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Objective:To investigate the protective effects and mechanism of Shenyan 1 Prescription on renal fibrosis of unilateral ureteral obstruction (UUO) rats through TGF- β 1/Smad homologous 3 (Smad3) pathway regulating ferroptosis.Methods:Totally 48 male SD rats were divided into four groups: sham-operation group, UUO model group, and Shenyan 1 Prescription low-(10 drug/kg) , and high-dosage (20 crude drug/kg) groups according to random number table method, with 12 rats in each group. The UUO model was induced by the method of unilateral ureteral obstruction except for those sham-operation group. After modeling, rats received corresponding drugs or normal saline by gavage for 4 weeks, once per day. After 4 weeks, the body mass and the left kidney weight were measured. The 24 h urine protein and the levels of serum albumin (ALB), alanine aminotransferase (ALT), serum creatinine (SCr) and blood urea nitrogen (BUN) were detected by biochemical analysis method; the ROS level in renal tissue was measured using a chemical fluorescence assay kit, and the SOD and MDA levels in left renal tissue of rats were measured using ELISA method; the morphology of renal tissue and the specific blue staining of hemosiderin were observed using HE and Prussian blue staining methods, respectively; the expressions of transforming growth factor-β1 (TGF-β1), Smad3, glutathione peroxidase 4 (GPX4), and solute carrier family 1 member 5 (SLC1A5) were detected by Western blot.Results:Compared with the model group, the 24 h urinary protein excretion in Shenyan 1 Prescription high-dosage group decreased ( P<0.05), the serum ALB level increased ( P<0.05), the ALT level decreased ( P<0.05), and the expression of SLC1A5 in renal tissue decreased ( P<0.05); the left kidney weight/body decreased in Shenyan 1 Prescription low- and high-dosage groups ( P<0.05); the levels of serum ROS and MDA decreased ( P<0.05), and the activity of SOD significantly increased ( P<0.05); the expressions of TGF-β1 and Smad3 in renal tissue decreased ( P<0.05), and the expression of GPX4 increased ( P<0.05), and the renal pathological injury and ion deposition were improved. Conclusion:Shenyan 1 Prescription has a protective effect on the structure and function of renal tissues in UUO rats through regulating ferroptosis via inhibition of the TGF-β1/ Smad3 pathway to inhibit renal fibrosis of UUO rats.
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AIM:Bone morphogenetic protein 7(BMP7)reduces the expression of Yes-related protein 1(YAP1)by down-regulating Ajuba level and decreasing extracellular matrix(ECM)deposition.This study aimed to inves-tigate the influence of these factors on modifying the degree of renal fibrosis in rats with diabetic nephropathy.METH-ODS:Eighteen Sprague-Dawley(SD)rats were randomly divided into three groups:the normal control(NC)group,the diabetes mellitus(DM)group,and the DM group treated with BMP7 overexpressing adeno-associated virus(DM+rAAV-BMP7).Each group consisted of six rats.Diabetic kidney disease(DKD)was established in the DM and DM+rAAV-BMP7 groups by injecting 55 mg/kg streptozotocin(STZ)via the tail vein.NRK-52E cells were divided into three groups:the normal glucose(NG)group,the high glucose(HG)group,and the high glucose group treated with recombinant hu-man BMP7(HG+rhBMP7)group.Pathological changes in renal tissues were observed using hematoxylin and eosin(HE)and Sirius red staining.Immunohistochemical staining was performed to examine the expression sites of Ajuba and YAP1 in the renal cortex.Western blot analysis was conducted to determine the expression levels of BMP7,Ajuba,YAP1,colla-gen type Ⅲ(Col-Ⅲ),and fibronectin(FN)in the rat renal cortex and NRK-52E cells.RT-qPCR was used to measure the mRNA levels of Ajuba and YAP1 in the rat renal cortex.RESULTS:Biochemical indices revealed significantly ele-vated levels of blood glucose,serum creatinine,triglycerides,total cholesterol,and 24-hour urinary protein in the DM group compared to the NC group(P<0.05).In the DM+rAAV-BMP7 group,the levels of serum creatinine,24-hour uri-nary protein,triglycerides,and total cholesterol were lower than those in the DM group(P<0.05).Pathological staining demonstrated that the renal interstitium of the DM group exhibited inflammatory cell infiltration,fibrous tissue,collagen fi-ber deposition,disordered renal tubule arrangement,atrophy,and vacuolar degeneration,which were ameliorated in the DM+rAAV-BMP7 group.Immunohistochemistry revealed that Ajuba and YAP1 were mainly expressed in the cytoplasm and nucleus,with high expression in the cytoplasm of the DM group,which was significantly decreased in the DM+rAAV-BMP7 group.Western blot results indicated that the protein levels of FN,Col-Ⅲ,Ajuba,and YAP1 were up-regulated in the DM and the HG groups(P<0.05),but significantly down-regulated in the DM+rAAV-BMP7 group(P<0.05).RT-qP-CR results demonstrated that the mRNA levels of Ajuba and YAP1 were higher in the DM group and significantly lower in the DM+rAAV-BMP7 group(P<0.05).CONCLUSION:The overexpression of BMP7 can ameliorate renal fibrosis in rats with DKD.This effect is likely mediated by the down-regulation of Ajuba,reduction of YAP1 expression,and subse-quent inhibition of ECM deposition.
