RESUMO
The discovery of insulin approximately a century ago greatly improved the management of diabetes, including many of its life-threatening acute complications like ketoacidosis. This breakthrough saved many lives and extended the healthy lifespan of many patients with diabetes. However, there is still a negative perception of ketone bodies stemming from ketoacidosis. Originally, ketone bodies were thought of as a vital source of energy during fasting and exercise. Furthermore, in recent years, research on calorie restriction and its potential impact on extending healthy lifespans, as well as studies on ketone bodies, have gradually led to a reevaluation of the significance of ketone bodies in promoting longevity. Thus, in this review, we discuss the emerging and hidden roles of ketone bodies in various organs, including the heart, kidneys, skeletal muscles, and brain, as well as their potential impact on malignancies and lifespan.
Assuntos
Diabetes Mellitus , Cetose , Humanos , Corpos Cetônicos , Longevidade , CoraçãoRESUMO
Adipose tissues accumulate excess energy as fat and heavily influence metabolic homeostasis. O-linked N-acetylglucosamine (O-GlcNAc) modification (O-GlcNAcylation), which involves the addition of N-acetylglucosamine to proteins by O-GlcNAc transferase (Ogt), modulates multiple cellular processes. However, little is known about the role of O-GlcNAcylation in adipose tissues during body weight gain due to overnutrition. Here, we report on O-GlcNAcylation in mice with high-fat diet (HFD)-induced obesity. Mice with knockout of Ogt in adipose tissue achieved using adiponectin promoter-driven Cre recombinase (Ogt-FKO) gained less body weight than control mice under HFD. Surprisingly, Ogt-FKO mice exhibited glucose intolerance and insulin resistance, despite their reduced body weight gain, as well as decreased expression of de novo lipogenesis genes and increased expression of inflammatory genes, resulting in fibrosis at 24 weeks of age. Primary cultured adipocytes derived from Ogt-FKO mice showed decreased lipid accumulation. Both primary cultured adipocytes and 3T3-L1 adipocytes treated with OGT inhibitor showed increased secretion of free fatty acids. Medium derived from these adipocytes stimulated inflammatory genes in RAW 264.7 macrophages, suggesting that cell-to-cell communication via free fatty acids might be a cause of adipose inflammation in Ogt-FKO mice. In conclusion, O-GlcNAcylation is important for healthy adipose expansion in mice. Glucose flux into adipose tissues may be a signal to store excess energy as fat.NEW & NOTEWORTHY We evaluated the role of O-GlcNAcylation in adipose tissue in diet-induced obesity using adipose tissue-specific Ogt knockout mice. We found that O-GlcNAcylation in adipose tissue is essential for healthy fat expansion and that Ogt-FKO mice exhibit severe fibrosis upon long-term overnutrition. O-GlcNAcylation in adipose tissue may regulate de novo lipogenesis and free fatty acid efflux to the degree of overnutrition. We believe that these results provide new insights into adipose tissue physiology and obesity research.
Assuntos
Acetilglucosamina , Ácidos Graxos não Esterificados , Animais , Camundongos , Acetilglucosamina/metabolismo , Obesidade/genética , Obesidade/metabolismo , Tecido Adiposo/metabolismo , Peso Corporal/genética , Aumento de Peso , N-Acetilglucosaminiltransferases/genética , N-Acetilglucosaminiltransferases/metabolismoRESUMO
Transient neurological events (TNEs) occur after bypass surgery in Moyamoya disease (MMD); however, their pathology remains unknown. To elucidate the pathophysiology of TNEs, we investigated their relationship with perioperative superficial temporal artery (STA) blood flow volume, which was evaluated using ultrasonography. Forty-nine patients with MMD, who underwent direct bypass surgery, were included and stratified into TNE and non-TNE groups, respectively. The STA blood flow volume was evaluated at four time points (preoperatively and 2-4, 7, and 10-14 days postoperatively), and a change in volume during the postoperative period was defined as a flow volume mismatch. We investigated the association between ultrasonographic findings of flow volume mismatch and TNEs and magnetic resonance imaging findings, such as the cortical hyperintensity belt (CHB) sign, using univariate and path analyses. The STA blood flow volume increased immediately postoperatively, gradually decreasing over time, in both groups. The TNE group showed a significant increase in blood flow volume 2-4 days postoperatively (P = 0.042). Flow volume mismatch was significantly larger in the TNE group than in the non-TNE group (P = 0.020). In the path analysis, STA flow volume mismatch showed a positive association with the CHB sign (P = 0.023) and TNEs (P = 0.000). Additionally, the CHB sign partially mediated the association between STA flow volume mismatch and TNEs. These results suggest that significantly high STA blood flow volume changes occurring during the acute postoperative period after direct bypass surgery in MMD are correlated with TNEs and the CHB sign, suggesting involvement in the pathophysiology of TNEs.
