Evaluation of epigenetic-related gene expression (DNMT, HDAC1) in Iranian patients with systemic lupus erythematosus.

BACKGROUND: Systemic lupus erythematosus (SLE) is an autoimmune disease in which the immune system abnormally reacts against cells and tissues leading to inflammation. Epigenetic alterations, including DNA methylation and histone modification, have critical effects on autoimmune disease and SLE pathogenesis via dysregulation of critical genes. AIMS: The purpose of this study was to evaluate the epigenetic-related gene expression of DNA methyltransferase (DNMT) and histone deacetylase 1 (HDAC1) in Iranian patients with SLE. METHODS: This matched case-control study included 16 people with SLE and 16 healthy people who were referred to the Rafsanjani rheumatology clinic, in southeast Iran. The expression of DNMT and HDAC1 genes was measured through a real-time PCR assay of blood samples. RESULTS: DNMT gene expression did not differ significantly between SLE and healthy groups (P=0.21). In contrast, HDAC1 gene expression was enhanced in the SLE group, but this enhancement failed to reach statistical significance (P=0.94). CONCLUSION: The results of this study suggest that overexpression of HDAC1 could serve as a diagnostic for SLE disease. Additional studies with larger sample sizes are required to confirm our findings. Evaluation of other genes related to SLE disease is essential and may help to make an accurate diagnosis of the disease.

Evaluation of epigenetic-related gene expression (DNMT, HDAC1) in Iranian patients with systemic lupus erythematosus / Evaluación de la expresión de los genes relacionados con la epigenética (DNMT, HDAC1) en pacientes iraníes con lupus eritematoso sistémico

Background: Systemic lupus erythematosus (SLE) is an autoimmune disease in which the immune system abnormally reacts against cells and tissues leading to inflammation. Epigenetic alterations, including DNA methylation and histone modification, have critical effects on autoimmune disease and SLE pathogenesis via dysregulation of critical genes. Aims: The purpose of this study was to evaluate the epigenetic-related gene expression of DNA methyltransferase (DNMT) and histone deacetylase 1 (HDAC1) in Iranian patients with SLE. Methods: This matched case–control study included 16 people with SLE and 16 healthy people who were referred to the Rafsanjani rheumatology clinic, in southeast Iran. The expression of DNMT and HDAC1 genes was measured through a real-time PCR assay of blood samples. Results: DNMT gene expression did not differ significantly between SLE and healthy groups (P=0.21). In contrast, HDAC1 gene expression was enhanced in the SLE group, but this enhancement failed to reach statistical significance (P=0.94). Conclusion: The results of this study suggest that overexpression of HDAC1 could serve as a diagnostic for SLE disease. Additional studies with larger sample sizes are required to confirm our findings. Evaluation of other genes related to SLE disease is essential and may help to make an accurate diagnosis of the disease.(AU)

Antecedentes: El lupus eritematoso sistémico (LES) es una enfermedad autoinmune, en la cual el sistema inmunitario reacciona de manera anormal frente a las células y tejidos causantes de la inflamación. Las alteraciones epigenéticas, incluyendo la metilación del ADN y la modificación de la histona, tienen efectos críticos en la enfermedad autoinmune y la patogenia del LES, a través de la desregulación de los genes críticos. Objetivo: El objetivo de este estudio fue evaluar la expresión del gen relacionado con la epigenética de ADN metiltransferasa (DNMT) e histona deacetilasa 1 (HDAC1) en los pacientes iraníes afectados de LES. Métodos: Este estudio pareado caso-control incluyó 16 personas con LES y 16 personas sanas, derivadas a la clínica de reumatología de Rafsanjan, en el sudeste de Irán. La expresión de los genes DNMT y HDAC1 se midió mediante una PCR a tiempo real de muestras de sangre.Resultados: La expresión del gen DNMT no difirió significativamente entre los grupos de pacientes de LES y de controles sanos (p=0,21). Por contra, la expresión del gen HDAC1 se incrementó en el grupo LES, aunque dicho incremento no alcanzó significación estadística (p=0,94). Conclusión: Los resultados de este estudio sugieren que la sobreexpresión de HDAC1 podría servir para diagnosticar el LES. Son necesarios estudios adicionales con muestras de mayor tamaño para confirmar nuestros hallazgos. Es esencial la evaluación de otros genes relacionados con el LES, pudiendo ayudar a realizar un diagnóstico preciso de la enfermedad.(AU)

Evaluation of H19, Mest, Meg3, and Peg3 genes affecting growth and metabolism in umbilical cord blood cells of infants born to mothers with gestational diabetes and healthy mothers in Rafsanjan City, Iran.

