RESUMO
AIMS: High glucose (HG)-induced pancreatic ß-cell apoptosis may be a major contributor to the progression of diabetes mellitus (DM). NADPH oxidase (NOX2) has been considered a crucial regulator in ß-cell apoptosis. This study was designed to evaluate the impact of GLP-1 receptor agonist (GLP-1Ra) liraglutide on pancreatic ß-cell apoptosis in diabetes and the underlying mechanisms involved. METHODS: The diabetic rat models induced by streptozotocin (STZ) and a high fat diet (HFD) received 12â¯weeks of liraglutide treatment. Hyperglycemic clamp test was carried out to evaluate ß-cell function in vivo. Flow cytometry analysis was used to measure apoptosis rates in vitro. DCFH-DA method was used to detected ROS level in vivo and in vitro. RESULTS: Liraglutide significantly improved islet function and morphology in diabetic rats and decreased cell apoptosis rates. Thr183/Thr185 p-JNK1/2 and NOX2 levels reduced in diabetic rats and HG-induced INS-1 cell following liraglutide treatment. In addition, liraglutide upregulated the phosphorylation of AMPKα (p-AMPKα), which prevented NOX2 activation and alleviated HG-induced ß-cell apoptosis. CONCLUSION: The p-AMPKα/NOX2/JNK1/2 pathway is essential for liraglutide to attenuate HG-induced ß-cell apoptosis, which further proves that GLP-1Ras may become promising therapeutics for diabetes mellitus.
Assuntos
Apoptose/efeitos dos fármacos , Diabetes Mellitus Experimental/patologia , Células Secretoras de Insulina/efeitos dos fármacos , Liraglutida/farmacologia , NADPH Oxidase 2/metabolismo , Animais , Células Cultivadas , Citoproteção/efeitos dos fármacos , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/metabolismo , Regulação para Baixo/efeitos dos fármacos , Células Secretoras de Insulina/fisiologia , Liraglutida/uso terapêutico , Masculino , Camundongos , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , EstreptozocinaRESUMO
BACKGROUND Protective effects of reduced beta 2 glycoprotein I (Rb2GPI) against vascular injury of diabetes mellitus have been extensively investigated. However, the effects of Rb2GPI on liver injury in diabetic animals have not been reported. MATERIAL AND METHODS A diabetic rat model of was produced by systemic injection of streptozotocin (STZ). Rats were divided into a normal control group, a model group, and an Rb2GPI treatment group (N=6 in each group). After treatments, blood serum and liver tissue were collected to test the protection of Rb2GPI. AMP-activated protein kinase (AMPK) was detected by immunohistochemistry and Western blotting. RESULTS Our results revealed that Rß2GPI reduced blood glucose, serum creatinine, and urea nitrogen levels, as well as serum inflammation cytokines, including interleukin (IL)-6, tumor necrosis factor (TNF)-α and C-reactive protein in the diabetic rats. Importantly, Rß2GPI prevented liver injury in the diabetic rats as confirmed by hematoxylin-eosin (H&E) staining, alanine transaminase, aspartate transaminase, and gamma-glutamyl transferase. Reactive oxygen species (ROS) were promoted by diabetic modeling and were attenuated by Rß2GPI administration. Moreover, Rß2GPI significantly reduced liver catalase, malondialdehyde, and superoxide dismutase levels in the diabetic rats. Rß2GPI reduced liver glycolipid storage in STZ diabetic rats. Both immunohistochemistry and Western blotting demonstrated that Rß2GPI promoted AMPK phosphorylation in the diabetic rats. CONCLUSIONS Our data proved that Rß2GPI prevented liver injury in diabetic rats, likely through activating the AMPK signaling pathway.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , beta 2-Glicoproteína I/metabolismo , beta 2-Glicoproteína I/farmacologia , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Glicemia/análise , Proteína C-Reativa/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Creatinina/análise , Creatinina/sangue , Complicações do Diabetes/tratamento farmacológico , Complicações do Diabetes/metabolismo , Diabetes Mellitus Experimental/metabolismo , Modelos Animais de Doenças , Interleucina-6/metabolismo , Fígado/patologia , Masculino , Estresse Oxidativo/efeitos dos fármacos , Fosforilação , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Estreptozocina/farmacologia , Superóxido Dismutase/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Serum Mac-2-binding protein glycosylation isomer (M2BPGi) level was found to be a useful prognostic marker for hepatitis B e antigen (HBeAg)-positive chronic hepatitis B (CHB) patients treated with nucleoside/nucleotide analogs (NUCs) therapy, and the aim of our study is to evaluate the clinical implementation of M2BPGi level in the prediction of antiviral responses to pegylated-interferon-α (PEG-IFN-α) treatment in HBeAg-positive CHB patients. Ninety-six CHB patients who received PEG-IFN-α treatment for at least 48 weeks were recruited. The serum M2BPGi, alanine aminotransferase (ALT), hepatitis B surface antigen (HBsAg), HBeAg, and HBV DNA levels at baseline, weeks 4, 12, and 24 after PEG-IFN-α treatment were determined and their associations with antiviral responses were evaluated and the virological response (VR) rate and serological response (SR) rate after 48 weeks of treatment were 65.6% and 35.4%, respectively. Baseline serum M2BPGi level was significantly different between VR and non-VR (P = 0.002) or SR and non-SR groups (P = 0.012). Multivariate analyses suggested that baseline serum M2BPGi level was independently associated with VR and SR of PEG-IFN-α treatment at week 48. The area under the ROC curve (AUC) of baseline M2BPGi was 0.682 in predicting VR, which was superior to HBsAg (AUC = 0.566) or HBV DNA (AUC = 0.567). The AUC of baseline M2BPGi in predicting SR was 0.655, which was also higher than that of HBsAg (AUC = 0.548) or HBV DNA (AUC = 0.583). These results suggested that baseline serum M2BPGi level was a novel predictor of VR and SR for PEG-IFN-α treatment in HBeAg-positive CHB patients.
