been established that the birthing experiences and outcomes of rural women are shaped by poverty, isolation, limited economic opportunities, and diminishing maternity services. We lack research into how these dynamics are compounded by intersecting forms of oppression faced by Aboriginal women, to impact on their birthing experiences and outcomes. The findings of this study of rural Aboriginal maternity care in 4 communities in British Columbia show how diminishing local birthing choices and women's struggles to exert power, choice, and control are influenced by centuries of colonization. The research questions focus on rural Aboriginal women's experiences of birthing and maternity care in this neocolonial context and their desire for supportive birthing environments. A community-based participatory and ethnographic design was employed. Individual interviews, focus groups, and participant observation were the primary data sources. Although the women's experiences in each community were shaped by distinct histories and traditions, economics, politics, and geographies, the impacts of colonization and medical paternalism and the struggle for control of women's bodies during birth intersect, placing additional stress on women. The implications for nurses of accounting for the intersecting dynamics that shape Aboriginal women's experiences and birth outcomes are discussed.


Effects of BMP-2 dose and delivery of microvascular fragments on healing of bone defects with concomitant volumetric muscle loss.
Traumatic composite bone-muscle injuries, such as open fractures, often require multiple surgical interventions and still typically result in long-term disability. Clinically, a critical indicator of composite injury severity is vascular integrity; vascular damage alone is sufficient to assign an open fracture to the most severe category. Challenging bone injuries are often treated with bone morphogenetic protein 2 (BMP-2), an osteoinductive growth factor, delivered on collagen sponge. Previous studies in a composite defect model found that a minimally bridging dose in the segmental defect model was unable to overcome concomitant muscle damage, but the effect of BMP dose on composite injuries has not yet been studied. Here, we test the hypotheses that BMP-2-mediated functional regeneration of composite extremity injuries is dose dependent and can be further enhanced via co-delivery of adipose-derived microvascular fragments (MVF), which have been previously shown to increase tissue vascular volume. Although MVF did not improve healing outcomes, we observed a significant BMP-2 dose-dependent increase in regenerated bone volume and biomechanical properties. This is the first known report of an increased BMP-2 dose improving bone healing with concomitant muscle damage. While high dose BMP-2 delivery can induce heterotopic ossification (HO) and increased inflammation, the maximum 10 μg dose used in this study did not result in HO and was associated with a lower circulating inflammatory cytokine profile than the low dose (2.5 μg) group. These data support the potential benefits of an increased, though still moderate, BMP-2 dose for treatment of bone defects with concomitant muscle damage. Published 2019. This article is a U.S. Government work and is in the public domain in the USA. J Orthop Res.


Extremely elevated room-temperature kinetic isotope effects quantify the critical role of barrier width in enzymatic C-H activation.
The enzyme soybean lipoxygenase (SLO) has served as a prototype for hydrogen-tunneling reactions, as a result of its unusual kinetic isotope effects (KIEs) and their temperature dependencies. Using a synergy of kinetic, structural, and theoretical studies, we show how the interplay between donor-acceptor distance and active-site flexibility leads to catalytic behavior previously predicted by quantum tunneling theory. Modification of the size of two hydrophobic residues by site-specific mutagenesis in SLO reduces the reaction rate 10(4)-fold and is accompanied by an enormous and unprecedented room-temperature KIE. Fitting of the kinetic data to a non-adiabatic model implicates an expansion of the active site that cannot be compensated by donor-acceptor distance sampling. A 1.7 Å resolution X-ray structure of the double mutant further indicates an unaltered backbone conformation, almost identical side-chain conformations, and a significantly enlarged active-site cavity. These findings show the compelling property of room-temperature hydrogen tunneling within a biological context and demonstrate the very high sensitivity of such tunneling to barrier width.


WNK1 controls endosomal trafficking through TRIM27-dependent regulation of actin assembly.
The protein kinase WNK1 (with-no-lysine 1) influences trafficking of ion and small-molecule transporters and other membrane proteins as well as actin polymerization state. We investigated the possibility that actions of WNK1 on both processes are related. Strikingly, we identified the E3 ligase tripartite motif-containing 27 (TRIM27) as a binding partner for WNK1. TRIM27 is involved in fine tuning the WASH (Wiskott-Aldrich syndrome protein and SCAR homologue) regulatory complex which regulates endosomal actin polymerization. Knockdown of WNK1 reduced the formation of the complex between TRIM27 and its deubiquitinating enzyme USP7 (ubiquitin-specific protease 7), resulting in significantly diminished TRIM27 protein. Loss of WNK1 disrupted WASH ubiquitination and endosomal actin polymerization, which are necessary for endosomal trafficking. Sustained receptor tyrosine kinase (RTK) expression has long been recognized as a key oncogenic signal for the development and growth of human malignancies. Depletion of either WNK1 or TRIM27 significantly increased degradation of the epidermal growth factor receptor (EGFR) following ligand stimulation in breast and lung cancer cells. Like the EGFR, the RTK AXL was also affected similarly by WNK1 depletion but not by inhibition of WNK1 kinase activity. This study uncovers a mechanistic connection between WNK1 and the TRIM27-USP7 axis and extends our fundamental knowledge about the endocytic pathway regulating cell surface receptors.


Involvement of CitCHX and CitDIC in developmental-related and postharvest-hot-air driven citrate degradation in citrus fruits.
Citrate is the predominant organic acid associated with taste in citrus fruit. Although citrate metabolism has been widely studied in recent years, the potential contributions of transport proteins to citrate content remain unclear. In the present study, high-acid citrus fruit Gaocheng ('GC', Citrus sp.) and low-acid citrus fruit Satsuma mandarin ('SM', Citrus unshiu Marc.) were selected for study, and the degradation of citrate was deduced to be the main cause of the difference in acidity in fully mature fruits. RNA-seq analysis was carried out on 'GC' and 'SM' fruit samples over the same time course, and the results indicated that citrate degradation occurred mainly through the glutamine pathway, catalyzed by CitAco3-CitGS2-CitGDU1, and also two transport-related genes, CitCHX and CitDIC, were shown to be associated with citrate degradation. These results were confirmed by real-time PCR. In postharvest 'GC' fruit, the expressions of these two transport-related genes were induced by 2-fold under hot air treatment, accompanied by a reduction of 7%-9% in total acid degradation. Transient expression of CitCHX and CitDIC in tobacco leaves was performed, and the citrate content was reduced by 62%, 75% and 78% following CitCHX, CitDIC and CitCHX plus CitDIC treatments, respectively, as compared with expression of an empty vector. Overall, these data indicated that two transport proteins, CitCHX and CitDIC, are not only involved in citrate degradation during fruit development, but also involved in postharvest hot air triggered citrate reduction.