Hyperglycemia during the first trimester leads to an increased risk of innate malformations as well as death at times close to delivery dates. The methylated genes include those from paternal H19 and PEG3 and those from maternal MEST and MEG3 that are necessary for the growth and regulation of the human fetus and its placenta. The aim of this study was to evaluate and compare the expression of these genes in the cord blood of healthy infants born to mothers with gestational diabetes mellitus (GDM) and healthy mothers.This case-control study was conducted on the cord blood of 40 infants born to mothers with GDM and 35 infants born to healthy mothers. Mothers were identified by measuring oral glucose tolerance in the 24th-26th week of pregnancy. Cord blood was obtained post-delivery, and cord blood mononuclear cells were immediately extracted, using Ficoll solution. Then, RNA extraction and cDNA synthesis were performed, and gene expression of MEG3, PEG3, H19, and MEST was assessed through quantitative real-time PCR.Findings show that the expression levels of MEG3, PEG3, H19, and MEST genes were significantly decreased in mononuclear cord blood cells of infants born to mothers with GDM when compared to those of the healthy control group.These findings reveal that the reduction of imprinted genes in mothers with GDM is most likely due to changes in their methylation by an epigenetic process. Considering the importance of GDM due to its high prevalence and its side effects both for mother and fetus, recognizing their exact mechanisms is of high importance. This has to be studied more widely.

Academic bullying and diversity: challenges and solutions.

The academy remains disproportionally white despite decades of efforts to diversify the professoriate. Recruitment of faculty from underrepresented populations continues to challenge and little attention has been placed on retention. Bullying threatens all workplaces and is particularly insidious in higher education where the practice of tenure grants bad actors immunity from many corrective measures. Strategies and approaches summarized here have been proposed and should be adopted to provide a better workplace for all.

The role of sex as a biological variable in the efficacy and toxicity of therapeutic nanomedicine.

Males and females have physiological, hormonal, and genetic differences that can cause different responses to medicinal treatments. The role of sex in the pharmacokinetics and pharmacodynamics of drugs is well established in the literature. However, researchers have yet to robustly and consistently consider the impact of sex differences on the pharmacokinetics and pharmacodynamics of nanomedicine formulations when designing nanomedicine therapeutics and/or constructing clinical trials. In this review, we highlight the physiological and anatomical differences between sexes and discuss how these differences can influence the therapeutic efficacy, side effects, and drug delivery safety of nanomedicine products. A deep understanding of the effects of sex on nano-based drug delivery agents will robustly improve the risk assessment process, resulting in safer formulations, successful clinical translation, and improved therapeutic efficacies for both sexes.

The File Drawer Problem in Nanomedicine.

The role of the 'file drawer' problem in nanomedicine, which partly drives the current limited clinical success of therapeutic nanoparticles, has been poorly investigated. We propose an integrated functioning of all stakeholders as the only effective way to address the file drawer problem in an efficient and timely manner.

BDNF infusion into the MPN mag is sufficient to restore copulatory behavior in the castrated Syrian hamster.

Testosterone plays a key role in the expression of male sex behavior by influencing cellular activity and synapses within the magnocellular medial preoptic nucleus (MPN mag), a sub-nucleus of the medial preoptic area (MPOA) in the Syrian hamster. Although the mechanisms underlying hormonally-induced synaptic plasticity in this region remain elusive, the data suggests that an increase in synaptic density may mediate testosterone's effects on copulation. As brain derived neurotrophic factor (BDNF) plays an integral role in regulating synaptic plasticity and gonadal steroids regulate the levels of BDNF, we hypothesize that BDNF may mediate the effects of gonadal hormones on copulatory behavior. To test this hypothesis, we infused BDNF or controls into the MPN mag of long-term castrates. Our results indicate that BDNF, but not the controls, restored copulatory behavior in castrated male Syrian hamsters. Furthermore, the rise of BDNF expression in the MPOA preceded the rise of synaptophysin following testosterone replacement in castrated males. These data are consistent with our hypothesis, implicating a role for BDNF in mediating testosterone's action on copulation and suggest that the delay in testosterone's restoration of copulation is, in part, due to the delay in the increase of BDNF and synaptophysin.

