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1.
Am J Chin Med ; 48(6): 1435-1454, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32907363

RESUMO

Endoplasmic reticulum stress (ER stress) plays a main role in pancreatic [Formula: see text]-cell dysfunction and death because of intracellular Ca[Formula: see text] turbulence and inflammation activation. Although several drugs are targeting pancreatic [Formula: see text]-cell to improve [Formula: see text]-cell function, there still lacks agents to alleviate [Formula: see text]-cell ER stress conditions. Therefore we used thapsigargin (THAP) or high glucose (HG) to induce ER stress in [Formula: see text]-cell and aimed to screen natural molecules against ER stress-induced [Formula: see text]-cell dysfunction. Through screening the Traditional Chinese drug library ([Formula: see text] molecules), luteolin was finally discovered to improve [Formula: see text]-cell function. Cellular viability results indicated luteolin reduced the THAP or HG-induced [Formula: see text]-cell death and apoptosis through MTT and flow cytometry assay. Moreover, luteolin improved [Formula: see text]-cell insulin secretion ability under ER stress conditions. Also ER stress-induced intracellular Ca[Formula: see text] turbulence and inflammation activation were inhibited by luteolin treatment. Mechanically, luteolin inhibited HNF4[Formula: see text] signaling, which was induced by ER stress. Moreover, luteolin reduced the transcriptional level of HNF4[Formula: see text] downstream gene, such as Asnk4b and HNF1[Formula: see text]. Conversely HNF4[Formula: see text] knockdown abolished the effect of luteolin on [Formula: see text]-cell using siRNA. These results suggested the protective effect of luteolin on [Formula: see text]-cell was through HNF4[Formula: see text]/Asnk4b pathway. In conclusion, our study discovered that luteolin improved [Formula: see text]-cell function and disclosed the underlying mechanism of luteolin on [Formula: see text]-cell, suggesting luteolin is a promising agent against pancreatic dysfunction.


Assuntos
Sobrevivência Celular/efeitos dos fármacos , Medicamentos de Ervas Chinesas/química , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Estresse do Retículo Endoplasmático/genética , Fator 4 Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/metabolismo , Células Secretoras de Insulina/patologia , Células Secretoras de Insulina/fisiologia , Luteolina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Tapsigargina/efeitos adversos , Apoptose/efeitos dos fármacos , Cálcio/metabolismo , Morte Celular/efeitos dos fármacos , Células Cultivadas , Avaliação Pré-Clínica de Medicamentos , Estresse do Retículo Endoplasmático/fisiologia , Glucose/efeitos adversos , Células Secretoras de Insulina/metabolismo , Luteolina/isolamento & purificação
2.
PLoS One ; 15(8): e0237667, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32833960

RESUMO

BACKGROUND AND AIMS: This is the first time that obesity and diabetes mellitus (DM) as protein conformational diseases (PCD) are reported in children and they are typically diagnosed too late, when ß-cell damage is evident. Here we wanted to investigate the level of naturally-ocurring or real (not synthetic) oligomeric aggregates of the human islet amyloid polypeptide (hIAPP) that we called RIAO in sera of pediatric patients with obesity and diabetes. We aimed to reduce the gap between basic biomedical research, clinical practice-health decision making and to explore whether RIAO work as a potential biomarker of early ß-cell damage. MATERIALS AND METHODS: We performed a multicentric collaborative, cross-sectional, analytical, ambispective and blinded study; the RIAO from pretreated samples (PTS) of sera of 146 pediatric patients with obesity or DM and 16 healthy children, were isolated, measured by sound indirect ELISA with novel anti-hIAPP cytotoxic oligomers polyclonal antibody (MEX1). We carried out morphological and functional studied and cluster-clinical data driven analysis. RESULTS: We demonstrated by western blot, Transmission Electron Microscopy and cell viability experiments that RIAO circulate in the blood and can be measured by ELISA; are elevated in serum of childhood obesity and diabetes; are neurotoxics and works as biomarkers of early ß-cell failure. We explored the range of evidence-based medicine clusters that included the RIAO level, which allowed us to classify and stratify the obesity patients with high cardiometabolic risk. CONCLUSIONS: RIAO level increases as the number of complications rises; RIAOs > 3.35 µg/ml is a predictor of changes in the current indicators of ß-cell damage. We proposed a novel physio-pathological pathway and shows that PCD affect not only elderly patients but also children. Here we reduced the gap between basic biomedical research, clinical practice and health decision making.


