Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 16.997
Filtrar
1.
Nat Commun ; 11(1): 4912, 2020 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-32999275

RESUMO

Most signals detected by genome-wide association studies map to non-coding sequence and their tissue-specific effects influence transcriptional regulation. However, key tissues and cell-types required for functional inference are absent from large-scale resources. Here we explore the relationship between genetic variants influencing predisposition to type 2 diabetes (T2D) and related glycemic traits, and human pancreatic islet transcription using data from 420 donors. We find: (a) 7741 cis-eQTLs in islets with a replication rate across 44 GTEx tissues between 40% and 73%; (b) marked overlap between islet cis-eQTL signals and active regulatory sequences in islets, with reduced eQTL effect size observed in the stretch enhancers most strongly implicated in GWAS signal location; (c) enrichment of islet cis-eQTL signals with T2D risk variants identified in genome-wide association studies; and (d) colocalization between 47 islet cis-eQTLs and variants influencing T2D or glycemic traits, including DGKB and TCF7L2. Our findings illustrate the advantages of performing functional and regulatory studies in disease relevant tissues.


Assuntos
Glicemia/genética , Diabetes Mellitus Tipo 2/genética , Predisposição Genética para Doença , Ilhotas Pancreáticas/metabolismo , Locos de Características Quantitativas , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Glicemia/metabolismo , Linhagem Celular Tumoral , Estudos de Coortes , Diabetes Mellitus Tipo 2/sangue , Diacilglicerol Quinase/genética , Diacilglicerol Quinase/metabolismo , Elementos Facilitadores Genéticos , Feminino , Regulação da Expressão Gênica , Estudo de Associação Genômica Ampla , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , RNA-Seq , Análise de Sequência de DNA , Proteína 2 Semelhante ao Fator 7 de Transcrição/genética , Proteína 2 Semelhante ao Fator 7 de Transcrição/metabolismo , Adulto Jovem
2.
Adv Exp Med Biol ; 1255: 143-152, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32949397

RESUMO

The pancreas is a complex organ composed of an endocrine (pancreatic islets) and an exocrine portion. This mixed cell population has resulted in an implacable barrier to exploring the detailed mechanism and function of each cell type in previous investigative approaches. In recent years, single-cell RNA sequencing (scRNA-seq) technologies have provided in-depth analysis of cell heterogeneity in the pancreas and in pancreatic cancer. It is especially effective in cell-type-specific molecule identification and detection of interactions between cancer cells and the stromal microenvironment. To date, numerous reports have described the application of scRNA-seq in studies of pancreatic islets and pancreatic cancer. The aim of this paper is to review recent advances of pancreatic transcriptomics and pancreatic cancer using scRNA-seq strategies.


Assuntos
Ilhotas Pancreáticas/metabolismo , Neoplasias Pancreáticas/genética , RNA-Seq , Análise de Célula Única , Humanos , Microambiente Tumoral
3.
Am J Physiol Regul Integr Comp Physiol ; 319(3): R255-R263, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32667834

RESUMO

Fetal conditions associated with placental insufficiency and intrauterine growth restriction (IUGR) chronically elevate plasma norepinephrine (NE) concentrations. Our objective was to evaluate the effects of chronically elevated NE on insulin-stimulated glucose metabolism in normally grown, non-IUGR fetal sheep, which are independent of other IUGR-related reductions in nutrients and oxygen availability. After surgical placement of catheters, near-term fetuses received either a saline (control) or NE intravenous infusion with controlled euglycemia. In NE fetuses, plasma NE concentrations were 5.5-fold greater than controls, and fetal euglycemia was maintained with a maternal insulin infusion. Insulin secretion was blunted in NE fetuses during an intravenous glucose tolerance test. Weight-specific fluxes for glucose were measured during a euinsulinemic-euglycemic clamp (EEC) and a hyperinsulinemic-euglycemic clamp (HEC). Plasma glucose and insulin concentrations were not different between groups within each clamp, but insulin concentrations increased 10-fold between the EEC and the HEC. During the EEC, rates of glucose uptake (umbilical uptake + exogenous infusion) and glucose utilization were 47% and 35% lower (P < 0.05) in NE fetuses compared with controls. During the HEC, rates of glucose uptake were 28% lower (P < 0.05) in NE fetuses than controls. Glucose production was undetectable in either group, and glucose oxidation was unaffected by the NE infusion. These findings indicate that chronic exposure to high plasma NE concentrations lowers rates of net glucose uptake in the fetus without affecting glucose oxidation rates or initiating endogenous glucose production. Lower fetal glucose uptake was independent of insulin, which indicates insulin resistance as a consequence of chronically elevated NE.