TrkB is necessary for male copulatory behavior in the Syrian Hamster (Mesocricetus auratus).

The magnocellular medial preoptic nucleus (MPN mag), a subdivision of the medial preoptic area (MPOA), plays a critical role in the regulation of copulation in the male Syrian hamster; in part by mediating the effects of gonadal steroids. For example, ablation of the MPN mag eliminates mating and testosterone placed in the MPN mag restores mating in castrated males. Furthermore, testosterone treatment enhances synaptic density and dendritic spines in the MPN mag. Thus, copulatory behaviors are correlated with increases in synaptic morphology in the MPN mag. As brain derived neurotrophic factor (BDNF) and its receptor, tyrosine receptor kinase-B (TrkB), effect neuronal growth and synaptic plasticity, this study explored the role of TrkB and BDNF in mediating testosterone's effects on the MPN mag and behavior. Testosterone treatment increased BDNF expression and conversely lowered TrkB expression in the MPOA. siRNA-mediated TrkB knockdown in the MPN mag eliminated copulation two-days post injection and the behavior was restored one week later. These data indicate that testosterone influences the expression of BDNF and TrkB in the MPOA and that expression of copulation is dependent on the presence of TrkB. Taken together our findings support a role for TrkB and BDNF in mediating the effects of testosterone on copulatory behavior in the Syrian hamster.

Sexual dimorphic expression of TrkB, TrkB-T1, and BDNF in the medial preoptic area of the Syrian hamster.

Neurotrophins regulate many aspects of neuronal function and activity. Specifically, the binding of Brain-derived neurotrophic factor (BDNF) to Tyrosine receptor kinase-B (TrkB) or its truncated version, TrkB-T1, can cause growth and differentiation or dominant inhibition of receptor signaling, respectively. There is evidence that these neurotropic effects on nervous tissue, in both the central and peripheral nervous system, behave differently between the sexes. This study used western blots to examine the expression of these neurotrophins in the medial preoptic area (MPOA), a sexually dimorphic region of the hamster brain that controls male sex behavior. We report that TrkB-FL and BDNF show greater expression in male MPOA tissue, when compared to female. On the contrary, TrkB-T1 is expressed in greater abundance in the female MPOA. Our results indicate a clear sexual dimorphism of neurotrophins in the MPOA of the Syrian hamster. Furthermore, the greater expression of TrkB-FL and BDNF in the male MPOA suggests that these neurotrophins could be promoting synaptic growth to facilitate male-typical copulation. In contrast, the greater TrkB-T1 expression in the female MPOA suggests a possible inhibition of synaptic growth, and may contribute to the lack of male-typical copulation. Altogether, our data suggests that neurotrophins may play a larger role sexual differentiation than previously thought.

Childhood long-term conditions in primary care: a qualitative study of practitioners' views.

BACKGROUND: Improving child health and wellbeing in England was the key focus of the Chief Medical Officer's Annual Report 2012, which recommended that all children with long-term conditions (LTCs) have a named GP responsible for their care. Little is known, however, about practitioners' views and experiences of supporting children with LTCs in primary care. AIM: To explore practitioners' views of supporting children with LTCs and their families in primary care. DESIGN AND SETTING: Qualitative interview study in primary care settings in South Yorkshire, England. METHOD: Interviews explored practitioners' views and experiences of supporting children with asthma, cystic fibrosis, type 1 diabetes, and epilepsy. Interviews were audiotaped, transcribed verbatim, and analysed using the framework approach. RESULTS: Nineteen practitioners were interviewed: 10 GPs, five practice nurses, and four nurse practitioners. The GPs' clinical roles included prescribing and concurrent illness management; nurse practitioners held minor illness clinics; and practice nurses conduct asthma clinics and administer immunisations. GPs were coordinators of care and provided a holistic service to the family. GPs were often unsure of their role with children with LTCs, and did not feel they had overall responsibility for these patients. Confidence was dependent on experience; however, knowledge of GPs' own limits and accessing help were felt to be more important than knowledge of the condition. CONCLUSION: Primary care has a valuable role in the care of children with LTCs and their families. This study suggests that improving communication between services would clarify roles and help improve the confidence of primary care practitioners.