Use of human airway smooth muscle in vitro and ex vivo to investigate drugs for the treatment of chronic obstructive respiratory disorders.
Isolated airway smooth muscle has been extensively investigated since 1840 to understand the pharmacology of airway diseases. There has been often poor predictability from murine experiments to drugs evaluated in patients with asthma or chronic obstructive pulmonary disease (COPD). However, the use of isolated human airways represents a sensible strategy to optimise the development of innovative molecules for the treatment of respiratory diseases. This review aims to provide updated evidence on the current uses of isolated human airways in validated in vitro methods to investigate drugs in development for the treatment of chronic obstructive respiratory disorders. This review also provides historical notes on the pioneering pharmacological research on isolated human airway tissues, the key differences between human and animal airways, as well as the pivotal differences between human medium bronchi and small airways. Experiments carried out with isolated human bronchial tissues in vitro and ex vivo replicate many of the main anatomical, pathophysiological, mechanical and immunological characteristics of patients with asthma or COPD. In vitro models of asthma and COPD using isolated human airways can provide information that is directly translatable into humans with obstructive lung diseases. Regardless of the technique used to investigate drugs for the treatment of chronic obstructive respiratory disorders (i.e. isolated organ bath systems, videomicroscopy, wire myography), the most limiting factors to produce high-quality and repeatable data remain closely tied to the manual skills of the researcher conducting experiments and the availability of suitable tissue.


Knowledge and attitude of secondary school students in Jos, Nigeria on sickle cell disease.
Knowledge about sickle cell disease among youths could constitute an important variable that influences their premarital attitude and behaviour. The study is to determine the knowledge and attitude on Sickle Cell Disease among selected secondary school students in Jos metropolis, Nigeria. A cross sectional descriptive study involving 137 Secondary School Students within Jos metropolis selected by a multistage stratified sampling technique, using self administered structured questionnaire. Data were analyzed using SPSS version 17. A total of 137 students were interviewed, Christians 88%, modal age range 15-20 years (72%) and males (51%). Majority (83.2%) of the respondents were aware of SCDs, as an inherited disorder (80.0%), affecting the red blood cells (83.0%) but only half (54%) knew that the disease can only be diagnosed through blood test. Also, only 59% knew their genotype and 11. 1% claimed AS genotype. More than one fourth (25.5%) had wrong belief that SCD is caused by evil spirit while 76% showed wrong attitude involving stigmatization towards individuals with sickle cell disease. Comprehensive knowledge about SCD was found to be low despite good awareness among respondents, but only few knew their haemoglobin genotype. If sickle cell disease control strategies must yield any significant results, there is a need to raise awareness about SCD, especially among students in secondary institutions in Nigeria is recommended.


Cyberlindnera sylvatica sp. nov., a yeast species isolated from forest habitats.
Five yeast strains isolated from forest habitats in Hungary and Germany were characterized phenotypically and by sequencing of the D1/D2 domain of the large subunit rRNA gene and the ITS1-5.8S-ITS2 (ITS) region of the rRNA gene. The strains have identical D1/D2 domain and ITS region sequences. By sequence comparisons, Candida mycetangii and Candida maritima were identified as the closest relatives among the currently recognized yeast species. The DNA sequences of the investigated strains differ by 1.2 % (six substitutions) in the D1/D2 domain and by 3.5 % (12 substitutions and eight indels) in the ITS region from the type strain of C. mycetangii (CBS 8675T) while by 1.2 % (six substitutions and one indel) in the D1/D2 domain and by 7 % (32 substitutions and seven indels) in the ITS region from the type strain of C. maritima (CBS 5107T). Because the intraspecies heterogeneity seems to be very low and the distance to the most closely related species is above the commonly expected level for intraspecies variability Cyberlindnera sylvatica sp. nov. (holotype, CBS 16335T; isotype, NCAIM Y.02233T; MycoBank no., MB 835268) is proposed to accommodate the above-noted five yeast strains. Phenotypically the novel species can be distinguished from C. mycetangii and C. maritima by the formation of ascospores. Cyberlindnera sylvatica forms one or two hat-shaped ascospores per ascus on many different media as well as well-developed pseudohyphae and true hyphae. Additionally, we propose the transfer of three anamorphic members of the Cyberlindnera americana sub-clade to the genus Cyberlindnera as the following new taxonomic combinations Cyberlindnera maritima f.a., comb. nov., Cyberlindnera mycetangii f.a., comb. nov. and Cyberlindnera nakhonratchasimensis f.a., comb. nov.


How translational accuracy influences reading frame maintenance.
Most missense errors have little effect on protein function, since they only exchange one amino acid for another. However, processivity errors, frameshifting or premature termination result in a synthesis of an incomplete peptide. There may be a connection between missense and processivity errors, since processivity errors now appear to result from a second error occurring after recruitment of an errant aminoacyl-tRNA, either spontaneous dissociation causing premature termination or translational frameshifting. This is clearest in programmed translational frameshifting where the mRNA programs errant reading by a near-cognate tRNA; this error promotes a second frameshifting error (a dual-error model of frameshifting). The same mechanism can explain frameshifting by suppressor tRNAs, even those with expanded anticodon loops. The previous model that suppressor tRNAs induce quadruplet translocation now appears incorrect for most, and perhaps for all of them. We suggest that the 'spontaneous' tRNA-induced frameshifting and 'programmed' mRNA-induced frameshifting use the same mechanism, although the frequency of frameshifting is very different. This new model of frameshifting suggests that the tRNA is not acting as the yardstick to measure out the length of the translocation step. Rather, the translocation of 3 nucleotides may be an inherent feature of the ribosome.


Influence of polymerization mode on degree of conversion and micropush-out bond strength of resin core systems using different adhesive systems.
The aim of this study was to evaluate the influence of different polymerization modes on degree of conversion and micropush-out bond strength of dual-polymerized resin core systems to dentin by using two different adhesive systems. Two resin core systems, Rebilda DC and Build-It FR, were used in combination with total-etch (Adper Scotchbond Multi-Purpose) and self-etch (Clearfil SE Bond) adhesive systems. After treatment of dentin surfaces, resin core systems were applied into the cavities and subjected to different polymerization modes as follows: (1) chemical polymerization; (2) dual polymerization with standard mode of LED (Elipar FreeLight 2 LED); or (3) dual polymerization with exponential mode of LED. The cavities (n=10 per group) were tested in a universal testing machine. Degree of monomer conversion (DC%) was determined by Fourier transform infrared spectroscopy (n=5 cavities per group). ANOVA revealed that resin core (p = 0.002), adhesive system (p < 0.001), and polymerization mode (p < 0.001) had significant effects on bond strength values. The degree of conversion for resin cores decreased significantly (p < 0.001) when only chemical polymerization was employed.