Assuntos
Antígenos de Neoplasias/sangue , Antivirais/administração & dosagem , Biomarcadores/sangue , Hepatite B Crônica/tratamento farmacológico , Interferon-alfa/administração & dosagem , Glicoproteínas de Membrana/sangue , Polietilenoglicóis/administração & dosagem , Adulto , Alanina Transaminase/sangue , DNA Viral/sangue , Feminino , Seguimentos , Antígenos de Superfície da Hepatite B/sangue , Antígenos E da Hepatite B/sangue , Hepatite B Crônica/diagnóstico , Hepatite B Crônica/patologia , Humanos , Masculino , Prognóstico , Curva ROC , Proteínas Recombinantes/administração & dosagem , Estudos Retrospectivos , Soro/química , Resultado do TratamentoRESUMO
Reduced ß2 glycoprotein I (ß2GPI) has been demonstrated to exhibit a beneficial effect in diabetic atherosclerosis and retinal neovascularization. However, the effect of reduced ß2GPI on vascular disorders in diabetic mellitus (DM) remains to be elucidated. The present study established a high glucoseinduced injury model using human umbilical cords veins (HUVECs) and evaluated the protective effects of reduced ß2GPI against the injury. The data demonstrated that a low concentration of reduced ß2GPI (0.5 µM) mitigated high glucoseinduced cell loss, decreased nitric oxide (NO) production and resulted in calcium overloading. Mechanically, reduced ß2GPI additionally reversed high glucoseinduced phosphatase and tensin homolog (PTEN) accumulation, decrease of protein kinase B phosphorylation and nitric oxide synthase activity, and increase of cyclooxygenase2 activity. It was further confirmed that PTEN inhibitorbpV (1 µM) exhibited similar effects to those resulting from reduced ß2GPI. Overall, the data revealed that reduced ß2GPI exerts protective effects from glucoseinduced injury in HUVECs, potentially via decreasing PTEN levels. The present study suggests reduced ß2GPI may act as a novel therapeutic strategy for the treatment of vascular disorders in DM.
Assuntos
Glucose/toxicidade , Células Endoteliais da Veia Umbilical Humana/patologia , beta 2-Glicoproteína I/farmacologia , Adulto , Cálcio/metabolismo , Morte Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Feminino , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Espaço Intracelular/metabolismo , Masculino , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase/metabolismo , Oxirredução , PTEN Fosfo-Hidrolase/metabolismo , Adulto JovemRESUMO
BACKGROUND & AIMS: Accurate evaluation of liver fibrosis is crucial for predicting progression of chronic hepatitis B virus (HBV) infection. We assessed the utility of a novel fibrosis glycobiomarker Wisteria floribunda agglutinin-positive Mac-2-binding protein (WFA+ -M2BP) for evaluating liver fibrosis and disease progression in patients with chronic HBV infection. METHODS: We enrolled 774 patients with chronic HBV infection, with or without fibrosis, diagnosed by liver biopsy/FibroScan. Patients who underwent liver biopsy (n = 297) were divided into training (n = 221) and validation (n = 76) groups. Serum WFA+ -M2BP values were measured and compared with FIB-4 index, aspartate aminotransferase (AST)-to-platelet ratio (APRI) and AST-to-alanine aminotransferase ratio (AAR) using receiver-operating characteristic (ROC) analysis. RESULTS: Serum WFA+ -M2BP levels increased significantly with fibrosis progression (P < 0.0001). Area under the ROC curve of WFA+ -M2BP for diagnosing significant fibrosis was higher than that of FIB-4 (P = 0.198), APRI (P = 0.017) and AAR (P < 0.001), with sensitivity and specificity in the training set of 60.5% and 79.8% and validation set of 59.5% and 82.1%, respectively. Serum WFA+ -M2BP levels were significantly correlated with FibroScan values (P < 0.0001) and improved the accuracy of FibroScan in assessing significant fibrosis. Changes in WFA+ -M2BP levels were parallel with those in FibroScan values during nucleot(s)ide analogues therapy in patients with chronic HBV infection. CONCLUSIONS: WFA+ -M2BP is an accurate serum indicator for assessing early stages of liver fibrosis and may monitor regression of fibrosis during the treatment of chronic HBV infection. WFA+ -M2BP provides a simple and reliable alternative or complementary method to liver biopsy and FibroScan.
Assuntos
Antígenos de Neoplasias/sangue , Hepatite B Crônica/sangue , Hepatite B Crônica/complicações , Cirrose Hepática/sangue , Glicoproteínas de Membrana/sangue , Adolescente , Adulto , Idoso , Biomarcadores/sangue , China , Progressão da Doença , Técnicas de Imagem por Elasticidade , Feminino , Humanos , Modelos Lineares , Fígado/patologia , Cirrose Hepática/patologia , Masculino , Pessoa de Meia-Idade , Lectinas de Plantas , Curva ROC , Receptores de N-Acetilglucosamina , Estudos Retrospectivos , Adulto JovemRESUMO
We explored the redox status of beta 2 glycoprotein I (ß2GPI) in different stages of diabetic angiopathy. Type 2 diabetes mellitus (T2DM) had a significantly lower proportion of reduced ß2GPI as compared to healthy controls (p < 0.05). There was a trend that the mild coronal atherosclerosis heart disease (CAD) had higher proportion of reduced ß2GPI than non-CAD and severe-CAD groups, however without significances (p > 0.05). The mild-A-stenosis group and mild-diabetic retinopathy (DR) groups had higher proportion of reduced ß2GPI than their severely affected counterparts. The mild-slow nerve conduction velocity (NCVS) group had higher proportion of reduced ß2GPI than normal nerve conduction velocity (NCVN group) and severe-NCVS groups. The proportion of reduced ß2GPI was in positive correlation with 24 h urine microalbumin and total urine protein, and the proportion of reduced ß2GPI was in negative correlation with serum and skin advanced glycation end products (AGEs). Taken together, our data implicate that the proportion of reduced ß2GPI increased in the early stage of angiopathy and decreased with the aggravation of angiopathy.