Assuntos
Diabetes Mellitus Tipo 1/patologia , Diabetes Mellitus Tipo 2/patologia , Células Secretoras de Insulina/patologia , Polipeptídeo Amiloide das Ilhotas Pancreáticas/metabolismo , Obesidade/patologia , Estrutura Quaternária de Proteína , Adolescente , Animais , Linhagem Celular , Sobrevivência Celular , Células Cultivadas , Criança , Pré-Escolar , Estudos Transversais , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/complicações , Humanos , Polipeptídeo Amiloide das Ilhotas Pancreáticas/sangue , Polipeptídeo Amiloide das Ilhotas Pancreáticas/toxicidade , Polipeptídeo Amiloide das Ilhotas Pancreáticas/ultraestrutura , Microscopia Eletrônica de Transmissão , Neurônios/efeitos dos fármacos , Obesidade/sangue , Obesidade/complicações , Projetos Piloto , Cultura Primária de Células , Multimerização Proteica , Ratos , Testes de Toxicidade Aguda
3.
PLoS One ; 15(8): e0237669, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32810137

RESUMO

Pancreatic beta cell death is a hallmark of type 1 and 2 diabetes (T1D/T2D), but the underlying molecular mechanisms are incompletely understood. Key proteins of the DNA damage response (DDR), including tumor protein P53 (P53, also known as TP53 or TRP53 in rodents) and Ataxia Telangiectasia Mutated (ATM), a kinase known to act upstream of P53, have been associated with T2D. Here we test and compare the effect of ATM and P53 ablation on beta cell survival in the rat beta cell line Ins1E. We demonstrate that ATM and P53 differentially regulate beta cell apoptosis induced upon fundamentally different types of diabetogenic beta cell stress, including DNA damage, inflammation, lipotoxicity and endoplasmic reticulum (ER) stress. DNA damage induced apoptosis by treatment with the commonly used diabetogenic agent streptozotocin (STZ) is regulated by both ATM and P53. We show that ATM is a key STZ induced activator of P53 and that amelioration of STZ induced cell death by inhibition of ATM mainly depends on P53. While both P53 and ATM control lipotoxic beta cell apoptosis, ATM but not P53 fails to alter inflammatory beta cell death. In contrast, tunicamycin induced (ER stress associated) apoptosis is further increased by ATM knockdown or inhibition, but not by P53 knockdown. Our results reveal differential roles for P53 and ATM in beta cell survival in vitro in the context of four key pathophysiological types of diabetogenic beta cell stress, and indicate that ATM can use P53 independent signaling pathways to modify beta cell survival, dependent on the cellular insult.


Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Sobrevivência Celular/genética , Células Secretoras de Insulina/patologia , Transdução de Sinais/genética , Proteína Supressora de Tumor p53/metabolismo , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Proteínas Mutadas de Ataxia Telangiectasia/genética , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Diabetes Mellitus/patologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Estresse do Retículo Endoplasmático/genética , Técnicas de Silenciamento de Genes , Humanos , Células Secretoras de Insulina/efeitos dos fármacos , Inibidores de Proteases/farmacologia , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , RNA Interferente Pequeno/metabolismo , Ratos , Estreptozocina/toxicidade , Tunicamicina/toxicidade
4.
Diab Vasc Dis Res ; 17(4): 1479164120945675, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32722929

RESUMO

Activation of the prostaglandin E2 receptor EP4 alters polarization of adipose tissue macrophages towards the anti-inflammatory M2 phenotype to suppress chronic inflammation. However, the role of EP4 signalling in pancreatic macrophages that affect insulin secretion is unclear. We examined the role of EP4 signalling in islet inflammation in vitro and in vivo. Obese diabetic db/db mice were treated with an EP4-selective agonist or vehicle for 4 weeks. Islet morphology did not significantly differ and glucose-stimulated insulin secretion was increased, whereas the pancreatic M1/M2 ratio was decreased in the EP4 agonist-treated group compared to the vehicle group. Because EP4 activation in MIN6 cells did not affect insulin secretion, we used a MIN6/macrophage co-culture system to evaluate the role of EP4 signalling in islet inflammation and subsequent inhibition of insulin release. Co-culture with M1-polarized macrophages markedly suppressed insulin expression in MIN6 cells; however, modulation of M1 polarization by the EP4 agonist significantly reversed the negative impact of co-cultivation on insulin production. The enhanced expression levels of pro-inflammatory cytokines in co-cultured MIN6 cells were markedly inhibited by EP4 agonist treatment of M1 macrophages. Thus, EP4 activation may suppress islet inflammation and protect ß-cell function by altering inflammatory macrophages in the diabetic pancreas.


Assuntos
Plasticidade Celular , Diabetes Mellitus Tipo 2/metabolismo , Inflamação/metabolismo , Células Secretoras de Insulina/metabolismo , Macrófagos Peritoneais/metabolismo , Obesidade/metabolismo , Receptores de Prostaglandina E Subtipo EP4/metabolismo , Animais , Linhagem Celular Tumoral , Técnicas de Cocultura , Citocinas/metabolismo , Diabetes Mellitus Tipo 2/patologia , Modelos Animais de Doenças , Inflamação/patologia , Mediadores da Inflamação/metabolismo , Insulina/metabolismo , Células Secretoras de Insulina/patologia , Ativação de Macrófagos , Macrófagos Peritoneais/patologia , Camundongos , Obesidade/patologia , Fenótipo , Via Secretória , Transdução de Sinais
5.
BMC Med Genet ; 21(1): 91, 2020 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-32375679