Assuntos
Glicemia/metabolismo , Feto/metabolismo , Norepinefrina/sangue , Insuficiência Placentária/metabolismo , Animais , Feminino , Retardo do Crescimento Fetal/metabolismo , Insulina/sangue , Resistência à Insulina/fisiologia , Ilhotas Pancreáticas/metabolismo , Gravidez , Ovinos
4.
Life Sci ; 258: 118145, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32717270

RESUMO

AIMS: Studies on diabetes mellitus have shown that the phosphoinositide 3-kinase (PI3K)/serine threonine kinase (AKT)/forkhead box protein O1 (FoxO1) signaling pathway can regulate insulin secretion by modulating the expression of pancreatic and duodenal homeobox-1 (PDX-1). Therefore, it was hypothesized that the pathway also played an important role in functional abnormalities of pancreatic islets after severe burns. This study aimed to explore the role and mechanism of the PI3K/AKT/FoxO1/PDX-1 signaling pathway in functional changes of pancreatic islets in rats post severe burns. MAIN METHODS: Rats were grouped, subjected to full thickness burn injuries involving 50% total body surface area (TBSA), and injected intraperitoneally with BPV (HOpic) (0.6 mg/kg) or DMSO (0.55 mg/kg) once a day for three days. Glucose metabolism related indexes were measured by the glucometer, transmission electron microscopy (TEM) and enzyme-linked immunosorbent assay (ELISA). PI3K/AKT/FoxO1/PDX-1 signaling pathway related indexes were detected through immunofluorescence, western blot and RT-qPCR analyses. KEY FINDINGS: Dysglycemia and impaired insulin secretion occurred in rats, the activity of the PI3K/AKT/FoxO1/PDX-1 signaling pathway in the islets fell, and PDX-1 was translocated from the nucleus to the cytoplasm post severe burns. When BPV (HOpic) was used, glucose metabolism and insulin secretion were improved, the activity of the PI3K/AKT/FoxO1/PDX-1 signaling pathway in the islets was up-regulated, and PDX-1 was redistributed from the cytoplasm to the nucleus. SIGNIFICANCE: The activity of the PI3K/AKT/FoxO1/PDX-1 signaling pathway declined following severe burns. When its activity was up-regulated, insulin secretion could be improved, thus ameliorating hyperglycemia.


Assuntos
Queimaduras/patologia , Proteínas de Homeodomínio/metabolismo , Ilhotas Pancreáticas/patologia , Proteínas do Tecido Nervoso/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transativadores/metabolismo , Animais , Queimaduras/metabolismo , Secreção de Insulina , Ilhotas Pancreáticas/metabolismo , Masculino , Ratos , Ratos Wistar , Transdução de Sinais
5.
Phys Rev Lett ; 124(20): 208101, 2020 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-32501061

RESUMO

Network flows often exhibit a hierarchical treelike structure that can be attributed to the minimization of dissipation. The common feature of such systems is a single source and multiple sinks (or vice versa). In contrast, here we study networks with only a single source and sink. These systems can arise from secondary purposes of the networks, such as blood sugar regulation through insulin production. Minimization of dissipation in these systems leads to vascular shunting, a single vessel connecting the inlet and outlet. We show instead how optimizing the transport time yields network topologies that match those observed in the insulin-producing pancreatic islets. These are patterns of periphery-to-center and center-to-periphery flows. The obtained flow networks are broadly independent of how the flow velocity depends on the flow flux, but continuous and discontinuous phase transitions appear at extreme flux dependencies. Lastly, we show how constraints on flows can lead to buckling of the branches of the network, a feature that is also observed in pancreatic islets.