Application of Randomly Amplified Polymorphic DNA Fingerprinting for Species Identification of Bacteria Isolated from Bovine Milk.
A polymerase chain reaction-based DNA fingerprinting system for species identification of bacteria in milk was developed using randomly amplified polymorphic DNA. A total of 108 organisms including 24 ATCC reference strains and 84 wild-type isolates belonging to gram-negative, Staphylococcus , Enterococcus , and Streptococcus species were used to develop the system. Organisms included in the study were those that are isolated frequently from milk. Forty primers from two commercially available primer kits were evaluated to determine the "ideal" primer that could be used for several bacterial species. Over 960 DNA fingerprint patterns were analyzed by laser densitometry. Seven of the 40 primers met criteria established for primer selection. However, only primers OPE-4 (5' GTGACATGCC-3') and OPE-20 (5'-AACGGTGACC-3') allowed differentiation between all 19 ATCC bacterial species included in the study. The other five primers were restricted to either gram-negative bacteria (OPA-7, OPA-14), Staphylococcus species (OPA-13, OPA-14, OPA-18), or Streptococcus species (OPA-3). Primers OPE-4 and OPE-20 were further evaluated using 84 wild-type isolates. A bacterial species identification scheme was developed based on characteristic polymorphic DNA fragments obtained with primers OPE-4 and OPE-20. Results of this study suggest that RAPD fingerprinting has the potential for being developed into a rapid and accurate method for species identification of bacteria in milk.


Gene polymorphisms and drug-drug interactions determine the metabolic profile of blonanserin.
This study aimed to evaluate the effects of cytochrome P450 3A4 (CYP3A4) gene polymorphism and drug interaction on the metabolism of blonanserin. Human recombinant CYP3A4 was prepared using the Bac-to-Bac baculovirus expression system. A microsomal enzyme reaction system was established, and drug-drug interactions were evaluated using Sprague-Dawley rats. Ultra-performance liquid chromatography-tandem mass spectrometry was used to detect the concentrations of blonanserin and its metabolite. Compared with wild-type CYP34A, the relative clearance of blonanserin by CYP3A4.29 significantly increased to 251.3%, while it decreased notably with CYP3A4.4, 5, 7, 8, 9, 10, 12, 13, 14, 16, 17, 18, 23, 24, 28, 31, 33, and 34, ranging from 6.09% to 63.34%. Among 153 tested drugs, nimodipine, felodipine, and amlodipine were found to potently inhibit the metabolism of blonanserin. Moreover, the inhibitory potency of nimodipine, felodipine, and amlodipine varied with different CYP3A4 variants. The half-maximal inhibitory concentration and enzymatic kinetics assay demonstrated that the metabolism of blonanserin was noncompetitively inhibited by nimodipine in rat liver microsomes, and was inhibited in a mixed manner by felodipine and amlodipine in both rat liver microsomes and human liver microsomes. When nimodipine and felodipine were co-administered with blonanserin, the AUC(0-t), AUC(0- ∞), and Cmax of blonanserin increased. When amlodipine and blonanserin were combined, the Cmax of blonanserin C increased remarkably. The vast majority of CYP3A4 variants have a low ability to catalyze blonanserin. With combined administration of nimodipine, felodipine, and amlodipine, the elimination of blonanserin was inhibited. This study provides the basis for individualized clinical use of blonanserin. Significance Statement The enzyme kinetics of novel CYP3A4 enzymes for metabolizing blonanserin were investigated. Clearance of blonanserin by CYP3A4.4, 5, 7-10, 12-14, 16-18, 23-24, 28, 31, 33, and 34 decreased notably, but increased with CYP3A4.29. Additionally, we established a drug interaction spectrum for blonanserin, in which nimodipine, felodipine, and amlodipine kinetics exhibited mixed inhibition. Moreover, their inhibitory potencies decreased with CYP3A4.4 and 5 compared to CYP3A4.1. This study provides essential data for personalized clinical use of blonanserin.


Multi-quantum-well nanowire heterostructures for wavelength-controlled lasers.
Rational design and synthesis of nanowires with increasingly complex structures can yield enhanced and/or novel electronic and photonic functions. For example, Ge/Si core/shell nanowires have exhibited substantially higher performance as field-effect transistors and low-temperature quantum devices compared with homogeneous materials, and nano-roughened Si nanowires were recently shown to have an unusually high thermoelectric figure of merit. Here, we report the first multi-quantum-well (MQW) core/shell nanowire heterostructures based on well-defined III-nitride materials that enable lasing over a broad range of wavelengths at room temperature. Transmission electron microscopy studies show that the triangular GaN nanowire cores enable epitaxial and dislocation-free growth of highly uniform (InGaN/GaN)n quantum wells with n=3, 13 and 26 and InGaN well thicknesses of 1-3 nm. Optical excitation of individual MQW nanowire structures yielded lasing with InGaN quantum-well composition-dependent emission from 365 to 494 nm, and threshold dependent on quantum well number, n. Our work demonstrates a new level of complexity in nanowire structures, which potentially can yield free-standing injection nanolasers.


Idiopathic hypereosinophilic syndrome with cutaneous involvement: a comparative review of 32 cases.
Although idiopathic hypereosinophilic syndrome (HES) is uncommon, we studied the clinical characteristics of this disorder in patients with cutaneous involvement. We chronicle the case of a patient with diffuse skin rash due to idiopathic HES from our clinical experience. Furthermore, a systematic literature search of the medical databases PubMed and Google Scholar was conducted. A total of 32 cases fulfilled the inclusion criteria. The data on patients' characteristics, epidemiology, clinical features, diagnosis, treatment and outcome were collected and analysed. This review illustrates that physicians should maintain a high index of clinical suspicion for idiopathic HES in patients presenting with dermatological lesions and hypereosinophilia, without an obvious cause. Randomised clinical trials are warranted to outline a generalised and efficient therapeutic approach in these patients. Additionally, this paper highlights the need for population-based studies to delineate the magnitude and scope of this association.


[Frequency of incidentalomas on computed tomography scans in the hospitalised elderly].
Incidentaloma is the result of the medical problem created by the development of imaging. Without a universal definition, incidentaloma describes any incidental finding revealed by a medical examination performed for another indication, and which will be the origin of a questioning intended to give it a clinical meaning. The frequency of incidentalomas is very variable depending on the imaging technique, the organ affected and the definition used. The aim of this study was to investigate the frequency of incidentalomas on computed tomography (CT) scans in a geriatric hospitalised population. In an observational, retrospective, monocentric study, we analysed, for a full year, all the planned CT scans performed, during a hospitalisation in acute and subacute care service of a Parisian geriatric hospital in search of incidentalomas. 113 patients were included in the study, with a mean age of 87,4 years. The frequency of incidentalomas on CT scans in this population was 53 %, with an average of two incidentalomas per patient. Eight percent of the incidentalomas required further examination, specialist advice or treatment. We found incidentalomas in half of the brain and abdomino-pelvic CT scans and in a quarter of the chest CT scans. Age was not associated with the presence of incidentalomas. Incidentaloma has become an important part of current medical practice. The geriatrician must know how to anticipate it and propose to his patient an adapted management.


