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Technological growth in production and efficient utilization of input factors are the two biggest contributors to total factor productivity (TFP). TFP of the four major forest industries (logging, pulp and paper, sawmill, and veneer and plywood industries) of Ontario are compared by analyzing their production structures using duality theory in production and costs. The study uses annual data of output and four inputs — labour, capital, energy and materials — from 1967 to 2003. Different restrictions on the translog cost function are applied to each industry to determine the cost function that best describes each industry’s technology, which is further used to estimate Morishima elasticities of substitution, own-price and cross-price elasticities, rate of technological change, and TFP. The production structure of sawmill and veneer and plywood industries is found to be linear homogeneous and homothetic, and that of logging and pulp and paper industries is non-homothetic. Further, Hicks neutral technological change for all four industries is rejected, indicating that the production structure in all four industries is biased in favour of certain inputs and against others. This suggests that policies that improve the efficiency of each industry should focus on input-saving factors of that industry, thereby improving its competitive position.

Air polluted with methanol vapours was treated in a biofilter and a biotrickling filter, both packed with inert materials. The effects of the nitrogen concentration present in the nutrient solution, the empty bed residence time, and the methanol inlet load, on the biofilter and biotrickling filter performance were all examined and compared. The elimination capacity, the biomass and the carbon dioxide production rates all increased with the increase of the parameters tested. The maximum elimination capacity for the biotrickling filter was 240 g·m–3·h–1 with corresponding removal efficiency of 75% and carbon dioxide production rate of 10 g·m–3·h–1, whereas the maximum elimination capacity for the biofilter was 80 g·m–3·h–1 with corresponding removal efficiency of 35% and carbon dioxide production rate of 70 g·m–3·h–1. The biomass production rate was similar for both the biofilter and the biotrickling filter. The carbon dioxide production rate was higher by a factor of 2 to 9 for the biofilter compared to the biotrickling filter.

This position paper is the result of discussions between the Associate Editors and Editors of the Canadian Journal of Physiology and Pharmacology, which took place over the summer of 2013 in response to the need to have some guidelines regarding publishing research findings that involved compounds or extracts of natural products that have not yet been fully characterized. Some journals have adopted the policy that they will not publish such studies. In the instruction for authors for the journal Basic & Clinical Pharmacology & Toxicology it states as follows: “For all compounds used in the studies, the chemical constitution and composition should be known, also for natural compounds” (http://onlinelibrary.wiley.com/journal/10.1111/%28ISSN%291742–7843/ homepage/ForAuthors.html). However, I will argue that to do so is not only a form of censorship, but it also ignores the need for disclosure of information that may be critical to the evaluation of the neutral, beneficial, or detrimental properties of such compounds. Failure to expose these studies to full scientific scrutiny abandons the primary tenant of science: to dispassionately examine all of the facts. Further, failure to do somay put individuals at undue risk, and thus is ethically questionable. Although it is much easier to evaluate the potential when there is a “defined chemical”, and certainly easier to find reviewers, extracts of native species may not have reached that level of purity. Indeed, the American College of Cardiology ACCF Complementary Medicine Experts Consensus publication on complementary andalternativemedicines concluded that “FewU.S. products benefit from rigorous characterization and standardization necessary for clinical study” (Vogel et al. 2005). Nevertheless, 40% to 50% of Americans are using complementary medicines together with prescription drugs (Vogel et al. 2005). Thus, to ignore the potential interactions of nonprescription usage is putting a very high proportion of the population at risk (Zipes 2001; Tachjian et al. 2010). To restrict CJPP publication to only fully chemically characterized compounds, removes from scientific scrutiny some early and potentially important discoveries. For example, there is the classical description in 1785 by William Withering, an English botanist, chemist, geologist, and physician, of the benefits of extracts from the foxglove (Digitalis purpurea) (Goldthorp 2009). Withering's description has been described as the beginning of modern therapeutics. Subsequently, there was the determination of the active compound digitalis, which is still in common clinical use today. Another example is a naturally occurring biological product, the long term search for the elusive endothelium-derived relaxation factor “EDRF”. It took decades to identify this as the gas nitric oxide (NO) (Moncada 2006). There also is the example provided by my colleague Dr. Ed Lui, (pers. comm.) of the purification of rosy periwinkle (Cathanthus roseus) extract. The best knownmedical agents from rosy periwinkle arose from investigation into the plant's use in Jamaican folk medicine. Known as periwinkle tea in Jamaica, the well-known folk-use of rosy periwinkle led researchers to delve into its natural chemical properties in the 1950s. Scientific analysis of rosy periwinkle led to the discovery of 2 previously unknown compounds, vincristine and vinblastine, which have been subsequently developed into potent medicines to save lives from leukemia (Bucur et al. 2013) and Hodgkin's lymphoma (Skoetz et al. 2013), respectively. One final example is the recent publication by Cheng (Yale University, New Haven, Connecticut, USA), of a nonpurified product based on a traditional Chinese formula, and its ability to reduce chemotherapy-induced gastrointestinal toxicity (Lam et al. 2010). Of course not all studies show beneficial responses: toxicity is equally important to document. A neutral response is also important as the sale of “natural products” in North America has been stated as reaching 30 billion dollars in 2001 (Vogel et al. 2005). CJPP is not considering an open door policy for undisciplined science. Rather, it is proposing a set of guidelines to assist in maintaining an ethical and scientifically rigorous approach to publishing articles that use natural products. The approach in this set of guidelines attempts to reduce potential misinformation; however, at the same time, following the guidelines would permit some early exploratory descriptions and knowledge of/alert to potentially beneficial/harmful compounds being assessed. Two keys to the publication of the results include (i) clear and detailed descriptions of the methods used, and (ii), to completely as possible describe the way in which the product was obtained. It would be necessary to have these “keys” in the Instructions to Authors, and it would be important for the journal to screen articles for compliance prior to sending to the Associate Editors for review assignment. In conclusion, exclusion from publication simply because a study uses a natural product is not only a form of censorship, but it removes the important disclosure of information that may be critical to the evaluation of the neutral, beneficial, or detrimental properties of such compounds. It removes these studies from rigorous scientific scrutiny of all the facts. A lack of rigorous scientific scrutiny may put individuals at undue risk and thus, in my opinion, is ethically questionable.

Physical activity has established mental and physical health benefits, but related adverse events have not received attention. The purpose of this paper was to review the documented adverse events occurring from physical activity participation among individuals with psychological or cognitive conditions. Literature was identified through electronic database (e.g., MEDLINE, psychINFO) searching. Studies were eligible if they described a published paper examining the effect of changes on physical activity behaviour, included a diagnosed population with a cognitive or psychological disorder, and reported on the presence or absence of adverse events. Quality of included studies was assessed, and the analyses examined the overall evidence by available subcategories. Forty trials passed the eligibility criteria; these were grouped (not mutually exclusively) by dementia (n = 5), depression (n = 10), anxiety disorders (n = 12), eating disorders (n = 4), psychotic disorders (n = 4), and intellectual disability (n = 15). All studies displayed a possible risk of bias, ranging from moderate to high. The results showed a relatively low prevalence of adverse events. Populations with dementia, psychological disorders, or intellectual disability do not report considerable or consequential adverse events from physical activity independent of associated comorbidities. The one exception to these findings may be Down syndrome populations with atlantoaxial instability; in these cases, additional caution may be required during screening for physical activity. This review, however, highlights the relative paucity of the reported presence or absence of adverse events, and finds that many studies are at high risk of bias toward reporting naturally occurring adverse events.