Assuntos
Ilhotas Pancreáticas/irrigação sanguínea , Ilhotas Pancreáticas/metabolismo , Modelos Cardiovasculares , Transporte Biológico , Glucagon/sangue , Glucagon/metabolismo , Insulina/sangue , Insulina/metabolismo , Transdução de Sinais
6.
Metabolism ; 109: 154290, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32522488

RESUMO

BACKGROUND: Males absent on the first (Mof) is implicated in gene control of diverse biological processes, such as cell growth, differentiation, apoptosis and autophagy. However, the relationship between glucose regulation and Mof-mediated transcription events remains unexplored. We aimed to unravel the role of Mof in glucose regulation by using global and pancreatic α-cell-specific Mof-deficient mice in vivo and α-TC1-6 cell line in vitro. METHODS: We used tamoxifen-induced temporal Mof-deficient mice first to show Mof regulate glucose homeostasis, islet cell proportions and hormone secretion. Then we used α-cell-specific Mof-deficient mice to clarify how α-cell subsets and ß-cell mass were regulated and corresponding hormone level alterations. Ultimately, we used small interfering RNA (siRNA) to knockdown Mof in α-TC1-6 and unravel the mechanism regulating α-cell mass and glucagon secretion. RESULTS: Mof was mainly expressed in α-cells. Global Mof deficiency led to lower glucose levels, attributed by decreased α/ß-cell ratio and glucagon secretion. α-cell-specific Mof-deficient mice exhibited similar alterations, with more reduced prohormone convertase 2 (PC2)-positive α-cell mass, responsible for less glucagon, and enhanced prohormone convertase 1 (PC1/3)-positive α-cell mass, leading to more glucagon-like peptide-1 (GLP-1) secretion, thus increased ß-cell mass and insulin secretion. In vitro, increased DNA damage, dysregulated autophagy, enhanced apoptosis and altered cell fate factors expressions upon Mof knockdown were observed. Genes and pathways linked to impaired glucagon secretion were uncovered through transcriptome sequencing. CONCLUSION: Mof is a potential interventional target for glucose regulation, from the aspects of both α-cell subset mass and glucagon, intra-islet GLP-1 secretion. Upon Mof deficiency, Up-regulated PC1/3 but down-regulated PC2-positive α-cell mass, leads to more GLP-1 and insulin but less glucagon secretion, and contributed to lower glucose level.


Assuntos
Glicemia/metabolismo , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Células Secretoras de Glucagon/citologia , Glucagon/metabolismo , Histona Acetiltransferases/fisiologia , Homeostase , Animais , Linhagem Celular , Histona Acetiltransferases/deficiência , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Camundongos , Pró-Proteína Convertase 1/metabolismo , Pró-Proteína Convertase 2/metabolismo
7.
PLoS One ; 15(6): e0235052, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32584858

RESUMO

INTRODUCTION: 3D printing is being used more extensively in modern biomedicine. One of the problems is selecting a proper crosslinking method of bioprinted material. Amongst currently used techniques we can distinguish: physical crosslinking (e.g. Ca2+ and Sr2+) and chemical crosslinking-the UV light crosslinking causing the biggest discussion. UV radiation is selectively absorbed by DNA, mainly in the UV-B region but also (to some extent) in UV-A and UV-C regions. DNA excitement results in typical photoproducts. The amount of strand breaks may vary depending on the period of exposition, it can also differ when cells undergo incubation after radiation. AIM: The aim of this study was to show whether and how the time of irradiation with 405 nm and 365 nm wavelengths affect DNA damage in cell lines and micro-organs (pancreatic islets). MATERIALS AND METHODS: The degree of DNA damage caused by different wavelengths of radiation (405 nm and 365 nm) was evaluated by a comet assay. The test was performed on fibroblasts, alpha cells, beta cells and porcine pancreatic islets after 24 hours incubation period. Samples without radiation treatment were selected as a control group. Results analysis consisted of determining the percent of cells with damaged DNA and the tail intensity evaluation. RESULTS: The degree of DNA damage in pancreatic islets after exposure to 405 nm wavelength oscillated between 2% and 6% depending on the tested time period (10 - 300 seconds). However, treating islets using 365 nm wavelength resulted in damage up to 50%. This clearly shows significantly less damage when using 405 nm wavelength. Similar results were obtained for the tested cell lines. CONCLUSIONS: Crosslinking with 405 nm is better for pancreatic islets than crosslinking with 365 nm UV light.