Contribution of Diabetes to the Incidence and Prevalence of Comorbid Conditions (Cancer, Periodontal Disease, Fracture, Impaired Cognitive Function, and Depression): A Systematic Review of Epidemiological Studies in Japanese Populations.
Several epidemiological studies have determined the relationship between diabetes and the incidence and/or prevalence of recently identified comorbid conditions (cancer, periodontal disease, fracture, cognitive impairment, and depression). These relationships may vary by country or race/ethnicity. We aimed to systematically review studies in this field conducted with the Japanese population because such a review in the Japanese population has never been undertaken. We conducted systematic literature searches in PubMed and Ichushi-Web databases for studies published until December 2016. Studies comparing the incidence and/or prevalence of the comorbidities among the Japanese population were included. The studies were classified as integrated analyses, cohort studies, case-control studies, or cross-sectional studies. We identified 33 studies (cancer: 17, periodontal disease: 5, fracture: 5, cognitive impairment: 4, and depression: 2). Although several cohort studies and meta-analyses had assessed the development of cancer in diabetes, there was scant epidemiological evidence for the other conditions. Indeed, only one cohort study each had been conducted for periodontal disease, fracture, and cognitive impairment, whereas other evidence was cross-sectional, some of which was induced from baseline characteristic tables of studies designed for other purposes. In Japan, there is insufficient evidence about the relationship between diabetes and the incidence/prevalence of periodontal disease, fracture, cognitive impairment, and depression. By contrast, several cohort studies and integrated analyses have been conducted for the relationship with cancer. Further studies should be undertaken to estimate the contribution of diabetes on the incidence/prevalence of comorbidities that may be specific to the Japanese population.


Effects of stuffer DNA on transgene expression from helper-dependent adenovirus vectors.
We have analyzed transgene (lacZ) expression from a first-generation adenovirus (Ad) vector in comparison to helper-dependent (hd) Ads deleted for various portions of the viral coding sequences and generated by using the Cre/loxP helper-dependent system (R. J. Parks et al., Proc. Natl. Acad. Sci. USA 93:13565-13570, 1996). An hd vector deleted for approximately 70% of the Ad genome (AdRP1001) provided levels and durations of transgene expression similar to those of a control first generation Ad vector containing an identical expression cassette. Deletion of all Ad sequences from the hdAd and replacement with a approximately 22-kb fragment of lambda DNA resulted in a decrease in the level and duration of lacZ expression which could not be reversed by the inclusion of a matrix attachment region. However, substitution of the lambda stuffer in the fully deleted hdAd with sequences from the human hypoxanthine-guanine phosphoribosyltransferase gene resulted in significantly improved transgene expression. In vitro assays for cytotoxic T lymphocytes (CTL) directed against putative peptides encoded by the vector backbone showed that, although CTL were generated against the vector containing the lambda DNA, no such CTL were generated against the vector containing the hypoxanthine-guanine phosphoribosyltransferase (HPRT) sequences. Surprisingly, the rate of loss of the HPRT- and lambda-containing vectors from mouse liver was similar, despite the differences in expression kinetics, indicating that the lambda stuffer-directed CTL were inefficient at eliminating the transduced cells. Thus, the nature of the DNA backbone of hdAds can have important effects on the functioning of the vector. Since most fully deleted vectors require "stuffer" DNA as part of the vector backbone to maintain optimum vector size, these observations must be taken into account in the design of hdAd vectors.


Early evolution of vertebrate skeletal tissues and cellular interactions, and the canalization of skeletal development.
The stratigraphically earliest and the most primitive examples of vertebrate skeletal mineralization belong to lineages that are entirely extinct. Therefore, palaeontology offers a singular opportunity to address the patterns and mechanisms of evolution in the vertebrate mineralized skeleton. We test the two leading hypotheses for the emergence of the four skeletal tissue types (bone, dentine, enamel, cartilage) that define the present state of skeletal tissue diversity in vertebrates. Although primitive vertebrate skeletons demonstrate a broad range of tissues that are difficult to classify, the first hypothesis maintains that the four skeletal tissue types emerged early in vertebrate phylogeny and that the full spectrum of vertebrate skeletal tissue diversity is explained by the traditional classification system. The opposing hypothesis suggests that the early evolution of the mineralized vertebrate skeleton was a time of plasticity and that the four tissue types did not emerge until later. On the basis of a considerable, and expanding, palaeontological dataset, we track the stratigraphic and phylogenetic histories of vertebrate skeletal tissues. With a cladistic perspective, we present findings that differ substantially from long-standing models of tissue evolution. Despite a greater diversity of skeletal tissues early in vertebrate phylogeny, our synthesis finds that bone, dentine, enamel and cartilage do appear to account for the full extent of this variation and do appear to be fundamentally distinct from their first inceptions, although why a higher diversity of tissue structural grades exists within these types early in vertebrate phylogeny is a question that remains to be addressed. Citing recent evidence that presents a correlation between duplication events in secretory calcium-binding phosphoproteins (SCPPs) and the structural complexity of mineralized tissues, we suggest that the high diversity of skeletal tissues early in vertebrate phylogeny may result from a low diversity of SCPPs and a corresponding lack of constraints on the mineralization of these tissues.


The effect of dopamine therapy on ventral and dorsal striatum-mediated cognition in Parkinson's disease: support from functional MRI.
The central aim of our study was to elucidate functions mediated by the ventral and dorsal striatum, respectively, to better understand the cognitive effects of dopamine replacement in Parkinson's disease. We proposed that the ventral striatum underlies general learning of stimulus associations, whereas the dorsal striatum promotes integration of various influences on selecting. In Parkinson's disease, dopamine depletion is substantially less notable in the ventral relative to the dorsal striatum, and therefore greater improvements are expected for dorsal striatum-mediated functions with dopamine replacement. Using a simple selection task, we found that dopamine replacement impaired encoding and facilitation of consistent stimulus-stimulus relations across trials. This finding was in line with our contention that ventral striatum mediates learning stimulus associations, even when explicit feedback or reward is not provided. In contrast, dopamine replacement enhanced interference related to assimilating conflicting influences on selection across trials, consistent with our hypothesis that the dorsal striatum supports deciding in ambiguous contexts. We further confirmed these separable roles for the ventral and dorsal striatum in our selection task with healthy young volunteers using functional magnetic resonance imaging. In summary, we present a within-subject, double dissociation of the effects of dopamine replacement in patients with Parkinson's disease for ventral striatum-mediated facilitation and dorsal striatum-mediated interference, confirmed in a separate functional magnetic resonance imaging experiment. Defining the distinct functions of the ventral and dorsal striatum will have direct clinical implications. Titration of therapy in Parkinson's disease is generally geared towards optimizing dorsal striatum-mediated motor symptoms, possibly at the expense of ventral striatum operations, a consequence that is only beginning to be recognized. Enhanced awareness of these different processes will translate into medication strategies that take into account those symptoms that dopamine replacement might hinder, as well as improve. Here, we show impairments in learning new stimulus associations compared with improvements in integrating varied influences related to selection. Ultimately, this knowledge will lead clinicians to survey a broader range of symptoms in determining optimal therapy based on individual patient priorities.