RESUMO

BACKGROUND: Renal hypouricemia (RHUC) is a hereditary disorder where mutations in SLC22A12 gene and SLC2A9 gene cause RHUC type 1 (RHUC1) and RHUC type 2 (RHUC2), respectively. These genes regulate renal tubular reabsorption of urates while there exist other genes counterbalancing the net excretion of urates including ABCG2 and SLC17A1. Urate metabolism is tightly interconnected with glucose metabolism, and SLC2A9 gene may be involved in insulin secretion from pancreatic ß-cells. On the other hand, a myriad of genes are responsible for the impaired insulin secretion independently of urate metabolism. CASE PRESENTATION: We describe a 67 year-old Japanese man who manifested severe hypouricemia (0.7 mg/dl (3.8-7.0 mg/dl), 41.6 µmol/l (226-416 µmol/l)) and diabetes with impaired insulin secretion. His high urinary fractional excretion of urate (65.5%) and low urinary C-peptide excretion (25.7 µg/day) were compatible with the diagnosis of RHUC and impaired insulin secretion, respectively. Considering the fact that metabolic pathways regulating urates and glucose are closely interconnected, we attempted to delineate the genetic basis of the hypouricemia and the insulin secretion defect observed in this patient using whole exome sequencing. Intriguingly, we found homozygous Trp258* mutations in SLC22A12 gene causing RHUC1 while concurrent mutations reported to be associated with hyperuricemia were also discovered including ABCG2 (Gln141Lys) and SLC17A1 (Thr269Ile). SLC2A9, that also facilitates glucose transport, has been implicated to enhance insulin secretion, however, the non-synonymous mutations found in SLC2A9 gene of this patient were not dysfunctional variants. Therefore, we embarked on a search for causal mutations for his impaired insulin secretion, resulting in identification of multiple mutations in HNF1A gene (MODY3) as well as other genes that play roles in pancreatic ß-cells. Among them, the Leu80fs in the homeobox gene NKX6.1 was an unreported mutation. CONCLUSION: We found a case of RHUC1 carrying mutations in SLC22A12 gene accompanied with compensatory mutations associated with hyperuricemia, representing the first report showing coexistence of the mutations with opposed potential to regulate urate concentrations. On the other hand, independent gene mutations may be responsible for his impaired insulin secretion, which contains novel mutations in key genes in the pancreatic ß-cell functions that deserve further scrutiny.


Assuntos
Complicações do Diabetes/genética , Proteínas Facilitadoras de Transporte de Glucose/genética , Transportadores de Ânions Orgânicos/genética , Proteínas de Transporte de Cátions Orgânicos/genética , Erros Inatos do Transporte Tubular Renal/genética , Cálculos Urinários/genética , Idoso , Complicações do Diabetes/complicações , Complicações do Diabetes/patologia , Glucose/metabolismo , Fator 1-alfa Nuclear de Hepatócito/genética , Heterozigoto , Proteínas de Homeodomínio/genética , Homozigoto , Humanos , Insulina/biossíntese , Insulina/genética , Secreção de Insulina/genética , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Masculino , Mutação/genética , Erros Inatos do Transporte Tubular Renal/complicações , Erros Inatos do Transporte Tubular Renal/patologia , Ácido Úrico/metabolismo , Cálculos Urinários/complicações , Cálculos Urinários/patologia , Sequenciamento Completo do Exoma
6.
Nat Commun ; 11(1): 2538, 2020 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-32439909

RESUMO

Compromised ß-cell identity is emerging as an important contributor to ß-cell failure in diabetes; however, the precise mechanism independent of hyperglycemia is under investigation. We have previously reported that mTORC1/Raptor regulates functional maturation in ß-cells. In the present study, we find that diabetic ß-cell specific Raptor-deficient mice (ßRapKOGFP) show reduced ß-cell mass, loss of ß-cell identity and acquisition of α-cell features; which are not reversible upon glucose normalization. Deletion of Raptor directly impairs ß-cell identity, mitochondrial metabolic coupling and protein synthetic activity, leading to ß-cell failure. Moreover, loss of Raptor activates α-cell transcription factor MafB (via modulating C/EBPß isoform ratio) and several α-cell enriched genes i.e. Etv1 and Tspan12, thus initiates ß- to α-cell reprograming. The present findings highlight mTORC1 as a metabolic rheostat for stabilizing ß-cell identity and repressing α-cell program at normoglycemic level, which might present therapeutic opportunities for treatment of diabetes.


Assuntos
Diferenciação Celular , Plasticidade Celular , Diabetes Mellitus/patologia , Células Secretoras de Insulina/patologia , Proteína Regulatória Associada a mTOR/metabolismo , Animais , Glicemia/metabolismo , Diferenciação Celular/genética , Plasticidade Celular/genética , Diabetes Mellitus/genética , Diabetes Mellitus/metabolismo , Regulação da Expressão Gênica , Células Secretoras de Glucagon/metabolismo , Células Secretoras de Glucagon/patologia , Insulina/metabolismo , Células Secretoras de Insulina/metabolismo , Fator de Transcrição MafB/genética , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos , Camundongos Knockout , Proteína Regulatória Associada a mTOR/genética , Transdução de Sinais
8.
Life Sci ; 255: 117724, 2020 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-32360624