Assuntos
Dano ao DNA , Ilhotas Pancreáticas/metabolismo , Raios Ultravioleta/efeitos adversos , Animais , Linhagem Celular Tumoral , Humanos , Ilhotas Pancreáticas/patologia , Camundongos , Suínos
8.
Nutr Metab Cardiovasc Dis ; 30(7): 1216-1226, 2020 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-32482454

RESUMO

BACKGROUND AND AIMS: Successful islet transplantation as a promising treatment of diabetes type 1 is threatened with the loss of islets during the pre-transplant culture due to hypoxia and oxidative stress-induced apoptosis. Therefore, optimization of culture in order to preserve the islets is a critical point. In this study, we investigated the effect of resveratrol, as a cytoprotective agent, on the cultured human islets. METHODS AND RESULTS: Isolated islets were treated with different concentrations of resveratrol for 24 and 72 h. Islets' viability, apoptosis, apoptosis markers, and insulin and C-peptide secretion, along with the production of reactive oxygen species (ROS), hypoxia inducible factor 1 alpha (HIF-1α), and its target genes in the islets were investigated. Our findings showed that the islets were exposed to hypoxia and oxidative stress after isolation and during culture. This insult induced apoptosis and decreased viability during 72 h. The presence of resveratrol significantly attenuated HIF-1α and ROS production, reduced apoptosis, promoted the VEGF secretion, and increased the insulin and C-peptide secretion. In this regard, resveratrol improved the islet's survival and function in the culture period. CONCLUSIONS: Using resveratrol can attenuate the stressful condition for the islets in the pre-transplant culture and subsequently ameliorate their viability and functionality that lead to successful outcome after clinical transplantation.


Assuntos
Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Ilhotas Pancreáticas/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Resveratrol/farmacologia , Adulto , Idoso , Peptídeo C/metabolismo , Hipóxia Celular , Sobrevivência Celular/efeitos dos fármacos , Citoproteção , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Pessoa de Meia-Idade , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Técnicas de Cultura de Tecidos , Fator A de Crescimento do Endotélio Vascular/metabolismo
9.
Nat Commun ; 11(1): 2742, 2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32488111

RESUMO

Next generation sequencing studies have highlighted discrepancies in ß-cells which exist between mice and men. Numerous reports have identified MAF BZIP Transcription Factor B (MAFB) to be present in human ß-cells postnatally, while its expression is restricted to embryonic and neo-natal ß-cells in mice. Using CRISPR/Cas9-mediated gene editing, coupled with endocrine cell differentiation strategies, we dissect the contribution of MAFB to ß-cell development and function specifically in humans. Here we report that MAFB knockout hPSCs have normal pancreatic differentiation capacity up to the progenitor stage, but favor somatostatin- and pancreatic polypeptide-positive cells at the expense of insulin- and glucagon-producing cells during endocrine cell development. Our results describe a requirement for MAFB late in the human pancreatic developmental program and identify it as a distinguishing transcription factor within islet cell subtype specification. We propose that hPSCs represent a powerful tool to model human pancreatic endocrine development and associated disease pathophysiology.


Assuntos
Células Secretoras de Insulina/metabolismo , Fator de Transcrição MafB/genética , Fator de Transcrição MafB/metabolismo , Células Estreladas do Pâncreas/metabolismo , Animais , Sistemas CRISPR-Cas , Diferenciação Celular , Feminino , Edição de Genes , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Silenciamento de Genes , Glucagon/metabolismo , Células Secretoras de Glucagon/metabolismo , Humanos , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Células-Tronco , Transcriptoma
10.
Ecotoxicol Environ Saf ; 201: 110802, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32531573

RESUMO

Extended exposure to inorganic arsenic through contaminated drinking water has been linked with increased incidence of diabetes mellitus. The most common exposure occurs through the consumption of contaminated drinking water mainly through geogenic sources of inorganic arsenic. Epigenetic modifications are important mechanisms through which environmental pollutants could exert their toxic effects. Bisulfite sequencing polymerase chain reaction method followed by Sanger sequencing was performed for DNA methylation analysis. Our results showed that sodium arsenite treatment significantly reduced insulin secretion in pancreatic islets. It was revealed that the methylation of glucose transporter 2 (Glut2) gene was changed at two cytosine-phosphate-guanine (CpG) sites (-1743, -1734) in the promoter region of the sodium arsenite-treated group comparing to the control. No changes were observed in the methylation status of peroxisome proliferator-activated receptor-gamma (PPARγ), pancreatic and duodenal homeobox 1 (Pdx1) and insulin 2 (Ins2) CpG sites in the targeted regions. Measuring the gene expression level showed increase in Glut2 expression, while the expression of insulin (INS) and Pdx1 were significantly affected by sodium arsenite treatment. This study revealed that exposure to sodium arsenite changed the DNA methylation pattern of Glut2, a key transporter of glucose entry into the pancreatic beta cells (ß-cells). Our data suggested possible epigenetic-mediated toxicity mechanism for arsenite-induced ß-cells dysfunction. Further studies are needed to dissect the precise epigenetic modulatory activity of sodium arsenite that affect the biogenesis of insulin.