Assuntos
Revascularização Cerebral , Doença de Moyamoya , Humanos , Doença de Moyamoya/diagnóstico por imagem , Doença de Moyamoya/cirurgia , Complicações Pós-Operatórias , Imageamento por Ressonância Magnética/métodos , Artérias Temporais/diagnóstico por imagem , Artérias Temporais/cirurgia , Revascularização Cerebral/métodos , Artéria Cerebral Média/cirurgiaRESUMO
OBJECTIVE: Direct bypass surgery by superficial temporal artery (STA) - middle cerebral artery anastomosis is an established procedure for moyamoya disease (MMD). However, some patients may develop cerebral infarction (CI) due to the watershed shift phenomenon after the surgery. This study sought to investigate the correlation between the postoperative changes of STA flow as well as cerebral blood flow (CBF) and the incidence of CI after direct bypass surgery for MMD. METHODS: We conducted a retrospective study of 62 hemispheres in 50 subjects who underwent direct bypass surgery for MMD. All subjects underwent pre- and post-operative MR imaging, ultrasound evaluation of STA, and single-photon emission computed tomography. The presence of CI was correlated with preoperative CBF, the delta difference of each value of the STA between before and after the surgery, and the postoperative increase ratio of CBF. RESULTS: All bypass procedures were patent, and CI was observed in 4 cases (6.4%). There was no significant association between the incidence of CI and both pre- and post-operative CBF. However, there was a significant difference in delta pulsatility index (PI) of the STA between cases with or without CI (-0.38±0.22 and -0.87±0.63, respectively, p=0.03). Whereas, other factors did not show any significant differences between those with or without CI. CONCLUSIONS: A relatively high postoperative PI of the STA was significantly associated with the incidence of CI after direct bypass surgery for MMD. A larger study is needed to confirm these findings.
RESUMO
Diabetes mellitus (DM) causes injury to tissues and organs, including to the heart and kidney, resulting in increased morbidity and mortality. Thus, novel potential therapeutics are continuously required to minimize DM-related organ damage. We have previously shown that dipeptidyl peptidase III (DPPIII) has beneficial roles in a hypertensive mouse model, but it is unknown whether DPPIII has any effects on DM. In this study, we found that intravenous administration of recombinant DPPIII in diabetic db/db mice for 8 weeks suppressed the DM-induced cardiac diastolic dysfunctions and renal injury without alteration of the blood glucose level. This treatment inhibited inflammatory cell infiltration and fibrosis in the heart and blocked the increase in albuminuria by attenuating the disruption of the glomerular microvasculature and inhibiting the effacement of podocyte foot processes in the kidney. The beneficial role of DPPIII was, at least in part, mediated by the cleavage of a cytotoxic peptide, named Peptide 2, which was increased in db/db mice compared with normal mice. This peptide consisted of nine amino acids, was a digested fragment of complement component 3 (C3), and had an anaphylatoxin-like effect determined by the Miles assay and chemoattractant analysis. The effect was dependent on its interaction with the C3a receptor and protein kinase C-mediated RhoA activation downstream of the receptor in endothelial cells. In conclusion, DPPIII plays a protective role in the heart and kidney in a DM animal model through cleavage of a peptide that is a part of C3.
Assuntos
Cardiomiopatias Diabéticas/tratamento farmacológico , Nefropatias Diabéticas/tratamento farmacológico , Dipeptidil Peptidases e Tripeptidil Peptidases/uso terapêutico , Coração/efeitos dos fármacos , Rim/efeitos dos fármacos , Substâncias Protetoras/uso terapêutico , Animais , Diabetes Mellitus/metabolismo , Diabetes Mellitus/fisiopatologia , Cardiomiopatias Diabéticas/metabolismo , Cardiomiopatias Diabéticas/fisiopatologia , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/fisiopatologia , Dipeptidil Peptidases e Tripeptidil Peptidases/metabolismo , Terapia Enzimática , Coração/fisiopatologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Rim/metabolismo , Rim/fisiopatologia , Masculino , Camundongos Endogâmicos C57BL , Substâncias Protetoras/metabolismo , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/uso terapêuticoRESUMO
Lipoprotein lipase (LPL) is an enzyme that catalyzes the hydrolysis of circulating triglyceride and the transport of fatty acids into cells. Its activity is positively regulated by insulin, and insulin resistance is associated with low LPL activity and subsequent hypertriglyceridemia. The involvement of hypertriglyceridemia in chronic kidney disease (CKD) is still under the debate in a clinical setting. Therefore, we aimed to study the role of hypertriglyceridemia in the disease using mice with systemic or renal-specific LPL deficiency. Systemic LPL deficiency was characterized by hypertriglyceridemia, but not renal injury or dyslipidemia-related conditions, such as fatty liver. Furthermore, the LPL deficiency-induced hypertriglyceridemia was not associated with a worsening of the CKD phenotype or atherosclerosis, even when CKD was induced by 5/6 nephrectomy. Next, because LPL-mediated fatty acid uptake may be important for energy metabolism in proximal tubular epithelial cells (PTECs), the role of renal LPL in renal physiology was studied by generating mice lacking LPL specifically in PTECs. These mice showed no abnormalities in their histology or renal reabsorption of micro molecules. These findings suggest that systemic and renal lipid abnormalities caused by LPL deficiency do not cause or worsen the development of renal injury, and provide novel insight regarding the potential role of lipotoxicity in the pathogenesis of obesity-related kidney injury.