Dietary patterns have been used to summarize diet consumption and to evaluate how diet is associated with diseases in epidemiological research. However, there are many issues surrounding both the use and interpretation of dietary patterns. These issues include how to collect, summarize, and create dietary patterns, as well as how to interpret the results. Because labels are given arbitrarily to dietary patterns to help characterize the pattern in a meaningful way, it is often not clear from the literature as to the consistency of results among studies. Additionally, the utilization of dietary patterns as a tool for public health messages is a topic that is unresolved. These issues are discussed in this paper.

Physical activity is becoming increasingly acknowledged as an integral component of in the multidisciplinary management of cancer patients. Intensive inquiry in this area is likely to increase further over the next decade; however, cancer-specific, evidence-based risk assessment and recommendations for physical activity are not available. A systematic literature review was performed of all studies conducting an exercise training intervention and (or) any form of objective exercise test among adults diagnosed with cancer. Studies were assessed according to evaluation criteria developed by a panel of experts. A total of 118 studies involving 5529 patients were deemed eligible. Overall, the results suggest that exercise training and maximal and submaximal exercise testing are relatively safe procedures with a total nonlife-threatening adverse event rate of

Physical activity (PA) has potential benefits after stroke or spinal cord injury (SCI), especially in improving efficiency and functional capacity in activities of daily living. Currently, many who could benefit from PA may be routinely excluded from participation because of myths related to functional capacity and the concern for harm. The purpose of this review was to evaluate the literature for reports of adverse events during exercise after stroke or SCI, and to provide recommendations regarding exercise participation in supervised and unsupervised environments. Studies were evaluated for quality, and the summary level and quality of evidence were evaluated using the AGREE rubric, modified to address the main outcome measure of adverse events. Levels of exercise stress were evaluated for aerobic activities, using an established rubric. Included in the current analysis were 32 studies for stroke and 4 for SCI. In aggregate, this yielded a total of 730 experimental participants with stroke and 143 with SCI. It should be noted that almost all studies were not designed to examine naturally occurring adverse events from PA. Significant contraindications to unsupervised exercise include manifestation of autonomic dysreflexia in SCI and cardiovascular comorbidity after stroke. There are clear benefits of exercise training on physiological outcomes in stroke and SCI, but the relation between outcomes and safety remains unclear. However, taken on balance, the risk-to-benefit ratio favors the recommendation of exercise. This recommendation is based on studies in which participants were almost universally screened for participation in supervised environments. Thus, the grading of evidence for finding adverse events to support this conclusion is inadequate.

Physical activity (PA) is one of the most powerful treatment options for persons with prediabetes or diabetes. However, some elevation in risk occurs with increased PA, at least initially, and certain precautions need to be made to lower these risks, particularly if these persons are unaccustomed to exercise. We conducted a standardized search of all adverse events associated with increased PA in persons with prediabetes or diabetes (type 1 or type 2) and provided evidence-based guidelines on PA screening in these apparently high-risk individuals. A systematic literature review was performed of all studies reporting on adverse events in persons with prediabetes or diabetes. Studies included were from all designs (retrospective and prospective including randomized controlled trials) and were assessed according to evaluation criteria adapted by a consensus panel. A total of 47 studies, involving >8000 individuals, were deemed eligible. A number of these studies identified a range of mild to severe acute risks with exercise (musculoskeletal injury, hypoglycemia, foot ulceration, proliferative retinopathy, hypotension, sudden death) but the overall prevalence was low. Based on several randomized controlled trials and prospective studies in which prescribed exercise was performed at a wide range of intensities, it appears that increased PA is a relatively safe procedure with no evidence of a loss of life. Based on our assessment of the available literature, we provide a new PA risk algorithm for persons with prediabetes and diabetes and comment on the role of the patient, the qualified exercise professional, and the patient’s physician in the risk screening process.

Recent feedback from physical activity (PA) participants, fitness professionals, and physicians has indicated that there are limitations to the utility and effectiveness of the existing PAR-Q and PARmed-X screening tools for PA participation. The aim of this study was to have authorities in exercise and chronic disease management to work with an expert panel to increase the effectiveness of clearance for PA participation using an evidence-based consensus approach and the well-established Appraisal of Guidelines for Research and Evaluation (AGREE) Instrument. Systematic reviews were conducted to develop a new PA clearance protocol involving risk stratification and a decision-tree process. Evidence-based support was sought for enabling qualified exercise professionals to have a direct role in the PA participation clearance process. The PAR-Q+ was developed to use formalized probes to clarify problematic responses and to explore issues arising from currently diagnosed chronic disease or condition. The original PARmed-X tool is replaced with an interactive computer program (ePARmed-X+) to clear prospective PA participants for either unrestricted or supervised PA or to direct them to obtain medical clearance. Evidence-based validation was also provided for the direct role of highly qualified university-educated exercise professionals in the PA clearance process. The risks associated with exercise during pregnancy were also evaluated. The systematic review and consensus process, conforming to the AGREE Instrument, has provided a sound evidence base for enhanced effectiveness of the clearance process for PA participation of both asymptomatic populations and persons with chronic diseases or conditions.

Increased physical activity (PA) is associated with improved health and quality of life in the general population. A dose–response effect is evident between increasing levels of PA participation and a lower relative risk for cardiovascular disease and all-cause mortality. However, there is also clear evidence that PA acutely increases the risk of an adverse cardiovascular (CV) event and sudden cardiac death (SCD) significantly above levels expected at rest. Adverse CV events during PA may be triggered acutely by the physiological stress of exercise. This investigation will review the available literature describing the CV risks of exercise testing and PA participation in apparently healthy individuals. A systematic review of the literature was performed using electronic databases, including Medline, CINAHL, SPORT discus, EMBASE, Cochrane DSR, ACP Journal Club, and DARE; additional relevant articles were hand-picked and the final grouping was used for the review using the AGREE process to assess the impact and quality of the selected articles. Six hundred and sixteen relevant articles were reviewed with 51 being identified as describing adverse CV events during exercise and PA. Data suggests the risks of fatal and nonfatal events during maximal exercise testing in apparently healthy individuals rarely occur (approximately

The Physical Activity Readiness Questionnaire (PAR-Q) and the Physical Activity Readiness Medical Evaluation (PARmed-X) are internationally known preparticipation screening tools developed on the basis of expert opinion. The primary purposes of this consensus document were to seek evidence-based support for the PAR-Q and PARmed-X forms, to identify whether further revisions of these instruments are warranted, to determine how people responding positively to questions on the PAR-Q can be safely cleared without medical referral, and to develop exercise clearance procedures appropriate for various clinical conditions across the human lifespan. Seven systematic reviews were conducted, examining physical-activity-related risks and effective risk-stratification procedures for various prevalent chronic conditions. An additional systematic review assessed the risks associated with exercise testing and training of the general population. Two gap areas were identified and evaluated systematically: the role of the qualified exercise professional and the requisite core competencies required by those working with various chronic conditions; and the risks associated with physical activity during pregnancy. The risks associated with being physically inactive are markedly higher than transient risks during and following an acute bout of exercise in both asymptomatic and symptomatic populations across the lifespan. Further refinements of the PAR-Q and the PARmed-X (including online versions of the forms) are required to address the unique limitations imposed by various chronic health conditions, and to allow the inclusion of individuals across their entire lifespan. A probing decision-tree process is proposed to assist in risk stratification and to reduce barriers to physical activity. Qualified exercise professionals will play an essential role in this revised physical activity clearance process.

This systematic review examines critically “best practices” in the training of qualified exercise professionals. Particular attention is given to the core competencies and educational requirements needed for working with clinical populations. Relevant information was obtained by a systematic search of 6 electronic databases, cross-referencing, and through the authors’ knowledge of the area. The level and grade of the available evidence was established. A total of 52 articles relating to best practices and (or) core competencies in clinical exercise physiology met our eligibility criteria. Overall, current literature supports the need for qualified exercise professionals to possess advanced certification and education in the exercise sciences, particularly when dealing with “at-risk” populations. Current literature also substantiates the safety and effectiveness of exercise physiologist supervised stress testing and training in clinical populations.