Assuntos
Arsenitos/toxicidade , Metilação de DNA/efeitos dos fármacos , Transportador de Glucose Tipo 2/genética , Insulina/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Compostos de Sódio/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Epigênese Genética/efeitos dos fármacos , Proteínas de Homeodomínio/genética , Técnicas In Vitro , Insulina/genética , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Masculino , Regiões Promotoras Genéticas , Ratos , Ratos Wistar , Transativadores/genética
11.
Am J Physiol Gastrointest Liver Physiol ; 319(1): G36-G42, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32463335

RESUMO

After 50% proximal small bowel resection (SBR) in mice, we have demonstrated hepatic steatosis, impaired glucose metabolism without insulin resistance, and increased pancreatic islet area. We sought to determine the consequences of SBR on pancreatic ß-cell morphology, proliferation, and expression of a key regulatory hormone, glucagon-like peptide-1 (GLP-1). C57BL/6 mice underwent 50% SBR or sham operation. At 10 wk, pancreatic insulin content and secretion was measured by ELISA. Immunohistochemistry was performed to determine structural alterations in pancreatic α-and ß-cells. Western blot analysis was used to measure GLP-1R expression, and immunoassay was used to measure plasma insulin and GLP-1. Experiments were repeated by administering a GLP-1 agonist (exendin-4) to a cohort of mice following SBR. After SBR, there was pancreatic islet hypertrophy and impaired glucose tolerance. The proportion of α and ß cells was not grossly altered. Whole pancreas and pancreatic islet insulin content was not significantly different; however, SBR mice demonstrated decreased insulin secretion in both static incubation and islet perfusion experiments. The expression of pancreatic GLP-1R was decreased approximately twofold after SBR, compared with sham and serum GLP-1, was decreased. These metabolic derangements were mitigated after administration of the GLP-1 agonist. Following massive SBR, there is significant hypertrophy of pancreatic islet cells with morphologically intact α- and ß-cells. Significantly reduced pancreatic insulin release in both static and dynamic conditions demonstrate a perturbed second phase of insulin secretion. GLP-1 is a key mediator of this amplification pathway. Decreased expression of serum GLP-1 and pancreatic GLP-1R in face of no change in insulin content presents a novel pathway for enteropancreatic glucose regulation following SBR.NEW & NOTEWORTHY Metabolic changes occur following intestinal resection; however, the effects on pancreatic function are unknown. Prior studies have demonstrated that glucagon-like protein-1 (GLP-1) signaling is a crucial player in the improved insulin sensitivity after bariatric surgery. In this study, we explore the effect of massive small bowel resection on gut hormone physiology and provide novel insights into the enteropancreatic axis.


Assuntos
Peptídeo 1 Semelhante ao Glucagon/metabolismo , Intestinos/lesões , Ilhotas Pancreáticas/metabolismo , Pâncreas/metabolismo , Animais , Glucagon/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/metabolismo , Insulina/sangue , Células Secretoras de Insulina/metabolismo , Camundongos Endogâmicos C57BL , Pâncreas Exócrino/metabolismo
12.
Am J Hum Genet ; 106(6): 846-858, 2020 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-32470372

RESUMO

The burden of several common diseases including obesity, diabetes, hypertension, asthma, and depression is increasing in most world populations. However, the mechanisms underlying the numerous epidemiological and genetic correlations among these disorders remain largely unknown. We investigated whether common polymorphic inversions underlie the shared genetic influence of these disorders. We performed an inversion association analysis including 21 inversions and 25 obesity-related traits on a total of 408,898 Europeans and validated the results in 67,299 independent individuals. Seven inversions were associated with multiple diseases while inversions at 8p23.1, 16p11.2, and 11q13.2 were strongly associated with the co-occurrence of obesity with other common diseases. Transcriptome analysis across numerous tissues revealed strong candidate genes for obesity-related traits. Analyses in human pancreatic islets indicated the potential mechanism of inversions in the susceptibility of diabetes by disrupting the cis-regulatory effect of SNPs from their target genes. Our data underscore the role of inversions as major genetic contributors to the joint susceptibility to common complex diseases.


