Endocrine Development Research Articles

Identification and characterization of light-responsive genes in pre-settlement eyed veligers of Mytilus coruscus

Mytilus coruscus is one of large bivalve mollusks that is wildly distributed along the coasts of Korea, Japan and China and as a potential economic shellfish candidate. Efforts to propagate and culture M. coruscus have expanded worldwide recent years. The larval stage of bivalves is a vital life-history phase, of which the eyed veligers larva with a pigmented eyespot is the pre-settlement phase of M. coruscus. However, information on light conditions for the breeding of its larval during the eye-spotting period is still limited. In our study, based on the transcriptome analysis of the different light cycles from M. coruscus were performed with the DNBSEQ to screen for light-response genes. Comparative transcriptome analysis identified 853 differentially expressed genes (DEGs) in the 12 h vs 0 h group and 696 genes were differentially expressed between 12 h vs 24 h group. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses revealed that DEGs were assigned to 1078 and 969 pathways, respectively. The DEGs were enriched in organismal systems pathways such as immune system pathway, endocrine system pathway, development pathway, and environmental adaptation pathway. Further analysis revealed that DEGs, including CRY and CLOCK, were significantly overrepresented in the circadian rhythm pathway. We also identified a visual OPIN gene, which affects the detection of relevant visual signals. Quantitative real-time PCR (qRTPCR) showed that CRY and CLOCK were down-regulated, whereas OPIN was up-regulated. The data from this study provides important insights into the light response of M. coruscus, which will aid future investigations into the management of M. coruscus breeding in Bivalvia.

SAT427 Effects Of Polychlorinated Biphenyls (PCBs) On Transcription Factor NFκB Activation In Neonatal Rat Microglia Primary Culture

Abstract Disclosure: C.E. Dressel: None. G.M. Valdez: None. M.R. Bell: None. Polychlorinated biphenyls (PCBs) are a class of persistent organic pollutants that are lipophilic and accumulate in the food chain. Despite bans on PCB production, human exposure continues and can cause disruptions in endocrine, nervous, and immune system functions. As such, PCBs may interfere with the actions of microglia, the innate immune cells of the central nervous system. Healthy microglia are critical for neural development, maintenance of neural functions, and responses to stress, injury, or infection. Initial studies of primary neonatal rat microglial cultures indicate that PCBs alter gene expression of proinflammatory cytokine TNF-α and IL-1β in response to a secondary inflammatory lipopolysaccharide (LPS) challenge. Production of these cytokines is, in part, regulated by transcription factor NFκB. This experiment tests the hypothesis that PCBs alter NFκB activation in neonatal microglia. The forebrains of male and female neonatal Sprague-Dawley rats were dissected to remove meninges, homogenized with collagenase and gentle trituration, and plated on coverslips in DMEM FBS+ media to establish a primary mixed glial culture. After one week, microglia were subcultured using mild trypsin treatment and maintained in conditioned media for 24 hours. The microglia were then exposed to either PCB 153 (0.1 µM) or DMSO vehicle control (0.036%) for 18 hours. After 18 hours, the cells were treated with either LPS (0.5 µg/mL) or PBS vehicle control for 4 hours and then fixed. Immunocytochemistry was used to visualize NFκB and identify microglia, with anti-p65 and Ox42 antibodies, respectively. Coverslips were mounted with DAPI-containing media and the cells were imaged via confocal microscopy. In response to immune activation, NFκB typically translocates from the cytosol to the nucleus. Therefore, we assessed NFκB activation by quantifying p65 fluorescent pixel intensity and percentage of area covered in nuclear or cytoplasmic regions according to DAPI and Ox42 channel masks using ImageJ. A two-way ANOVA was used to identify the effects of PCB vs. DMSO exposure and LPS vs. PBS challenge. Preliminary data indicated that LPS increased the ratio of nuclear to cytosolic NFκB, as expected. PCB exposed cells tended to have lower NFκB presence in the nucleus and cell as a whole, but a greater proportion of NFκB staining localized within the nucleus relative to the rest of the cell. With the current sample (n=4-6 per group, 2-3 males and females each), no interactions between PCBs and LPS were observed. Continued work will increase sample size to explore sex differences and the time course of NFκB response to PCBs, which may help broaden our understanding of the mechanisms by which PCBs affect normal brain activity and disrupt healthy endocrine, immune, and neural development. Presentation: Saturday, June 17, 2023