Assuntos
Hiperlipoproteinemia Tipo I , Hipertrigliceridemia , Insuficiência Renal Crônica , Animais , Rim/metabolismo , Lipase Lipoproteica/metabolismo , Camundongos , Insuficiência Renal Crônica/etiologia , Triglicerídeos/metabolismoRESUMO
The early stages of age-related macular degeneration (AMD) are characterized by the accumulation of basal laminar deposits (BLamDs). The mechanism for BLamDs accumulating between the retinal pigment epithelium (RPE) and its basal lamina remains elusive. Here we examined the role in AMD of lysosome-associated membrane protein-2 (LAMP2), a glycoprotein that plays a critical role in lysosomal biogenesis and maturation of autophagosomes/phagosomes. LAMP2 was preferentially expressed by RPE cells, and its expression declined with age. Deletion of the Lamp2 gene in mice resulted in age-dependent autofluorescence abnormalities of the fundus, thickening of Bruch's membrane, and the formation of BLamDs, resembling histopathological changes occurring in AMD. Moreover, LAMP2-deficient mice developed molecular signatures similar to those found in human AMD-namely, the accumulation of APOE, APOA1, clusterin, and vitronectin-adjacent to BLamDs. In contrast, collagen 4, laminin, and fibronectin, which are extracellular matrix proteins constituting RPE basal lamina and Bruch's membrane were reduced in Lamp2 knockout (KO) mice. Mechanistically, retarded phagocytic degradation of photoreceptor outer segments compromised lysosomal degradation and increased exocytosis in LAMP2-deficient RPE cells. The accumulation of BLamDs observed in LAMP2-deficient mice was eventually followed by loss of the RPE and photoreceptors. Finally, we observed loss of LAMP2 expression along with ultramicroscopic features of abnormal phagocytosis and exocytosis in eyes from AMD patients but not from control individuals. Taken together, these results indicate an important role for LAMP2 in RPE function in health and disease, suggesting that LAMP2 reduction may contribute to the formation of BLamDs in AMD.
Assuntos
Envelhecimento/genética , Membrana Basal/patologia , Proteína 2 de Membrana Associada ao Lisossomo/genética , Retina/patologia , Envelhecimento/patologia , Animais , Lâmina Basilar da Corioide/patologia , Exocitose , Humanos , Proteína 2 de Membrana Associada ao Lisossomo/metabolismo , Lisossomos/metabolismo , Degeneração Macular/genética , Degeneração Macular/metabolismo , Camundongos , Camundongos Knockout , Fagocitose , Epitélio Pigmentado da Retina/metabolismoRESUMO
Ketone bodies have a strong negative image in medicine because of ketoacidosis, a life-threatening complication in diabetes. However, Fang et al. report that ketone bodies exert antisenescent effects in podocytes under diabetic conditions, via activation of the nuclear factor E2-related factor 2-related antioxidative stress pathway. With recent progression of research on longevity, the beneficial effects of ketone bodies are being clarified, and a positive image of ketone bodies is gradually beginning to develop in many research fields including nephrology.