Assuntos
Biomarcadores/metabolismo , Diabetes Mellitus Tipo 2/fisiopatologia , Angiopatias Diabéticas/etiologia , Infarto do Miocárdio/complicações , beta 2-Glicoproteína I/metabolismo , Estudos de Casos e Controles , Angiopatias Diabéticas/metabolismo , Angiopatias Diabéticas/patologia , Seguimentos , Humanos , Estadiamento de Neoplasias , Oxirredução , PrognósticoRESUMO
Oxidized low-density lipoprotein (oxLDL) can bind to ß2-glycoprotein I (ß2GPI) and C-reactive protein (CRP) to form stable complexes, which exert certain effects in diabetic cardiovascular disease. A previous study by our group has confirmed that the resulting complexes promote atherosclerosis in diabetic BALB/c mice. The present study was designed to investigate the effects and potential mechanisms of oxLDL complexes on lipid accumulation and inflammatory reactions in RAW264.7 macrophages cultured in a hyperglycemic environment. Cultured cells were divided into seven groups, which were treated with phosphatebuffered saline (control), CRP, ß2GPI, oxLDL, CRP/oxLDL, oxLDL/ß2GPI or CRP/oxLDL/ß2GPI. The results revealed the formation of foam cells in the oxLDL, CRP/oxLDL, oxLDL/ß2GPI as well as CRP/oxLDL/ß2GPI groups. Compared with oxLDL, the three complexes induced less lipid accumulation (P<0.05) through inhibiting the expression of CD36 mRNA and promoting the expression of and ABCG1 mRNA (P<0.05 vs. oxLDL). Furthermore, the levels of inflammatory factors interleukin (IL)1ß, IL6 and tumor necrosis factorα were elevated in the CRP/oxLDL and CRP/oxLDL/ß2GPI groups (P>0.05 vs. oxLDL), and obvious effects on p38/mitogenactivated protein kinase and nuclear factor (NF)κB phosphorylation were also observed in these groups (P<0.05 vs. oxLDL). These results suggested that CRP/oxLDL/ßG2P1 complexes may induce lipid accumulation and inflammation in macrophages via the p38/MAPK and NFκB signaling pathways. However, some differences were observed between the complexes, which may be attributed to the property of each constituent; therefore, further studies are required.
Assuntos
Proteína C-Reativa/imunologia , Inflamação/imunologia , Lipídeos/imunologia , Lipoproteínas LDL/imunologia , Macrófagos/imunologia , NF-kappa B/imunologia , beta 2-Glicoproteína I/imunologia , Animais , Colesterol/análise , Colesterol/imunologia , Lipídeos/análise , Sistema de Sinalização das MAP Quinases , Camundongos , Proteínas Quinases Ativadas por Mitógeno/imunologia , Células RAW 264.7 , Transdução de SinaisRESUMO
OBJECTIVES: The aim of this study was to determine the effect of obesity, gestational diabetes mellitus (GDM) on circulating chemerin concentrations and chemerin gene expression of adipotissue in pregnancy women. MATERIAL AND METHODS: Totally 42 normal glucose tolerant (NGT) women and 48 women with GDM were included in this study. Their clinical features and biochemical parameters were analyzed. The NGT and GDM women were subgrouped by prepregnancy BMI as normal-weight group, overweight group, and obese group, respectively. Serum chemerin and tumor necrosis factor α (TNF-α) of these individuals were determined by ELISA methods, and subcutaneous adipose tissues' mRNA expressions of chemerin and CMKLR1 (encoding the receptor of chemerin) were analyzed by real-time PCR. RESULTS: Serum chemerin in obese-NGT group and normal-weight-GDM group was significantly higher than that of normal-weight-NGT group. Chemerin and CMKLR1 mRNA expression of subcutaneous adipose tissue was lower in the obese-NGT group than normal-weight-NGT group. There was no significant difference of CMKLR1 mRNA expression between normal-weight-NGT and normal-weight-GDM group. Serum chemerin significantly and positively correlated with triglycerides (TG) and homeostasis model assessment of insulin resistance (HOMA-IR) assessed both by uni- and multivariate. CONCLUSIONS: Gestational obesity, GDM may contribute to the elevating serum chemerin. Serum chemerin in pregnancy was associated with insulin resistance and triglycerides. Chemerin gene may play a role both in obese and GDM patients. CMKLR1 might exert its action only in obese individuals, not in GDM patients before delivery.