RESUMO

AIMS: Type 1 diabetes (T1D) is the most common autoimmune disease that affects a global scale. Accumulating evidence has indicated, nuclear factor kappa B (NF-κB) and some microRNAs (miRNAs) as important biomarkers participating in the development of T1D. Thus, we aimed to determine the role of NF-κB and miR-150 in the development of T1D and to unravel the molecular mechanism. MAIN METHODS: Non-obese diabetic mice were used for the T1D model establishment by injecting with streptozotocin. Besides, pancreatic islet ß cells, separated from T1D mice, were induced by interferon-γ and tumor necrosis factor-α for 3 days to mimic T1D damage. The expression of NF-κB p65, miR-150, and p53 up-regulated modulator of apoptosis (PUMA) was evaluated by RT-qPCR, while the expression of PUMA, p65, and apoptotic proteins in pancreatic islet ß cells were determined by western blot analysis. Besides, inflammatory factors IL-17A, IL-2, IFN-γ, and IL-4 were detected by ELISA. The relationship among NF-κB, miR-150, and PUMA was analyzed by the dual-luciferase reporter gene, chromatin- and RNA-immunoprecipitation assays, respectively. KEY FINDINGS: Restoration of NF-κB reduced the incidence of T1D in mice. Over-expressed NF-κB inhibited the release of inflammatory factors and apoptosis in pancreatic islet ß cells. PUMA was confirmed to be a potential target gene of miR-150. miR-150 suppressed PUMA to inhibit the T1D-induced inflammation and ß cell apoptosis whereas NF-κB activated the miR-150 expression by binding to the miR-150 promoter, thereby preventing the T1D-induced inflammation and ß cell apoptosis. SIGNIFICANCE: NF-κB/miR-150/PUMA may serve as potential therapeutic targets for T1D.


Assuntos
Proteínas Reguladoras de Apoptose/genética , Diabetes Mellitus Experimental/fisiopatologia , Diabetes Mellitus Tipo 1/fisiopatologia , MicroRNAs/genética , NF-kappa B/genética , Proteínas Supressoras de Tumor/genética , Animais , Apoptose/genética , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Tipo 1/genética , Inflamação/genética , Inflamação/patologia , Células Secretoras de Insulina/patologia , Camundongos , Camundongos Endogâmicos NOD , Regulação para Cima
9.
Nat Commun ; 11(1): 2238, 2020 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-32382089

RESUMO

In type1 diabetes (T1D) autoreactive T-cells infiltrate the islets of Langerhans, depleting insulin-secreting ß-cells (insulitis). Insulitis arises during an asymptomatic phase, prior to clinical diagnosis of T1D. Methods to diagnose insulitis and ß-cell mass changes during this asymptomatic phase are limited, precluding early therapeutic intervention. During T1D the islet microvasculature increases permeability, allowing nanoparticles to access the microenvironment. Contrast enhanced ultrasound (CEUS) uses shell-stabilized gas bubbles to provide acoustic backscatter in vasculature. Here, we report that sub-micron sized 'nanobubble' ultrasound contrast agents can be used to measure increased islet microvasculature permeability and indicate asymptomatic T1D. Through CEUS and histological analysis, pre-clinical models of T1D show accumulation of nanobubbles specifically within pancreatic islets, correlating with insulitis. Importantly, accumulation is detected early in disease progression and decreases with successful therapeutic intervention. Thus, sub-micron sized nanobubble ultrasound contrast agents provide a predicative marker for disease progression and therapeutic reversal early in asymptomatic T1D.


Assuntos
Meios de Contraste , Diabetes Mellitus Tipo 1/diagnóstico por imagem , Diabetes Mellitus Tipo 1/patologia , Animais , Feminino , Humanos , Células Secretoras de Insulina/patologia , Ilhotas Pancreáticas/diagnóstico por imagem , Ilhotas Pancreáticas/patologia , Camundongos , Pâncreas/diagnóstico por imagem , Pâncreas/patologia , Ultrassonografia
10.
PLoS One ; 15(5): e0233797, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32470042

RESUMO

The incidence of metabolic diseases, such as type 2 diabetes, has increased steadily worldwide. Diet, beverages, and food texture can all markedly influence these metabolic disorders. However, the combined effects of food texture and beverages on energy metabolism remains unclear. In the present study, we examined the effect of food texture on energy metabolism in mice administered high-fructose corn syrup (HFCS). Mice were fed a soft or hard diet along with 4.2% HFCS or tap water. Body weight and total caloric intake were not affected by food texture irrespective of HFCS consumption. However, caloric intake from HFCS (i.e., drinking volume) and diet were higher and lower, respectively, in the hard food group than in the soft food group. The hard food group's preference for HFCS was absent in case of mice treated with the µ-opioid receptor antagonist naltrexone. Despite increased HFCS consumption, blood glucose levels were lower in the hard-diet group than in the soft-diet group. In HFCS-fed mice, insulin levels after glucose stimulation and insulin content in the pancreas were higher in the hard food group than the soft food group, whereas insulin tolerance did not differ between the groups. These food texture-induced differences in glucose tolerance were not observed in mice fed tap water. Thus, food texture appears to affect glucose tolerance by influencing pancreatic ß-cell function in HFCS-fed mice. These data shed light on the combined effects of eating habits and food texture on human health.