Assuntos
Inversão Cromossômica/genética , Diabetes Mellitus/genética , Predisposição Genética para Doença , Hipertensão/genética , Obesidade/complicações , Obesidade/genética , Polimorfismo Genético , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Cromossomos Humanos Par 16/genética , Cromossomos Humanos Par 8/genética , Conjuntos de Dados como Assunto/normas , Diabetes Mellitus/patologia , Europa (Continente)/etnologia , Feminino , Perfilação da Expressão Gênica , Haplótipos , Humanos , Hipertensão/complicações , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único/genética , Reprodutibilidade dos Testes , Adulto Jovem
14.
Nat Commun ; 11(1): 2338, 2020 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-32393754

RESUMO

Single-cell RNA sequencing (scRNA-seq) can characterize cell types and states through unsupervised clustering, but the ever increasing number of cells and batch effect impose computational challenges. We present DESC, an unsupervised deep embedding algorithm that clusters scRNA-seq data by iteratively optimizing a clustering objective function. Through iterative self-learning, DESC gradually removes batch effects, as long as technical differences across batches are smaller than true biological variations. As a soft clustering algorithm, cluster assignment probabilities from DESC are biologically interpretable and can reveal both discrete and pseudotemporal structure of cells. Comprehensive evaluations show that DESC offers a proper balance of clustering accuracy and stability, has a small footprint on memory, does not explicitly require batch information for batch effect removal, and can utilize GPU when available. As the scale of single-cell studies continues to grow, we believe DESC will offer a valuable tool for biomedical researchers to disentangle complex cellular heterogeneity.


Assuntos
Análise por Conglomerados , Aprendizado Profundo , RNA-Seq , Análise de Célula Única , Algoritmos , Animais , Medula Óssea/metabolismo , Regulação da Expressão Gênica , Humanos , Ilhotas Pancreáticas/metabolismo , Leucócitos Mononucleares/metabolismo , Macaca , Camundongos , Monócitos/metabolismo , Retina/metabolismo
15.
Gene ; 753: 144785, 2020 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-32445922

RESUMO

OBJECTIVE: Type 1 diabetes onset is preceded by a pre-inflammatory stage leading to insulitis and followed by targeted destruction of the insulin-producing beta cells of the pancreas. Osteopontin (OPN) is a secreted phosphoprotein with cytokine properties, implicated in many physiological and pathological processes, including infection and autoimmunity. We have previously identified up-regulated osteopontin transcripts in the pancreatic lymph nodes of the NOD (Non-Obese Diabetic) mouse at the pre-diabetic stages. Investigating the underlined disease initiating mechanisms may well contribute to the development of novel preventive therapies. Our aim was to construct opn null mice in a NOD autoimmune-prone genetic background and address the pathogenic or protective role of the osteopontin molecule in the early stages of type 1 diabetes. METHODS: We generated opn null mutant mice in a NOD genetic background by serial backcrossing to the existing C57BL/6 opn knockout strain. The presence of opn wild type or null alleles in the congenic lines was evaluated by PCR amplification. We used NOD opn-null mice to assess the phenotypic evolution of type 1 diabetes. The presence of OPN in the serum was evaluated by ELISA and by immunostaining on the mouse tissues. The primary gene structure of the NOD opn encoding gene and protein sequences were compared to the known alleles of other mouse strains. Evaluation of Single Nucleotide Polymorphisms (SNPs) variation between opn alleles of the opn gene is reported. RESULTS: In the absence of OPN, type 1 diabetes is accelerated, suggesting a protective role of this cytokine on the insulin-producing cells of the pancreatic islets. Conversely, in the presence of the opn gene, an increase of the OPN protein in the serum of young NOD mice indicates that this molecule might be involved in the immune regulatory events taking place at early stages, prior to disease onset. Our data support that OPN acts as a positive regulator of the early islet autoimmune damage, possibly by a shift of the steady-state of T1D pathogenesis. We report that the OPN protein structure of the NOD/ShiLtJ strain corresponds to the a-type allele of the osteopontin gene. Comparative analysis of the single nucleotide polymorphisms between the a-type and b-type alleles indicates that the majority of variations are within the non-coding regions of the gene. CONCLUSIONS: The construction of opn null mice in an autoimmune genetic background (NOD.B6.Cg-spp1-/-) provides important tools for the study of the implication of the OPN in type 1 diabetes, offering the possibility to address the significance of this molecule as an early marker of the disease and as a therapeutic agent in preclinical studies.