We systematically reviewed the safety of physical activity (PA) for people with arthritis, osteoporosis, and low back pain. We searched PubMed, MEDLINE, Sport Discus, and the Cochrane Central Register of Controlled Trials (1966 through March 2008) for relevant articles on PA and adverse events. A total of 111 articles met our inclusion criteria. The incidence for adverse events during PA was 3.4%–11% (0.06%–2.4% serious adverse events) and included increased joint pain, fracture, and back pain for those with arthritis, osteoporosis, and low back pain, respectively. Recommendations were based on the Appraisal of Guidelines for Research and Evaluation, which applies Levels of Evidence based on type of study ranging from high-quality randomized controlled trials (Level 1) to anecdotal evidence (Level 4) and Grades from A (strong) to C (weak). Our main recommendations are that (i) arthritic patients with highly progressed forms of disease should avoid heavy load-bearing activities, but should participate in non-weight-bearing activities (Level 2, Grade A); and (ii) patients with osteoporosis should avoid trunk flexion (Level 2, Grade A) and powerful twisting of the trunk (Level 3, Grade C); (iii) patients with acute low back pain can safely do preference-based PA (i.e., PA that does not induce pain), including low back extension and flexion (Level 2, Grade B); (iv) arthritic patients with stable disease without progressive joint damage and patients with stable osteoporosis or low back pain can safely perform a variety of progressive aerobic or resistance-training PAs (Level 2, Grades A and B). Overall, the adverse event incidence from reviewed studies was low. PA can safely be done by most individuals with musculoskeletal conditions.

Recommendations for physical activity during pregnancy have progressed significantly in the last 30 years and continue to evolve as an increasing body of scientific evidence becomes available. In addition, there is an increasing number of women who wish to either maintain physical fitness levels during the prenatal period or initiate exercise for a healthier lifestyle during pregnancy. As such, consistent evaluation of the risks associated with exercise during pregnancy is warranted for maternal and fetal well-being. The primary purpose of this systematic review was to evaluate the scientific information available regarding maternal and fetal responses as it relates to the occurrence of adverse exercise-related events in pregnant women without contraindications to exercise. A systematic and evidence-based approach was used to analyze critically the level of evidence for risks associated with acute and chronic exercise during pregnancy in healthy pregnant women. All articles were screened according to standardized evaluation criteria developed by a panel of experts. A total of 74 investigations that involved 3766 pregnant women were included in the analysis. Of the 74 studies, only 35 studies documented the presence or absence of adverse events. The serious adverse event rate for these studies was 1.4 per 10 000 h of exercise. The adverse event rate increased to 6.8 per 10 000 h of exercise when including the occurrence of more mild events and exercise-related fetal bradycardia and tachycardia. Previously inactive or active women (without contraindications) are at a low risk for adverse fetal or maternal events if they participate in routine physical activity during pregnancy.

The cyanobacterial Na+-dependent HCO3–transporter BicA is a member of the ubiquitous and important SulP/SLC26 family of anion transporters found in eukaryotes and prokaryotes. BicA is an important component of the cyanobacterial CO2concentrating mechanism, an adaptation that contributes to cyanobacteria being able to achieve an estimated 25% of global primary productivity, largely in the oceans. The human SLC26 members are involved in a range of key cellular functions involving a diverse range of anion transport activities including Cl–/HCO3–, I–/HCO3–, and SO42–/HCO3–exchange; mutations in SLC26 members are known to be associated with debilitating diseases such as Pendred syndrome, chondrodysplasias, and congenital chloride diarrhoea. We have recently experimentally determined the membrane topology of BicA using the phoA–lacZ reporter system and here consider some of the extrapolated implications for topology of the human SLC26 family and the Sultr plant sulphate transporters.

Bicarbonate is a waste product of mitochondrial respiration and one of the main buffers in the human body. Thus, bicarbonate transporters play an essential role in maintaining acid-base balance but also during fetal development as they ensure tight regulation of cytosolic and extracellular environments. Bicarbonate transporters belong to two gene families, SLC4A and SLC26A. Proteins from these two families are widely expressed, and thus mutations in their genes result in various diseases that affect bones, pancreas, reproduction, brain, kidneys, eyes, heart, thyroid, red blood cells, and lungs. In this minireview, we discuss the current state of knowledge regarding the effect of SLC4A and SLC26A mutants, with a special emphasis on mutants that have been studied in mammalian cell lines and how they correlate with phenotypes observed in mice models.

The Wilson disease protein (ATP7B) is a copper-transporting ATPase that is responsible for regulating copper homeostasis in human tissues. ATP7B is associated with cancer resistance to cisplatin, one of the most widely used anticancer drugs. This minireview discusses the possible mechanisms of tumor resistance to cisplatin mediated by ATP7B. Cisplatin binds to the N-terminal cytosolic domain of ATP7B, which contains multiple copper-binding sites. Active platinum efflux catalyzed by ATP7B is unlikely to significantly contribute to cisplatin resistance in vivo. Transient platinum sequestration in the metal-binding domain followed by transfer to an acceptor protein or a low molecular weight compound is proposed as an alternative mechanism of cisplatin detoxification in the cell.

Owing to the overlapping and redundant roles of the seven mammalian nucleoside transporters (NTs), which belong to two protein families (ENTs and CNTs), the physiological importance of individual NTs has been difficult to assess. Mice that have NT genes knocked out can be a valuable tool in gaining an understanding of the NT proteins. We have generated a strain of mice that is homozygous for a disruption mutation between exons 2 and 3 of the mouse equilibrative nucleoside transporter, mENT1. We have undertaken a quantitative survey of NT gene expression in 10 tissues, as well as microarray analysis of heart and kidney, from wild-type and mENT1 knockout mice. Rather than a consistent change in expression of NT genes in all tissues of mENT1 knockout mice, a complex pattern of changes was found. Some genes, such as those encoding mCNT1 and mCNT3 in colon tissue, exhibited increased expression, whereas other genes, such as those encoding mCNT2 and mENT4 in lung tissue, exhibited decreased expression. Although mCNT3 has been shown to be important in human and rat kidney tissue, we were unable to detect mCNT3 transcripts in the kidney of either the wild-type or mENT1 knockout mice, suggesting differences in renal nucleoside resorption between species.

The bicarbonate/chloride exchanger 1 (AE1, Band 3) is abundantly expressed in the red blood cell membrane, where it is involved in gas exchange and functions as a major site of cytoskeletal attachment to the erythrocyte membrane. A truncated kidney isoform (kAE1) is highly expressed in type A intercalated cells of the distal tubules, where it is vital for urinary acidification. Recently, kAE1 has emerged as a novel physiologically significant protein in the kidney glomerulus. This minireview will discuss the known interactions of kAE1 in the podocytes and the possible mechanisms whereby this important multispanning membrane protein may contribute to the function of the glomerular filtration barrier and prevent proteinuria.

Immediate-early genes have important roles in processes such as brain development, learning, and responses to drug abuse. Further, immediate-early genes play an essential role in cellular responses that contribute to long-term neuronal plasticity. Neuronal plasticity is a characteristic of the nervous system that is not limited to the first stages of brain development but persists in adulthood and seems to be an inherent feature of everyday brain function. The plasticity refers to the neuron’s capability of showing short- or long-lasting phenotypic changes in response to different stimuli and cellular scenarios. In this review, we focus on the immediate-early genes encoding transcription factors (AP-1 and Egr) that are relevant for neuronal responses. Our current understanding of the mechanisms involved in the induction of the immediate-early genes is presented.