Assuntos
Glicemia/efeitos dos fármacos , Alimentos Formulados , Xarope de Milho Rico em Frutose/efeitos adversos , Células Secretoras de Insulina/efeitos dos fármacos , Insulina/sangue , Doenças Metabólicas/induzido quimicamente , Animais , Peso Corporal/efeitos dos fármacos , Ingestão de Energia/efeitos dos fármacos , Metabolismo Energético/efeitos dos fármacos , Preferências Alimentares/efeitos dos fármacos , Células Secretoras de Insulina/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL
11.
Nat Rev Endocrinol ; 16(7): 349-362, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32398822

RESUMO

Loss of functional ß-cell mass is the key mechanism leading to the two main forms of diabetes mellitus - type 1 diabetes mellitus (T1DM) and type 2 diabetes mellitus (T2DM). Understanding the mechanisms behind ß-cell failure is critical to prevent or revert disease. Basic pathogenic differences exist in the two forms of diabetes mellitus; T1DM is immune mediated and T2DM is mediated by metabolic mechanisms. These mechanisms differentially affect early ß-cell dysfunction and eventual fate. Over the past decade, major advances have been made in the field, mostly delivered by studies on ß-cells in human disease. These advances include studies of islet morphology and human ß-cell gene expression in T1DM and T2DM, the identification and characterization of the role of T1DM and T2DM candidate genes at the ß-cell level and the endoplasmic reticulum stress signalling that contributes to ß-cell failure in T1DM (mostly IRE1 driven) and T2DM (mostly PERK-eIF2α dependent). Here, we review these new findings, focusing on studies performed on human ß-cells or on samples obtained from patients with diabetes mellitus.


Assuntos
Diabetes Mellitus Tipo 1/fisiopatologia , Diabetes Mellitus Tipo 2/fisiopatologia , Insuficiência Pancreática Exócrina/etiologia , Células Secretoras de Insulina/fisiologia , Animais , Diabetes Mellitus Tipo 1/etiologia , Diabetes Mellitus Tipo 1/patologia , Diabetes Mellitus Tipo 2/etiologia , Diabetes Mellitus Tipo 2/patologia , Insuficiência Pancreática Exócrina/fisiopatologia , Humanos , Células Secretoras de Insulina/patologia , Transdução de Sinais/fisiologia
12.
Metabolism ; 107: 154232, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32302619

RESUMO

AIMS: Parasympathetic nerve (PN) signaling plays a crucial role in the maintenance of pancreatic ß-cell volume density (Vß). PN may be pathologically affected in diabetic polyneuropathy (DPN). However, the association between the reduction of PNs in islets and Vß and the therapeutic effects of a DPP4 inhibitor (DPP4i) and an SGLT2 inhibitor (SGLT2i) in nonobese type 2 diabetes mellitus (T2DM) Goto-Kakizaki rats (GK) have not been investigated. MATERIALS AND METHODS: We divided 5-week old male GK and Wistar rats (W) into a DPP4i-treated group (GKTe), SGLT2i-treated group (GKCa), and combination-treated group (GKCaTe). After 25 weeks, the pancreata was pathologically evaluated. RESULTS: Vß in GK was significantly decreased (p < 0.01 vs. W), whereas Vß was the most well preserved in GKCaTe (p < 0.05 vs. GKTe), followed by GKTe (p < 0.05 vs. GK). The decreased amount of PNs in the islets and intraepidermal nerve fiber density (IENFD) in GK was significantly improved in the treated groups compared with GK (p < 0.05 vs. GKCa and GKTe and p < 0.01 vs. GKCaTe). PN density and IENFD were significantly correlated with Vß (r = 0.55, p < 0.01 and r = 0.54, p < 0.01, respectively). IENFD was identified as a surrogate marker for the prediction of Vß (cutoff value, 16.39). CONCLUSIONS: The combination therapy of DPP4i and SGLT2i improved Vß accompanied by PNs density and IENFD. IENFD was proportionally correlated with Vß. Therefore, the prevention of DPN development may be concurrently beneficial for the preservation of Vß in nonobese T2DM.


Assuntos
Canagliflozina/uso terapêutico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Neuropatias Diabéticas/tratamento farmacológico , Inibidores da Dipeptidil Peptidase IV/uso terapêutico , Hipoglicemiantes/uso terapêutico , Células Secretoras de Insulina/patologia , Pirazóis/uso terapêutico , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Tiazolidinas/uso terapêutico , Animais , Glicemia/metabolismo , Contagem de Células , Diabetes Mellitus Tipo 2/patologia , Quimioterapia Combinada , Hemoglobina A Glicada/análise , Ilhotas Pancreáticas/patologia , Masculino , Pâncreas/patologia , Sistema Nervoso Parassimpático/patologia , Ratos , Ratos Wistar
13.
Proc Natl Acad Sci U S A ; 117(16): 8912-8923, 2020 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-32253320

RESUMO

Pancreatic islets regulate glucose homeostasis through coordinated actions of hormone-secreting cells. What underlies the function of the islet as a unit is the close approximation and communication among heterogeneous cell populations, but the structural mediators of islet cellular cross talk remain incompletely characterized. We generated mice specifically lacking ß-cell primary cilia, a cellular organelle that has been implicated in regulating insulin secretion, and found that the ß-cell cilia are required for glucose sensing, calcium influx, insulin secretion, and cross regulation of α- and δ-cells. Protein expression profiling in islets confirms perturbation in these cellular processes and reveals additional targets of cilia-dependent signaling. At the organism level, the deletion of ß-cell cilia disrupts circulating hormone levels, impairs glucose homeostasis and fuel usage, and leads to the development of diabetes. Together, these findings demonstrate that primary cilia not only orchestrate ß-cell-intrinsic activity but also mediate cross talk both within the islet and from islets to other metabolic tissues, thus providing a unique role of cilia in nutrient metabolism and insight into the pathophysiology of diabetes.