Assuntos
Cetoacidose Diabética , Corpos Cetônicos , Humanos , Longevidade , Estresse OxidativoRESUMO
Autophagy-associated genes have been identified as susceptible loci for inflammatory bowel disease. We investigated the role of a core autophagy factor, Atg5, in the development of dextran sodium sulfate (DSS)-induced colitis. Intestinal epithelial cell (IEC)-specific Atg5 gene deficient mice (Atg5 ΔIEC mice) were generated by cross of Atg5-floxed mice (Atg5 fl/fl ) with transgenic mice expressing Cre-recombinase driven by the villin promotor. Mice were given three cycles of 1.5% DSS in drinking water for 5 days and regular water for 14 days over a 60-day period. The dysfunction of autophagy characterized by a marked accumulation of p62 protein, a substrate for autophagy degradation, was detected in epithelial cells in the non-inflamed and inflamed mucosa of inflammatory bowel disease patients. DSS-colitis was exacerbated in Atg5 ΔIEC mice compared to control Atg5 fl/fl mice. Phosphorylation of inositol-requiring transmembrane kinase/endonuclease1α (IRE1α), a sensor for endoplasmic reticulum stress, and c-Jun N-terminal kinase, a downstream target of IRE1α, were significantly enhanced in IECs in DSS-treated Atg5 ΔIEC mice. Accumulation of phosphorylated IRE1α was enhanced by the treatment with chloroquine, an autophagy inhibitor. Apoptotic IECs were more abundant in DSS-treated Atg5 ΔIEC mice. These findings suggest that Atg5 suppresses endoplasmic reticulum stress-induced apoptosis of IECs via the degradation of excess p-IRE1α.
RESUMO
To examine the cell-protective role of podocyte autophagy against glomerular endothelial dysfunction in diabetes, we analyzed the renal phenotype of tamoxifen (TM)-inducible podocyte-specific Atg5-deficient (iPodo-Atg5-/-) mice with experimental endothelial dysfunction. In both control and iPodo-Atg5-/- mice, high fat diet (HFD) feeding induced glomerular endothelial damage characterized by decreased urinary nitric oxide (NO) excretion, collapsed endothelial fenestrae, and reduced endothelial glycocalyx. HFD-fed control mice showed slight albuminuria and nearly normal podocyte morphology. In contrast, HFD-fed iPodo-Atg5-/- mice developed massive albuminuria accompanied by severe podocyte injury that was observed predominantly in podocytes adjacent to damaged endothelial cells by scanning electron microscopy. Although podocyte-specific autophagy deficiency did not affect endothelial NO synthase deficiency-associated albuminuria, it markedly exacerbated albuminuria and severe podocyte morphological damage when the damage was induced by intravenous neuraminidase injection to remove glycocalyx from the endothelial surface. Furthermore, endoplasmic reticulum stress was accelerated in podocytes of iPodo-Atg5-/- mice stimulated with neuraminidase, and treatment with molecular chaperone tauroursodeoxycholic acid improved neuraminidase-induced severe albuminuria and podocyte injury. In conclusion, podocyte autophagy plays a renoprotective role against diabetes-related structural endothelial damage, providing an additional insight into the pathogenesis of massive proteinuria in diabetic nephropathy.
Assuntos
Autofagia/fisiologia , Diabetes Mellitus Experimental/patologia , Células Endoteliais/patologia , Glomérulos Renais/patologia , Podócitos/patologia , Albuminúria/etiologia , Animais , Proteína 5 Relacionada à Autofagia/deficiência , Diabetes Mellitus Experimental/complicações , Nefropatias Diabéticas/complicações , Dieta Hiperlipídica , Camundongos , Proteinúria/etiologiaRESUMO
Diabetes mellitus causes systemic disorders. We previously demonstrated that diabetic condition forced bone marrow-derived cells (BMDCs) to express TNF-α, leading to the development of diabetic neuropathy in mice. Here, we hypothesized that these abnormal BMDCs are also involved in diabetic nephropathy. To test our hypothesis, mice were irradiated to receive total bone marrow (BM) from the transgenic mice expressing green fluorescent protein before diabetes was induced by streptozotocin. Confocal microscopy showed that the diabetic glomerulus had more BMDCs compared with the nondiabetic glomerulus. Most of these cells exhibited endothelial phenotypes, being negative for several markers, including podocin (a maker of podocyte), α8 integrin (mesangial cell), CD68, and F4/80 (macrophage). Next, the total BM of diabetic mice was transplanted into nondiabetic mice to examine if diabetic BM per se could cause glomerular injury. The recipient mice exhibiting normal glycemia developed albuminuria and mesangial expansion with an increase in capillary area. The number of BMDCs increased in the glomerulus of the recipient mice. These cells were found to exhibit the endothelial phenotype and to express TNF-α. These data suggest that diabetic BMDCs per se could initiate glomerular disease. Finally, eNOS knockout mice were used to examine if residential endothelial injury could attract BMDCs into the glomerulus. However, endothelial dysfunction due to eNOS deficiency failed to attract BMDCs into the glomerulus. In summary, BMDCs may be involved in the development of diabetic nephropathy.-Nobuta, H., Katagi, M., Kume, S., Terashima, T., Araki, S., Maegawa, H., Kojima, H., Nakagawa, T. A role for bone marrow-derived cells in diabetic nephropathy.