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AIM To explore the effect of Heidihuang Pills on renal fibrosis in a rat model of chronic renal failure(CRF)and its mechanism.METHODS Wistar rats were randomly divided into the blank group for normal feeding and the model group for the establishment of CRF rat models by 5/6 nephrectomy.Subsequently,the successfully established rat models were randomly divided into the model group,the Heidihuang Pills group(10.43 g/kg),and the Heidihuang Pills+IGF-1R blocker(JB1)group for a regimen of 7-day subcutaneous injection of 18 μg/kg JB1 followed by gavage of 10.43 g/kg Heidihuang Pills.Eight weeks after the administration,the rats had their serum levels of Scr and BUN detected;their pathological changes of renal tissue observed by HE and Masson staining;their renal protein expressions of TGF-β,HIF-1α and α-SMA detected by immunohistochemistry;their renal protein expressions of IGF-1R and TGF-β detected by Western blot;and their renal mRNA expressions of IGF-1R and TGF-β detected by RT-qPCR.RESULTS Compared with the blank group,the model group displayed increased serum levels of Scr and BUN(P＜0.05);increased,degree of renal fibrosis,and renal fibrosis area(P＜0.05);increased renal expressions of TGF-β,HIF-1α,α-SMA proteins and TGF-β mRNA(P＜0.05);and decreased expressions of IGF-1R mRNA and protein(P＜0.05).Compared with the model group,the Heidihuang Pills group displayed decreased serum Scr and BUN levels(P＜0.05);decreased inflammatory cells in renal interstitium and the fibrosis degree(P＜0.05);decreased renal expressions of TGF-β,HIF-1α,α-SMA proteins and TGF-β mRNA(P＜0.05);and increased expressions of IGF-1R mRNA and protein(P＜0.05).However,the administration of JB1 could weaken the improvement effect of Heidihuang Pills on renal fibrosis in CRF rats(P＜0.05).CONCLUSION Heidihuang Pills can inhibit the renal fibrosis in CRF rats,and the inhibition process is related to up-regulated IGF-1 expression and promoted combination of IGF-1 and IGF-1R.
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OBJECTIVE To investigate the effects of Taohong siwu decoction modified granules on podocyte epithelial- mesenchymal-transition （EMT） and renal fibrosis in diabetic kidney disease （DKD） model rats. METHODS Eight rats were selected as normal group （ordinary feed）； the remaining rats were given a high-glucose and high-fat diet combined with intraperitoneal injection of streptozotocin （35 mg/kg） to induce the DKD model. Model rats were randomly divided into model group， irbesartan group [positive control， 13.5 mg/（kg·d）] and modified Taohong siwu decoction group [6.48 g/（kg·d）]， with 8 rats in each group. All groups were given relevant medicine intragastrically， once a day， for 16 consecutive weeks. Twenty-four- hour urinary total protein （24 h UTP） was detected at the end of the 4th， 8th， 12th and 16th week of administration. After the last medication， the body mass， water intake， food intake， urine output， the levels of fasting blood glucose， serum creatinine （Scr） and blood urea nitrogen （BUN） as well as mRNA and protein expressions of P-cadherin， nephrin， α -smooth muscle actin （α-SMA）， Wilms’ tumor gene 1 （WT1）， transforming growth factor-β1（ TGF-β1） and type Ⅳ collagen （Col-Ⅳ） in renal tissue were determined. The pathological and morphological changes in renal tissue were observed and the thickness of the glomerular basement membrane was determined. RESULTS Compared with the model group， 24 h UTP of rats was significantly decreased in modified Taohong siwu decoction group since the 8th weekend （P＜0.05）； the body weight of rats increased significantly， but the amount of water intake and urine decreased significantly； Scr and BUN level， mRNA expression of α-SMA， mRNA and protein expressions of TGF-β1 and Col-Ⅳ were significantly reduced， while the mRNA expressions of P-cadherin， nephrin and WT1 were increased significantly （P＜0.05）； the protein deposition of α-SMA was reduced， protein depositions of P-cadherin， nephrin and WT1 were increased； the pathological damage and fibrosis of renal tissue were relieved； the thickness of glomerular basement membrane was decreased significantly （P＜0.05）. CONCLUSIONS Taohong siwu decoction modified granules can inhibit the EMT of podocyte in DKD model rats， and alleviate renal pathological damage and podocyte damage， thus protecting renal function， and delaying the process of renal fibrosis.