Assuntos
Diabetes Mellitus Tipo 1/genética , Osteopontina/genética , Alelos , Animais , Modelos Animais de Doenças , Regulação da Expressão Gênica/genética , Frequência do Gene/genética , Predisposição Genética para Doença/genética , Insulina/metabolismo , Células Secretoras de Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Pâncreas/metabolismo , Polimorfismo de Nucleotídeo Único/genética
16.
Chemistry ; 26(34): 7609-7621, 2020 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-32259327

RESUMO

The most common denominator of many of the neurodegenerative diseases is badly folded protein accumulation, which results in the formation of insoluble protein deposits located in different parts of the organism, causing cell death and tissue degeneration. Dendritic systems have turned out to be a promising new therapeutic approach for the treatment of these diseases due to their ability to modulate the folding of these proteins. With this perspective, and focused on type 2 diabetes (T2D), characterized by the presence of deposits containing the amyloidogenic islet amyloid polypeptide (IAPP), we demonstrate how different topologies of cationic carbosilane dendrimers inhibit the formation of insoluble protein deposits in pancreatic islets isolated from transgenic Tg-hIAPP mice. Also, the results obtained by the modification of dendritic carbosilane wedges with the chemical chaperone 4-phenylbutyric acid (4-PBA) at the focal point confirmed their potential as anti-amyloid agents with a concentration efficiency in their therapeutic action five orders of magnitude lower than that observed for free 4-PBA. Computational studies, which determined the main interaction between IAPP and dendrimers at the atomic level, support the experimental work.


Assuntos
Amiloidose/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Polipeptídeo Amiloide das Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/química , Fenilbutiratos/química , Silanos/química , Animais , Humanos , Polipeptídeo Amiloide das Ilhotas Pancreáticas/química , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/metabolismo , Camundongos , Camundongos Transgênicos
17.
Nat Commun ; 11(1): 1950, 2020 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-32327658

RESUMO

BDNF signaling in hypothalamic circuitries regulates mammalian food intake. However, whether BDNF exerts metabolic effects on peripheral organs is currently unknown. Here, we show that the BDNF receptor TrkB.T1 is expressed by pancreatic ß-cells where it regulates insulin release. Mice lacking TrkB.T1 show impaired glucose tolerance and insulin secretion. ß-cell BDNF-TrkB.T1 signaling triggers calcium release from intracellular stores, increasing glucose-induced insulin secretion. Additionally, BDNF is secreted by skeletal muscle and muscle-specific BDNF knockout phenocopies the ß-cell TrkB.T1 deletion metabolic impairments. The finding that BDNF is also secreted by differentiated human muscle cells and induces insulin secretion in human islets via TrkB.T1 identifies a new regulatory function of BDNF on metabolism that is independent of CNS activity. Our data suggest that muscle-derived BDNF may be a key factor mediating increased glucose metabolism in response to exercise, with implications for the treatment of diabetes and related metabolic diseases.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Secreção de Insulina , Células Secretoras de Insulina/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Animais , Fator Neurotrófico Derivado do Encéfalo/genética , Cálcio/metabolismo , Células Cultivadas , Glucose/metabolismo , Intolerância à Glucose , Humanos , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Camundongos Knockout , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Receptor trkB/química , Receptor trkB/genética , Receptor trkB/metabolismo , Transdução de Sinais
18.
Nature ; 581(7808): 303-309, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32214235

RESUMO

Single-cell analysis is a valuable tool for dissecting cellular heterogeneity in complex systems1. However, a comprehensive single-cell atlas has not been achieved for humans. Here we use single-cell mRNA sequencing to determine the cell-type composition of all major human organs and construct a scheme for the human cell landscape (HCL). We have uncovered a single-cell hierarchy for many tissues that have not been well characterized. We established a 'single-cell HCL analysis' pipeline that helps to define human cell identity. Finally, we performed a single-cell comparative analysis of landscapes from human and mouse to identify conserved genetic networks. We found that stem and progenitor cells exhibit strong transcriptomic stochasticity, whereas differentiated cells are more distinct. Our results provide a useful resource for the study of human biology.