Assuntos
Células da Medula Óssea/patologia , Nefropatias Diabéticas/patologia , Animais , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/genética , Antígenos de Diferenciação Mielomonocítica/metabolismo , Células da Medula Óssea/metabolismo , Cadeias alfa de Integrinas/genética , Cadeias alfa de Integrinas/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
BACKGROUND: Energy metabolism in proximal tubular epithelial cells (PTECs) is unique, because ATP production largely depends on lipolysis in both the fed and fasting states. Furthermore, disruption of renal lipolysis is involved in the pathogenesis of diabetic tubulopathy. Emerging evidence suggests that protein O-GlcNAcylation, an intracellular nutrient-sensing system, may regulate a number of metabolic pathways according to changes in nutritional status. Although O-GlcNAcylation in PTECs has been demonstrated experimentally, its precise role in lipolysis in PTECs is unclear. METHODS: To investigate the mechanism of renal lipolysis in PTECs-specifically, the role played by protein O-GlcNAcylation-we generated mice with PTECs deficient in O-GlcNAc transferase (Ogt). We analyzed their renal phenotypes during ad libitum feeding, after prolonged fasting, and after mice were fed a high-fat diet for 16 weeks to induce obesity and diabetes. RESULTS: Although PTEC-specific Ogt-deficient mice lacked a marked renal phenotype during ad libitum feeding, after fasting 48 hours, they developed Fanconi syndrome-like abnormalities, PTEC apoptosis, and lower rates of renal lipolysis and ATP production. Proteomic analysis suggested that farnesoid X receptor-dependent upregulation of carboxylesterase-1 is involved in O-GlcNAcylation's regulation of lipolysis in fasted PTECs. PTEC-specific Ogt-deficient mice with diabetes induced by a high-fat diet developed severe tubular cell damage and enhanced lipotoxicity. CONCLUSIONS: Protein O-GlcNAcylation is essential for renal lipolysis during prolonged fasting and offers PTECs significant protection against lipotoxicity in diabetes.
Assuntos
Regulação da Expressão Gênica , Túbulos Renais Proximais/metabolismo , Metabolismo dos Lipídeos/genética , Lipólise/genética , N-Acetilglucosaminiltransferases/genética , Animais , Apoptose/genética , Células Cultivadas , Diabetes Mellitus Experimental , Dieta Hiperlipídica , Modelos Animais de Doenças , Metabolismo Energético/genética , Jejum , Homeostase/genética , Túbulos Renais Proximais/citologia , Masculino , Camundongos , Camundongos Knockout , N-Acetilglucosaminiltransferases/metabolismo , Proteômica , Distribuição Aleatória , Valores de ReferênciaAssuntos
Rim , Transdução de Sinais , Inibidores do Transportador 2 de Sódio-Glicose , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Humanos , Rim/metabolismo , Rim/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Transportador 2 de Glucose-Sódio/metabolismo , Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/tratamento farmacológico , Nefropatias Diabéticas/etiologiaRESUMO
OBJECTIVES: The objective of this study was to investigate the usefulness of model-based iterative reconstruction (IR) for detecting neointimal formations after carotid artery stenting. METHODS: In a cervical phantom harbouring carotid artery stents, we placed simulated neointimal formations measuring 0.40, 0.60, 0.80 and 1.00 mm along the stent wall. The thickness of in-stent neointimal formations was measured on images reconstructed with filtered-back projection (FBP), hybrid IR (AIDR 3D), and model-based IR (FIRST). The clinical study included 43 patients with carotid stents. Cervical computed tomography (CT) images obtained on a 320-slice scanner were reconstructed with AIDR 3D and FIRST. Five blinded observers visually graded the likelihood of neointimal formations on AIDR 3D and AIDR 3D plus FIRST images. Carotid ultrasound images were the reference standard. We analysed results of visual grading by using a Jack-knife type receiver observer characteristics analysis software. RESULTS: In the phantom study, the difference between the measured and the true diameter of the neointimal formations was smaller on FIRST than FBP or AIDR 3D images. In the clinical study, the sensitivity, specificity, positive predictive value, negative predictive value and accuracy of AIDR 3D were 58%, 88%, 83%, 67% and 73%, respectively. For AIDR 3D plus FIRST images they were 84%, 78%, 80%, 82% and 81%, respectively. The mean area under the curve was significantly higher on AIDR 3D plus FIRST than AIDR 3D images (0.82 vs 0.72; p < 0.01). CONCLUSIONS: The model-based IR algorithm helped to improve diagnostic performance for the detection of neointimal formations after carotid artery stenting. KEY POINTS: ⢠Neointimal formations can be visualised more accurately with model-based IR. ⢠Model-based IR improves the detection of neointimal formations after carotid artery stenting. ⢠Model-based IR is suitable for follow up after carotid artery stenting.