Assuntos
Cílios/metabolismo , Diabetes Mellitus/patologia , Glucose/metabolismo , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Animais , Cálcio/metabolismo , Comunicação Celular/fisiologia , Cílios/genética , Cílios/patologia , Diabetes Mellitus/genética , Modelos Animais de Doenças , Metabolismo Energético/fisiologia , Feminino , Células Secretoras de Glucagon/metabolismo , Humanos , Secreção de Insulina , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/patologia , Masculino , Camundongos , Camundongos Knockout , Transdução de Sinais/fisiologia
14.
Exp Mol Pathol ; 114: 104432, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32243891

RESUMO

BACKGROUND: Type 1 diabetes (T1DM) severely threatens human health, and the dysfunction of insulin-secreting ß cells in islets is related to the reduced PDX-1 expression. It has been reported that long non-coding RNA MALAT1 regulates ß cell function, while the potential mechanism is unclear. METHODS: Islets were isolated from non-obese diabetic (NOD) mice and wild type (WT) mice. Mouse islets and ß cell line (Min6) were stimulated by IL-1ß. The expression of MALAT1 was determined using real-time PCR, while the PDX-1 protein expression was determined using western blotting. ChIP-qPCR was carried out to determine the histone acetylation of the PDX-1 promoter. RESULTS: In NOD islets and IL-1ß-stimulated Min6 cells, the expression of MALAT1 was increased, while the mRNA and protein levels of PDX-1 were decreased at an age/time-dependent manner. Overexpressing MALAT1 suppressed the H3 histone acetylation of the PDX-1 promoter, inhibiting both mRNA and protein expressions of PDX-1. Knocking down MALAT1 restored the decrease of the histone acetylation of the PDX-1 promoter, as well as the PDX-1 expression, which was reduced by IL-1ß stimulation. Under high glucose stimulation, the overexpression of PDX-1 alone restored the insulin secretion which was inhibited by the simultaneous overexpression of MALAT1 and PDX-1. Under high glucose and IL-1ß stimulation, the simultaneous knockdown of MALAT1 and PDX-1 reduced the enhancement of the insulin secretion which was raised by knocking down MALAT1 alone. CONCLUSION: MALAT1 induces the dysfunction of ß cells via reducing the H3 histone acetylation of the PDX-1 promoter and subsequently inhibiting the expression of PDX-1, thus suppressing the insulin secretion.


Assuntos
Diabetes Mellitus Tipo 1/genética , Proteínas de Homeodomínio/genética , Células Secretoras de Insulina/metabolismo , RNA Longo não Codificante/genética , Transativadores/genética , Acetilação , Animais , Linhagem Celular , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patologia , Modelos Animais de Doenças , Glucose/metabolismo , Histonas/genética , Humanos , Insulina/genética , Células Secretoras de Insulina/patologia , Camundongos , Camundongos Endogâmicos NOD , Regiões Promotoras Genéticas/genética
15.
Proc Natl Acad Sci U S A ; 117(16): 9022-9031, 2020 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-32284404

RESUMO

The vast majority of type 1 diabetes (T1D) genetic association signals lie in noncoding regions of the human genome. Many have been predicted to affect the expression and secondary structure of long noncoding RNAs (lncRNAs), but the contribution of these lncRNAs to the pathogenesis of T1D remains to be clarified. Here, we performed a complete functional characterization of a lncRNA that harbors a single nucleotide polymorphism (SNP) associated with T1D, namely, Lnc13 Human pancreatic islets harboring the T1D-associated SNP risk genotype in Lnc13 (rs917997*CC) showed higher STAT1 expression than islets harboring the heterozygous genotype (rs917997*CT). Up-regulation of Lnc13 in pancreatic ß-cells increased activation of the proinflammatory STAT1 pathway, which correlated with increased production of chemokines in an allele-specific manner. In a mirror image, Lnc13 gene disruption in ß-cells partially counteracts polyinosinic-polycytidylic acid (PIC)-induced STAT1 and proinflammatory chemokine expression. Furthermore, we observed that PIC, a viral mimetic, induces Lnc13 translocation from the nucleus to the cytoplasm promoting the interaction of STAT1 mRNA with (poly[rC] binding protein 2) (PCBP2). Interestingly, Lnc13-PCBP2 interaction regulates the stability of the STAT1 mRNA, sustaining inflammation in ß-cells in an allele-specific manner. Our results show that the T1D-associated Lnc13 may contribute to the pathogenesis of T1D by increasing pancreatic ß-cell inflammation. These findings provide information on the molecular mechanisms by which disease-associated SNPs in lncRNAs influence disease pathogenesis and open the door to the development of diagnostic and therapeutic approaches based on lncRNA targeting.