PAX4 loss of function increases diabetes risk by altering human pancreatic endocrine cell development

The coding variant (p.Arg192His) in the transcription factor PAX4 is associated with an altered risk for type 2 diabetes (T2D) in East Asian populations. In mice, Pax4 is essential for beta cell formation but its role on human beta cell development and/or function is unknown. Participants carrying the PAX4 p.His192 allele exhibited decreased pancreatic beta cell function compared to homozygotes for the p.192Arg allele in a cross-sectional study in which we carried out an intravenous glucose tolerance test and an oral glucose tolerance test. In a pedigree of a patient with young onset diabetes, several members carry a newly identified p.Tyr186X allele. In the human beta cell model, EndoC-βH1, PAX4 knockdown led to impaired insulin secretion, reduced total insulin content, and altered hormone gene expression. Deletion of PAX4 in human induced pluripotent stem cell (hiPSC)-derived islet-like cells resulted in derepression of alpha cell gene expression. In vitro differentiation of hiPSCs carrying PAX4 p.His192 and p.X186 risk alleles exhibited increased polyhormonal endocrine cell formation and reduced insulin content that can be reversed with gene correction. Together, we demonstrate the role of PAX4 in human endocrine cell development, beta cell function, and its contribution to T2D-risk.

PCOS and vitamin D: a clinical appraisal.

Polycystic ovary syndrome (PCOS) is the most common endocrine-reproductive disease linked not just to infertility but also to serious comorbidities. There is a reported association between low vitamin D levels and multiple health conditions including PCOS. This narrative review aims to analyze the role of vitamin D in PCOS development, use of the vitamin D in the treatment of PCOS, and the molecular basis of these observations. A Medline and PubMed research was performed, during the years 1990-2023, using a combination of keywords on such topic. According to the author's evaluation and target, papers were identified and included for a narrative review. There are associations between lower levels of vitamin D and PCOS, as well as with insulin resistance, metabolic syndrome, hyperandrogenemia, metabolic and endocrine disorders as well as the onset of oxidative stress and pro-inflammatory milieu, in PCOS women. Vitamin D has a role in pathologic changes linked to PCOS. Molecular and clinical investigations which give new information about the role of vitamin D in the development of PCOS and related endocrine and metabolic disturbance are further needed.

Aberrant metabolite trafficking and fuel sensitivity in human pluripotent stem cell-derived islets

Pancreatic islets regulate blood glucose homeostasis through the controlled release of insulin; however, current metabolic models of glucose-sensitive insulin secretion are incomplete. A comprehensive understanding of islet metabolism is integral to studies of endocrine cell development as well as diabetic islet dysfunction. Human pluripotent stem cell-derived islets (SC-islets) are a developmentally relevant model of human islet function that have great potential in providing a cure for type 1 diabetes. Using multiple 13C-labeled metabolic fuels, we demonstrate that SC-islets show numerous divergent patterns of metabolite trafficking in proposed insulin release pathways compared with primary human islets but are still reliant on mitochondrial aerobic metabolism to derive function. Furthermore, reductive tricarboxylic acid cycle activity and glycolytic metabolite cycling occur in SC-islets, suggesting that non-canonical coupling factors are also present. In aggregate, we show that many facets of SC-islet metabolism overlap with those of primary islets, albeit with a retained immature signature.

RNA atlas and competing endogenous RNA regulation in tissue-derived exosomes from luminal B and triple-negative breast cancer patients.

Luminal B and triple-negative breast cancer (TNBC) are malignant subtypes of breast cancer (BC), which can be attributed to the multifaceted roles of tissue-derived exosomes (T-exos). Competing endogenous RNA (ceRNA) networks can regulate gene expression post-transcriptionally. RNAs in T-exos from luminal B BC (n=8) and TNBC (n=8) patients were compared with those from persons with benign breast disease (n=8). The differentially expressed (DE) mRNA, microRNA (miRNA), and long noncoding RNA (lncRNA) target genes were annotated using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) to reveal the relevant biological processes.The ceRNA networks were constructed to show distinct regulation, and the mRNAs involved were annotated. The miRNAs involved in the ceRNA networks were screened with the Kaplan-Meier Plotter database to identify dysregulated ceRNAs with prognostic power. In total, 802 DE mRNAs, 441 DE lncRNAs, and 104 DE miRNAs were identified in luminal B BC T-exos, while 1699 DE mRNAs, 590 DE lncRNAs, and 277 DE miRNAs were identified in TNBC T-exos. Gene annotation revealed that the RAS-MAPK pathway was the primary biological process in luminal B BC T-exos, while endocrine system development and growth were the main processes in TNBC T-exos. Survival analysis established seven survival-related lncRNA/miRNA/mRNA regulations in luminal B BC T-exos, and nineteen survival-related lncRNA/miRNA/mRNA regulations in TNBC T-exos. In addition to survival-related ceRNA regulations, ceRNA regulation of RAS-MAPK in luminal B and endocrine system development and growth regulation in TNBC might contribute to the tumorigenesis of BC.

