Role of soluble and membrane-bound dipeptidyl peptidase-4 in diabetic nephropathy.

Diabetic nephropathy is one of the most frequent, devastating and costly complications of diabetes. The available therapeutic approaches are limited. Dipeptidyl peptidase type 4 (DPP-4) inhibitors represent a new class of glucose-lowering drugs that might also have reno-protective properties. DPP-4 exists in two forms: a plasma membrane-bound form and a soluble form, and can exert many biological actions mainly through its peptidase activity and interaction with extracellular matrix components. The kidneys have the highest DPP-4 expression level in mammalians. DPP-4 expression and urinary activity are up-regulated in diabetic nephropathy, highlighting its role as a potential target to manage diabetic nephropathy. Preclinical animal studies and some clinical data suggest that DPP-4 inhibitors decrease the progression of diabetic nephropathy in a blood pressure- and glucose-independent manner. Many studies reported that these reno-protective effects could be due to increased half-life of DPP-4 substrates such as glucagon-like peptide-1 (GLP-1) and stromal derived factor-1 alpha (SDF-1a). However, the underlying mechanisms are far from being completely understood and clearly need further investigations.

Low-protein diet supplemented with ketoacids ameliorates proteinuria in 3/4 nephrectomised rats by directly inhibiting the intrarenal renin-angiotensin system.

Low-protein diet plus ketoacids (LPD+KA) has been reported to decrease proteinuria in patients with chronic kidney diseases (CKD). However, the mechanisms have not been clarified. As over-activation of intrarenal renin-angiotensin system (RAS) has been shown to play a key role in the progression of CKD, the current study was performed to investigate the direct effects of LPD+KA on intrarenal RAS, independently of renal haemodynamics. In this study, 3/4 subtotal renal ablated rats were fed 18 % normal-protein diet (Nx-NPD), 6 % low-protein diet (Nx-LPD) or 5 % low-protein diet plus 1 % ketoacids (Nx-LPD+KA) for 12 weeks. Sham-operated rats fed NPD served as controls. The level of proteinuria and expression of renin, angiotensin II (AngII) and its type 1 receptors (AT1R) in the renal cortex were markedly higher in Nx-NPD group than in the sham group. LPD+KA significantly decreased the proteinuria and inhibited intrarenal RAS activation. To exclude renal haemodynamic impact on intrarenal RAS, the serum samples derived from the different groups were added to the culture medium of mesangial cells. It showed that the serum from Nx-NPD directly induced higher expression of AngII, AT1R, fibronectin and transforming growth factor-ß1 in the mesangial cells than in the control group. Nx-LPD+KA serum significantly inhibited these abnormalities. Then, proteomics and biochemical detection suggested that the mechanisms underlying these beneficial effects of LPD+KA might be amelioration of the nutritional metabolic disorders and oxidative stress. In conclusion, LPD+KA could directly inhibit the intrarenal RAS activation, independently of renal haemodynamics, thus attenuating the proteinuria in CKD rats.

Effects of herbal compound 861 on collagen synthesis and degradation in rat mesangial cells exposed to high glucose.

OBJECTIVE: To investigate the effects of Herbal Compound 861 (Cpd 861) on collagen synthesis and degradation in rat mesangial cells exposed to high glucose. METHODS: The third to fifth passage of rat mesangial cells were exposed to high glucose and Cpd 861 at a concentration of 0.25-4.00 g/L for 24, 48 and 72 h, respectively. Benazepril (10(-7)-10(-3) mmol/L) was selected as positive control. The methyl thiazolyl tetrazolium colorimetric assay was used to evaluate the effect of Cpd 861 on cell proliferation. After incubation with Cpd 861 at a concentration of 2.00 g/L for 48 h, the protein secretions of collagen type IV, matrix metallopeptidase 9 (MMP-9), tissue inhibitor of metalloproteinase-1 (TIMP-1), transforming growth factor beta 1 (TGF-ß1), and hepatocyte growth factor (HGF) were detected by enzyme-linked immunosorbent assay method. And rat mesangial cells were harvested to determine MMP-9, TIMP-1, TGF-ß1 and HGF mRNA expression by reverse transcription polymerase chain reaction. RESULTS: Cpd 861 inhibited cell proliferation induced by high glucose in a dose- and time-dependent manner. Compared with high glucose, collagen type IV production was decreased significantly by Cpd 861 (P<0.01). Cpd 861 increased the protein secretions and mRNA expressions of MMP-9 and HGF, whereas the protein secretions and mRNA expressions of TIMP-1 and TGF-ß1 were reduced markedly (P<0.05). The ratio of MMP-9 to TIMP-1 was enhanced by Cpd 861 significantly. There was no significant difference in all above-mentioned effects between Cpd 861 (2.00 g/L) and benazepril (10(-5) mmol/L). CONCLUSION: The anti-glomerulosclerosis mechanisms of Cpd 861 were partly attributed to its effects of inhibiting mesangial cell proliferation, decreasing collagen synthesis and enhancing collagen degradation.