Assuntos
Diabetes Mellitus Tipo 1/genética , Células Secretoras de Insulina/imunologia , RNA Longo não Codificante/metabolismo , Proteínas de Ligação a RNA/metabolismo , Fator de Transcrição STAT1/genética , Regiões 3' não Traduzidas/genética , Sobrevivência Celular/genética , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/virologia , Predisposição Genética para Doença , Células HEK293 , Humanos , Células Secretoras de Insulina/patologia , Células Secretoras de Insulina/virologia , Células Jurkat , Poli I-C/imunologia , Polimorfismo de Nucleotídeo Único , Cultura Primária de Células , Estabilidade de RNA/genética , RNA Mensageiro/metabolismo , RNA Viral/imunologia , Fator de Transcrição STAT1/imunologia , Fator de Transcrição STAT1/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Regulação para Cima/imunologia
16.
Artigo em Inglês | MEDLINE | ID: mdl-32292063

RESUMO

Both reactive nitrogen and oxygen species (RNS and ROS), such as nitric oxide, peroxynitrite, and hydrogen peroxide, have been implicated as mediators of pancreatic ß-cell damage during the pathogenesis of autoimmune diabetes. While ß-cells are thought to be vulnerable to oxidative damage due to reportedly low levels of antioxidant enzymes, such as catalase and glutathione peroxidase, we have shown that they use thioredoxin reductase to detoxify hydrogen peroxide. Thioredoxin reductase is an enzyme that participates in the peroxiredoxin antioxidant cycle. Peroxiredoxins are expressed in ß-cells and, when overexpressed, protect against oxidative stress, but the endogenous roles of peroxiredoxins in the protection of ß-cells from oxidative damage are unclear. Here, using either glucose oxidase or menadione to continuously deliver hydrogen peroxide, or the combination of dipropylenetriamine NONOate and menadione to continuously deliver peroxynitrite, we tested the hypothesis that ß-cells use peroxiredoxins to detoxify both of these reactive species. Either pharmacological peroxiredoxin inhibition with conoidin A or specific depletion of cytoplasmic peroxiredoxin 1 (Prdx1) using siRNAs sensitizes INS 832/13 cells and rat islets to DNA damage and death induced by hydrogen peroxide or peroxynitrite. Interestingly, depletion of peroxiredoxin 2 (Prdx2) had no effect. Together, these results suggest that ß-cells use cytoplasmic Prdx1 as a primary defense mechanism against both ROS and RNS.


Assuntos
Dano ao DNA , Peróxido de Hidrogênio/toxicidade , Células Secretoras de Insulina/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Peroxirredoxinas/metabolismo , Ácido Peroxinitroso/toxicidade , Animais , Morte Celular , Linhagem Celular Tumoral , Citoplasma/enzimologia , Citoproteção , Inibidores Enzimáticos/farmacologia , Células Secretoras de Insulina/enzimologia , Células Secretoras de Insulina/patologia , Masculino , Peroxirredoxinas/antagonistas & inibidores , Peroxirredoxinas/genética , Quinoxalinas/farmacologia , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais , Tiorredoxina Redutase 1/metabolismo
17.
Biochem Biophys Res Commun ; 525(3): 693-698, 2020 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-32139120

RESUMO

The levels of the anti-aging protein α-Klotho, in its soluble form (s-Klotho), are depressed in the circulation of patients with type 1 diabetes (T1D) or type 2 diabetes (T2D). Gene transfer experiments have suggested a protective role for ß-cell specific expression of α-Klotho in murine models of T1D and T1D, but these approaches are not easily translatable to clinical therapy. It is unknown whether systemic s-Klotho protein treatment ameliorates disease in T1D, which is characterized by autoimmune destruction of ß cells. We previously reported from in vitro experiments with ß cells that s-Klotho increases insulin secretion, reduces cells death and promotes ß-cell replication. Here, we investigated s-Klotho protein therapy in NOD mice, which have autoimmune T1D. We observed that diabetic NOD mice have significantly lower plasma levels of s-Klotho, compared to their non-diabetic counterparts. To examine in vivo effects of Klotho, we treated NOD mice with s-Klotho protein, or with a Klotho blocking antibody. Systemic treatment with s-Klotho ameliorated diabetes; notably increasing ß-cell replication and total ß-cell mass. Klotho expression was increased locally in the islets. s-Klotho also markedly reduced immune-cell infiltration of islets (insulitis). In contrast, administration of the Klotho antibody was detrimental, and aggravated the loss of ß-cell mass. Thus, s-Klotho has protective effects in this model of T1D, and this appears to depend on a combination of increased ß-cell replication and reduced insulitis. These findings suggest that s-Klotho might be effective as a new therapeutic agent for T1D.