Arginine 65 methylation of Neurogenin 3 by PRMT1 is required for pancreatic endocrine development of hESCs

Neurogenin 3 (NGN3) is a key transcription factor in the cell fate determination of endocrine progenitors (EPs) in the developing pancreas. Previous studies have shown that the stability and activity of NGN3 are regulated by phosphorylation. However, the role of NGN3 methylation is poorly understood. Here, we report that protein arginine methyltransferase-1 (PRMT1)-mediated arginine 65 methylation of NGN3 is required for the pancreatic endocrine development of human embryonic stem cells (hESCs) in vitro. We found that inducible PRMT1-knockout (P-iKO) hESCs did not differentiate from EPs into endocrine cells (ECs) in the presence of doxycycline. Loss of PRMT1 caused NGN3 accumulation in the cytoplasm of EPs and decreased the transcriptional activity of NGN3. We found that PRMT1 specifically methylates NGN3 arginine 65 and that this modification is a prerequisite for ubiquitin-mediated degradation. Our findings demonstrate that arginine 65 methylation of NGN3 is a key molecular switch in hESCs permitting their differentiation into pancreatic ECs.

85-OR: Swi/Snf Controls Transcriptional Programs Required for ß-Cell Development and Postnatal Islet Function

A key feature of diabetes is insufficient functional β-cell mass. Strategies to augment functional β-cell mass include directed differentiation of stem cells towards a β-cell fate, which requires extensive knowledge of transcriptional programs governing differentiation in vivo. Pdx1, an essential transcription factor involved in β-cell differentiation, dynamically recruits coregulators to drive different gene expression programs. A recent study identified the ATP-dependent Swi/Snf chromatin remodeling complex as an important Pdx1-interacting partner. In the developing pancreas, conditional removal of the Brg Swi/Snf ATPase subunit compromised final pancreas mass, and loss of both ATPase subunits, Brg1 and Brm, led to glucose dyshomeostasis. While Swi/Snf is essential during early pancreas development and mature β-cell function, the role of Swi/Snf during endocrine cell development has yet to be explored. Here, we generated a conditional knockout of either Brg1 (Brg1ΔendoBrm+/-), Brm (Brg1Δendo/+Brm-/-), or both subunits (DKOΔendo) during endocrine development using Neurogenin 3 Cre. Four-week-old Brg1ΔendoBrm+/- mice are glucose intolerant, hyperglycemic, and hypoinsulinemic, with no phenotype observed in Brg1Δendo/+Brm-/-mice. Impaired glucose homeostasis of Brg1ΔendoBrm+/- mice was due in part to reduced islet number, impaired insulin secretion, and altered gene expression programs. DKOΔendo mice have not been found at weaning; however, postnatal day 6 DKOΔendo mice are severely hyperglycemic with reduced plasma insulin levels. Single-cell RNA-Seq of embryonic day 15.5 lineage-labeled cells revealed reduced insulin and glucagon transcript in the β-cell and α-cell clusters, respectively, in both Brg1ΔendoBrm+/- and DKOΔendo mice and reduced Neurogenin 3­ transcript in DKOΔendo endocrine progenitor clusters. These data suggest Swi/Snf plays a critical role in governing gene-expression programs essential for endocrine cell development. Disclosure R.K.Davidson: Employee; Eli Lilly and Company. S.Kanojia: None. M.E.Osmulski: None. J.Spaeth: None. Funding National Institutes of Health (R01DK129287, R03DK127129)

290-LB: Single-Cell Analysis of Endocrine Development Identifies Novel Roles for GLIS3 in Pancreatic ß-Cell Fate