Assuntos
Quimiocinas/genética , Diabetes Gestacional/metabolismo , Expressão Gênica , Peptídeos e Proteínas de Sinalização Intercelular/genética , Obesidade/metabolismo , Complicações na Gravidez/metabolismo , Tecido Adiposo/química , Tecido Adiposo/metabolismo , Índice de Massa Corporal , Quimiocinas/sangue , Diabetes Gestacional/sangue , Feminino , Humanos , Resistência à Insulina , Peptídeos e Proteínas de Sinalização Intercelular/sangue , Obesidade/sangue , Gravidez , Complicações na Gravidez/sangue , RNA Mensageiro/análise , Triglicerídeos/sangue , Fator de Necrose Tumoral alfa/sangueRESUMO
Advanced glycation end products (AGEs) are a contributing factor in the angiogenesis that is characteristic of proliferative diabetic retinopathy. However, a previous study made a promising observation that domain IIV of ß2glycoprotein I (DIIV) inhibits angiogenesis in human umbilical vein cells. The present study aimed to confirm the inhibition of AGEinduced angiogenesis in retinal endothelial cells by DIIV and to investigate the potential underlying mechanisms. The RF/6A rhesus macaque choroidretinal vascular endothelial cell line was cultured in vitro and treated with AGEs in the presence or absence of different concentrations of DIIV. The proliferation, migration and tube formation of the RF/6A cells were evaluated using MTS assays, in vitro wound healing assays and in vitro Matrigel angiogenesis assays, respectively. The mRNA expression of vascular endothelial growth factor (VEGF), VEGF receptor (VEGFR) 2, VEGFR 1 and receptor for AGE (RAGE) were quantified by reverse transcription quantitative polymerase chain reaction. The expression of VEGFR1, VEGFR2 and the activation of protein kinase B (Akt) and extracellular signalregulated kinase (ERK) were also assessed by western blot analysis. The results indicated that AGEs promoted the migration, proliferation and tube formation of RF/6A cells in vitro (P<0.05), increased the expression of VEGF, VEGFR2 and RAGE (P<0.05) and increased the phosphorylation of Akt and ERK (P<0.05). DIIV inhibited the increase in VEGFR2 mRNA and protein, but did not inhibit the increase in VEGF or RAGE mRNAs. These results led to the conclusion that DIIV inhibited AGEinduced angiogenesis in the RF/6A cells, which was accompanied by a downregulation in the expression of VEGFR2 and its downstream phosphatidylinosol 3kinase/Akt and mitogenactivated protein kinase/ERK1/2 pathways. These findings provide further support towards the treatment of proliferative diabetic retinopathy by interventions that act via a mechanism similar to that of DIIV.
Assuntos
Produtos Finais de Glicação Avançada/farmacologia , Neovascularização Fisiológica/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , beta 2-Glicoproteína I/farmacologia , Animais , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Expressão Gênica , Humanos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Fosforilação , Domínios e Motivos de Interação entre Proteínas , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , beta 2-Glicoproteína I/químicaRESUMO
Accumulating evidence has implicated that liraglutide, one of the human glucagonlike peptide1 (GLP1) analogues, elicits protective effects on diabetic nephropathy; however, the mechanism has yet to be fully elucidated. The present study aimed to assess the effect and underlying mechanisms of liraglutide in diabetic nephropathy. Wistar rats with streptozotocininduced diabetes mellitus were subcutaneously injected with liraglutide or phosphate buffer for 12 weeks at a dose of 0.3 mg/kg/12 h. The biochemical parameters were determined, renal histological examination was performed by hematoxylin and eosin and periodic acid Schiff base staining, and the mRNA levels of nuclear factor κB (NFκB) and endothelial nitric oxide synthase (eNOS) were assessed by quantitative polymerase chain reaction. Furthermore, the protein expression of NFκB and eNOS as well as eNOS phosphorylation were examined by western blot analysis and the levels of inflammatory cytokines downstream of NFκB were evaluated by fluorescence-assisted cell sorting and finally, the eNOS activity and nitric oxide (NO) production were evaluated by ELISA. Liraglutide decreased the levels of total cholesterol, urine, 24-h urinary albumin, blood urea nitrogen, serum creatinine and histological damage. Liraglutide also reduced the expression of NFκB at mRNA and protein levels; the expression of tumor necrosis factorα, interferonγ, interleukin6 and monocyte chemoattractant protein1 were also reduced. By contrast, eNOS phosphorylation, eNOS activity and NO production appeared to have increased. Liraglutide may have a direct beneficial effect on diabetic nephropathy by improving eNOS activity by inhibiting the NFκB pathway without eliciting a glucose lowering effect.
Assuntos
Diabetes Mellitus Experimental/complicações , Nefropatias Diabéticas/prevenção & controle , Peptídeo 1 Semelhante ao Glucagon/análogos & derivados , NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Animais , Citocinas/metabolismo , Regulação para Baixo , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Peptídeo 1 Semelhante ao Glucagon/uso terapêutico , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Liraglutida , Masculino , Óxido Nítrico/metabolismo , Ratos Wistar , Transdução de Sinais , EstreptozocinaRESUMO
BACKGROUND & AIMS: HBV immune escape represents a challenge to prevention, diagnosis, and treatment of hepatitis B. Here, we analyzed the molecular and clinical characteristics of HBV immune escape mutants in a Chinese cohort of chronically infected patients. METHODS: Two hundred sixteen patients with HBsAg and anti-HBs were studied, with one hundred eighty-two HBV carriers without anti-HBs as a control group. Recombinant HBsAg bearing the most frequent N-glycosylation mutations were expressed in CHO and HuH7 cells. After confirming N-glycosylation at the most frequent sites (129 and 131), together with inserted mutations, functional analysis were performed to study antigenicity and secretion capacity. RESULTS: One hundred twenty-three patients had the wild-type HBs gene sequence, 93 patients (43%) had mutants in the major hydrophilic region (MHR), and 47 of the 93 patients had additional N-glycosylation mutations, which were transmitted horizontally to at least 2 patients, one of whom was efficiently vaccinated. The frequency of N-glycosylation mutation in the case group was much higher than that of the control group (47/216 vs. 1/182). Compared with wild-type HBsAg, HBsAg mutants reacted weakly with anti-HBs using a chemiluminescent microparticle enzyme immunoassay. Native gel analysis of secreted virion in supernatants of transfected HuH7 cells indicated that mutants had better virion enveloping and secretion capacity than wild-type HBV. CONCLUSIONS: Our results suggest that specific HBsAg MHR N-glycosylation mutations are implicated in HBV immune escape in a high endemic area.
Assuntos
Antígenos de Superfície da Hepatite B/genética , Evasão da Resposta Imune , Mutação , Adolescente , Adulto , Idoso , Feminino , Glicosilação , Antígenos de Superfície da Hepatite B/imunologia , Humanos , Interações Hidrofóbicas e Hidrofílicas , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: The aim of this study was to investigate the effect of C-reactive protein/oxidised low-density lipoprotein/ß2-glycoprotein I (CRP/oxLDL/ß2GPI) complex on atherosclerosis (AS) in diabetic BALB/c mice. METHODS: BALB/c mice were fed high-fat and normal diet. Eight weeks later, the mice fed with high-fat diet were injected with streptozotocin (STZ) to induce diabetes. The diabetic mice were respectively injected twice monthly with 20 µg oxLDL, 20 µg ß2GPI, 40 µg oxLDL/ß2GPI complex, 44 µg CRP/oxLDL/ß2GPI complex, and PBS. Aortas were stained with Sudan IV to investigate lipid plaque formation. The infiltration condition of smooth muscle cells (SMCs), macrophages, and T cells in the aortas were determined by immunohistochemistry (IH). The mRNA expressions of receptors associated with lipid metabolism were quantified by real-time PCR. The phosphorylation of p38 mitogen-activated protein kinase (p38MAPK) and MKK3/6 in aorta tissues were assessed by Western blot. The expression of inflammation cytokines was evaluated by protein chip. RESULTS: The lipid plaques were more extensive, the lumen area was obviously narrower, the ratio of intima and media thickness were increased, and the normal internal elastic lamia structure and endothelial cell disappeared (P < 0.05) in the oxLDL and CRP/oxLDL/ß2GPI groups (P < 0.05). CRP/oxLDL/ß2GPI complex dramatically promoted infiltration of SMCs, macrophages, and T cells, improved the mRNA expression of ABCA1 and ABCG1, but reduced the mRNA expression of SR-BI and CD36 and increased the phosphorylation of p38MAPK and MKK3/6 (all P < 0.05). The highest expression levels of IL-1, IL-9, PF-4, bFGF, and IGF-II were detected in the CRP/oxLDL/ß2GPI group (P < 0.05). CONCLUSIONS: CRP/oxLDL/ß2GPI complex aggravated AS in diabetic BALB/c mice by increasing lipid uptake, the mechanism of which may be mediated by the p38MAPK signal pathway.
Assuntos
Aterosclerose/etiologia , Aterosclerose/genética , Diabetes Mellitus Experimental/etiologia , Diabetes Mellitus Experimental/genética , Placa Aterosclerótica/etiologia , Placa Aterosclerótica/genética , Transdução de Sinais , Animais , Aorta/metabolismo , Aorta/patologia , Aterosclerose/metabolismo , Aterosclerose/patologia , Proteína C-Reativa/farmacologia , Citocinas/genética , Citocinas/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Dieta Hiperlipídica , Regulação da Expressão Gênica , Lipoproteínas LDL/farmacologia , MAP Quinase Quinase 3/genética , MAP Quinase Quinase 3/metabolismo , MAP Quinase Quinase 6/genética , MAP Quinase Quinase 6/metabolismo , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Placa Aterosclerótica/metabolismo , Placa Aterosclerótica/patologia , Estreptozocina , Linfócitos T/metabolismo , Linfócitos T/patologia , beta 2-Glicoproteína I/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
BACKGROUND: Cardiac failure is a leading cause of the mortality of diabetic patients. In part this is due to a specific cardiomyopathy, referred to as diabetic cardiomyopathy. Oxidative stress is widely considered to be one of the major factors underlying the pathogenesis of the disease. This study aimed to test whether the antioxidant alpha-lipoic acid (alpha-LA) could attenuate mitochondrion-dependent myocardial apoptosis through suppression of mitochondrial oxidative stress to reduce diabetic cardiomyopathy. METHODS: A rat model of diabetes was induced by a single tail intravenous injection of streptozotocin (STZ) 45 mg/kg. Experimental animals were randomly assigned to 3 groups: normal control (NC), diabetes (DM) and DM treated with alpha-LA (alpha-LA). The latter group was administered with alpha-LA (100 mg/kg ip per day), the remainder received the same volume vehicle. At weeks 4, 8, and 12 after the onset of diabetes, cardiac apoptosis was examined by TUNEL assay. Cardiomyopathy was evaluated by assessment of cardiac structure and function. Oxidative damage was evaluated by the content of malondialdehyde (MDA), reduced glutathione (GSH) and the activity of manganese superoxide diamutase (Mn-SOD) in the myocardial mitochondria. Expression of caspase-9 and caspase-3 proteins was determined by immunohistochemistry and mitochondrial cytochrome c release was detected by Western blotting. RESULTS: At 4, 8, and 12 weeks after the onset of diabetes, significant reductions in TUNEL-positive cells, caspase-9,-3 expression, and mitochondrial cytochrome c release were observed in the alpha-LA group compared to the DM group. In the DM group, the content of MDA in the myocardial mitochondria was significantly increased, and there was a decrease in both the mitochondrial GSH content and the activities of Mn-SOD. They were significantly improved by alpha-LA treatment. HE staining displayed structural abnormalities in diabetic hearts, while alpha-LA reversed this structural derangement. The index of cardiac function (+/-dp/dtmax) in the diabetes group was aggravated progressively from 4 weeks to 12 weeks, but alpha-LA delayed deterioration of cardiac function (P < 0.05). CONCLUSIONS: Our findings indicate that the antioxidant alpha-LA can effectively attenuate mitochondria-dependent cardiac apoptosis and exert a protective role against the development of diabetic cardiomyopathy. The ability of alpha-LA to suppress mitochondrial oxidative damage is concomitant with an enhancement of Mn-SOD activity and an increase in the GSH content of myocardial mitochondria.
Assuntos
Apoptose/efeitos dos fármacos , Cardiomiopatias/tratamento farmacológico , Mitocôndrias Cardíacas/metabolismo , Miocárdio/citologia , Ácido Tióctico/uso terapêutico , Animais , Caspase 3/análise , Caspase 9/análise , Glutationa/metabolismo , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Masculino , Malondialdeído/metabolismo , Mitocôndrias Cardíacas/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Wistar , Superóxido Dismutase/metabolismo , Ácido Tióctico/farmacologiaRESUMO
OBJECTIVE: To study the role of S6K1 in the induction of SREBP1c in mouse hepatic cell by high glucose stimulation. METHODS: S6K1 shRNA recombinant adenovirus (S6K1Ax) was injected into tail vein of db/db mice and then hepatic triglycerol content was analyzed. Liver specimen were stained with HE. After transfection with S6K1Ax or pU6Ax, mouse hepatic AML12 cells were treated with high glucose, insulin or glucose and insulin, the expression of mSREBP1c was detected by RT-PCR. S6K1 protein was detected by Western blot. RESULTS: Hepatic S6K1 protein in db/db mice was inhibited a week after S6K1Ax injection. Compared with the control group, hepatic triglycerol content of S6K1Ax group was decreased (0.65+/-0.02) mmol/L vs (0.56+/-0.01) mmol/L (t = 4.312, P less than 0.01), hepatocyte fat droplet and vaculor generation were also decreased, fatty liver was improved. The mSREBP1c expression in S6K1Ax transfected cells was lower than that in the control cells (0.03+/-0.01 vs 0.06+/-0.01, t = 5.624, P less than 0.01). Compared with the basal state, SREBP1c expression of both groups was increased on the insulin stimulation, S6K1Ax group was 0.06+/-0.02 (t = 8.452, P less than 0.01) and control group was 0.08+/-0.02 (t = 3.591, P less than 0.05). There is no difference between control and S6K1Ax group by glucose addition (P more than 0.05). CONCLUSION: S6K1 acts on fatty synthesis by regulating mSREBP1c expression.
Assuntos
Adenoviridae/genética , Ácido Graxo Sintases/metabolismo , Cirrose Hepática/patologia , Proteínas Quinases S6 Ribossômicas 90-kDa/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Animais , Linhagem Celular , Ácido Graxo Sintases/genética , Regulação da Expressão Gênica , Glucose/administração & dosagem , Insulina/administração & dosagem , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/etiologia , Cirrose Hepática/metabolismo , Camundongos , Camundongos Endogâmicos , Interferência de RNA , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas Quinases S6 Ribossômicas 90-kDa/metabolismo , Coloração e Rotulagem , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Transfecção , Triglicerídeos/metabolismoRESUMO
OBJECTIVE: To analyze mutation patterns in the RT region of hepatitis B virus (HBV) P gene in patients treated with nucleoside or nucleotide analogs. METHODS: Viral DNA was extracted from 227 serum samples of chronic hepatitis B patients from September, 2005 to March, 2007. The RT region of HBV P gene was PCR-amplified and sequenced. RESULTS: Known resistant mutations were found in 111 cases (48.9%) among 227 samples, 75 cases with clear therapy history. Novel mutations, including A222T, L229V and S256C, were also found. The incidence of multi-drug resistance in patients sequentially treated with lamivudine and adefovir was 25% (4/16), and none of the patients treated with lamivudine plus adefovir in combination shown multi-drug resistance. CONCLUSION: The patterns of mutation is complex in nucleotide analogue treated patients. Switching to adefovir in lamivudine resistant patients may lead to multi-drug resistance. Novel mutations identified in this study need further investigation.
