In the present study, a glucosamine-induced model of insulin-resistant skeletal muscle cells was established in order to investigate the effect of inhibition of phosphatase and tensin homolog (PTEN)/5'-adenosine monophosphate-activated protein kinase (AMPK) on these cells. The glucosamine-induced insulin-resistant skeletal muscle cells were produced and the rate of glucose uptake was measured using the glucose oxidase-peroxidase method. The expression levels of PTEN and phosphorylated PTEN (p-PTEN) were assessed using western blotting. Glucose transporter 4 (GLUT4) translocation was detected by immunofluorescence. Cell apoptosis was evaluated using flow cytometry. Following insulin stimulation, the rate of glucose uptake was significantly reduced in the cells with glucosamine-induced insulin-resistance in comparison with those in the control group. The expression and translocation of GLUT4 were reduced in the insulin-resistant muscle cells. By contrast, the expression of PTEN and p-PTEN as well as apoptosis were significantly increased. Following treatment with bisperoxopicolinatooxovanadate (BPV) or metformin in the insulin-resistant skeletal muscle cells, there was an increase in the rate of glucose uptake, an increase in GLUT4 expression and its translocation, a reduction in the expression of PTEN and p-PTEN, and a decrease in cell apoptosis compared with untreated insulin-resistant cells. Glucosamine may be used to produce an effective model of insulin-resistant skeletal muscle cells. Cells with glucosamine-induced insulin-resistance exhibited a reduced expression of GLUT4 and dysfunction in GLUT4 translocation, as well as increased activation of PTEN and increased cell apoptosis. Inhibition of PTEN or its upstream regulator, AMPK, protects glucosamine-induced insulin-resistant skeletal muscle cells from apoptosis.
Scutellarin inhibits hypoxia-induced and moderately high glucose-induced proliferation and vascular endothelial growth factor (VEGF) expression in human retinal endothelial cells (HRECs); thus, it could be a potential therapy for diabetic retinopathy. However, how scutellarin inhibits VEGF is unknown. In our study, HRECs were treated with high glucose and/or hypoxia-mimetic agent cobalt chloride to stimulate cell proliferation, migration, and angiogenesis, and the effects of scutellarin on these processes were analyzed through cell viability assay, Transwell migration assay and endothelial tube formation assay, respectively. The inhibition of angiogenic factor VEGF by scutellarin was confirmed by reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay. The mechanisms for VEGF inhibition were examined by luciferase reporter assay, Western blot, immunoprecipitation, and biochemical assays. We found that scutellarin not only concentration-dependently inhibited cell proliferation, migration, and tube formation in HRECs but also decreased their production of VEGF. The reduction of VEGF was due to increased ubiquitination and degradation of hypoxia-inducible factor (HIF)-1 alpha by scutellarin. Furthermore, scutellarin impaired the interaction of HIF-1 alpha with p300, which further decreased the transcriptional activity of HIF-1 alpha. As an inducer of HIF-1 alpha, oxidative stress was attenuated by scutellarin. Our data demonstrate that scutellarin exhibits an antiangiogenic effect via inhibition of oxidative stress, enhancement of HIF-1 alpha degradation, and reduction of VEGF secretion.
Objective To investigate the changes of bone mineral density (BMD) and the bone metabolism markers at different stages of diabetic nephropathy (DN) in patients with type 2 diabetes mellitus (T2DM).Methods Fifty-one T2DM patients with normal albuminuria, 40 DN patients with microalbuminuria, 28 DN patients with clinical albuminuria, and 20 DN patients with renal insufficiency were selected.BMD and the bone metabolism markers in all these patients were detected.Results The concentration of 25-hydroxy vitamin D3 (25-OH-D3) was the highest in normal albuminuria group, and it decreased gradually with the deterioration of renal function in DN groups ( P <0.05).The serum concentrations of bone γ-caboxyglutamic acid-containing protein (BGP) and total N-terminal propeptide of type I collagen ( T-PINP) in DN groups were lower than those in normal albuminuria group (P<0.05).The concentration ofβ-C-terminal telopeptides of type I collagen (βCTX) in DN group was higher than that in normal albuminuria group ( P <0.05).Serum level of parathyroid hormone ( PTH) in renal insufficiency was significantly higher than that in the other 3 groups (P<0.05), and no significant difference among the other 3 groups were observed (P>0.05).BMD of each part in the macroalbuminuria group and renal insufficiency was lower than that in normal albuminuria group (P<0.05).Multiple correlation analysis showed that BMD of the lumbar vertebrae and the femoral neck was negatively correlated with age, serum Cr, BUN, urinary MA/Cr,βCTX, and PTH.It was positively correlated with serum Ca, 25-OH-D3, BGP, and T-PINP.BMD of the lumbar vertebrae and the femoral neck in female patients was negatively correlated with the duration of menopause.Conclusion Along with the gradual decline of renal function in T2DM patients, the extent of bone loss gradually aggravates.The bone metabolic markers, such as T-PINP, BGP,βCTX, and 25-OH-D3, are more sensitive than BMD to reflect the changes of bone metabolism at the early stage of DN.BMD of DN patients is associated with various factors including age, duration of menopause, urinary MA/Cr, and 25-OH-D3.