Effects of Rest Interval Duration in Resistance Training on Measures of Muscular Strength: A Systematic Review.
Rest interval (RI) duration is an important resistance-training variable underlying gain in muscular strength. Recommendations for optimal RI duration for gains in muscular strength are largely inferred from studies examining the acute resistance training effects, and the generalizability of such findings to chronic adaptations is uncertain. The goals of this systematic literature review are: (i) to aggregate findings and interpret the studies that assessed chronic muscular strength adaptations to resistance training interventions involving different RI durations, and (ii) to provide evidence-based recommendations for exercise practitioners and athletes. The review was performed according to the PRISMA guidelines with a literature search encompassing five databases. Methodological quality of the studies was evaluated using a modified version of the Downs and Black checklist. Twenty-three studies comprising a total of 491 participants (413 males and 78 females) were found to meet the inclusion criteria. All studies were classified as being of good to moderate methodological quality; none of the studies were of poor methodological quality. The current literature shows that robust gains in muscular strength can be achieved even with short RIs (< 60 s). However, it seems that longer duration RIs (> 2 min) are required to maximize strength gains in resistance-trained individuals. With regard to untrained individuals, it seems that short to moderate RIs (60-120 s) are sufficient for maximizing muscular strength gains.


Hospital variability in use of anticoagulant strategies during acute myocardial infarction treated with an early invasive strategy.
During a myocardial infarction, no single best approach of systemic anticoagulation is recommended, likely due to a lack of comparative effectiveness studies and trade-offs between treatments. We investigated the patterns of use and site-level variability in anticoagulant strategies (unfractionated heparin [UFH] only, low-molecular-weight heparin [LMWH] only, UFH+LMWH, any bivalirudin) of 63 796 patients with a principal diagnosis of myocardial infarction treated with an early invasive strategy with percutaneous coronary intervention at 257 hospitals. About half (47%) of patients received UFH only, 6% UFH+LMWH, 7% LMWH only, and 40% bivalirudin. Compared with UFH, the median odds ratio was 2.90 for LMWH+UFH, 4.70 for LMWH only, and 3.09 for bivalirudin, indicating that 2 "identical" patients would have a 3- to 4-fold greater likelihood of being treated with anticoagulants other than UFH at one hospital compared with another. We then categorized hospitals as low- or high-users of LMWH and bivalirudin. Using hierarchical, multivariate regression models, we found that low bivalirudin-using hospitals had higher unadjusted bleeding rates, but the risk-adjusted and anticoagulant-adjusted bleeding rates did not differ across the hospital anticoagulation phenotypes. Risk-standardized mortality and risk-standardized length of stay also did not differ across hospital phenotypes. We found substantial site-level variability in the choice of anticoagulants for invasively managed acute myocardial infarction patients, even after accounting for patient factors. No single hospital-use pattern was found to be clinically superior. More studies are needed to determine which patients would derive the greatest benefit from various anticoagulants and to support consistent treatment of patients with the optimal anticoagulant strategy.


Prediction of gross energy and digestible energy in copra meal, palm kernel meal, and cassava root fed to pigs.
Many of the available prediction equations for feed energy value may not be applicable for ingredients such as copra (Cocos nucifera) meal (CM), palm kernel meal (PKM), and cassava (Manihot esculenta) root (CR). Therefore, we developed novel equations for estimating GE and DE concentrations in CM, PKM, CR, and diets containing these ingredients. Data for GE and DE concentrations were obtained from previous experiments in which the chemical composition in the ingredients and diets were determined. In addition, in vitro DM digestibility (IVDMD) values in 3 samples of ingredients including CM, PKM, and CR and 4 samples of diets including a corn (Zea mays)-soybean (Glycine max) meal-based diet and 3 diets containing CM, PKM, and CR were determined. Based on the model R(2), conceptual predictive criterion, and the P-value of the equation, the best equation for GE concentration (kcal/kg) was GE = 3313 + (24.81 × CP) + (9.83 × NDF) with R(2) = 0.93, root mean square error = 102, and P = 0.005 (CP and NDF values are percentages). Regression analysis was conducted between the DE:GE ratio and IVDMD (%). The DE:GE ratio was 0.81, 0.73, 0.83, 0.89, 0.84, 0.82, and 0.88 in CM, PKM, CR, a corn-soybean meal-based diet, and diets containing CM, PKM, or CR, respectively. The values for IVDMD were 70.3, 42.6, 88.2, 93.4, 86.7, 75.5, and 91.3%, respectively. The DE:GE ratio may be calculated by (0.0030 × IVDMD) + 0.5986 (R(2) = 0.91; P = 0.001). Using the estimated GE concentration and IVDMD, the prediction equation for DE concentration (kcal/kg) was DE = -1965 + (1.02 × GE) + (15.33 × IVDMD) with R(2) = 0.88 and P = 0.007. In conclusion, IVDMD values are useful in estimating energy digestibility in CM, PKM, CR, and diets containing these ingredients.


Speciation of new tri- and tetra-glucoconjugated tetrapyrrolic macrocycles (porphyrins and chlorins): an electronic molecular spectroscopy study.
In this work, we study the physicochemical properties of some newly developed glycoconjugated photosensitizers that can be used in photodynamic therapy (PDT) of cancers: meso-tri- and tetra-(meta-O-beta-D-glucosyloxyphenyl)porphyrins and meso-, tri-, and tetra-(meta-O-beta-D-glucosyloxyphenyl)chlorins. Their properties are compared to the non-glycosylated hydroxylated parent compounds meso-tetra-(meta-hydroxyphenyl) porphyrin and meso-tetra-(meta-hydroxyphenyl)chlorin. It was found that at the ground state, all porphyrins present, independent of the substitution, have the same mean ionization constant (pKa = 2.7), corresponding to two indistiguishable steps of protonation of tetrapyrrolic nitrogens. On the other hand, in the case of chlorins, one proton process can be observed and the corresponding nitrogen exhibits a slightly superior basicity (pKa = 3.0) with respect to porphyrins. Hydroxylated compounds present a second transition at high pH corresponding to the ionization of phenol groups (pKa = 10.5). Consequently, all photosensitizers are not charged at physiological pH (approximately 7.4), and so the ionization process does not influence their activity in biological media. Ionization induces very important variations in photosensitizer absorption and emission spectra. For example, absorption in the red region (band V), one of the most important characteristics of a good photosensitizer, is only important for diprotonated porphyrins and neutral chlorins. As far as fluorescence emission is concerned, neutral chlorins are almost six times more fluorescent than the corresponding neutral porphyrins (phi(chlorin)/phi(porphyrin) approximately = 6). It should be emphasized that the spectra modifications induced by pH variations can find interesting applications in the optimization of visible and fluorescence detection in high-performance liquid chromatography (HPLC) as well as in the development of direct, rapid fluorimetric analytical methods.