The objective of this study was to examine the effects of leptin on alpha (α) 1 (I) collagen gene expression in a human osteoblast-like MG63 cell line. MG63 cells were incubated with different doses of leptin (10−8, 10−7, and 10−6 mol·L–1) for 24, 48, and 72 h. α1 (I) collagen gene expression in MG63 cells was detected by real-time fluorescence quantitative polymerase chain reaction (FQ-PCR), with 17β-estradiol (17β-E2) as the positive control. Expression of the α1 (I) collagen gene, regulated by leptin, was dose and time dependent, with maximal expression in the 10−7 mol·L–1 group at 72 h of incubation. As a positive control, 17β-E2 reached its maximal effect in the 10−7mol·L–1 group at 24 h. We conclude that leptin has the ability to up-regulate α1 (I) collagen gene expression in MG63 cells, with a more potent effect but a less rapid response than 17β- E2.

There is a consensus that Gag-specific cytotoxic T lymphocyte (CTL) response plays a key role in the immune control of human immunodeficiency virus type 1 (HIV-1) infection. In this study, we analyzed all currently available gag sequences in the Los Alamos HIV sequence database and identified positive selection (PS) sites likely restricted by the host immune responses. We found that between 23.4% and 47.4% of PS sites were shared by clades A, B, and C of Gag, indicating similar positive selection pressure on Gag in different subtypes of HIV-1. Furthermore, a significant correlation was observed between the combined CTL and antibody responses and PS sites. The Gag regions of free from PS contained 9 CTL epitopes restricted by 11 HLA class I alleles associated with disease progression to acquired immune deficiency syndrome (AIDS). These analyses provide information important for the identification of cross-clade epitopes and development of a global HIV-1 vaccine.

Adenosine can exhibit cytotoxic activity in vivo and in vitro, though its mechanisms are still uncertain. In this study, we investigated the adenosine-mediated apoptotic signaling pathway and the role of NF-κB in human hepatocellular carcinoma HepG2 cells. HepG2 cells were treated with different concentrations of adenosine for 12–48 h, and the effect of adenosine on cell proliferation was evaluated by MTT assay. The cytotoxicity of adenosine alone or in combination with an NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC), was also evaluated by MTT assay and the mode of cell death was detected by Hoechst 33342 staining. Cell cycle progress was performed by flow cytometry with PI staining. The protein expressions of Bcl-2, p53, NF-κB subunit p65, and caspase-3 were assayed by Western blot. Caspase-3 activity was measured by spectrophotomteric assay. The results showed that adenosine significantly reduced the viability of HepG2 cells in a dose- and time-dependent manner, with IC50(24 and 48 h) of 2.52 and 1.89 mmol·L–1, respectively. The apoptotic index (percentage of sub-G1phase) of HepG2 cells in adenosine treatment alone for 12 and 24 h or in combination with PDTC were 8.30%, 22.32% and 20.18%, 30.89%, respectively. All of them were higher than that in the control group (0.81%, p < 0.01). The characteristic changes of cell apoptosis (chromatin condensation and sub-G1peak) were observed under fluorescent microscopy and flow cytometry. We also found that the apoptotic process triggered by adenosine was involved in G0–G1cell-cycle arrest, enhanced the activity of caspase-3, upregulated p53 and NF-κB p65 expression, and downregulated Bcl-2 expression. Inhibition of NF-κB by PDTC decreased NF-κB p65 expression, enhanced cell apoptosis ratio, and increased caspase-3 activity. NF-κB may play an anti-apoptosis role in adenosine-induced HepG2 cytotoxicity.

It is known that severe acute respiratory syndrome (SARS), a severe infectious illness, which caused an epidemic in Asia in 2003, has extensive and complex effects on human organ systems. It has been reported that the serum levels of prolactin (PRL), follicle stimulating hormone (FSH), and luteinizing hormone (LH) of SARS patients are significantly higher than those of control groups, while estradiol (E2), pregnancy hormone (P), and thyroid stimulating hormone (TSH) are considerably lower than those of normal controls. This phenomenon suggests that the adenohypophyseal endocrine cells in SARS patients may be damaged. However, up to now there has been no direct histological investigation on the endocrine cells of patients’ pituitary. Here we investigated the endocrine cells in the adenohypophysis obtained from autopsies of 5 SARS patients. The immunohistochemistry and quantitative image results showed that compared with control cases, both the number of positive cells and the staining intensity of immunoreactivity for growth hormone, TSH, and adrenocorticotrophic hormone in these cells were remarkably decreased, while that of PRL, FSH, and LH were significantly increased in all SARS cases studied. These findings indicated that alterations occurred in the patients’ adenohypophyseal endocrine cells, and these changes were consistent with the serum levels of relevant endocrine hormones reported previously. It appears that changes in these endocrine cells and their hormones are affected by the severity of this new infectious disease.

Haloperidol is a commonly used, typical, antipsychotic drug (APD) that acts strongly against positive symptoms, but has fewer therapeutic effects on, or may even aggravate, negative symptoms and cognitive deficits in patients with schizophrenia. Loss of oligodendrocytes has been suggested as a factor associated with the negative symptoms of schizophrenia. Recent study shows that chronic haloperidol treatment induced down-regulation of oligodendrocyte-related genes in certain brain regions of mouse. In this study, we used primary oligodendrocyte progenitor cell cultures from 1- to 3-day-postnatal rats to investigate the direct effects of haloperidol on the proliferation and differentiation of oligodendrocyte progenitor cells. Our results showed that (i) haloperidol (0–10.0 µmol·L–1) facilitated the proliferation of oligodendrocyte progenitor cells, (ii) chronic haloperidol (0.5 µmol·L–1) treatment decreased the number of myelin basic protein positive oligodendrocytes and reduced the oligodendrocytes cells possessing myelin-like membranes, resulting in inhibition of the terminal differentiation of oligodendrocytes, and (iii) D3 receptor mRNA was detected in oligodendrocyte progenitor cells, and haloperidol treatment induced a down-regulation of D3 receptor mRNA. These results suggest that the typical antipsychotic drug haloperidol affects the development of oligodendrocyte progenitor cells, and that D3 receptor down regulation may be involved. Our observations provide new insight into possible cellular mechanisms responsible for the side effects of typical antipsychotic drugs and support the concept that abnormality of oligodendrocytes may be involved in the pathogenesis of schizophrenia.

The pool of ovarian primordial follicles is established during embryonic development or at birth. During the development from primordial to primary, secondary, and antral follicles, only a small portion of follicles can mature and successfully ovulate; the others are destined to degenerate through apoptotic or atretic loss. As aging advances, females ultimately enter the cessation phase of the estrous cycle and are no longer capable of fertilization. The presumption is that if we can slow down the process of folliculogenesis or decrease follicle loss, females may have a larger ovarian follicular reserve and a longer reproductive lifespan. In our study, rats underwent intragastric administration with tea polyphenols, quercetin (meletin), genistein, or resveratrol, once a day for 4 months (from age 12 to 15 months), to test whether they have positive effects on follicular reserve or ovarian functions. The results showed that rats treated with tea polyphenols (27.8 ± 3.2) and quercetin (36.5 ± 4.1) had a comparable number of healthy follicles to those of controls (26.9 ± 3.8), although significantly fewer atretic follicles were observed in the tea polyphenol group (43.4 ± 5.9 vs 79.7 ± 7.5; p < 0.001). Remarkably, both genistein- and resveratrol-treated rats had more healthy follicles (respectively, 42.8 ± 3.9, p < 0.05; and 51.9 ± 6.4, p < 0.001) and fewer atretic follicles (respectively, 58.4 ± 8.0, p < 0.05; and 51.0 ± 6.2, p < 0.01) than controls. These results indicate that genistein and resveratrol can increase the ovarian follicular reserve and prolong the ovarian lifespan in rats, and their positive effects may be not only due to their intervention in the transition from primordial to primary follicle, but also due to the inhibiting effect on follicular atresia.