Assuntos
Células/citologia , Células/metabolismo , Análise de Célula Única/métodos , Adulto , Animais , Grupo com Ancestrais do Continente Asiático , Diferenciação Celular , Linhagem Celular , Separação Celular , China , Bases de Dados Factuais , Corpos Embrioides/citologia , Corpos Embrioides/metabolismo , Grupos Étnicos , Feto/citologia , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Humanos , Imunidade , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/metabolismo , Camundongos , Especificidade de Órgãos , RNA Mensageiro/análise , RNA Mensageiro/genética , Análise de Sequência de RNA , Análise de Célula Única/instrumentação , Processos Estocásticos
19.
Life Sci ; 247: 117458, 2020 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-32092333

RESUMO

AIMS: The use of natural agents with anti-diabetic effect in combination therapy adds further positive clinical implications in the management of diabetes mellitus. Interestingly, quercetin is one of the most potent naturally occurring antioxidant which possesses various pharmacological actions including anti-diabetic effect. Thus, this research was conducted to assess the efficiency of a new combination from gliclazide and quercetin on glycemic control as well as pancreatic islets and beta cells in STZ-experimental model of diabetes. MAIN METHODS: Diabetes has been induced by a single intraperitoneal injection of streptozotocin (STZ; 45 mg/kg) in adult male Wistar rats. For 3 consecutive weeks, diabetic rats were given orally either gliclazide (10 mg/kg), quercetin (50 mg/kg), or their combination. At the end of the experiment, histological, immunohistochemical and morphometrical examination of pancreatic tissues was performed. Furthermore, the changes in glucose metabolism, lipid profile, oxidative and inflammatory status were evaluated. KEY FINDINGS: Treatment with gliclazide alone decreased serum glucose, total cholesterol, triglycerides, malondialdehyde, tumor necrosis factor-alpha and nuclear factor kappa-Beta while increased serum C-peptide, superoxide dismutase, reduced glutathione and adiponectin levels. Combined administration of quercetin with gliclazide markedly augmented serum superoxide dismutase and reduced glutathione more than gliclazide alone and normalized all the above-mentioned parameters. Besides, this combination therapy restored immunostaining intensity, number of pancreatic islets and beta cells along with its area and perimeter. SIGNIFICANCE: Based on the aforementioned results, this combination could be considered a promising one in diabetes mellitus management.


Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Gliclazida/uso terapêutico , Hipoglicemiantes/uso terapêutico , Quercetina/uso terapêutico , Estreptozocina/metabolismo , Animais , Glicemia/metabolismo , Peptídeo C/metabolismo , Colesterol/metabolismo , Quimioterapia Combinada/métodos , Glutationa/metabolismo , Inflamação/metabolismo , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Masculino , Malondialdeído/metabolismo , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Pâncreas/efeitos dos fármacos , Ratos , Ratos Wistar , Superóxido Dismutase/metabolismo , Triglicerídeos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
20.
Transplantation ; 104(6): e144-e150, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32080160

RESUMO

BACKGROUND: Current histological methods cannot accurately determine the survival rate of human pancreatic islets following portal vein infusion. This is due, in part, to the low number of infused islets relative to the whole liver. In this study, we assessed the ability of confocal laser scanning microscopy (CLSM) to track human islets posttransplantation. METHODS: Immunodeficient mice were transplanted with human islets. Following engraftment, animals were euthanized, livers procured, and human islet ß cells immunofluorescently labeled with an insulin-specific antibody and evaluated by CLSM. A calibration curve comparing the area of insulin + hepatic islet ß cells to the number of human islets collected was developed. Levels of human C-peptide were measured in transplant recipients to determine islet function. RESULTS: The short-term survival rate of islet transplants was defined as y = 0.0422x + 2.7008, in which x is human islet number and y is liver islet ß cell area. Employing CLSM, human islets were detected in immunofluorescent labeled murine liver tissue sections posttransplantation. The ß cell-relative area of human islets in 500 islet equivalent (IEQ) specimens was 20.21 ± 1.16 mm and in 1000 IEQ specimens 39.4 ± 2.23 mm posttransplantation. Human islet posttransplant survival rates were 82.9 ± 5.50% (500 IEQ group) and 86.9 ± 5.28% (1000 IEQ group). CONCLUSIONS: These data indicate that CLSM can be employed to quantify and characterize pancreatic human islets after transplantation to murine livers.


Assuntos
Rejeição de Enxerto/diagnóstico , Microscopia Intravital/métodos , Transplante das Ilhotas Pancreáticas/efeitos adversos , Ilhotas Pancreáticas/diagnóstico por imagem , Microscopia Confocal/métodos , Animais , Peptídeo C/análise , Peptídeo C/metabolismo , Estudos de Viabilidade , Rejeição de Enxerto/etiologia , Sobrevivência de Enxerto , Humanos , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Modelos Animais , Transplante Heterólogo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA