[Molecular regulative mechanisms of NLRP3 inflammasome activation in diabetic nephropathy and interventional effects of Chinese herbal medicine].

The progression of renal damage in diabetic nephropathy(DN)is closely related to Nod-like receptor protein3(NLRP3)inflammasome activation. The characteristics of NLRP3 inflammasome activation include the changed expression and combination levels of NLRP3, apoptosis-associated speck-like protein(ASC)and pro-caspase-1, the increased expression levels of caspase-1, interleukin(IL)-1ß and IL-18 and the excessive release levels of the relative inflammatory mediators. Its molecular regulative mechanisms involve the activation of multiple signaling pathways including reactive oxygen species(ROS)/thioredoxin-interacting protein(TXNIP)pathway, nuclear factor(NF)-κB pathway, nuclear factor erythroid-related factor 2(Nrf2)pathway, long non-coding RNA(lncRNA)pathway and mitogen-activated protein kinases(MAPKs)pathway. In addition, more importantly, never in mitosis aspergillus-related kinase 7(Nek7), as a kinase regulator, could target-combine with NLRP3 at upstream to activate NLRP3 inflammasome. Some extracts of Chinese herbal medicines(CHMs)such as quercetin, curcumin, cepharanthine, piperine and salidroside, as well as Chinese herbal compound prescriptions such as Wumei Pills both could treat NLRP3 inflammasome to ameliorate inflammatory renal damage in DN. Therefore, accurately clarifying the targets of anti-inflammatory CHMs and Chinese herbal compound prescriptions delaying DN progression by targeting the molecular regulative mechanisms of NLRP3 inflammasome activation will be one of the development directions in the future.

[Molecular mechanisms of insulin resistance and interventional effects of Chinese herbal medicine].

It is considered that insulin resistance(IR)and its signaling pathway disorder are one of pathogenesis that causes insulin target-organs/issues lesions and their slow progression. The clinical diagnosis index of IR is the homeostatic model of insulin resistance(HOMA-IR)based on fasting blood-glucose and fasting serum insulin. Furthermore, the emerging IR biomarkers including adiponectin may be the references for clinical diagnosis. The influence factors of IR are obesity, chronic microinflammation and a lack of exercise. The major signaling pathways of IR include insulin receptor substrate 1(IRS1)/phosphatidylinositiol-3-kinase(PI3 K)/serine-threonine kinase(Akt)pathway, mitogen-activated protein kinase(MAPK)pathway and Smad3 pathway. In clinics, insulin sensibility and IR could be increased and improved via promoting insulin secretion and enhancing insulin signaling activation. At present, insulin sensitizers treating IR not only have the classic thiazolidinediones and its ramifications but also have the newly discovered metformin and vitamin D. In addition, it is reported that some extracts from single Chinese herbal medicine(CHM)and Chinese herbal compound prescription such as total flavone from the flowers of Abelmoschl manihot, berberine, astragalus polysaccharides and Huang-qi decoction also have the beneficial effects in ameliorating IR. In the field of chronic kidney disease, targeting a common insulin target-organs/issues lesion, the early renal damage in diabetic mellitus, the intervention studies regarding to regulating podocyte IR signaling pathways by CHM will be one of the significant directions in the future.

[Pathomechanisms of pericyte-myofibroblast transition in kidney and interventional effects of Chinese herbal medicine].

In the kidney, pericyte is the major source of myofibroblast (MyoF) in renal interstitium. It is reported that pericyte-myofibroblast transition（PMT）is one of the important pathomechanisms of renal interstitial fibrosis（RIF）. Among them, the main reasons for promoting RIF formation include pericyte recruitment, activation and isolation, as well as the lack of pericyte-derived erythropoietin. During the PMT startup process, pericyte activation and its separation from microvessels are controlled by multiple signal transduction pathways, such as transforming growth factor-ß（TGF-ß）pathway, vascular endothelial growth factor receptor (VEGFR) pathway and platelet derived growth factor receptor (PDGFR) pathway;Blocking of these signaling pathways can not only inhibit PMT, but also suppress renal capillaries reduction and further alleviate RIF. In clinic, many traditional Chinese medicine compound prescriptions, single traditional Chinese herbal medicine (CHM) and their extracts have the clear effects in alleviating RIF, and some of their intervention actions may be related to pericyte and its PMT. Therefore, the studies on PMT and its drug intervention will become the main development direction in the research field of anti-organ fibrosis by CHM.

Inhibition of Akt/mTOR/p70S6K Signaling Activity With Huangkui Capsule Alleviates the Early Glomerular Pathological Changes in Diabetic Nephropathy.

Huangkui capsule (HKC), a Chinese modern patent medicine extracted from Abelmoschus manihot (L.) medic, has been widely applied to clinical therapy in the early diabetic nephropathy (DN) patients. However, it remains elusive whether HKC can ameliorate the inchoate glomerular injuries in hyperglycemia. Recently the activation of phosphatidylinositol-3-kinase (PI3K)/serine-threonine kinase (Akt)/mammalian target of rapamycin (mTOR) signaling and its downstream regulator, 70-kDa ribosomal protein S6 kinase (p70S6K), play important roles in the early glomerular pathological changes of DN including glomerular hypertrophy, glomerular basement membrane (GBM) thickening and mild mesangial expansion. This study thereby aimed to clarify therapeutic effects of HKC during the initial phase of DN and its underlying mechanisms. Fifteen rats were randomly divided into 3 groups: the normal group, the model group and the HKC group. The early DN model rats were induced by unilateral nephrectomy combined with intraperitoneal injection of streptozotocin, and administered with either HKC suspension or vehicle after modeling and for a period of 4 weeks. Changes in the incipient glomerular lesions-related parameters in urine and blood were analyzed. Kidneys were isolated for histomorphometry, immunohistochemistry, immunofluorescence and Western blotting (WB) at sacrifice. In vitro, murine mesangial cells (MCs) were used to investigate inhibitory actions of hyperoside (HYP), a bioactive component of HKC, on cellular hypertrophy-associated signaling pathway by WB, compared with rapamycin (RAP). For the early DN model rats, HKC ameliorated micro-urinary albumin, body weight and serum albumin, but had no significant effects on renal function and liver enzymes; HKC improved renal shape, kidney weight and kidney hypertrophy index; HKC attenuated glomerular hypertrophy, GBM thickening and mild mesangial expansion; HKC inhibited the phosphorylation of Akt, mTOR and p70S6K, and the protein over-expression of transforming growth factor-ß1 in kidneys. In vitro, the phosphorylation of PI3K, Akt, mTOR and p70S6K in MCs induced by high-glucose was abrogated by treatment of HYP or RAP. On the whole, this study further demonstrated HKC safely and efficiently alleviates the early glomerular pathological changes of DN, likely by inhibiting Akt/mTOR/p70S6K signaling activity in vivo and in vitro, and provided the first evidence that HKC directly contributes to the prevention of the early DN.

[Low dose of triptolide ameliorates podocyte epithelial-mesenchymal transition induced by high dose of D-glucose via inhibiting Wnt3α/ß-catenin signaling pathway activation].