The Krüppel-like zinc finger transcription factor GLIS3 has been identified as a critical regulator of pancreatic beta cell development and function. Mice lacking a functional Glis3 gene fail to produce enough insulin and suffer from severe hyperglycemia soon after birth. This phenotype matches what has been observed in humans with genetic mutations in GLIS3 and who have been diagnosed with permanent neonatal diabetes within the first month of life. Additional and ongoing work has begun to identify some of the transcriptional mechanisms utilized by GLIS3, yet many questions remain unanswered. One such question is whether GLIS3 has roles in endocrine development prior to the formation of functional β-cells. To address this question, I have utilized single cell RNA-sequencing of mouse embryonic pancreas, at embryonic days (e)15.5 and e18.5, of both wild type (WT) and Glis3 knockout (KO) embryos. Analysis of this data has identified the expected developmental populations, from bipotent progenitor cells to pro-endocrine cells and ultimately hormone positive β-, α-, δ-, and ε-cells. Interestingly, there is little difference in gene expression between the WT and Glis3 KO pancreas in most of the populations observed (pro-endocrine, FEV+, α-/δ-/ε-cells). However, there are significant differences in gene expression between WT β-cells and KO β-cells. Surprisingly, most of the differences are in genes upregulated in the KO cells, with only a small number of genes downregulated. These upregulated genes are normally downregulated over the course of development from FEV+ to β-cells. These data indicate that GLIS3 plays a unique role in developing β-cells in that it primarily represses genes during the course of development but is also critical for the transcriptional activation of specific targets, particularly Ins2. Understanding how GLIS3 regulates β-cell generation, and how GLIS3 itself is regulated, could potentially provide new therapeutic avenues for the treatment of diabetes. Disclosure D. W. Scoville: None. S. Grimm: None. A. M. Jetten: None. Funding National Institutes of Health (Z01-ES-101585)

Pharmacological inhibition of human EZH2 can influence a regenerative β-like cell capacity with in vitro insulin release in pancreatic ductal cells

BackgroundTherapeutic replacement of pancreatic endocrine β-cells is key to improving hyperglycaemia caused by insulin-dependent diabetes . Whilst the pool of ductal progenitors, which give rise to the endocrine cells, are active during development, neogenesis of islets is repressed in the human adult. Recent human donor studies have demonstrated the role of EZH2 inhibition in surgically isolated exocrine cells showing reactivation of insulin expression and the influence on the H3K27me3 barrier to β-cell regeneration. However, those studies fall short on defining the cell type active in transcriptional reactivation events. This study examines the role of the regenerative capacity of human pancreatic ductal cells when stimulated with pharmacological inhibitors of the EZH2 methyltransferase.ResultsHuman pancreatic ductal epithelial cells were stimulated with the EZH2 inhibitors GSK-126, EPZ6438, and triptolide using a 2- and 7-day protocol to determine their influence on the expression of core endocrine development marker NGN3, as well as β-cell markers insulin, MAFA, and PDX1. Chromatin immunoprecipitation studies show a close correspondence of pharmacological EZH2 inhibition with reduced H3K27me3 content of the core genes, NGN3, MAFA and PDX1. Consistent with the reduction of H3K27me3 by pharmacological inhibition of EZH2, we observe measurable immunofluorescence staining of insulin protein and glucose-sensitive insulin response.ConclusionThe results of this study serve as a proof of concept for a probable source of β-cell induction from pancreatic ductal cells that are capable of influencing insulin expression. Whilst pharmacological inhibition of EZH2 can stimulate secretion of detectable insulin from ductal progenitor cells, further studies are required to address mechanism and the identity of ductal progenitor cell targets to improve likely methods designed to reduce the burden of insulin-dependent diabetes.

High Fat Diet Exaggerate Metabolic and Reproductive PCOS Features by Promoting Oxidative Stress: An Improved EV Model in Rats.

Background and Objectives: Polycystic ovary syndrome (PCOS) is a frequent multifactorial endocrinopathy affecting women in the reproductive period, often associated with infertility and metabolic disorders. The use of animal models helps to better understand etiopathogenesis, enabling the examination of the effects of certain drugs in order to discover the best possible therapeutic approach. We tried to investigate the additional effect of estradiol-valerate (EV) and high-fat diet (HFD) in female rats to explore PCOS-related alterations with special focus on oxidative stress. Materials and Methods: Animals were divided into three groups: control group (CTRL, n = 6), estradiol-valerate group (EV, n = 6), and estradiol-valerate group on HFD (EV + HFD, n = 6). PCOS was induced by single subcutaneous injection of long-acting EV in a dose of 4 mg/per rat. We tried to improve the metabolic characteristics of the PCOS animal model by adding HFD, so the CTRL and EV group had a regular diet, while the EV + HFD group had HFD during the induction period of 60 days. Results: We observed alterations of anthropometric parameters and hormonal disturbances, along with estrus cycle impairment reassembly to obese-type PCOS phenotype. Moreover, glucose metabolism was impaired after addition of HFD to EV protocol, contrary to EV administered alone. Histological analysis confirmed more numerous cystic follicles after the combination of EV and HFD protocol. The alterations of oxidative stress markers could be related to and serve as the mechanistic base for development of PCOS-related endocrine, reproductive, and metabolic properties. Conclusions: The additive effect of EV and HFD was obvious in the majority of the parameters observed. Our study strongly demonstrated metabolic as well as reproductive properties of PCOS in rats.

Sensing High 17β-Estradiol Concentrations Using a Planar Microwave Sensor Integrated with a Microfluidic Channel.

The global issue of pollution caused by endocrine-disrupting chemicals (EDCs) has been gaining increasing attention. Among the EDCs of environmental concern, 17β-estradiol (E2) can produce the strongest estrogenic effects when it enters the organism exogenously through various routes and has the potential to cause harm, including malfunctions of the endocrine system and development of growth and reproductive disorders in humans and animals. Additionally, in humans, supraphysiological levels of E2 have been associated with a range of E2-dependent disorders and cancers. To ensure environmental safety and prevent potential risks of E2 to human and animal health, it is crucial to develop rapid, sensitive, low cost and simple approaches for detecting E2 contamination in the environment. A planar microwave sensor for E2 sensing is presented based on the integration of a microstrip transmission line (TL) loaded with a Peano fractal geometry with a narrow slot complementary split-ring resonator (PF-NSCSRR) and a microfluidic channel. The proposed technique offers a wide linear range for detecting E2, ranging from 0.001 to 10 mM, and can achieve high sensitivity with small sample volumes and simple operation methods. The proposed microwave sensor was validated through simulations and empirical measurements within a frequency range of 0.5-3.5 GHz. The E2 solution was delivered to the sensitive area of the sensor device via a microfluidic polydimethylsiloxane (PDMS) channel with an area of 2.7 mm2 and sample value of 1.37 µL and measured by a proposed sensor. The injection of E2 into the channel resulted in changes in the transmission coefficient (S21) and resonance frequency (Fr), which can be used as an indicator of E2 levels in solution. The maximum quality factor of 114.89 and the maximum sensitivity based on S21 and Fr at a concentration of 0.01 mM were 1746.98 dB/mM and 40 GHz/mM, respectively. Upon comparing the proposed sensor with the original Peano fractal geometry with complementary split-ring (PF-CSRR) sensors without a narrow slot, several parameters were evaluated, including sensitivity, quality factor, operating frequency, active area, and sample volume. The results showed that the proposed sensor exhibited an increased sensitivity of 6.08% and had a 40.72% higher quality factor, while the operating frequency, active area, and sample volume showed decreases of 1.71%, 25%, and 28.27%, respectively. The materials under tests (MUTs) were analyzed and categorized into groups using principal component analysis (PCA) with a K-mean clustering algorithm. The proposed E2 sensor has a compact size and simple structure that can be easily fabricated with low-cost materials. With the small sample volume requirement, fast measurement with a wide dynamic range, and a simple protocol, this proposed sensor can also be applied to measure high E2 levels in environmental, human, and animal samples.

Peroxisomes during postnatal development of mouse endocrine and exocrine pancreas display cell-type- and stage-specific protein composition

Peroxisomal dysfunction unhinges cellular metabolism by causing the accumulation of toxic metabolic intermediates (e.g. reactive oxygen species, very -chain fatty acids, phytanic acid or eicosanoids) and the depletion of important lipid products (e.g. plasmalogens, polyunsaturated fatty acids), leading to various proinflammatory and devastating pathophysiological conditions like metabolic syndrome and age-related diseases including diabetes. Because the peroxisomal antioxidative marker enzyme catalase is low abundant in Langerhans islet cells, peroxisomes were considered scarcely present in the endocrine pancreas. Recently, studies demonstrated that the peroxisomal metabolism is relevant for pancreatic cell functionality. During the postnatal period, significant changes occur in the cell structure and the metabolism to trigger the final maturation of the pancreas, including cell proliferation, regulation of energy metabolism, and activation of signalling pathways. Our aim in this study was to (i) morphometrically analyse the density of peroxisomes in mouse endocrine versus exocrine pancreas and (ii) investigate how the distribution and the abundance of peroxisomal proteins involved in biogenesis, antioxidative defence and fatty acid metabolism change during pancreatic maturation in the postnatal period. Our results prove that endocrine and exocrine pancreatic cells contain high amounts of peroxisomes with heterogeneous protein content indicating that distinct endocrine and exocrine cell types require a specific set of peroxisomal proteins depending on their individual physiological functions. We further show that significant postnatal changes occur in the peroxisomal compartment of different pancreatic cells that are most probably relevant for the metabolic maturation and differentiation of the pancreas during the development from birth to adulthood.

USP7 controls NGN3 stability and pancreatic endocrine lineage development

Understanding the factors and mechanisms involved in beta-cell development will guide therapeutic efforts to generate fully functional beta cells for diabetes. Neurogenin 3 (NGN3) is the key transcription factor that marks endocrine progenitors and drives beta-cell differentiation. Here we screen for binding partners of NGN3 and identify the deubiquitylating enzyme USP7 as a key regulator of NGN3 stability. Mechanistically, USP7 interacts with, deubiquitinates and stabilizes NGN3. In vivo, conditional knockout of Usp7 in the mouse embryonic pancreas causes a dramatic reduction in islet formation and hyperglycemia in adult mice, due to impaired NGN3-mediated endocrine specification during pancreatic development. Furthermore, pharmacological inhibition of USP7 during endocrine specification in human iPSC models of beta-cell differentiation decreases NGN3 expressing progenitor cell numbers and impairs beta cell differentiation. Thus, the USP7-NGN3 axis is an essential mechanism for driving endocrine development and beta-cell differentiation, which can be therapeutically exploited.

Presence of immunogenic alternatively spliced insulin gene product in human pancreatic delta cells

Aims/hypothesisTranscriptome analyses revealed insulin-gene-derived transcripts in non-beta endocrine islet cells. We studied alternative splicing of human INS mRNA in pancreatic islets.MethodsAlternative splicing of insulin pre-mRNA was determined by PCR analysis performed on human islet RNA and single-cell RNA-seq analysis. Antisera were generated to detect insulin variants in human pancreatic tissue using immunohistochemistry, electron microscopy and single-cell western blot to confirm the expression of insulin variants. Cytotoxic T lymphocyte (CTL) activation was determined by MIP-1β release.ResultsWe identified an alternatively spliced INS product. This variant encodes the complete insulin signal peptide and B chain and an alternative C-terminus that largely overlaps with a previously identified defective ribosomal product of INS. Immunohistochemical analysis revealed that the translation product of this INS-derived splice transcript was detectable in somatostatin-producing delta cells but not in beta cells; this was confirmed by light and electron microscopy. Expression of this alternatively spliced INS product activated preproinsulin-specific CTLs in vitro. The exclusive presence of this alternatively spliced INS product in delta cells may be explained by its clearance from beta cells by insulin-degrading enzyme capturing its insulin B chain fragment and a lack of insulin-degrading enzyme expression in delta cells.Conclusions/interpretationOur data demonstrate that delta cells can express an INS product derived from alternative splicing, containing both the diabetogenic insulin signal peptide and B chain, in their secretory granules. We propose that this alternative INS product may play a role in islet autoimmunity and pathology, as well as endocrine or paracrine function or islet development and endocrine destiny, and transdifferentiation between endocrine cells. INS promoter activity is not confined to beta cells and should be used with care when assigning beta cell identity and selectivity.Data availabilityThe full EM dataset is available via www.nanotomy.org (for review: http://www.nanotomy.org/OA/Tienhoven2021SUB/6126-368/). Single-cell RNA-seq data was made available by Segerstolpe et al [13] and can be found at https://sandberglab.se/pancreas. The RNA and protein sequence of INS-splice was uploaded to GenBank (BankIt2546444 INS-splice OM489474).Graphical abstract

IDF2022-0498 PRMT1-mediated arginine methylation of Neurogenin 3 is required for pancreatic endocrine development from hESCs

IDF2022-0498 PRMT1-mediated arginine methylation of Neurogenin 3 is required for pancreatic endocrine development from hESCs

Sex-Dependent Differences in Mouse Placental Gene Expression following a Maternal High-Fat Diet.