Chlamydia trachomatis, an obligate intracellular bacterial pathogen, is the major cause of sexually transmitted diseases worldwide. Although a variety of strategies have been taken to promote the development of a protective vaccine, no ideal vaccine has been generated so far. In this study, we transfected dendritic cells (DCs) with recombinant adenovirus carrying C. trachomatis serovar E major outer membrane protein gene (Ad-MOMP), and investigated their ability to induce specific protection against genital tract chlamydial challenge infection. The results showed that when DCs were transfected with Ad-MOMP in vitro, the DCs exhibited increased expression of CD80 and MHC-II molecules as well as enhanced IL-12 secretion and were able to stimulate T-cell proliferation. The level of IFN-γ secreted by stimulated T cells was also up-regulated significantly. When the Ad-MOMP transfected DCs were adoptively transferred intravenously to naive mice, they generated Th1-biased cytokine production and mucosal IgA responses specific for C. trachomatis. More importantly, the mice immunized with Ad-MOMP-DC mounted protection against genital tract challenge infection, shown by lower body mass loss, lower chlamydial loads, and less severe pathological changes. In conclusion, Ad-MOMP transfected DCs are capable of inducing effective protective immune responses against C. trachomatis genital infection.

The aim of this study was to explore the histogenesis and carcinogenesis of pulmonary cancer induced by N-nitrosopiperidine (NPIP) in mice. NPIP is a form of N-nitrosamine found in tobacco smoke, which has been shown to be a genotoxic chemical as well as a mutagenic compound for inducing chromosome aberrations and severe clastogenicity. In this study, 80 BALB/C strain mice were injected with 0.2 mmol/kg NPIP intraperitoneally for 8 weeks, and experiments were conducted for a further 16 weeks. For the control group, 40 mice were injected with an equal volume of 0.9% NaCl. Pulmonary tissues and tumors in the NPIP-treated group were examined by light microscopy and transmission electron microscopy and compared with the control group at 4-week intervals. The mRNA levels of p53 (mutant), bcl-2, c-myc, ras, and subunits of telomerase — telomerase reverse transcriptase (TERT) and an RNA component, TR — were assayed by mPCR or RT–PCR. Twenty-two mice in the experimental group were found to develop pulmonary tumors, but none in the control group. All tumors found in the experimental group originated from alveolar type II epithelial cells. In addition, 6 of the 22 mice also developed tumors of bronchogenic origin. The expression of p53, bcl-2, c-myc, ras, and the subunits of telomerase were found to increase in all pulmonary tissues and tumors formed thereafter upon NPIP treatment. In summary, NPIP-induced mouse lung tumors exhibited morphological changes during carcinogenesis, which may be the consequence of overexpression of some genes associated with the development of carcinoma and changes in subunits of telomerase. This mouse model of lung tumor formation may be a useful tool to delineate the histogenesis and carcinogenesis of human pulmonary cancer.

The sirtuin proteins are nicotinamide adenine dinucleotide dependent deacetylases and adenosine diphosphate (ADP)-ribosyl transferases associated with metabolic balance and lifespan extension. Sirtuin 1 (SIRT1) and sirtuin 4 (SIRT4) have been reported to regulate insulin secretion, but their association with the development of insulin resistance and nonalcoholic fatty liver disease remain undefined. The aim of this study was to determine the expression of SIRT1 and SIRT4 in the liver and pancreas of rats fed with different diets and analyze the association of these proteins with insulin resistance and nonalcoholic fatty liver disease. Male Sprague–Dawley rats were randomly divided into the following 4 diet treatment groups: normal control (NC), calorie restriction (CR), high-fat (HFa), and high-fructose (HFr), and these groups were maintained for 12 weeks. Blood biochemical analysis and histopathology indicated that HFa and HFr groups were insulin resistant and developed nonalcoholic fatty livers. SIRT1 was present in the nucleus and cytoplasm of the pancreatic β-cells, while SIRT4 was located in the cytoplasm. Treatment with the CR diet increased the expression of SIRT1 in both the pancreas and liver, while treatment with the HFa and HFr diets caused a decrease. SIRT4 was upregulated in the liver of rats treated with the HFa diet, but did not change with the CR diet treatment. These data suggest that SIRT1 and SIRT4 were both involved in the development of insulin resistance and nonalcoholic fatty liver disease.

Barth syndrome is a rare X-linked disease in which mild hypocholesterolemia is observed in some patients. We investigated cholesterol biosynthesis in lymphoblasts from a normal and age-matched Barth syndrome patient. Control and Barth syndrome (ΔTAZ1) lymphoblasts were incubated in the presence or absence of serum to induce cholesterol synthesis and hydroxymethylglutaryl-coenzyme A reductase activity and expression, and cholesterol biosynthesis from radioactive precursors was determined. Cholesterol biosynthesis from [2-14C]pyruvate was stimulated 2-fold in control cells, but was unchanged in ΔTAZ1 lymphoblasts, and from [1-14C]acetate was stimulated 77% in control but only 26% in ΔTAZ1 lymphoblasts upon serum removal, indicating a lower ability of ΔTAZ1 cells to upregulate cholesterol biosynthesis. The reason was an inability to increase hydroxymethylglutaryl-coenzyme A reductase activity, which was already near maximum in ΔTAZ1 lymphoblasts, in response to serum removal, compared with control cells. The reduced ability to increase hydroxymethylglutaryl-coenzyme A reductase enzyme activity in ΔTAZ1 lymphoblasts was due to a decrease in hydroxymethylglutaryl-coenzyme A reductase messenger RNA. Although total cholesterol levels are similar under standard culture conditions, ΔTAZ1 lymphoblasts have a diminished capacity to respond to increased demand for cholesterol biosynthesis because of an already elevated level of synthesis under standard culture conditions.

Autologous or allogeneic mesenchymal stem cells (MSCs) have been used as one of the potential cell sources for cellular cardiomyoplasty. The adverse microenvironment in acute myocardial infarction, however, is considered a deleterious factor for MSC transplantation and cell survival. Tumor necrosis factor (TNF)-α is an inflammatory mediator produced during ischemia that may affect the survival of MSCs. In this study, we investigated the enhancement of MSC survival by transfecting cells with the TNF receptor (TNFR) gene, leading to the overproduction of TNFR and the binding of TNF-α. Rats with acute myocardial infarction, induced by the occlusion of the left coronary artery, were transplanted with MSC or MSC-TNFR. After 2 weeks of acute myocardial infarction, cardiac function was assessed. Engrafted MSC survival and localization of TNF-α protein in infarction myocardium were evaluated. The levels of TNF-α and TNFR in the infarction zone were assessed. The results indicate that MSC-TNFR transplantation (1) improved left ventricular function; (2) enhanced engrafted MSC survival in the infarcted myocardium; (3) attenuated the level of TNF-α in serum and cardiac tissue; and (4) increased TNFR protein production in the infarcted myocardium. Our results showed that MSC modified by the TNFR gene improved cell viability and thereby has the potential to improve the efficiency of MSC transplantation therapy in the ischemic heart.

Choline kinase (CK) was discovered in 1953. Progress in understanding the function of CK was slow until its purification in 1984. The subsequent cloning and expression of the cDNA led to the description of the gene structures. Two genes encode choline kinase, Chka and Chkb, and 3 isoforms of the enzyme have been identified — CKα-1, CKα-2, and CKβ — and the active form of CK is a hetero- or homo-dimer. More recently, gene-disrupted mice have been described. Mice that lack CKα die early in embryogenesis. In contrast, mice that lack CKβ survive to adulthood, but develop hindlimb muscular dystrophy and forelimb bone deformity. It has been shown that this hindlimb muscular dystrophy is due to decreased biosynthesis of phosphatidylcholine and increased catabolism of phosphatidylcholine in the hindlimbs, but not the forelimbs, of mice. CK and its product phosphocholine have also been implicated in development of numerous cancers. Thus, a possible treatment for some kinds of cancer may involve drug inhibition of CK or targeting the expression of CK with RNA interference. In the mid 1950s it was clear that CK was important for the biosynthesis of phosphatidylcholine, but no one predicted a role for CK in muscular dystrophy, bone deformities, or cancer.