Using professional interpreters in undergraduate medical consultation skills teaching.

The ability to work with interpreters is a core skill for UK medical graduates. At the University of Sheffield Medical School, this teaching was identified as a gap in the curriculum. Teaching was developed to use professional interpreters in role-play, based on evidence that professional interpreters improve health outcomes for patients with limited English proficiency. Other principles guiding the development of the teaching were an experiential learning format, integration to the core consultation skills curriculum, and sustainable delivery. The session was aligned with existing consultation skills teaching to retain the small-group experiential format and general practitioner (GP) tutor. Core curricular time was found through conversion of an existing consultation skills session. Language pairs of professional interpreters worked with each small group, with one playing patient and the other playing interpreter. These professional interpreters attended training in the scenarios so that they could learn to act as patient and family interpreter. GP tutors attended training sessions to help them facilitate the session. This enhanced the sustainability of the session by providing a cohort of tutors able to pass on their expertise to new staff through the existing shadowing process. Tutors felt that the involvement of professional interpreters improved student engagement. Student evaluation of the teaching suggests that the learning objectives were achieved. Faculty evaluation by GP tutors suggests that they perceived the teaching to be worthwhile and that the training they received had helped improve their own clinical practice in consulting through interpreters. We offer the following recommendations to others who may be interested in developing teaching on interpreted consultations within their core curriculum: 1) consider recruiting professional interpreters as a teaching resource; 2) align the teaching to existing consultation skills sessions to aid integration; and 3) invest in faculty development for successful and sustainable delivery.

Testosterone regulates the density of dendritic spines in the male preoptic area.

Male-typical behavior is dependent on testosterone. Castrated males gradually stop mating and engaging in sexual behaviors. Castrates treated with testosterone regain motivation and sex behaviors over time. Although this effect is well characterized, the specific mechanisms by which testosterone treatment recovers sexual behaviors remain unknown. The medial preoptic area (MPOA) is a likely site for testosterone's action on copulation. The integrity of the area is essential for the expression of male sex behavior; and the MPOA is densely populated with receptors for gonadal steroids. Moreover testosterone appears to regulate synaptic efficacy in the MPOA. Exposure to sexually relevant stimuli stimulates the MPOA but only in the presence of circulating testosterone. Sites afferent to the area respond to similar exposure independent of the hormonal milieu suggesting that testosterone mediates communication between the MPOA and its afferents. The protracted time course suggests that the effects of steroidal manipulation are mediated by structural changes. The present experiment evaluated this hypothesis by comparing dendritic spine density among Syrian hamsters that were castrated, castrated and treated with testosterone, or were left gonadally intact. Brains were sectioned and stained using the rapid Golgi stain protocol (FD Neurotechnologies, Baltimore), and the spine density, dendrite length, and the number of branches were compared among groups. Intact and testosterone replaced animals had more spines and greater spine density but did not differ in dendrite length and branching from castrated animals. These results suggest that existing dendrites increase the number of spines available for synapse formation but do not extend their dendrites in response to testosterone treatment.

Exposure to female pheromones stimulates a specific type of neuronal population in the male but not female magnocellular division of the medial preoptic nucleus (MPN mag) of the Syrian hamster.

The magnocellular division of the medial preoptic area (MPN mag) integrates pheromonal and hormonal signals to play a critical role in the expression of male typical sex behavior. The MPN mag contains two morphologically distinct neuronal populations; the percentage of each type within the nucleus is sex specific. Males have more neurons with a single nucleolus whereas females have more with multiple nucleoli. To determine which neuronal subtype mediates pheromonal induction of copulation, tissue from male and female hamsters exposed to female pheromones was immunolabeled for the immediate early protein (EGR-1). Subsequently the tissue was counterstained and the number of ERG-1 neurons with one or two nuclei was determined. The results indicate that pheromones stimulate neurons with single nucleoli in males but fail to stimulate either neuronal subtype in females suggesting that synaptic input to the MPN mag is sexually differentiated.

Neuronal composition of the magnocellular division of the medial preoptic nucleus (MPN mag) is sex specific in the Syrian hamster (Mesocricetus auratus).