In utero fetal exposures may have sex-specific placental gene responses. Our objective was to measure sex-based differences in placental gene expression from dams fed high-fat diet (HFD) versus control diet (CD). We fed timed pregnant Friend virus B-strain dams either a CD (n = 5) or an HFD (n = 5). We euthanized dams on embryonic day 17.5 to collect placentas. We extracted placental RNA and hybridized it to a customized 96-gene Nanostring panel focusing on angiogenic, inflammatory, and growth genes. We compared normalized gene expression between CD and HFD, stratified by fetal sex, using analysis of variance. Pathway analysis was used to further interpret the genomic data. Pups from HFD-fed dams were heavier than those from CD-fed dams (0.97 ± 0.06 vs. 0.84 ± 0.08 g, p < 0.001). Male pups were heavier than females in the HFD (0.99 ± 0.05 vs. 0.94 ± 0.06 g, p = 0.004) but not CD (0.87 ± 0.08 vs. 0.83 ± 0.07 g, p = 0.10) group. No sex-based differences in placental gene expression in CD-fed dams were observed. Among HFD-fed dams, placentas from female pups exhibited upregulation of 15 genes (q = 0.01). Network analyses identified a cluster of genes involved in carbohydrate metabolism, cellular function and maintenance, and endocrine system development and function (p = 1 × 10-23). The observed female-specific increased gene expression following in utero HFD exposure was predicted to be regulated by insulin (p = 5.79 × 10-13). In female compared with male pups, in utero exposure to HFD upregulated placental gene expression in 15 genes predicted to be regulated by insulin. Sex-specific differences in placental expression of these genes should be further investigated. · Male pups were heavier than female pups at the time of sacrifice when dams were fed an HFD.. · HFD was associated with upregulated gene expression in female placentas.. · Female-specific increased gene was predicted to be regulated by insulin..

ВОЗМОЖНОСТИ УЛЬТРАЗВУКОВОЙ ДИАГНОСТИКИ В ПРОГНОСТИЧЕСКОЙ ОЦЕНКЕ ТОЛЩИНЫ ЭПИКАРДИАЛЬНОЙ ЖИРОВОЙ ТКАНИ ПРИ СЕРДЕЧНО-СОСУДИСТОЙ ПАТОЛОГИИ

Recent studies have shown that different visceral fat depots can make a special contribution in development of endocrine and cardiovascular diseases. Epicardial adipose tissue (EAT) is a unique visceral fat depot, closely anatomically and physiologically related to the myocardium. This visceral fat depot has a transcriptome and secret that are very different from fat depots of other localizations. In healthy individuals, the EAT performs a buffer and cardioprotective function, but with the development of pathology, the EAT begins to secrete pro-inflammatory factors, and the synthesis of anti-inflammatory cytokines decreases. This fact is confirmed by numerous studies. A promising area of modern cardiology is the study of the prognostic value of epicardial fat in assessing the risk of cardiovascular disasters and survival of patients with various comorbid conditions. It has been proven, that people even with a normal body mass index have a risk of cardiovascular diseases and complications. People with normal body weight and the presence of visceral obesity have the cardiovascular risk in 2.75 higher, and the risk of death from all causes in 2.08 higher than in people with normal body weight without visceral obesity. Thus, only the use of anthropometric examination methods in practice may be insufficient for the diagnosis of visceral obesity. The most affordable, reproducible, cost-effective method of visualization of epicardial fat in clinic is echocardiography doppler methods. This review is devoted to the possibilities of using ultrasound diagnostics in the assessment of epicardial fat, as well as the possibility of improving the stratification of cardiovascular and metabolic risks based on an additional analysis of the epicardial fat thickness.