Assuntos
Artéria Carótida Primitiva/cirurgia , Estenose das Carótidas/cirurgia , Angiografia por Tomografia Computadorizada/métodos , Imageamento Tridimensional , Tomografia Computadorizada Multidetectores/métodos , Stents , Procedimentos Cirúrgicos Vasculares/métodos , Idoso , Idoso de 80 Anos ou mais , Algoritmos , Artéria Carótida Primitiva/diagnóstico por imagem , Estenose das Carótidas/diagnóstico , Feminino , Humanos , Masculino , Neointima/diagnóstico por imagem , Imagens de Fantasmas , Curva ROCRESUMO
O-GlcNAcylation is a post-translational modification that is characterized by the addition of N-acetylglucosamine (GlcNAc) to proteins by O-GlcNAc transferase (Ogt). The degree of O-GlcNAcylation is thought to be associated with glucotoxicity and diabetic complications, because GlcNAc is produced by a branch of the glycolytic pathway. However, its role in skeletal muscle has not been fully elucidated. In this study, we created skeletal muscle-specific Ogt knockout (Ogt-MKO) mice and analyzed their glucose metabolism. During an intraperitoneal glucose tolerance test, blood glucose was slightly lower in Ogt-MKO mice than in control Ogt-flox mice. High fat diet-induced obesity and insulin resistance were reversed in Ogt-MKO mice. In addition, 12-month-old Ogt-MKO mice had lower adipose and body mass. A single bout of exercise significantly reduced blood glucose in Ogt-MKO mice, probably because of higher AMP-activated protein kinase α (AMPKα) protein expression. Furthermore, intraperitoneal injection of 5-aminoimidazole-4-carboxamide ribonucleotide, an AMPK activator, resulted in a more marked decrease in blood glucose levels in Ogt-MKO mice than in controls. Finally, Ogt knockdown by siRNA in C2C12 myotubes significantly increased protein expression of AMPKα, glucose uptake and oxidation. In conclusion, loss of O-GlcNAcylation facilitates glucose utilization in skeletal muscle, potentially through AMPK activation. The inhibition of O-GlcNAcylation in skeletal muscle may have an anti-diabetic effect, through an enhancement of glucose utilization during exercise.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Glucose/metabolismo , Contração Muscular/fisiologia , Músculo Esquelético/fisiologia , N-Acetilglucosaminiltransferases/metabolismo , Esforço Físico/fisiologia , Acilação/fisiologia , Animais , Glicemia/metabolismo , Ativação Enzimática/fisiologia , Regulação Enzimológica da Expressão Gênica/fisiologia , Masculino , Camundongos , Camundongos Knockout , Condicionamento Físico Animal/métodosRESUMO
A single nucleotide polymorphism (SNP) within the acetyl CoA carboxylase (ACC) ß gene (ACACB), rs2268388, has been shown to be associated with susceptibility to development of proteinuria in patients with type 2 diabetes. To investigate the biological roles of ACCß in the pathogenesis of diabetic nephropathy, we examined the effects of overexpression of ACACB using podocyte-specific ACACB-transgenic mice or ACACB-overexpressing murine podocytes. Podocyte-specific ACACB-transgenic mice or littermate mice were treated with streptozotocin (STZ) to induce diabetes, and 12 weeks after induction of diabetes, we examined the expression of podocyte markers to evaluate the degree of podocyte injury in these mice. We also examined the effects of ACCß on podocyte injury in ACACB- or LacZ-overexpressing murine podocytes. Podocyte-specific ACACB overexpression did not cause visible podocyte injury in non-diabetic mice. In STZ-induced diabetic mice, ACACB-transgenic mice showed a significant increase in urinary albumin excretion, accompanied by decreased synaptopodin expression and podocin mislocalization in podocytes, compared with wild-type mice. In cultured murine podocytes, overexpression of ACACB significantly decreased synaptopodin expression and reorganized stress fibers under high glucose conditions, but not in normal glucose conditions. The decrease of synaptopodin expression and reorganized stress fibers observed in ACACB overexpressing cells cultured under high glucose conditions was reversed by a treatment of 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside (AICAR), activator of AMP-activated protein kinase (AMPK). The excess of ACCß might contribute to exacerbation of podocyte injury in the kidney of an animal model for diabetes mellitus, and the AMPK/ACCß pathway may be a novel therapeutic target for the prevention of diabetes-related podocyte injury.