The magnocellular division of the medial Preoptic nucleus (MPN mag) plays a critical role in the regulation of male sexual behavior in the hamster. Results from previous studies indicated that the number of neurons in the MPN mag is greater in males than females but failed to find significant differences in the volume of the nucleus suggesting that other elements in the nucleus may be greater in the female. The results of the present study, using NeuN to identify neurons, are in line with this hypothesis. The data show that (1) neurons in the MPN mag display two distinct phenotypes, those with a single nucleolus and those with multiple nucleoli; (2) the percentage of each phenotype is sex specific, differing over the course of development and (3) there is no sex difference in the number of glial cells at any age. Sex differences in the numbers of each type are correlated with developmental milestones and suggest that morphological changes are influenced by changes in circulating gonadal steroids during development.

Fluctuations in cellular proliferation across the light/dark cycle in the subgranular zone of the dentate gyrus in the adult male Syrian hamster.

A lifelong persistent neurogenesis occurs in the dentate gyrus of the mammalian hippocampus. Research in peripheral cell tissue has shown that the timing of cellular division of these cells coincide with the light/dark cycle, however it remains unclear as to whether there is an association between the time of day and cellular proliferation in the brain. The timing of cellular division can be studied through the use of a cellular proliferation marker, such as 5-bromo-2-deoxyuridine (BrdU), which is taken up by the DNA of dividing cells during replication. The goal of this study was to determine whether the time of day affects the number of BrdU labeled cells in the subgranular zone of the dentate gyrus of adult male Syrian hamsters. Adult males received a single systemic injection of BrdU (300 mg/kg) at either the end of the light (ZT-13) or dark phase (ZT-23) of a 14:10 LD cycle and were sacrificed 24h or 3 days later. Sections through the hippocampus were immunolabeled for BrdU. Cellular proliferation fluctuated across the light/dark cycle during the expansion phase rather than during initial cellular proliferation. A twofold increase in number was expected between 24 and 72h following a single BrdU injection, but this increase was only seen in the population of cells injected at the end of the light phase.

Hamster sperm-associated alpha-L-fucosidase functions during fertilization.

Sperm-associated alpha-L-fucosidases have been identified in diverse organisms. Their wide phylogenetic distribution and known properties support the likelihood that L-fucose and alpha-L-fucosidase have fundamental function(s) during gamete interaction. This is consistent with the substantial evidence in the literature documenting the importance of carbohydrate moieties during fertilization. Direct enzyme assays were employed to evaluate the functional distribution of alpha-L-fucosidase in preparations of hamster sperm. In vitro fertilization was performed using Syrian hamster sperm and eggs to identify the functional role of hamster sperm-associated alpha-L-fucosidase during zona pellucida binding/penetration, sperm-egg membrane fusion, and postfusion events. Results reported here document the presence of hamster sperm-associated alpha-L-fucosidase and demonstrate that it functions during fertilization at the stage of sperm-oocyte membrane interaction and/or postfusion events within the zygote. Understanding the role of alpha-L-fucosidase during human fertilization could lead to development of improved infertility treatments.

Temperature, peroxide concentration, and immunohistochemical staining method affects staining intensity, distribution, and background.

The techniques used to label neural tissue for specific antigens can vary significantly. Some immunostaining methods use free-floating tissue sections, whereas others use tissue sections mounted on slides. Mounting sections on glass slides before labeling the tissue with antigens is preferred method for neonatal tissue; processing young tissue by free-floating methods often destroy it. Surprisingly optimal temperature for storing tissue can vary with age. This study describes parameters developed to obtain robust staining of both young and old tissue. Our results show the most robust staining was found in tissue that was (1) stored at very low temperatures (-20 degrees C and -80 degrees C), (2) pretreated with 0.01% peroxide, and (3) entirely immersed in the staining solutions during immunohistochemistry.

The MPN mag: introducing a critical area mediating pheromonal and hormonal regulation of male sexual behavior.

Mating behavior in male hamsters is regulated by a chemosensory pathway that converges on the bed nucleus of the stria terminalis (BST) and the medial nucleus of the amygdala (Me). Both the BST and the Me project to the lateral part of the medial preoptic area. Lesion studies have identified a small group of large cells referred to as the magnocellular medial preoptic nucleus (MPN mag) whose integrity is required for normal mating behavior. Our data, summarized within, indicate that the MPN mag is a sexually differentiated nucleus in a large steroid-responsive network that relays pheromonal signals from the sensory systems to the motor areas to affect behavior.