Consequences of parental preconceptional irradiation: Endocrine-metabolic pathology in offspring

Epidemiological studies of late effects of preconceptional (prior to conception) radiation exposure are necessary in order to understand the risks of tumor and non-tumor pathology in next generations. Research of possible relation between parental preconceptional exposure and development of endocrine and metabolic disorders in the offspring is one of the topical issues of radiation hygiene. The objective of the study: comparative analysis of endocrine and metabolic pathology in children of the individuals exposed to long term preconceptional external gamma-radiation at the Mayak Production Association – at the first atomic industry facility on Russia. Materials and methods: A retrospective research was performed based on the Ozyorsk Children’s Health Registry. Ozyorsk – the city where Mayak Production Association is the enterprise forming the city. Analysis of endocrine and metabolic disorders was performed for 13880 children of 1949-1973 years of birth who were born and resided in Ozyorsk; parents of 9321 children of this number had accumulated doses of prolonged external gamma-radiation exposure at the production facility prior to conception. A comparative analysis of the structure and incidence of “Endocrine, nutritional and metabolic diseases” and nosological forms of this class of pathology was performed. Characteristic features of parental preconceptional exposure were analyzed in relation to thyroid pathology in children. Relative risk of endocrine and metabolic disorders among the children of exposed and unexposed parents was calculated for the whole disease class and for certain nosologies. Results: A significant predominance of thyroid diseases, particularly, iodine deficiencies, was detected at an earlier age among the offspring of Mayak Production Association workers compared to the control group; these cases were mostly related to low dose parental preconceptional exposure. Thyroid cancer was further detected in 0.16% (15/9321) of the studied cohort of the offspring of exposed parents with average manifestation age of 42.6. Relative risk of thyroid diseases was significantly higher among the exposed parents’ offspring compared to the controls: 2.0 among boys (95% confidence interval 1.38-2.9), 1.59 among girls (1.25-2.02) and in the group as a whole – 1.64 (1.34- 2.01). Relative risk of thyroid diseases related to iodine deficiency among the offspring of Mayak Production Association personnel was almost twice higher than the in the control group: 1.92 (1.3-2.84) among boys, 1.68 (1.29-2.2) among girls, 1.7 (1.36-2.12) in the whole group. Discussion and conclusion: A statistically significant overweight of non-tumor thyroid pathology in the offspring of Mayak Production Association personnel cannot be explained through external reasons taking into account similar climate and geographic conditions, possible technogenic exposure and the same unified medical survey of the followed children. We cannot exclude increased sensibility if thyroid tissue to iodine deficiency and its predisposition to hypertrophy and hyperplasia among the offspring of individuals exposed to long term occupational contact with ionizing radiation sources; that should be taken into account in the course of medical monitoring of these patients. Further research of transgenerational effects in the offspring of personnel of radiation hazardous production facilities are needed for complete understanding of the role of parental occupational exposure in health risks for further generations.

Loss of RREB1 in pancreatic beta cells reduces cellular insulin content and affects endocrine cell gene expression

Aims/hypothesisGenome-wide studies have uncovered multiple independent signals at the RREB1 locus associated with altered type 2 diabetes risk and related glycaemic traits. However, little is known about the function of the zinc finger transcription factor Ras-responsive element binding protein 1 (RREB1) in glucose homeostasis or how changes in its expression and/or function influence diabetes risk.MethodsA zebrafish model lacking rreb1a and rreb1b was used to study the effect of RREB1 loss in vivo. Using transcriptomic and cellular phenotyping of a human beta cell model (EndoC-βH1) and human induced pluripotent stem cell (hiPSC)-derived beta-like cells, we investigated how loss of RREB1 expression and activity affects pancreatic endocrine cell development and function. Ex vivo measurements of human islet function were performed in donor islets from carriers of RREB1 type 2 diabetes risk alleles.ResultsCRISPR/Cas9-mediated loss of rreb1a and rreb1b function in zebrafish supports an in vivo role for the transcription factor in beta cell mass, beta cell insulin expression and glucose levels. Loss of RREB1 also reduced insulin gene expression and cellular insulin content in EndoC-βH1 cells and impaired insulin secretion under prolonged stimulation. Transcriptomic analysis of RREB1 knockdown and knockout EndoC-βH1 cells supports RREB1 as a novel regulator of genes involved in insulin secretion. In vitro differentiation of RREB1KO/KO hiPSCs revealed dysregulation of pro-endocrine cell genes, including RFX family members, suggesting that RREB1 also regulates genes involved in endocrine cell development. Human donor islets from carriers of type 2 diabetes risk alleles in RREB1 have altered glucose-stimulated insulin secretion ex vivo, consistent with a role for RREB1 in regulating islet cell function.Conclusions/interpretationTogether, our results indicate that RREB1 regulates beta cell function by transcriptionally regulating the expression of genes involved in beta cell development and function.Graphical abstract