Assuntos
Acetil-CoA Carboxilase/metabolismo , Nefropatias Diabéticas/enzimologia , Nefropatias Diabéticas/patologia , Podócitos/enzimologia , Podócitos/patologia , Animais , Células Cultivadas , Regulação Enzimológica da Expressão Gênica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Regulação para CimaRESUMO
AIMS/HYPOTHESIS: O-GlcNAcylation is characterised by the addition of N-acetylglucosamine to various proteins by O-GlcNAc transferase (OGT) and serves in sensing intracellular nutrients by modulating various cellular processes. Although it has been speculated that O-GlcNAcylation is associated with glucose metabolism, its exact role in whole body glucose metabolism has not been fully elucidated. Here, we investigated whether loss of O-GlcNAcylation globally and in specific organs affected glucose metabolism in mammals under physiological conditions. METHODS: Tamoxifen-inducible global Ogt-knockout (Ogt-KO) mice were generated by crossbreeding Ogt-flox mice with R26-Cre-ERT2 mice. Liver, skeletal muscle, adipose tissue and pancreatic beta cell-specific Ogt-KO mice were generated by crossbreeding Ogt-flox mice with Alb-Cre, Mlc1f-Cre, Adipoq-Cre and Pdx1 PB-CreER™ mice, respectively. Glucose metabolism was evaluated by i.p. glucose and insulin tolerance tests. RESULTS: Tamoxifen-inducible global Ogt-KO mice exhibited a lethal phenotype from 4 weeks post injection, suggesting that O-GlcNAcylation is essential for survival in adult mice. Tissue-specific Ogt deletion from insulin-sensitive organs, including liver, skeletal muscle and adipose tissue, had little impact on glucose metabolism under physiological conditions. However, pancreatic beta cell-specific Ogt-KO mice displayed transient hypoglycaemia (Ogt-flox 5.46 ± 0.41 vs Ogt-ßKO 3.88 ± 0.26 mmol/l) associated with about twofold higher insulin secretion and accelerated adiposity, followed by subsequent hyperglycaemia (Ogt-flox 6.34 ± 0.32 vs Ogt-ßKO 26.4 ± 2.37 mmol/l) with insulin depletion accompanied by beta cell apoptosis. CONCLUSIONS/INTERPRETATION: These findings suggest that O-GlcNAcylation has little effect on glucose metabolism in insulin-sensitive tissues but plays a crucial role in pancreatic beta cell function and survival under physiological conditions. Our results provide novel insight into O-GlcNAc biology and physiology in glucose metabolism.
Assuntos
Insulina/metabolismo , N-Acetilglucosaminiltransferases/metabolismo , Animais , Apoptose/fisiologia , Glucose/metabolismo , Resistência à Insulina/fisiologia , Células Secretoras de Insulina/metabolismo , Camundongos , Camundongos Knockout , Processamento de Proteína Pós-TraducionalRESUMO
BACKGROUND: O-linked ß- N -acetylglucosamine modification O-GlcNAcylation) is a post-translational modification of intracellular proteins, serving as a nutrient sensor. Growing evidence has demonstrated its physiological and pathological importance in various mammalian tissues. This study examined the physiological role of O-GlcNAcylation in podocyte function and development. METHODS: O-GlcNAc transferase (Ogt) is a critical enzyme for O-GlcNAcylation and resides on the X chromosome. To abrogate O-GlcNAcylation in podocytes, we generated congenital and tamoxifen (TM)-inducible podocyte-specific Ogt knockout mice (Podo-Ogt y/- and TM-Podo-Ogt y/- , respectively) and analyzed their renal phenotypes. RESULTS: Podo-Ogt y/- mice showed normal podocyte morphology at birth. However, they developed albuminuria at 8 weeks of age, increasing progressively until age 32 weeks. Glomerular sclerosis, proteinuria-related tubulointerstitial lesions and markedly altered podocyte foot processes, with decreased podocin expression, were observed histologically in 32-week-old Podo-Ogt y/- mice. Next, we induced adult-onset deletion of the Ogt gene in podocytes by TM injection in 8-week-old TM-Podo-Ogt y/- mice. In contrast to Podo-Ogt y/- mice, the induced TM-Podo-Ogt y/- mice did not develop albuminuria or podocyte damage, suggesting a need for O-GlcNAcylation to form mature foot processes after birth. To test this possibility, 3-week-old Podo-Ogt y/- mice were treated with Bis-T-23, which stimulates actin-dependent dynamin oligomerization, actin polymerization and subsequent foot process elongation in podocytes. Albuminuria and podocyte damage in 16-week-old Podo-Ogt y/- mice were prevented by Bis-T-23 treatment. CONCLUSIONS: O-GlcNAcylation is necessary for maturation of podocyte foot processes, particularly after birth. Our study provided new insights into podocyte biology and O-GlcNAcylation.