Uncoupling endothelial nitric oxide synthase is ameliorated by green tea in experimental diabetes by re-establishing tetrahydrobiopterin levels.

The current study investigated the potential of green tea (GT) to improve uncoupling of endothelial nitric oxide synthase (eNOS) in diabetic conditions. In rats with streptozotocin-induced diabetes, nitric oxide (NO) bioavailability was reduced by uncoupling eNOS, characterized by a reduction in tetrahydrobiopterin (BH(4)) levels and a decrease in the eNOS dimer-to-monomer ratio. GT treatment ameliorated these abnormalities. Moreover, immortalized human mesangial cells (ihMCs) exposed to high glucose (HG) levels exhibited a rise in reactive oxygen species (ROS) and a decline in NO levels, which were reversed with GT. BH(4) and the activity of guanosine triphosphate cyclohydrolase I decreased in ihMCs exposed to HG and was normalized by GT. Exogenous administration of BH(4) in ihMCs reversed the HG-induced rise in ROS and the decline in NO production. However, coadministration of GT with BH(4) did not result in a further reduction in ROS production, suggesting that reduced ROS with GT was indeed secondary to uncoupled eNOS. In summary, GT reversed the diabetes-induced reduction of BH(4) levels, ameliorating uncoupling eNOS, and thus increasing NO bioavailability and reducing oxidative stress, two abnormalities that are involved in the pathogenesis of diabetic nephropathy.

[Dracorhodin perchlorate inhibit high glucose induce serum and glucocorticoid induced protein kinase 1 and fibronectin expression in human mesangial cells].

OBJECTIVE: To investigate the effect of dracorhodin perchlorate (DP) on inhibiting high glucose-induced serum and glucocorticoid induced protein kinase 1 (SGK1) and fibronectin (FN) expression in human mesangial cells (HMC), and its mechanism of prevention and treatment on renal fibrosis in diabetic nephropathy (DN) . METHOD: The HMC were divided into normal glucose group (NG group, 5.5 mmol x L(-1) D-glucose), normal glucose +low DP group (NG + LDP group, 5.5 mmol x L(-1) D-glucose +7.5 micromol x L(-1) DP), normal glucose +high DP group (NG + HDP group, 5.5 mmol x L(-1) D-glucose + 15 micromol x L(-1) DP), high glucose group (HG group,25 mmol x L(-1) D-glucose), high glucose +low DP group (HG + LDP group, 25 mmol x L(-1) D-glucose + 7.5 micromol x L(-1) DP)and high glucose +high DP group (HG +HDP group, 25 mmol x L(-1) D-glucose + 15 micromol x L(-1) DP). Each group was examined at 24 hours. The levels of SGK1 and FN mRNA was detected by real-time fluorescence quantitative PCR,and the expression of SGK1 and FN protein was detected by Western blot or indirect immunofluorescence. RESULT: A basal level of SGK1 and FN in HMC were detected in NG group, and the level of SGK1 and FN mRNA and protein were not evidently different compared to that of NG group adding 7.5 micromol x L(-1) DP for 24 hours. On the other hand, the levels of SGK1 and FN mRNA and protein were obviously decreased by adding 15 micromol x L(-1) DP for 24 hours. Compared to NG group, the levels of SGK1 and FN mRNA and protein were increased in HG group after stimulating for 24 hours (P < 0.01). Compared to HG group, the level of SGK1 and FN mRNA and protein were evidently reduced in HG + LDP and HG + HDP groups by adding 7.5 micromol x L(-1) DP and 15 micromol x L(-1) DP for 24 hours (P < 0.01). CONCLUSION: Dracorhodin perchlorate can inhibit high glucose-induced serum and glucocorticoid induced protein kinase 1 (SGK1) and fibronectin(FN) expression in human mesangial cells, and this may be part of the mechanism of preventing and treating renal fibrosis of DN.

Protective effect of Lycium barbarum polysaccharide 4 on kidneys in streptozotocin-induced diabetic rats.

Lycium barbarum polysaccharide (LBP) has been shown to have hypoglycemic and antioxidative properties, although its mode of action is yet unknown. Because oxidative stress is implicated in the pathogenesis of diabetic nephropathy, we evaluated the protective effect of LBP-4, the major active component of Lycium barbarum, on the defensive antioxidative mechanism in kidneys in a streptozotocin-induced diabetic rat model. Moreover, we investigated the effects of LBP-4 on the activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) in isolated mesangial cells. The role of protein kinase C (PKC)-dependent and -independent pathways in LBP-4-reduced ERK1/2 was studied by bisindolylmaleimide (BIM) IV, an inhibitor of PKC. Diabetic rats treated with LBP-4 (10 mg/kg) for 8 weeks showed increased activity of antioxidant enzymes and increased scavenging of oxygen radicals, while the activity of PKC in the renal cortex was maintained at a physiological level. The decreased activation of ERK1/2 in mesangial cells, through the involvement of PKC, could explain the protective mechanism in kidneys of diabetic rats treated with LBP-4.

Saireito probably prevents mesangial cell proliferation in HIGA mice via PDGF-BB tyrosine kinase inhibition.

BACKGROUND: In this study, we administered saireito to high serum IgA (HIGA) mice and investigated its inhibitory effect on platelet-derived growth factor (PDGF) receptor tyrosine kinase (which causes mesangial proliferation) as one of the possible antinephritic mechanisms of saireito. METHODS: Female HIGA/NscSlc mice, aged 10 weeks, were divided into five groups (each, n = 12; a control group, three saireito-mixed feed groups, and a captopril-mixed feed group) so that the plasma IgA levels were comparable among the groups. After the grouping, the animals were administered the saireito or captopril, mixed in the feed, until the age of 45 weeks. RESULTS: At the age of 45 weeks, the glomerular cell number was 47.8 +/- 3.9 / cross section in the HIGA mice in the control group, but 41.6 +/- 2.3 / cross section in the 1.3% saireito-mixed feed group and 38.7 +/- 3.5 / cross section in the captopril-mixed feed group, being significantly lower in both these treatment groups than in the control group. At the age of 45 weeks, the sclerosis score in the HIGA mice in the control group was 0.92 +/- 0.23. However, the sclerosis scores in the 0.26% (0.59 +/- 0.26) and 1.3% (0.58 +/- 0.16) saireito-mixed feed groups were significantly lower than that in the control group. In the captopril-mixed feed group, the sclerosis score was 0.64 +/- 0.34, significantly lower than that in the control group. It was clarified that saireito suppressed mesangial cell proliferation without showing any cytotoxicity. Furthermore, as a result of investigating the mesangial cell proliferation-suppressing effect similarly with the 23 substances constituting saireito, a proliferation-suppressing effect was recognized with isoliquiritigenin (a component of Glycyrrhizae Radix) and oroxylin A (a component of Scutellariae Radix). Oroxylin A and isoliquiritigenin showed an inhibitory effect on PDGF receptor tyrosine kinase. Furthermore, the inhibitory effects of oroxylin A and isoliquiritigenin on tyrosine kinase were found to be specific to the PDGF receptor, and showed no influence on the tyrosine kinase activities of other growth-factor receptors examined. CONCLUSION: These results suggest that the antinephritic effects of saireito in HIGA mice may be partly due to the inhibiton of PDGF tyrosine kinase by oroxylin A and isoliquiritigenin, components of saireito.