To explore the effects and molecular mechanisms of triptolide（TP）on improving podocyte epithelial-mesenchymal transition（EMT）induced by high dose of D-glucose（HG）, the immortalized podocytes of mice in vitro were divided into the normal group（N）, the high dose of D-glucose group（HG）, the low dose of TP group（L-TP）, the high dose of TP group（H-TP）and the mannitol group（MNT）, and treated by the different measures respectively. More specifically, the podocytes in each group were separately treated by D-glucose（DG, 5 mmol·L⁻¹ï¼‰or HG（25 mmol·L⁻¹ï¼‰or HG（25 mmol·L⁻¹ï¼‰+ TP（3 µg·L⁻¹ï¼‰or HG（25 mmol·L⁻¹ï¼‰+ TP（10 µg·L⁻¹ï¼‰or DG（5 mmol·L⁻¹ï¼‰+ MNT（24.5 mmol·L⁻¹ï¼‰. After the intervention for 24, 48 and 72 hours, firstly, the activation of podocyte proliferation was investigated. Secondly, the protein expression levels of the epithelial markers in podocytes such as nephrin and podocin, the mesenchymal markers such as desmin and collagen Ⅰ and the EMT-related mediators such as snail were detected respectively. Finally, the protein expression levels of Wnt3α and ß-catenin as the key signaling molecules in Wnt3α/ß-catenin pathway were examined severally. The results indicated that, HG could cause the low protein expression levels of nephrin and podocin and the high protein expression levels of desmin, collagen Ⅰ and snail in podocytes, and inducing podocyte EMT. On the other hand, HG could cause the high protein expression levels of Wnt3α and ß-catenin in podocytes, and activating Wnt3α/ß-catenin signaling pathway. In addition, L-TP had no effect on the activation of podocyte proliferation, the co-treatment of L-TP and HG could significantly recover the protein expression levels of nephrin and podocin, inhibit the protein expression levels of desmin, collagen I and snail in podocytes, thus, further improving podocyte EMT. And that, the co-treatment of L-TP and HG could obviously decrease the high protein expression levels of Wnt3α and ß-catenin induced by HG in podocytes, and inhibit Wnt3α/ß-catenin signaling pathway activation. On the whole, HG can induce podocyte EMT by activating Wnt3α/ß-catenin signaling pathway; L-TP can ameliorate podocyte EMT through inhibiting Wnt3α/ß-catenin signaling pathway activation, which may be one of the effects and molecular mechanisms in vitro.

[Tea polyphenols delays human glomerular mesangial cells senescence induced by high glucose via regulating STAT3/miR-126/telomere signaling pathway activation].

The aim of this paper was to explore the effects and possible mechanisms in vitro of tea polyphenols (TP) delaying human glomerular mesangial cells (HGMCs) senescence induced by high glucose (HG). HGMCs were cultured in vitro and divided into the normal group (N, 5.5 mmol·L⁻¹ glucose), the mannitol group(MNT, 5.5 mmol·L⁻¹ glucose plus 24.5 mmol·L⁻¹ mannitol), the high dose of D-glucose group (HG, 30 mmol·L⁻¹ glucose), the low dose of TP group (L-TP, 30 mmol·L⁻¹ glucose plus 5 mg·L⁻¹ TP) and the high dose of TP group (H-TP, 30 mmol·L⁻¹ glucose plus 20 mg·L⁻¹ TP), which were cultured in 5% CO2 at 37 °C, respectively. Firstly, the effects of TP on the cell morphology of HGMCs were observed after 72 h-intervention. Secondly, the cell cycle, the positive rate of senescence-associated-ß-galactosidase (SA-ß-gal) staining and the telomere length were detected, respectively. Finally, the protein expressions of p53, p21 and Rb in the p53-p21-Rb signaling pathway were investigated, respectively. And the expressions of p-STAT3 and miR-126 were examined severally. The results indicated that HG not only arrested the cell cycle in G1 phase but also increased the positive rate of SA-ß-gal staining, and shortened the telomere length. HG led to the protein over-expressions of p53, p21 and Rb and HGMCs senescence by activating the p53-p21-Rb signaling pathway. In addition, L-TP delayed HGMCs senescence by improving the cell cycle G1 arrest, reducing SA-ß-gal staining positive rate and lengthening the telomere length. L-TP reduced the protein over-expressions of p53, P21 and Rb induced by HG and inhibited the telomere-p53-p21-Rb signaling pathway. Moreover, the expression of p-STAT3 was increased and the expression of miR-126 was decreased in HGMCs induced by HG. L-TP reduced the expression of p-STAT3 and increased the expression of miR-126 in HGMCs. In conclusion, HG could induce HGMCs senescence by activating the telomere-p53-p21-Rb signaling pathway in vitro. L-TP could delay HGMCs senescence through regulating STAT3/miR-126 expressions and inhibiting the telomere-p53-p21-Rb signaling pathway activation. These findings could provide the effective interventions in clinic for preventing and treating renal cell senescence in diabetic kidney disease.

[Pathogenesis and treatment of insulin resistance in chronic kidney disease and interventional effects of Chinese herbal medicine].

The kidney is the target organ of insulin with abundant insulin receptors. Thereinto,the renal intrinsic cells including glomerular podocytes,endothelial cells,mesangial cells,renal tubular epitheliums and collecting duct epithelial cells are all highly sensitive to insulin as the effector cells. Furthermore,the structural and functional abnormalities of these cells are closely related to insulin and its receptors activity. It is reported that the chronic kidney disease（CKD）patients have systemic or renal insulin resistance（IR）. IR is not only the pathogenic factor of CKD but also one of the mechanisms of CKD progression. The pathogenic factors of IR in the CKD patients include the systemic factors and the local factors in muscles and fat cells. The pathogenesis of IR is related to glomeruli,proximal tubules,collecting ducts and corresponding renal intrinsic cells such as podocytes,mesangial cells,renal tubular epitheliums and collecting duct epithelial cells. IR-related signaling pathways include insulin receptor substrate（IRS）/phosphatidylinositol 3 kinase（PI3K）/serine threonine kinase（Akt）pathway,adenosine monophosphate activated protein kinase（AMPK）pathway,glucose transporter4（GLUT4）pathway,nuclear factor（NF）-κB pathway and mitogen activated protein kinase（MAPK）pathway. Among them,IRS1/PI3K/Akt2 is the main signaling pathway of IR in podocytes of glomeruli, thus intervening its activity can improve podocyte injury. In clinic,some classical oral hypoglycemic agents and diuretic including metformin,rosiglitazone,glibenclamide,thiazolidinedione and spironolactone,as well as some extracts from Chinese herbal medicines including astragalus polysaccharides,quercetin,puerarin,emodin,berberine,curcumin and geniposide can both affect insulin and its receptor activity,and regulate IR-related signaling pathways,thereby ameliorating IR and CKD progression. Overall,the pharmacological studies based on IR-related signaling pathways in the renal intrinsic cells of CKD will become one of the developmental directions in the future.

[Pathomechanism and treatment of gut microbiota dysbiosis in chronic kidney disease and interventional effects of Chinese herbal medicine].

The gut microbiota dysbiosis is one of the risk factors in the progression from the advanced chronic kidney disease（CKD）to uremia, characterized by the reduction of probiotics and the increase of opportunistic pathogens including urease-related microbes, endotoxin-related microbes and toxin-related microbes, which can produce uremic toxins. According to the core point of "the gut-kidney axis" theory and "the chronic kidney disease-colonic axis" concept, the gut microbiota dysbiosis aggravates renal damage by accumulating uremic toxins and inducing the systemic micro-inflammation. The preliminary clinical trials and animal experiments show that the probiotics biologicals from Lactobacillus acidophilus or Bifidobacterium, and the prebiotics including inulin and galactooligosaccharides, as well as lubiprostone and activated carbon adsorbents can be used for improving dysfunction of CKD patients with the gut microbiota dysbiosis via reducing uremic toxins and inhibiting the systemic micro-inflammation. But not only that, it is reported that, to some extent, a number of the single Chinese herbal medicine（CHM）, the CHM prescriptions and the CHM extracts（emodin, etc.）with oral or enema administration can also regulate the gut microbiota dysbiosis, protect the intestinal epithelial barrier, reduce uremic toxins accumulation and delay CKD progression. Thereinto, Dahuang Gancao Decoction（the concentrated granule TJ-84）, a classical CHM prescription of rhubarb, can ameliorate uremic toxins accumulation in the animal models with renal failure probably through targeting the gut-kidney axis triggered from gut microbiota, but not targeting the kidney. Based on these results, the interventional studies targeting the gut microbiota-related pathological factors such as tight junction proteins, helper T cells and regulatory T cells in the intestinal tract of the advanced CKD patients will become one of the key development directions in the future.

Multi-glycoside of Tripterygium wilfordii Hook. f. attenuates glomerulosclerosis in a rat model of diabetic nephropathy by exerting anti-microinflammatory effects without affecting hyperglycemia.