Assuntos
Acetilglucosamina/química , Pé/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/metabolismo , N-Acetilglucosaminiltransferases/fisiologia , Podócitos/metabolismo , Processamento de Proteína Pós-Traducional , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
AIMS/HYPOTHESIS: The efficacy of a low-protein diet (LPD) on diabetic nephropathy is controversial. We aimed to investigate the renoprotective effects of an LPD and the underlying molecular mechanism in a rat model of type 2 diabetes and obesity. METHODS: Diabetic male Wistar fatty (fa/fa) rats (WFRs) were treated with a standard diet (23.84% protein) or an LPD (5.77% protein) for 20 weeks from 24 weeks of age. We investigated the effect of the LPD on renal function, fibrosis, tubular cell damage, inflammation, mitochondrial morphology of proximal tubular cells (PTCs), apoptosis, autophagy and activation of mammalian target of rapamycin complex 1 (mTORC1). RESULTS: Kidney weight, albuminuria, excretion of urinary liver-type fatty acid binding protein, levels of plasma cystatin C and changes in renal histology, including fibrosis, tubular cell damage and inflammation, were aggravated in WFRs compared with non-diabetic Wistar lean rats (WLRs). Fragmented and swelling mitochondria accumulated in PTCs and apoptosis were enhanced in the kidney of WFRs. Immunohistochemical staining of p62 and p-S6 ribosomal protein (p-S6RP) in the tubular lesions of WFRs was increased compared with WLRs. The LPD intervention clearly ameliorated damage as shown by the assessment of renal function and histology, particularly tubulointerstitial damage in diabetic kidneys. Additionally, the 5.77% LPD, but not the 11.46% LPD, significantly suppressed p-S6RP levels and increased microtubule-associated protein light chain 3-II levels in the renal cortex. The LPD intervention partially decreased HbA1c levels in WFRs, and no differences in mean BP were observed among any of the groups. CONCLUSIONS/INTERPRETATION: A very-low-protein diet improved advanced diabetic renal injuries, including tubulointerstitial damage, by restoring autophagy through the suppression of the mTORC1 pathway.
Assuntos
Autofagia/fisiologia , Diabetes Mellitus Tipo 2/dietoterapia , Diabetes Mellitus Tipo 2/metabolismo , Nefropatias Diabéticas/dietoterapia , Nefropatias Diabéticas/metabolismo , Dieta com Restrição de Proteínas , Complexos Multiproteicos/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Animais , Autofagia/genética , Diabetes Mellitus Tipo 2/patologia , Nefropatias Diabéticas/patologia , Modelos Animais de Doenças , Imuno-Histoquímica , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina , Microscopia Eletrônica de Transmissão , Complexos Multiproteicos/genética , Obesidade/dietoterapia , Obesidade/metabolismo , Obesidade/patologia , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Serina-Treonina Quinases TOR/genéticaRESUMO
Hypoxia causes proximal tubular cell damage in diabetes, even though proximal tubular cells have an adaptive system to combat hypoxia involving induction of hypoxia factor-1 (HIF-1) and inhibition of mechanistic target of rapamycin complex 1 (mTORC1). Here, we examined the interference effect of altered glucose and lipid metabolism on the hypoxia responses in proximal tubular cells. In culture, hypoxia alone induced HIF-1 and inhibited mTORC1, preventing death in proximal tubular cells. However, hypoxia with high glucose and palmitate increased mTORC1 activity and promoted apoptosis in proximal tubular cells, which was inhibited by pharmacological and genetic inactivation of mTORC1. Since inhibition of all mTORC1's physiological functions regulated by growth factors including insulin causes various adverse effects, we screened for a microRNA that can inhibit only pro-apoptotic effects of mTORC1 to discover a safe therapeutic target. This screen found microRNA-148b-3p was able to specifically inhibit mTORC1-dependent apoptosis in hypoxic proximal tubular cells exposed to high glucose and palmitate, without affecting insulin-dependent mTORC1 activation. Furthermore, tumor necrosis factor receptor (TNFR) 2 was the target of microRNA-148b-3p and its suppression inhibited apoptosis. Finally, enhanced apoptosis with TNFR2 overexpression was found in hypoxic and mTORC1-activated proximal tubular cells in diabetic rats. Thus, diabetes activated mTORC1 even in hypoxic proximal tubular cells, leading to apoptosis by reducing microRNA-148b-3p expression. Modulating this pathogenic pathway may be a novel therapy for proximal tubular cell damage in diabetes.