Assuntos
Farmacorresistência Viral , Produtos do Gene pol/genética , Variação Genética , Vírus da Hepatite B/genética , Hepatite B Crônica/tratamento farmacológico , Inibidores da Transcriptase Reversa/uso terapêutico , Adolescente , Adulto , Idoso , Antivirais/farmacologia , Antivirais/uso terapêutico , Análise Mutacional de DNA , DNA Viral/genética , Feminino , Genes Virais , Vírus da Hepatite B/efeitos dos fármacos , Hepatite B Crônica/genética , Hepatite B Crônica/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Inibidores da Transcriptase Reversa/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Adulto JovemRESUMO
BACKGROUND: Currently, there are still divergent opinions about the mechanisms of the impaired neovascularization in diabetic subjects. Due to the remarkable therapeutic effect of angiotensin-converting enzyme inhibititors (ACEIs) on the reduction of blood pressure and the protection of target organs, the clinical application of this kind of drugs is very widespread. However, it is still not clear about the role and related molecular pathway of this kind of drugs in the limbs' postischemic revascularization. It is of major therapeutic importance to resolve these questions. This study aimed to investigate the reasons of the impaired angiogenesis in the hind limbs of rats with diabetic ischemia, the role and related molecular mechanisms of ACEI in postischemic revascularization. METHODS: Hind limbs ischemia was induced in diabetic rats by right femoral artery excision. Diabetic rats were randomly allocated to one of the following treatments for 4 weeks: ACEI by perindopril; perindopril in combination with a nitric oxide synthase (NOS) inhibitor; perindopril in combination with bradykinin (BK)-B1 receptor (B1R) antagonist or saline. The differences of angiogenesis, the mRNA and protein expression of endothelial nitric oxide synthase (eNOS), vascular endothelial growth factor (VEGF) and basic fibroblast (bFGF), constitutive nitric oxide synthase (cNOS) activity and nitric oxide (NO) content were observed after treatment. RESULTS: In non-ischemic hind limbs, no significant changes in capillary density, or the mRNA and protein expression of eNOS, VEGF and bFGF, or the NO content and the cNOS activity were observed among all groups. On the contrary, in ischemic hind limbs, the capillary density in diabetic rats decreased by 27% when compared with the control rats, so did the mRNA and protein expression of eNOS, VEGF and bFGF, or the NO content and the cNOS activity (P < 0.05). The capillary density was increased by 1.65-fold in the perindopril treatment group in reference to untreated diabetic rats. Moreover, administration of perindopril enhanced the mRNA expression of eNOS, VEGF, and bFGF by 1.45-, 1.44-, and 1.33-fold, increased the protein content of the above indices by 1.55-, 1.30- and 1.50-fold compared with the untreated diabetic rats respectively. Perindopril also increased NO content and cNOS activity to 1.33- and 1.38-fold of that in untreated diabetic rats. The combination of BK-B1R antagonist significantly decreased the above indices (P < 0.05). In contrast, the combination of NOS inhibitor decreased the expression of eNOS and bFGF, the NO content and the cNOS activity, while the expression of VEGF did not change. CONCLUSIONS: Diabetes mellitus reduces the neovascularization, related growth factors expression and activity in the diabetic rat ischemic legs model. Treatment of perindopril improves postischemic revascularization. This effect is mediated, at least in part, by the BK-B1R-related pathway, and the activation of VEGF/eNOS/bFGF signals may be involved in the pro-angiogenic effect.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/farmacologia , Diabetes Mellitus Experimental/fisiopatologia , Extremidades/irrigação sanguínea , Neovascularização Fisiológica/efeitos dos fármacos , Perindopril/farmacologia , Animais , Glicemia/análise , Fator 2 de Crescimento de Fibroblastos/genética , Óxido Nítrico/análise , Óxido Nítrico Sintase Tipo III/metabolismo , RNA Mensageiro/análise , Ratos , Ratos Wistar , Estreptozocina , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
OBJECTIVE: To investigate the effects of lisinopril, an angiotensin-converting enzyme inhibitor, on diabetic peripheral neuropathy (DNP). METHODS: Twenty-five Wistar rats underwent intravenous injection of streptozocin to establish diabetes models and 10 rats were injected with sodium citrate solution as normal controls. The diabetic rats were randomly divided into 2 groups: lisinopril group treated with gastric perfusion of lisinopril daily for 8 weeks, and diabetic control group. The diabetic controls and normal controls were treated with gastric perfusion of water. Sciatic nerve electrode penetration method was used to measure the motor nerve conduction velocity (MNCV) and sensory nerve conduction velocity (SNCV). Light and heat pain measuring apparatus was used to measure the pain threshold. Then the sciatic nerves were isolated. Electron microscopy was used to observe the ultra-structure. The contents of superoxide dismutase (SOD) and malonyldialdehyde (MDA), and Na(+)K(+)-ATPase activity were detected by chemical colorimetry. Immunohistochemistry was used to detect the CD34 in the sciatic nerve. The capillary density of sciatic nerve was calculated. RESULTS: The MNCV and SNCV levels of the lisinopril group were both lower than those of the 2 control groups (all P < 0.01). The potency of heat pain leg retraction response of the lisinopril group was significantly shorter than that pf the diabetic control group (P < 0.05). The pathological changes of the lisinopril group were milder than those of the other groups. The SOD level and the Na(+)K(+)-ATPase activity of the diabetic group were significantly lower than those of the normal control group, and the MDA of the diabetic group was significantly higher than those of the other 2 groups (all P < 0.05). And the SOD level and Na(+)K(+)-ATPase activity of the lisinopril group were significantly higher than those of the diabetic control group, and the MDA level of the lisinopril group was significantly lower than that of the diabetic control group. The capillary density of sciatic nerve of the diabetic control group was lower and the normal control group, and that of the lisinopril group was significantly higher than that of the diabetic control group (P < 0.01). CONCLUSION: ACE inhibitors effectively prevent and treat DPN.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Diabetes Mellitus Experimental/tratamento farmacológico , Neuropatias Diabéticas/prevenção & controle , Lisinopril/uso terapêutico , Animais , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/fisiopatologia , Masculino , Malondialdeído/metabolismo , Nervo Fibular/metabolismo , Nervo Fibular/fisiopatologia , Ratos , Ratos Wistar , Nervo Isquiático/irrigação sanguínea , ATPase Trocadora de Sódio-Potássio/metabolismo , Superóxido Dismutase/metabolismoRESUMO
The hepatitis C virus (HCV) alternate reading frame protein or F protein of the HCV 1b genotype is a double-frameshift product of the HCV core protein. In order to assess the presence of antibodies specific for F protein and their clinical relevance in sera from HCV patients, we produced recombinant F protein and core protein of the HCV 1b genotype in Escherichia coli. An enzyme-linked immunosorbent assay was developed using purified recombinant HCV core, F protein, and a 99-residue synthetic F peptide (F99). The seroprevalences of anticore, anti-F protein, and anti-F99 synthetic peptide were 95%, 68%, and 36%, respectively, in 168 HCV patients. The prevalence of anti-F antibodies did not correlate with viral load, genotype, or alanine aminotransferase level. Interferon combination therapy induced a decline in the level of anti-F antibodies in 55 responders (P < 0.01). Thirteen responders (24%) lost their anti-F recombinant protein antibodies, and 17 (31%) lost their anti-F synthetic peptide antibodies, whereas no decrease was observed for the 17 nonresponders. These changes were significant between responders and nonresponders (P < 0.05). Meanwhile, no change was found in the anticore antibody titer of the 72 treated patients. The percentage of anti-F-protein-negative patients (15/15 [100%]) who achieved a sustained virological response (SVR) was higher than that of the anti-F-positive patients (70%) (P < 0.05). Based on these findings, HCV F protein elicits a specific antibody response other than the anticore protein response. Our data also suggest that the presence and level of anti-F antibody responses might be influenced by the treatment (interferon plus ribavirin) and associated with an SVR in Chinese hepatitis C patients.