Neuregulin-1 (NRG-1) is a member of the epidermal growth factor family. Our previous study showed that NRG-1 protected neurons from apoptosis following focal cerebral ischemia by the inhibition of caspase-3 and TNF-α expression. However, the molecular signaling mechanisms for this action of NRG-1 following cerebral ischemia are not fully understood. Presently, activation of the PI3K/Akt pathway has been implicated as a major contributor to neuronal survival after an ischemic insult. In the present study, we investigated whether NRG-1 modulates the activation of Akt and its downstream targets Bad and Bcl-2 expression after transient focal cerebral ischemia by intraluminal blockade of the middle cerebral artery. Western blot was employed to analyze the change of phosphorylated Akt (p-Akt) expression; reverse transcription and polymerization chain reaction (RT-PCR) were used to measure changes of Bcl-2 mRNA. The level of phosphorylation of Bad (p-Bad) was determined using an enzyme-linked immunosorbent assay (ELISA). Our results showed that recombinant human NRG-1(3.0 ng·kg–1) significantly increased the expression of p-Akt protein, Bcl-2 mRNA, and the level of p-Bad, respectively, whereas administration of LY294002, a specific inhibitor of PI3K, significantly decreased the expression of p-Akt, p-Bad, and Bcl-2 induced by NRG-1 after a 60 min ischemic insult, followed by 24 h of reperfusion. These results indicate that NRG-1 may be involved in regulating the expression of Bcl-2 and p-Bad through the PI3K/Akt pathway after transient focal cerebral ischemia.

The cystic fibrosis transmembrane conductance regulator (CFTR) is an important protein that acts as a chloride channel and regulates many physiological functions, including salt transport and fluid flow. Mutations in the gene encoding the CFTR protein cause cystic fibrosis. CFTR is expressed in the epithelial cells of the lungs, pancreas, intestines, and other organs. In the peripheral and central nervous system, CFTR expression has been detected in the neurons of rat brains, ganglion cells of rat hearts, human hypothalamus, human spinal cord, and human spinal and sympathetic ganglia. However, CFTR has not been identified in other parts of the nervous system. In this study, we used immunohistochemistry, in situ hybridization, and laser-assisted microdissection (LMD) followed by reverse transcriptase (RT) PCR to identify CFTR proteins and messenger RNA in human and rat paracervical ganglion cells. CFTR and its gene expression were both detected in paracervical ganglion cells, a finding that might link this important protein to the neuronal regulation of female urogenital function. These findings could provide new insights into the symptoms related to the reproductive system frequently observed in female cystic fibrosis patients.

The 2009 H1N1 and H5N1 influenza viruses are newly (re-) emerged influenza A viruses (2009 A(H1N1) and A(H5N1), respectively) that have recently posed tremendous health threats in many regions worldwide. With the 2009 outbreak of H1N1 influenza A, the world witnessed the first influenza pandemic of the 21st century. The disease has rapidly spread across the entire globe, and has resulted in hundreds of thousands of cases with confirmed infection. Although characterized by high transmissibility, the virulence and fatality of the 2009 A(H1N1) influenza virus have thus far remained relatively low. The reverse holds true for A(H5N1) influenza; at a fatality rate that exceeds 60%, it is known to cause severe damage to the human respiratory system, but is not presently capable of efficient transmission from human to human. Apart from the clear differences between the two types of influenza, there are some significant similarities that warrant attention. In particular, the more severe and fatal 2009 A(H1N1) influenza cases have shown symptoms similar to those reported in cases of A(H5N1) influenza. Histopathological findings for these cases, to the extent available, also appear to have similarities for both diseases in terms of damage and severity. Here we review important recent publications in this area, and we discuss some of the key commonalities and contrasts between the two influenza A types in terms of their biology, origins, clinical features, pathology and pathogenesis, and receptors and transmissibility.

Celecoxib is a selective inhibitor of cyclooxygenase-2 (COX-2). It may reduce the risk of cancer formation by affecting the metabolism of arachidonic acid (AA), which has been implicated in the development of cancer. Accordingly, this study was designed to determine the effects of celecoxib on the AA pathway in mouse hepatoma H22 cells. Celecoxib was found to inhibit the proliferation of H22 cells in a dose- and time-dependent manner. Low doses (50 and 100 µmol·L–1) of celecoxib caused an increase in the expression of cytosolic phospholipase A2 (cPLA2), but did not affect the expression of COX-2 in terms of the mRNA and protein levels. Surprisingly, the amount of AA was elevated and the level of prostaglandin E2 (PGE2) was unaltered in the culture supernatant. At higher celecoxib doses (200 and 400 µmol·L–1), the mRNA and protein of both COX-2 and cPLA2 were inhibited. The concentration of AA was increased, and PGE2 level was depressed in H22 cells. The ratio of AA to PGE2 was increased in a dose-dependent manner. Our findings suggest that the imbalance between AA and PGE2, characterized by increased AA at a low dosage and decreased PGE2 at a high dosage of celecoxib, was an important indicator of cytotoxicity of celecoxib on H22 cells.

Diabetic nephropathy (DN) is one of the most important complications of diabetic patients and is characterized histologically by an accumulation of extracellular matrix (ECM) protein in the glomerular mesangium. Therefore, mesangial cells likely play an important role in the development of diabetic nephropathy. Here, we employed proteomic techniques to investigate the protein profile of rat mesangial cells under high-glucose culture conditions. Primary isolated rat glomerular mesangial cells were cultured under different concentrations of glucose (5.4 mmol·L–1for normal control and 30 mmol·L–1for high glucose) for 0, 8, 16, and 72 h, as well as for 25 days. Cellular total proteins were isolated from these cells and employed for two-dimensional gel electrophoresis (2-DE). Differentially expressed proteins were identified by matrix-assisted laser desorption – ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and some of these proteins were documented in rat models of diabetes by Western blot. Rat mesangial cells were successfully isolated in the laboratory and their proliferation rates were significantly inhibited by high glucose. Two-dimensional gel electrophoresis analyses revealed 28 differentially expressed protein spots between the normal and high-glucose groups. After MALDI-TOF-MS analysis, all 28 protein spots were successfully identified with the peptide mass fingerprint (PMF) method. Representatively, SOD1, PCBP1 and PSMA6 were validated by Western blot analysis following protein extractions from the normal and high-glucose groups. Abundance of these proteins was consistent with that found in 2-DE. Moreover, expression of SOD1, PCBP1, and PSMA6 in renal cortex was further examined in two rat models of diabetes (streptozotocin-induced and spontaneous OLETF diabetic models). Abundance of SOD1 and PCBP1 proteins did not show any significant difference between normal control and diabetic rats. However, abundance of the PSMA6 protein was significantly reduced in the renal cortex of both STZ-induced and spontaneous OLETF diabetic rats. Proteomic analysis identified 28 differentially expressed proteins in primary isolated rat mesangial cells between normal and high glucose treatments. Expression of one identified protein was found to be consistent with expression in the renal cortex of two rat diabetic models. Therefore, identification of protein expression patterns in mesangial cells can be employed to develop a therapeutic target for treatment of diabetic nephropathy.

This study was designed to explore the effects of amiloride, a Na+–H+exchange (NHE) inhibitor, on vessel stenosis by observing the expression of NHE-1 protein in vascular smooth muscle (VSM) after balloon injury and the effects of amiloride on VSM cell proliferation, migration, and excretion of extracellular matrices (ECMs). A total of 32 adult male New Zealand white rabbits were randomly divided into a balloon injury group (BG), an amiloride-treated group (AG), and a sham-operated group (SG). The left iliac artery was injured by inflating a 2.5 mm × 20 mm Foley catheter in BG and AG rabbits; in SG rabbits, the Foley catheter was inserted but not inflated. Amiloride (5 mg·kg–1·d–1) was injected intraperitoneally in AG and the same volume of distilled water was used in BG 3 days before balloon injury and for 28 days after the injury. The left iliac artery was stained by hematoxylin–eosin, α-actin, and Masson's trichrome to observe the vessel cava, neointima, media layer, and ECMs. NHE-1 proteins of the VSM were detected by Western blotting. A narrowing of the arterial cava, neointima formation, and thickened VSM layer were observed 28 days after balloon injury in BG and AG. However, in AG, the vessel cava was not as narrowed as that of BG and the intimal areas were to a lesser extent than in BG. In AG, the α-actin-positive areas and the ECM areas in the neointima were increased compared with SG, but to a lesser extent than in BG. The expression of NHE-1 protein in VSM was increased in BG and AG after balloon injury; however, the levels in AG were significantly less than in BG. In conclusion, VSM cell proliferation, migration, and excretion of ECMs contributed to vessel stenosis in the BG and AG rabbits. The expression of NHE-1 protein in VSM increased after balloon injury. Amiloride, an inhibitor of NHE-1, can limit the development of vessel stenosis through inhibition of VSM cell proliferation, migration, and excretion of ECMs.

Early growth response 1 (Egr-1) over-expression has been demonstrated in myocardial ischemia–reperfusion injury, which is closely associated with endothelial dysfunction. In the present study we investigated the expression of Egr-1 on cultured cardiac microvascular endothelial cells (CMECs) to help define the mechanism of myocardial ischemia–reperfusion injury. A model of cultured CMECs exposed to hypoxia–reoxygenation was developed in which synthesized Egr-1 sense and antisense oligodeoxyribonucleotide were transfected into the cells. The expression of Egr-1 was examined by Western blot analysis. Lactate dehydrogenase, malondialdehyde, superoxide dismutase, tumor necrosis factor α, and intercellular adhesion molecule 1 were measured after hypoxia–reoxygenation to assess cell function and injury. Cell morphology, cell viability, and neutrophil adhesion to the CMECs were measured to assess the degree of injury and inflammation. Only cells transfected with Egr-1 antisense oligodeoxyribonucleotide showed a significant reduction in Egr-1 protein expression following hypoxia–reoxygenation. Consistent with the down-regulation of Egr-1 expression, other forms of cell injury were significantly reduced in this group of cells, as evidenced by less alteration in cell morphology, a decrease in expression of tumor necrosis factor α and intercellular adhesion molecule 1, improved cell survival, and reduced neutrophil adhesion.

Saccharomyces cerevisiae remains an ideal organism for studying the cell biological roles of lipids in vivo, as yeast has phospholipid metabolic pathways similar to mammalian cells, is easy and economical to manipulate, and is genetically tractable. The availability of isogenic strains containing specific genetic inactivation of each non-essential gene allowed for the development of a high-throughput method, called synthetic genetic analysis (SGA), to identify and describe precise pathways or functions associated with specific genes. This review describes the use of SGA to aid in elucidating the function of two lipid-binding proteins that regulate vesicular transport, Sec14 and Kes1. Sec14 was first identified as a phosphatidylcholine (PC) – phosphatidylinositol (PI) transfer protein required for viability, with reduced Sec14 function resulting in diminished vesicular transport out of the trans-Golgi. Although Sec14 is required for cell viability, inactivating the KES1 gene that encodes for a member of the oxysterol binding protein family in cells lacking Sec14 function results in restoration of vesicular transport and cell growth. SGA analysis identified a role for Kes1 and Sec14 in regulating the level and function of Golgi PI-4-phosphate (PI-4-P). SGA also determined that Sec14 not only regulates vesicular transport out of the trans-Golgi, but also transport from endosomes to the trans-Golgi. Comparing SGA screens in databases, coupled with genetic and cell biological analyses, further determined that the PI-4-P pool affected by Kes1 is generated by the PI 4-kinase Pik1. An important biological role for Sec14 and Kes1 revealed by SGA is coordinate regulation of the Pik1-generated Golgi PI-4-P pool that in turn is essential for vesicular transport into and out of the trans-Golgi.

Venlafaxine, a novel antidepressant, inhibits serontonin and norepinephrine reuptake in the presynaptic cleft. Unlike typical selective serontonin reuptake inhibitors (SSRIs), venlafaxine may have modulatory effects on nerve terminals and neuronal plasticity. Our preliminary data found that 5 mg·kg–1·d–1 of venlafaxine treatment prevented decreased synaptophysin (SYP) in the hippocampus, which results from chronic restrained stress in the rat model. The present study investigates whether venlafaxine regulates alterations of synaptophysin and neuronal cell adhesion molecule (NCAM) in a post-stroke depression mouse model. We compared the expression level of SYP and NCAM in the hippocampus of global cerebral ischemic (GCI) mice treated with different doses of venlafaxine using immunohistological and Western blot analysis. Pre-treatment with intraperitoneal injection of venlafaxine (2.5 and 5.0 mg·kg–1·d–1) for 14 days significantly prevented the decrease of SYP in the hilus area of the hippocampus in vehicle-treated GCI mice. NCAM was significantly higher in the hippocampus of vehicle-treated GCI mice, and pretreatment with venlafaxine prevented alterations of NCAM, with the high-dose venlafaxine group comparable with vehicle-sham mice. The results suggest the alteration of neuronal remodeling proteins in the hippocampus may be an underlying mechanism of venlafaxine in treating post-stroke depression.

Cleft lip and cleft palate are common congenital craniofacial birth defects in humans. Phenytoin (PHT) is a risk factor of cleft palate formation; however, the molecular mechanisms by which phenytoin exerts its teratogenic effects resulting in cleft palate remain unknown. The Satb2 gene mutation is associated with cleft palate. Satb2-deficient mice exhibit cleft palate deformity and an up-regulation of Hoxa2 in the fronto-nasal region. In this study, phenytoin was administered intraperitoneally to pregnant C57BL/6 mice on the 10th day of gestation. Real-time PCR results showed that the expressions of Satb2 and Hoxa2 in craniofacial tissues of mouse embryos were obviously different at different time points. The Satb2 gene was down-regulated and the Hoxa2 gene was up-regulated in phenytoin-treated mouse embryonic craniofacial tissue. We conclude that phenytoin may regulate the expression of these two genes in C57BL/6 mice and it may also be involved in the formation of cleft palate.

Previous studies have proposed that the forkhead transcription factor FOXO3a is involved in cell cycle arrest and apoptosis and that it may also repress follicular development by inducing cell cycle arrest in ovaries. We have recently demonstrated that FOXO3a induces oocyte apoptosis of neonatal rat ovaries under in vitro conditions. In the present study, we evaluated the role of FOXO3a in oocyte apoptosis under in vivo conditions. Ovaries from rats were obtained from newborns on postnatal day (PD) 1, 2, 3, and 4. TUNEL assay results showed that oocyte apoptosis occurred mainly on PD 1 and 2. Immunohistochemical staining of FOXO3a, Bim, Fas ligand (FasL), p27KIP1, caspase-8, and caspase-3 showed that they were all expressed mainly in naked oocytes on PD 1 and 2. The percentage of positive FOXO3a staining of oocytes reached peak levels in the ovaries of 2-day-old rats, which was consistent with the rate of the apoptotic profiles determined by TUNEL. The percentage between TUNEL-positive and FOXO3a-positive oocytes in the nucleus showed no statistical differences within the 4-day-old rat ovaries. Furthermore, the positive oocyte percentage of the target factors of FOXO3a (Bim, p27KIP1, and FasL) and pro-apoptotic proteins (caspase-3 and caspase-8) also reached peak levels in the ovaries of 2-day-old rats, which was similar to the rate of FOXO3a-positive oocytes. These results suggest that FOXO3a in the oocyte nucleus is involved in oocyte apoptosis; that is, FOXO3a-positive oocytes may be the apoptotic cells. To verify this, rat oocytes were subjected to TUNEL and immunofluorescent double-labeling assays. We found that TUNEL-positive cells were also FOXO3a-, Bim-, or FasL-positive. To identify the downstream target of FOXO3a, double immunofluorescent staining with antibodies to Bim and FasL was performed. We found that FOXO3a-positive cells were also Bim- and FasL-positive. We conclude that the overexpression of FOXO3a in the oocyte nucleus of neonatal rat ovaries may play an important role in the apoptosis of naked oocytes, and that Bim, FasL, and p27KIP1 are the key downstream factors of FOXO3a.

GroEL is a chaperone thought of as essential for bacterial life. However, some species of Mollicutes are missing GroEL. We use phylogenetic analysis to show that the presence of GroEL is polyphyletic among the Mollicutes, and that there is evidence for lateral gene transfer of GroEL to Mycoplasma penetrans from the Proteobacteria. Furthermore, we propose that the presence of GroEL in Mycoplasma may be required for invasion of host tissue, suggesting that GroEL may act as an adhesin–invasin.

Enteric bacteria such as Escherichia coli have acquired a wide array of acid stress response systems to counteract the extreme acidity encountered when invading the host’s digestive or urinary tracts. These acid stress response systems are both enzyme and chaperone based. The 3 main enzyme-based acid resistance pathways are glutamate-, arginine-, and lysine-decarboxylase pathways. They are under a complex regulatory network allowing the bacteria to fine tune its response to the external environment. HdeA and HdeB are the main chaperones involved in acid stress response. The decarboxylase systems are also found in Vibrio cholera, Vibrio vulnifus, Shigella flexneri, and Salmonella typhimurium, although some differences exist in their functional mechanism and regulation.

More than 30 years of research on nuclear RNA polymerases (RNAP I, II, and III) has uncovered numerous factors that regulate the activity of these enzymes during the transcription reaction. However, very little is known about the machinery that regulates the fate of RNAPs before or after transcription. In particular, the mechanisms of biogenesis of the 3 nuclear RNAPs, which comprise both common and specific subunits, remains mostly uncharacterized and the proteins involved are yet to be discovered. Using protein affinity purification coupled to mass spectrometry (AP–MS), we recently unraveled a high-density interaction network formed by nuclear RNAP subunits from the soluble fraction of human cell extracts. Validation of the dataset using a machine learning approach trained to minimize the rate of false positives and false negatives yielded a high-confidence dataset and uncovered novel interactors that regulate the RNAP II transcription machinery, including a set of proteins we named the RNAP II-associated proteins (RPAPs). One of the RPAPs, RPAP3, is part of an 11-subunit complex we termed the RPAP3/R2TP/prefoldin-like complex. Here, we review the literature on the subunits of this complex, which points to a role in nuclear RNAP biogenesis.

The Core protein of hepatitis C virus is involved in several interactions other than the encapsidation of viral RNA. We recently proposed that this is related to the fact that the N-terminal half of this protein (C82) is an intrinsically unstructured protein (IUP) domain. IUP domains can adopt a secondary structure when they are interacting with another molecule, such as a nucleic acid or a protein. It is also possible to mimic these conditions by modifying the environment of the protein. We investigated the propensity of this protein to fold as a function of salt concentration, detergent, pH, and 2,2,2-trifluoro-ethanol (TFE); only the addition of TFE resulted in a structural change. The effect of TFE addition was studied by circular dichroism, structural, and dynamic data obtained by NMR. The data indicate that C82 can adopt an α-helical structure; this conformation is likely relevant to one of the functional roles of the HCV Core protein.

Cdc48p/p97 is a highly conserved essential AAA protein that is required for many cellular processes, and is identified as a causative gene for an autosomal dominant human disorder, inclusion body myopathy associated with Paget’s disease of the bone and frontotemporal dementia (IBMPFD). Cdc48p/p97 is composed of an N-terminal domain, followed by two AAA domains (D1 and D2) whose ATPase activities have been characterized extensively. In this study, effects of mutations on the essential functions of yeast Cdc48p/p97 in vivo were systematically analyzed. IBMPFD-related mutations do not affect the essential functions of Cdc48p/p97. Loss of ATPase activity of D2 leads to loss of function of the protein in vivo. In contrast, ATPase activity of D1 per se is not essential, but a mutation locking D1 in an ATP-bound form is exceptionally lethal. Site-directed and random mutagenesis analyses suggest that the ATP-bound form of D1 changes an inter-domain interaction, thereby perturbing an essential function of Cdc48p/p97.

To initiate chromosomal DNA replication, specific proteins bind to the replication origin region and form multimeric and dynamic complexes. Bacterial DnaA, the eukaryotic origin recognition complex (ORC), and Cdc6 proteins, most of which include an AAA+(-like) motif, play crucial roles in replication initiation. The importance of ATP binding and hydrolysis in these proteins has recently become recognized. ATP binding of Escherichia coli DnaA is required for the formation of the activated form of a DnaA multimer on the replication origin. The ATP–DnaA multimer can unwind duplex DNA in an origin-dependent manner, which is supported by various specific functions of several AAA+ motifs. DnaA–ATP hydrolysis is stimulated after initiation, repressing extra initiations, and sustaining once-per-cell cycle replication. ATP binding of ORC and Cdc6 in Saccharomyces cerevisiae is required for heteromultimeric complex formation and specific DNA binding. ATP hydrolysis of these proteins is important for the efficient loading of the minichromosome maintenance protein complex, a component of the putative replicative helicase. In this review, we discuss the roles of DnaA, ORC, and Cdc6 in replication initiation and its regulation. We also summarize the functional features of the AAA+ domains of these proteins, and the functional divergence of ORC in chromosomal dynamics.

Lon protease, also known as protease La, is an ATP-dependent serine protease. Despite the presence of a proteolytic Ser–Lys dyad, the enzyme only catalyzes protein degradation in the presence of ATP. Lon possesses an intrinsic ATPase activity that is stimulated by protein and certain peptide substrates. Through sequence alignment and analysis, it is concluded that Lon belongs to the AAA+ protein family. Previous kinetic characterization of the ATPase domain of Escherichia coli Lon protease implicates a half-site reactivity model in which only 50% of the ATP bound to Lon are hydrolyzed to yield ADP; the remaining ATPase sites remain bound with ATP and are considered non-catalytic. In this model, it is implied that ATP hydrolysis is irreversible. To further evaluate the proposed half-site reactivity model, the reversibility of the ATPase activity of E. coli Lon was evaluated by positional isotope exchange experiments. The ATPase reactions were conducted in the18O-enriched buffer such that the extent of18O incorporation into inorganic phosphate generated from ATP hydrolysis could be used to evaluate the extent of reversibility in ATP hydrolysis. Collectively, our experimental data reveal that the ATPase reaction catalyzed by E. coli Lon in the presence and absence of peptide substrate that stimulated the enzyme’s ATPase activity is irreversible. Therefore, the half-site ATPase reactivity of E. coli Lon is validated, and can be used to account for the kinetic mechanism of the ATP-dependent peptidase activity of the enzyme.