Assuntos
Glucuronidase/uso terapêutico , Células Secretoras de Insulina/patologia , Animais , Proliferação de Células , Feminino , Glucuronidase/sangue , Humanos , Camundongos Endogâmicos NOD , Tamanho do Órgão
18.
Diabetes ; 69(4): 499-507, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32198193

RESUMO

In type 2 diabetes, ß-cells endure various forms of cellular stress, including oxidative stress and endoplasmic reticulum stress, secondary to increased demand for insulin production and extracellular perturbations, including hyperglycemia. Chronic exposure to stress causes impaired insulin secretion, apoptosis, and loss of cell identity, and a combination of these processes leads to ß-cell failure and severe hyperglycemia. Therefore, a better understanding of the molecular mechanisms underlying stress responses in ß-cells promises to reveal new therapeutic opportunities for type 2 diabetes. In this perspective, we discuss posttranscriptional control of gene expression as a critical, but underappreciated, layer of regulation with broad importance during stress responses. Specifically, regulation of mRNA translation occurs pervasively during stress to activate gene expression programs; however, the convenience of RNA sequencing has caused translational regulation to be overlooked compared with transcriptional controls. We highlight the role of RNA binding proteins in shaping selective translational regulation during stress and the mechanisms underlying this level of regulation. A growing body of evidence indicates that RNA binding proteins control an array of processes in ß-cells, including the synthesis and secretion of insulin. Therefore, systematic evaluations of translational regulation and the upstream factors shaping this level of regulation are critical areas of investigation to expand our understanding of ß-cell failure in type 2 diabetes.


Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Estresse do Retículo Endoplasmático/fisiologia , Células Secretoras de Insulina/metabolismo , Estresse Oxidativo/fisiologia , Processamento Pós-Transcricional do RNA/fisiologia , Proteínas de Ligação a RNA/metabolismo , Animais , Apoptose/fisiologia , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patologia , Regulação da Expressão Gênica , Humanos , Secreção de Insulina/fisiologia , Células Secretoras de Insulina/patologia , Processamento de Proteína Pós-Traducional/fisiologia , Proteínas de Ligação a RNA/genética
19.
Oxid Med Cell Longev ; 2020: 8609213, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32215179

RESUMO

Type 2 diabetes mellitus (T2DM) is the most prevalent metabolic disorder characterized by chronic hyperglycemia and an inadequate response to circulatory insulin by peripheral tissues resulting in insulin resistance. Insulin resistance has a complex pathophysiology, and it is contributed to by multiple factors including oxidative stress. Oxidative stress refers to an imbalance between free radical production and the antioxidant system leading to a reduction of peripheral insulin sensitivity and contributing to the development of T2DM via several molecular mechanisms. In this review, we present the molecular mechanisms by which the oxidative milieu contributes to the pathophysiology of insulin resistance and diabetes mellitus.


Assuntos
Diabetes Mellitus/metabolismo , Resistência à Insulina , Estresse Oxidativo , Animais , Antioxidantes/metabolismo , Antioxidantes/uso terapêutico , Diabetes Mellitus/classificação , Diabetes Mellitus/tratamento farmacológico , Diabetes Mellitus/patologia , Transportador de Glucose Tipo 4/genética , Transportador de Glucose Tipo 4/metabolismo , Humanos , Inflamação/metabolismo , Insulina/metabolismo , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Mitocôndrias/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais
20.
Dalton Trans ; 49(15): 4732-4740, 2020 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-32207493

RESUMO

Global rates of diabetes mellitus are increasing, and treatment of the disease consumes a growing proportion of healthcare spending across the world. Pancreatic ß-cells, responsible for insulin production, decline in mass in type 1 and, to a more limited degree, in type 2 diabetes. However, the extent and rate of loss in both diseases differs between patients resulting in the need for the development of novel diagnostic tools, which could quantitatively assess changes in mass of ß-cells over time and potentially lead to earlier diagnosis and improved treatments. Exendin-4, a potent analogue of glucagon-like-peptide 1 (GLP-1), binds to the receptor GLP-1R, whose expression is enriched in ß-cells. GLP-1R has thus been used in the past as a means of targeting probes for a wide variety of imaging modalities to the endocrine pancreas. However, exendin-4 conjugates designed specifically for MRI contrast agents are an under-explored area. In the present work, the synthesis and characterization of an exendin-4-dota(ga)-Gd(iii) complex, GdEx, is reported, along with its in vivo behaviour in healthy and in ß-cell-depleted C57BL/6J mice. Compared to the ubiquitous probe, [Gd(dota)]-, GdEx shows selective uptake by the pancreas with a marked decrease in accumulation observed after the loss of ß-cells elicited by deleting the microRNA processing enzyme, DICER. These results open up pathways towards the development of other targeted MRI contrast agents based on similar chemistry methodology.


Assuntos
Meios de Contraste/química , Complexos de Coordenação/química , Exenatida/química , Gadolínio/química , Células Secretoras de Insulina/patologia , Imagem por Ressonância Magnética , Pâncreas/diagnóstico por imagem , Compostos Radiofarmacêuticos/química , Animais , Meios de Contraste/síntese química , Complexos de Coordenação/síntese química , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Estrutura Molecular , Compostos Radiofarmacêuticos/síntese química
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