Assuntos
Antivirais/uso terapêutico , Anticorpos Anti-Hepatite C/sangue , Hepatite C/tratamento farmacológico , Hepatite C/imunologia , Interferons/uso terapêutico , Ribavirina/uso terapêutico , Proteínas do Core Viral/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Alanina Transaminase/sangue , Criança , China , Ensaio de Imunoadsorção Enzimática/métodos , Feminino , Hepacivirus/classificação , Hepacivirus/genética , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas Recombinantes , Estudos Soroepidemiológicos , Estatística como Assunto , Carga ViralRESUMO
OBJECTIVE: To investigate the prevalence of peripheral arterial disease (PAD) in China type 2 diabetic patients and to demonstrate the relationships between putative risk factors and PAD. METHODS: In total 1,397 type 2 diabetic patients aged 50 years and older were enrolled and determined ankle-brachial index (ABI) and brachial-ankle pulse wave velocity (baPWV) in 15 Class III Grade A hospitals in 7 major cities of China. RESULTS: Mean patient age was 63.7 +/- 8.2 years and mean duration of diabetes mellitus was 9.39 +/- 7.4 years. Two hundreds and seventy-two (19.47%) patients were diagnosed as PAD by ABI < 0.9, 122 (18.37%) in male and 150 (20.46%) in female. PAD patients had a significantly longer duration of diabetes mellitus, higher hemoglobin A1c, and a significantly lower mean body mass index than non-PAD ones. Aging, smoking, and systolic blood pressure were found to be positively related with the prevalence of PAD. In terms of lipid profiles, no variable was found to relate with PAD. Notably, baPWV showed as the same significant guiding index for PAD, almost matched with ABI. CONCLUSIONS: PAD is a common complication in China type 2 diabetic patients. Therefore, PAD screening and treatment should be emphasized for diabetic patients with high risk factors.
Assuntos
Angiopatias Diabéticas/epidemiologia , Doenças Vasculares Periféricas/epidemiologia , Idoso , China/epidemiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prevalência , Fatores de Risco , População Urbana/estatística & dados numéricosRESUMO
OBJECTIVE: To analyze the peripheral arterial obstructive disease (PAD) related factors among diabetic population aged > or = 50 in China. METHODS: The clinical data of 1397 diabetic patients aged > or = 50 with at least one of the following risk factors: smoking, hypertension, and hyperlipidemia, from 15 Class III Grade A hospitals in 7 major cities of China were collected. Diagnosis of PAD was based on the ankle brachial index (ABI) < 0.9, and diagnosis of arteriosclerosis was based on pulse wave velocity (PWV) > 1400 cm/s. Regression studies were made to analyze the relations among PAD and various risk factors: age, sex, body mass index (BMI), smoking, hypertension, hyperlipidemia, history of cerebral vascular disease (CVD), history of ischemia heart disease (IHD) etc. RESULTS: The current prevalence rate of PAD was 19.47% among the 1397 patients, 18.3% (122/664) among the male patients, and 20.4% (150/733) among the female patients. The prevalence of PAD in the patients aged > or = 70 was as high as 31.9%. The duration of diabetes course was positively correlated with the prevalence of PAD (chi2 = 11.9, P = 0.0026). The ABI abnormality rate was 15.78% among those with a diabetic course of 5 years and was 23.84% among those with a diabetic course of 10 years. The abnormal ABI rate of the patients with CVD was 30.57%, significantly higher than that of hose without CVD (17.29%, chi2 = 21.49, P < 0.0001). The abnormal ABI rate of the patients with IHD was 24.64%, significantly higher than that of the patients without IHD (18.20%, chi2 = 5.85, P = 0.0155). The HbA1c value of the PAD patients was significantly higher than that of the patients without PAD (chi2 = 5.10, P = 0.0239) Odd risk analysis showed that age increase of 10 years increased the PAD risk by 1.64 times (OR = 1.6444, P = 0.0001). The PAD risk of the smokers was 1.68 times higher than that of the non-smokers (OR = 1.6852, P = 0.0001). Increase of 10 mm Hg in systolic blood pressure (SBP) increased the PAD risk by 1.19 times (OR = 1.1926, P = 0.01). The PAD risk of the patients with abnormal HbAlc was 2.44 times higher than that of the patients with normal HbA1c (OR = 2.4473, P = 0.0001). One-year's increase of the hypertension course increased the PAD risk by 1.02 times (OR = 1.0194, P = 0.03). Logistic analysis indicated that the relations among PWV and the risk factors were almost the same among ABI abnormality and the risk factors. CONCLUSION: Approximately one fifth of diabetic patients aged > or = 50 in China suffer from PAD. Age, course of diabetes, blood glucose level, SBP, IHD, and CVD are risk factors for PAD. Early intervention and treatment of hypertension and hyperglycemia, and quitting smoking are important in reducing the occurrence of PAD. ABI and PWV are not only diagnostic means for PAD, but also alarm guide indexes for cerebral vascular disease (CVD).