Multi-glycoside of Tripterygium wilfordii Hook. f. (GTW) has been proven to be clinically effective in relieving microinflammation in patients with early diabetic nephropathy (DN). However, the therapeutic mechanisms involved in vivo remain unclear. In the process of early DN, microinflammation and activation of p38 mitogen­activated protein kinase (MAPK) and canonical nuclear factor (NF)-κB signaling pathways are the important mechanisms by which hyperglycemia contributes to glomerulosclerosis (GS). Therefore, this study aimed to examine the ameliorative effects of GTW on GS, and then to clarify its anti­microinflammatory mechanisms by inhibiting p38 MAPK and NF-κB signaling activities in the kidney. All rats were divided into 4 groups: the sham group, the sham + GTW group, the vehicle group and the GTW group. The suitable dose of GTW and vehicle were daily administered for 8 weeks after the induction of DN by unilateral nephrectomy combined with intraperitoneal injections of streptozotocin (STZ). The general status of the rats, biochemical parameters, renal histological changes and macrophages in glomeruli, as well as expression of the key proteins in the p38 MAPK and canonical NF-κB signaling pathways and inflammatory cytokines including tumor necrosis factor (TNF)-α, interleukin (IL)-1ß and transforming growth factor (TGF)-ß1 in the kidney were examined, respectively. The results revealed that, GTW improved the general cond-ition and biochemical parameters of the rats, but did not lower blood glucose; GTW attenuated GS and suppressed glomerular microinflammation including the infiltration of ED1+ cells in glomeruli and the protein overexpression of TNF-α, IL-1ß and TGF-ß1 in the kidney; GTW inhibited the protein overexpression of key signaling molecules of p38 MAPK and canonical NF-κB pathways in the kidney including phosphorylated p38 MAPK, phosphorylated inhibitor protein IκB and NF-κB (p65). On the whole, we expounded that GTW, as a natural regulator in vivo, alleviates GS without affecting hyperglycemia, by exerting anti-microinflammatory effects, including reducing macrophage infiltration in glomeruli, suppressing TNF-α, IL-1ß and TGF-ß1 overexpression in the kidney and inhibiting p38 MAPK and NF-κB signaling activities.

[Effects and mechanisms of Shenkang injection promoting extracellular matrix degradation via regulating ERK1/2/MMPs signaling pathway in renal failure rats].

This study aimed to clarify preliminarily the effects and mechanisms of Shenkang injection (SKI) promoting extracellular matrix(ECM)degradation via regulating extracellular-signal regulated protein kinase(ERK)1/2/matrix metalloproteinases(MMPs)signaling pathway in renal failure rats. Twenty rats were randomly divided into 4 groups：the Sham group,the Model group,the SKI group and the Enalapril maleate(EM)group. The model rats with renal failure were induced by intragastric administration of adenine and unilateral ureteral obstruction(UUO). After modeling, the rats in SKI group and EM group were intervened by intraperitoneal injection of SKI or intragastric administration of the EM suspension,while the rats in Sham group and Model group were administrated with distilled water respectively for 3 weeks. The 24 h urinary protein excretion(Upro)and urinary N-acety1-ß-D-glucosaminidase(UNAG)in all rats were tested after drug administration. All rats were sacrificed after drug administration for 3 weeks,blood and kidney were collected,renal morphological characteristics were observed. Furthermore,serum biochemical indices and the protein expressions of collagen type IV(CIV),MMP-2,MMP-9,tissue inhibitors of metalloproteinase(TIMP)-1,ERK1/2 and phosphorylated-ERK1/2(p-ERK1/2)in the kidney were evaluated respectively. The results indicated that,after the intervention of SKI,serum creatinine(Scr),blood urea nitrogen(BUN),uric acid(UA),albumin(Alb),Upro,UNAG and renal morphological change in model rats were improved at different levels,respectively. Moreover,these actions were similar to EM. In addition to these,SKI adjusted the protein expressions of MMP-2,MMP-9 and TIMP-1,and down-regulated the protein expressions of p-ERK1/2 in the kidney. Moreover,these actions were different from EM. In conclusion,SKI promotes ECM degradation and delays the progression of renal failure possibly through regulating ERK1/2 signaling pathway activation in the kidney and intervening MMPs/TIMP-1 expressions in vivo.