Cognitive memory control in borderline personality disorder patients.
It has been demonstrated that the mechanism of cognitive memory control in humans is sustained by the hippocampus and prefrontal cortices, which have been found to be structurally and functionally abnormal in borderline personality disorder (BPD). We investigated whether the memory control mechanism is affected in BPD. Nineteen Diagnostic and Statistical Manual of Mental Disorders (DSM)-IV BPD patients and 19 matched healthy controls (HC) performed a specific think/no-think paradigm exploring the capacity of remembering and suppressing pair of words previously learned. After the think-no think phase, the second member of each word pair has to be remembered either when subjects are presented with the cue word showed at the beginning of the test (Same Probe Test; SPT) or when they are presented with an extra-list categorical word (Independent Probe Test; IPT). We evaluated the effect of suppression and of retrieval activity on later retention of words. Both on the SPT and on the IPT, HC showed the expected improvement of memory retrieval on to-be-remembered words, unlike BPD patients. On the SPT, HC, but not BPD patients, correctly recalled significantly more words among remembered words (RW) than among suppressed words (SW). Similarly to HC, subjects with BPD without a history of childhood abuse showed a significantly higher percentage of correctly recalled words among RW than among SW. The mechanism of active retrieval of memories and of improvement through repetition is impaired in BPD, particularly in those who experienced traumatic experiences. This impairment might play an important role, possibly resulting in the emergence of unwanted memories and dissociative symptoms.


Transcription start site profiling of 15 anatomical regions of the Macaca mulatta central nervous system.
Rhesus macaque was the second non-human primate whose genome has been fully sequenced and is one of the most used model organisms to study human biology and disease, thanks to the close evolutionary relationship between the two species. But compared to human, where several previously unknown RNAs have been uncovered, the macaque transcriptome is less studied. Publicly available RNA expression resources for macaque are limited, even for brain, which is highly relevant to study human cognitive abilities. In an effort to complement those resources, FANTOM5 profiled 15 distinct anatomical regions of the aged macaque central nervous system using Cap Analysis of Gene Expression, a high-resolution, annotation-independent technology that allows monitoring of transcription initiation events with high accuracy. We identified 25,869 CAGE peaks, representing bona fide promoters. For each peak we provide detailed annotation, expanding the landscape of 'known' macaque genes, and we show concrete examples on how to use the resulting data. We believe this data represents a useful resource to understand the central nervous system in macaque.


Photolithographic fabrication method of computer-generated holographic interferograms.
We consider the fabrication of high-quality interferogram-type diffractive optical elements with conventional photolithographic techniques and compare the results with those achievable with electron-beam lithography. The fringes associated with the phase transfer function of the binary phase holographic interferogram are approximated with rectangles, which can be realized at submicron accuracy using a pattern generator and step-and-repeat camera. The effects of the rectangle quantization are analyzed both numerically and experimentally with the aid of diffraction patterns produced by simple focusing elements. Both resolution and diffraction efficiency of the best holograms approach their theoretical values.


Clinical Utility of the Electrocardiographic P-Wave Axis in Patients with Chronic Obstructive Pulmonary Disease.
The vertical P-wave axis on electrocardiography (ECG) is a useful criterion for screening patients with chronic obstructive pulmonary disease (COPD). This study aimed to investigate the clinical characteristics of patients with COPD with a vertical P-wave axis as they have not yet been elucidated. Keio University and its affiliated hospitals conducted an observational COPD cohort study over 3 years. We analyzed 201 patients using ECG and chest computed tomography. The severity of airflow limitation was higher in patients with a P-wave axis >75° than in those with a P-wave axis ≤75°. Patients with a P-wave axis >75° exhibited significantly higher total COPD assessment test scores and increased St. George's Respiratory Questionnaire total, activity, and impact scores than those with a P-wave axis ≤75°. The incidence of exacerbations over 1 and 3 years was significantly higher in patients with a P-wave axis >75° than in those with a P-wave axis ≤75°. The optimal cutoff for the P-wave axis for a percentage of the predicted forced expiratory volume in 1 s <50% and future exacerbations over 3 years was 70° (the areas under the curve [AUC]: 0.788; sensitivity: 65.3%; specificity: 78.3%) and 79° (AUC: 0.642; sensitivity: 36.7%; specificity: 92.6%). The ratio of the low attenuation area was also significantly higher in patients with a P-wave axis >75° than in those with a P-wave axis ≤75°. However, the ratio of the airway wall area did not differ between the 2 groups. Patients with COPD with a vertical P-wave axis exhibited severe airflow limitation and emphysema, a worse health status, and more frequent exacerbation than patients without a vertical P-wave. Detection of the vertical P-wave axis by ECG is beneficial for the management of patients with COPD.


Importance of contralateral aldosterone suppression during adrenal vein sampling in the subtype evaluation of primary aldosteronism.
Adrenal vein sampling (AVS) is the standard criterion for the subtype diagnosis in primary aldosteronism (PA). Although lateralized index (LI) ≥4 after cosyntropin stimulation is the commonly recommended cut-off for unilateral aldosterone hypersecretion, many of the referral centres in the world use LI cut-off of <4 without sufficient evidence for its diagnostic accuracy. The aim of the study was to establish the diagnostic significance of contralateral (CL) aldosterone suppression for the subtype diagnosis in patients with LI <4 in AVS. A retrospective multicentre study was conducted in Japan. Of 124 PA patients subjected to unilateral adrenalectomy after successful AVS with cosyntropin administration, 29 patients with LI < 4 were included in the study. The patients were divided into Group A with CL suppression (n = 16) and Group B (n = 13) without CL suppression. Three outcome indices were assessed after 6 months postoperatively: normalization/significant improvement of hypertension, normalization of the aldosterone to renin ratio (ARR) and normalization of hypokalaemia. The normalization/significant improvement of hypertension was 81% in Group A and 54% in Group B (P = 0·2). The normalization of ARR was 100% in Group A and 46% in Group B (P = 0·004). Hypokalaemia was normalized in all patients of both groups. The overall cure rate of PA based on meeting all the three criteria was 81% in Group A and 31% in Group B (P = 0·01). In patients with PA, where the LI is <4 on AVS, CL suppression of aldosterone is an accurate predictor of a unilateral source of aldosterone excess. CL suppression data should be interpreted in conjunction with computed tomographic adrenal imaging findings to guide surgical management.


Glomerular localization of platelet cationic proteins after immune complex-induced platelet activation.
Synthetic polycations bind to glomerular polyanions (GPA) and increase permeability to macromolecules and immune complexes. Platelet factor 4 and other platelet cationic proteins also bind to GPA and may play a role in immune complex deposition. Here we examine the potential of locally released cationic proteins to bind to GPA after immune complex-induced platelet activation within the renal microvasculature. Rabbits were immunized against bovine serum albumin (BSA), and BSA (2 or 4 mg/ml in buffered saline) was infused into the left renal artery to deliver 8 or 16 mg of BSA over 20 minutes. Thirty minutes later, kidneys were removed and tissue processed for light, immunofluorescence, and electron microscopy for the assessment of glomerular alterations and the localization of immune complexes (IgG and BSA), platelet factor 4, and platelet cationic proteins. GPA was measured by quantitative ultrastructural assessment of polyethyleneimine binding sites. Glomerular capillaries contained large intraluminal immune complexes and platelet aggregates. Also, numerous deposits were observed within subendothelial and subepithelial aspects of the glomerular basement membrane (GBM). Immunofluorescence and immunocytochemistry revealed prominent localization of platelet factor 4, platelet cationic proteins, IgG and BSA within peripheral capillary walls of glomeruli concomitant with a reduction in GPA. Glomeruli of controls or contralateral kidneys did not show GBM localization of immune complexes or platelet proteins. Thus, nascent formation of immune complexes in capillaries was associated with platelet activation and deposition of endogenous cationic proteins in the GBM. This mechanism may be involved in neutralization of GPA and mediation of increased permeability, which leads to GBM deposition of immune complexes.


Maternal parenting behaviors and child coping in African American families.
The purpose of the current study was to examine the impact of three parental influences (i.e., socialization of coping, modeling of coping, and the parent-child relationship) on coping strategies of African American children, as well as child gender as a moderator of these associations. Participants were 83 African American children (mean age = 11.2, SD = 1.44) and their maternal caregivers (mean age = 40.45, SD = 9.55). Both children and parents completed measures of coping behaviors, parental socialization of coping, and maternal support. Regression analyses demonstrated that child reports of their mothers' behavior were better predictors of child coping than mothers' self-reports, with child reports of maternal support and socialization of coping predicting child coping. Results also revealed that child gender moderated the association between maternal parenting behavior and child coping. Specifically, maternal parenting behaviors were more important for girls' coping strategies than for boys' coping strategies. Our results add to the literature on the effects of parent-child relationships on children's responses to stress.


Results and follow-up of locally advanced cancer of the exocrine pancreas treated with radiochemotherapy.
In locally advanced carcinoma of the exocrine pancreas combined radiochemotherapy has been established as a standard treatment. Two different treatment schemes have been consecutively used. Between 1/1994 and 12/2001, a total of 110 patients with locally advanced adenocarcinoma of the pancreas were treated with hyperfractionated accelerated radiotherapy to a total dose of 44.8 Gy combined with 5-fluorouracil (5-FU) (600 mg/m2) and folinic acid (FA) (300 mg/m2) injection. Chemotherapy was repeated monthly in non-progressive disease. From 1/2002 to 11/2003, in another 15 consecutive patients, chemotherapy was changed to gemcitabine (Gem) (300 mg/m2) and cisplatinum (Cis) (30 mg/m2), followed by gemcitabine (1000 mg/m2) every 2 weeks in non-progressive patients. Median survival in the 5-FU/FA group was 10.3 months with a 1-year survival of 46.6% and a 2-year survival of 20.1%. Median time to progression was 8.6 months. Treatment was well tolerated with nausea/vomiting grade I/II in 58.2%, grade III/IV in 14.5%, diarrhea grade I/II in 27.3%, leucopenia/thrombopenia grade I/II in 21.8%, grade III/IV in 7.2%, and mucositis grade III/IV in 7.2%. In the Gem/Cis group, median survival was 13.8 months with a 1-year survival of 54.9% and a 2-year survival of 24.4%. The toxicity data also revealed comparable feasibility: nausea/vomiting grade I/II in 46.7%, grade III/IV in 20%, diarrhea grade I/II in 20%, leucopenia/thrombopenia grade I/II in 26. 7%, and grade III/IV in 13.3%. Radiochemotherapy in locally advanced pancreatic cancer is an effective and well-tolerated treatment. The long-term efficacy concerning survival is limited. The integration of predictive factors and new chemotherapeutic agents like gemcitabine in the multimodality treatment may give a more promising perspective. Because of the narrow therapeutic index of gemcitabine-based radiochemotherapy schemes, a feasible combination of radiotherapy treatment volume and gemcitabine dose must be found.


Matrix vesicles from chondrocytes and osteoblasts: Their biogenesis, properties, functions and biomimetic models.
Matrix vesicles (MVs) are released from hypertrophic chondrocytes and from mature osteoblasts, the cells responsible for endochondral and membranous ossification. Under pathological conditions, they can also be released from cells of non-skeletal tissues such as vascular smooth muscle cells. MVs are extracellular vesicles of approximately 100-300nm diameter harboring the biochemical machinery needed to induce mineralization. The review comprehensively delineates our current knowledge of MV biology and highlights open questions aiming to stimulate further research. The review is constructed as a series of questions addressing issues of MVs ranging from their biogenesis and functions, to biomimetic models. It critically evaluates experimental data including their isolation and characterization methods, like lipidomics, proteomics, transmission electron microscopy, atomic force microscopy and proteoliposome models mimicking MVs. MVs have a relatively well-defined function as initiators of mineralization. They bind to collagen and their composition reflects the composition of lipid rafts. We call attention to the as yet unclear mechanisms leading to the biogenesis of MVs, and how minerals form and when they are formed. We discuss the prospects of employing upcoming experimental models to deepen our understanding of MV-mediated mineralization and mineralization disorders such as the use of reconstituted lipid vesicles, proteoliposomes and, native sample preparations and high-resolution technologies. MVs have been extensively investigated owing to their roles in skeletal and ectopic mineralization. MVs serve as a model system for lipid raft structures, and for the mechanisms of genesis and release of extracellular vesicles.


Retinoic acid receptor gamma is required for proliferation of pancreatic cancer cells.
Despite the expectation that retinoic acid receptor could be the potential therapeutic targets for pancreatic cancers, there has been the lack of information about the role and the impact of retinoic acid receptor gamma (RARγ, RARG) on pancreatic cancer, unlike other two RARs. Herein, we applied TCGA and GEO database to show that the expression and prognosis of RARG is closely related to pancreatic cancer, which demonstrates that RARG is commonly overexpressed in human pancreatic cancer and is an independent diagnostic marker predicting the poor prognosis of pancreatic cancer patients. In addition, we demonstrated that the reduction in the expression of RARG in human pancreatic cancer cells dramatically suppress their proliferation and tumor growth in vivo, partially attributable to the downregulation of tumor-supporting biological processes such as cell proliferation, antiapoptosis and metabolism and the decreased expression of various oncogenes like MYC and STAT3. Mechanistically, RARG binds on the promoters of MYC, STAT3, and SLC2A1 which is distinguished from well-known conventional Retinotic acid response elements (RAREs) and that the binding is likely to be responsible for the epigenetic activation in the level of chromatin, assessed by the measurement of deposition of the gene activation marker histone H3 K27 acetylation (H3K27ac) using ChIP-qPCR. In this study, we reveal that RARG plays important role in the tumorigenesis of pancreatic cancer and represents new therapeutic targets for human pancreatic cancer.