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OBJECTIVE To investigate the effects of Taohong siwu decoction modified granules on podocyte epithelial- mesenchymal-transition （EMT） and renal fibrosis in diabetic kidney disease （DKD） model rats. METHODS Eight rats were selected as normal group （ordinary feed）； the remaining rats were given a high-glucose and high-fat diet combined with intraperitoneal injection of streptozotocin （35 mg/kg） to induce the DKD model. Model rats were randomly divided into model group， irbesartan group [positive control， 13.5 mg/（kg·d）] and modified Taohong siwu decoction group [6.48 g/（kg·d）]， with 8 rats in each group. All groups were given relevant medicine intragastrically， once a day， for 16 consecutive weeks. Twenty-four- hour urinary total protein （24 h UTP） was detected at the end of the 4th， 8th， 12th and 16th week of administration. After the last medication， the body mass， water intake， food intake， urine output， the levels of fasting blood glucose， serum creatinine （Scr） and blood urea nitrogen （BUN） as well as mRNA and protein expressions of P-cadherin， nephrin， α -smooth muscle actin （α-SMA）， Wilms’ tumor gene 1 （WT1）， transforming growth factor-β1（ TGF-β1） and type Ⅳ collagen （Col-Ⅳ） in renal tissue were determined. The pathological and morphological changes in renal tissue were observed and the thickness of the glomerular basement membrane was determined. RESULTS Compared with the model group， 24 h UTP of rats was significantly decreased in modified Taohong siwu decoction group since the 8th weekend （P＜0.05）； the body weight of rats increased significantly， but the amount of water intake and urine decreased significantly； Scr and BUN level， mRNA expression of α-SMA， mRNA and protein expressions of TGF-β1 and Col-Ⅳ were significantly reduced， while the mRNA expressions of P-cadherin， nephrin and WT1 were increased significantly （P＜0.05）； the protein deposition of α-SMA was reduced， protein depositions of P-cadherin， nephrin and WT1 were increased； the pathological damage and fibrosis of renal tissue were relieved； the thickness of glomerular basement membrane was decreased significantly （P＜0.05）. CONCLUSIONS Taohong siwu decoction modified granules can inhibit the EMT of podocyte in DKD model rats， and alleviate renal pathological damage and podocyte damage， thus protecting renal function， and delaying the process of renal fibrosis.
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ObjectiveTo observe the effects of Hirudo， Notoginseng Radix et Rhizoma， and drug pair on renal pathological morphology and protein phosphatase 2A （PP2A）/adenylate activated protein kinase （AMPK）/mammalian target of rapamycin （mTOR） signal pathway in rats with chronic renal failure （CRF）. MethodThe 55 male SD rats were randomly divided into a normal group （n=11） and a modeling group （n=44）. The normal group was fed conventionally， and the modeling group was given 0.25 g·kg-1·d-1 adenine by gavage for 28 days to replicate the CRF model. After successful modeling， rats were randomly divided into model group， Hirudo group （3 g·kg-1·d-1）， Notoginseng Radix et Rhizoma group （3 g·kg-1·d-1）， and Hirudo + Notoginseng Radix et Rhizoma group （3 g·kg-1·d-1）， with 9 rats in each group. The normal group and model group were given a constant volume of normal saline by intragastric administration for 30 days. At the end of the experiment， the levels of serum creatinine （SCr） and urea nitrogen （BUN） in all groups were measured. The renal pathological morphology changes were observed by hematoxylin-eosin （HE） staining， Masson staining， and electron microscopy. The mRNA expressions of PP2A， AMPK， and mTOR were detected by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. The protein expression levels of PP2A， AMPK， phosphorylation（p）-AMPK， mTOR， and p-mTOR in renal tissue were detected by Western blot. ResultCompared with the normal group， the renal pathological structure changes were obvious， and the levels of SCr and BUN were significantly increased. The mRNA expression of PP2A， protein expression of PP2A， and p-mTOR/mTOR expression were significantly increased， and the p-AMPK/AMPK was significantly decreased in the model group （P<0.05）. Compared with the model group， the renal pathological morphology changes were significantly improved， and the levels of SCr and BUN were significantly decreased. The mRNA expression of PP2A， protein expression of PP2A， and p-mTOR/mTOR expression in the renal tissue were significantly decreased， and the p-AMPK/AMPK was significantly increased （P<0.05） in all groups after drug intervention. In addition， the effect in the Hirudo+Notoginseng Radix et Rhizoma group was better. The mRNA expression levels of AMPK and mTOR in the renal tissue were not significantly different among the normal group， model group， and other groups. ConclusionThe efficacy of Hirudo and Notoginseng Radix et Rhizoma pairs in improving renal fibrosis in rats with CRF is significantly better than that of the single drug， and its improvement on renal fibrosis in rats with CRF may be related to the regulation of PP2A/AMPK/mTOR signaling pathway.