AR1 is an integral part of the adenovirus type 2 E1A-CR3 transactivation domain.
We have previously shown that the nonconserved carboxy-terminal exon of the adenovirus type 2 E1A-289R protein contains two interchangeable sequence elements, auxiliary region (AR) 1 and AR2, that are required for efficient CR3-mediated transcriptional activation of the viral E4 promoter (M. Bondesson, C. Svensson, S. Linder, and G. Akusjärvi, EMBO J. 11:3347-3354, 1992). Here we show that CR3-mediated transactivation of all adenovirus early promoters and the HSP70 promoter requires the AR1 element. We further show that AR2 can substitute for AR1 only when artificially juxtaposed to CR3. AR1 consists of six tandem glutamic acid-proline (EP) repeats and is positioned immediately downstream of CR3. Genetic dissection of AR1 showed that the number of EP repeats in AR1 is critical for CR3 function. Thus, reducing or increasing the number of EP repeats reduces the CR3 transactivation capacity. Furthermore, the introduction of amino acid substitutions into AR1 suggested that the net negative charge in AR1 is of critical importance for its function as an enhancer of CR3-mediated transcriptional activation. Using an in vitro binding approach, we showed that the AR1 element is not part of the CR3 promoter localization signal mediating contact with the Sp1, ATF-2, or c-Jun upstream-binding transcription factors. Previous studies have suggested that the 49-amino-acid sequence constituting CR3 represents the minimal domain required for E1A-induced activation of viral early promoters. Since AR1 was required for efficient CR3-mediated transcriptional activation of all tested promoters, we suggest that the carboxy-terminal boundary for the CR3 transactivation domain should be extended to include the AR1 element.


Recombinatorial biases and convergent recombination determine interindividual TCRβ sharing in murine thymocytes.
Overlap of TCR repertoires among individuals provides the molecular basis for public T cell responses. By deep-sequencing the TCRβ repertoires of CD4+CD8+ thymocytes from three individual mice, we observed that a substantial degree of TCRβ overlap, comprising ∼10-15% of all unique amino acid sequences and ∼5-10% of all unique nucleotide sequences across any two individuals, is already present at this early stage of T cell development. The majority of TCRβ sharing between individual thymocyte repertoires could be attributed to the process of convergent recombination, with additional contributions likely arising from recombinatorial biases; the role of selection during intrathymic development was negligible. These results indicate that the process of TCR gene recombination is the major determinant of clonotype sharing between individuals.


Cerebral arterial blood flow and aneurysm surgery. Part 2: Induced hypotension and autoregulatory capacity.
A study of 21 patients was conducted to clarify the autoregulatory capacity in patients subjected to induced hypotension during intracranial surgery for saccular aneurysms. Trimethaphan camsylate (Arfonad) was used for induced hypotension and arterial blood flow was measured with an electromagnetic flow probe on the internal carotid artery or one of its main intracranial branches. In Grade I and II patients the control arterial blood pressure (ABP) ranged from a mean of 90 to 135 mm Hg (average 110 mm Hg), with a lower level of autoregulation (LLAR) from 35 to 85 mm Hg (average 62 mm Hg). Grade III patients had a control ABP of between 105 and 145 mm Hg (average 124 mm Hg) and the LLAR was found to be between 60 and 95 mm Hg (average 76 mm Hg). There was a significant difference between the two groups with regard to both the control ABP and the LLAR. A surprising result obtained from these data was that the average lower autoregulatory range (the difference between control ABP and LLAR) is practically the same in the two groups. A systematic investigation of the upper limit of autoregulation was not possible for ethical reasons. In those few patients in whom spontaneous increase in the ABP made such observations possible, upper limits up to 150 mm Hg with a total autoregulatory capacity of about 75 mm Hg were observed. In some patients, however, lower limits and cing that the upper limit of autoregulation is markedly influenced by several factors.


Long-term cell-mediated protein release from calcium phosphate ceramics.
Efficient delivery of growth factors from carrier biomaterials depends critically on the release kinetics of the proteins that constitute the carrier. Immobilizing growth factors to calcium phosphate ceramics has been attempted by direct adsorption and usually resulted in a rapid and passive release of the superficially adherent proteins. The insufficient retention of growth factors limited their bioavailability and their efficacy in the treatment of bone regeneration. In this study, a coprecipitation technique of proteins and calcium phosphate was employed to modify the delivery of proteins from biphasic calcium phosphate (BCP) ceramics. To this end, tritium-labeled bovine serum albumin ([(3)H]BSA) was utilized as a model protein to analyze the coprecipitation efficacy and the release kinetics of the protein from the carrier material. Conventional adsorption of [(3)H]BSA resulted in a rapid and passive release of the protein from BCP ceramics, whereas the coprecipitation technique effectively prevented the burst release of [(3)H]BSA. Further analysis of the in vitro kinetics demonstrated a sustained, cell-mediated release of coprecipitated [(3)H]BSA from BCP ceramics induced by resorbing osteoclasts. The coprecipitation technique described herein, achieved a physiologic-like protein release, by incorporating [(3)H]BSA into its respective carriers, rendering it a promising tool in growth factor delivery for bone healing.


Real-time 4D phase unwrapping applied to magnetic resonance elastography.
Phase amplitude is a source of signal in magnetic resonance elastography (MRE) experiments but its exploitation in experimental design has been limited due to the challenges of phase wrap. This study addressed this aspect of MRE through new developments in algorithms, heuristic strategy, and user interface. A test dataset with systematic variation of three parameters-nested wrap, gradient, and noise level-was developed to choose phase-unwrapping algorithms and to analyze their performance. A new application, PhaseTools, was developed that implemented three phase-unwrapping algorithms that adhere to a "real-time" criterion of less than 3 min for a four-dimensional MRE acquisition. Two of the algorithms extend previously published algorithms and one was newly developed. The algorithms were then applied to five datasets from MRE, two typical cases and three edge cases that were particularly challenging in one of the three parameters. The performance of the PhaseTools algorithms on the test dataset was comparable to two widely cited algorithms that take hours or days to complete. Guidelines for the optimal use of each algorithm are established. PhaseTools enables the substantial increase of signal-to-noise in MRE experiments at negligible additional computational cost. PhaseTools is freely released with this study, making robust real-time phase unwrapping available to any group using phase-based imaging.


Speciation in Passerina buntings: introgression patterns of sex-linked loci identify a candidate gene region for reproductive isolation.
Sex-chromosomes are thought to play an important role in speciation, but few studies of non-model organisms have investigated the relative influence of multiple sex-linked markers on reproductive isolation. We collected 222 individuals along a geographical transect spanning the hybrid zone between Passerina amoena and P. cyanea (Aves: Cardinalidae). Using maximum-likelihood cline fitting methods, we estimated locus-specific introgression rates for 10 z-linked markers. Although the cline width estimates ranged from 2.8 to 584 km, eight of 10 loci had cline widths between 224 and 271 km. We also used coalescent-based estimates of locus-specific divergence times between P. amoena and P. cyanea to test a recently proposed hypothesis of an inverse relationship between divergence time and cline width but did not find a significant association. The narrow width (2.8 km) of the cline estimated from the VLDLR9 locus indicates strong selection retarding introgression of alleles at this locus across the hybrid zone. Interestingly, a mutation in the very low density lipoprotein receptor (VLDLR) gene, in which VLDLR9 is an intron, is known to reduce the egg-laying ability of some chickens, suggesting a possible link between this gene region and reproductive isolation between P. amoena and P. cyanea. These results underscore the importance of sampling multiple loci to investigate introgression patterns across a chromosome or genome and support previous findings of the importance of sex-linked genes in speciation.