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Renal allograft fibrosis is one of the common and severe complications after kidney transplantation, which seriously affects the function and survival rate of renal allograft, and may even lead to organ failure and patient death. At present, the researches on renal allograft fibrosis are highly complicated, including immunity, ischemia-reperfusion injury, infection and drug toxicity, etc. The diagnosis and treatment of renal allograft fibrosis remain extremely challenging. In this article, the latest research progress was reviewed and the causes, novel diagnosis and treatment strategies for renal allograft fibrosis were investigated. By improving diagnostic accuracy and optimizing treatment regimen, it is expected to enhance clinical prognosis of kidney transplant recipients, aiming to provide reference for clinicians to deliver proper management for kidney transplant recipients.
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Renal fibrosis is a common pathological change from development to end-stage renal diseases in all progressive chronic kidney diseases. Renal fibrosis after kidney transplantation will severely affect the renal graft function. Macrophages are characterized with high heterogeneity and plasticity. During the process of kidney injury, macrophages are recruited, activated and polarized by local microenvironment, and participate in the process of renal tissue injury, repair and fibrosis through multiple mechanisms. Recent studies have shown that macrophages may transit into myofibroblasts and directly participate in the formation of renal fibrosis. This process is known as macrophage-myofibroblast transition. Nevertheless, the regulatory mechanism remains elusive. In this article, the role of macrophages in renal fibrosis, the characteristics of macrophage-myofibroblast transition and the possible regulatory mechanism were reviewed, aiming to provide reference for relevant research of renal fibrosis.
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This paper investigates the effect of myricetin (MYR) on renal fibrosis induced by unilateral ureteral obstruction (UUO) and common bile duct ligation (CBDL) in mice and its mechanism. The animal experiment has been approved by the Ethics Committee of China Pharmaceutical University (NO: 2022-10-020). Thirty-five ICR mice were divided into control, UUO, UUO+MYR, CBDL and CBDL+MYR groups. H&E and Masson staining were used to detect pathological changes in kidney tissues. Western blot (WB) was used to detect the expression of fibrosis-related proteins in renal tissue, and total superoxide dismutase (SOD) activity detection kit (WST-8) was used to detect the changes of total SOD in renal tissue of CBDL mice. In vitro, HK-2 cells and transforming growth factor beta 1 (TGF-β1, 10 ng·mL-1) were used to induce fibrotic model, and high glucose (30 mmol·L-1) was used to induce oxidative stress model, and then treated with different concentrations of MYR, WB was used to detect the expression of fibrosis and oxidative stress-related proteins, while NIH/3T3 cells were treated with different concentrations of MYR, and their effects on cell proliferation were detected by 5-bromo-2′-deoxyuridine (Brdu). The results showed that the renal lesions in UUO group and CBDL group were severe, collagen deposition was obvious, the expression of collagen-Ⅰ (COL-Ⅰ), α-smooth muscle actin (α-SMA), fibronectin (FN), vimentin and plasminogen activator inhibitor-1 (PAI-1) protein was up-regulated, and the activity of SOD enzyme in CBDL group was significantly decreased. MYR partly reversed the above changes after treatment. MYR inhibited the proliferation of NIH/3T3 cells but had no effect on the proliferation of HK-2 cells, and decreased the upregulation of PAI-1, FN and vimentin in HK-2 cells stimulated by TGF-β1. MYR can also up-regulate the down-regulation of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) in HK-2 cells stimulated by high glucose. To sum up, MYR can improve renal fibrosis in vivo and in vitro, probably by inhibiting the proliferation of fibroblasts and activating Nrf2/HO-1 signal pathway to inhibit oxidative stress.
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Diabetic kidney disease （DKD） is a common complication of diabetes and a leading cause of end-stage kidney disease. Its pathogenesis is complex， and it presents a significant challenge in treatment， gradually becoming a major global public health issue. One of the main pathological changes in DKD is tubulointerstitial fibrosis， clinically characterized by proteinuria and declining kidney function， which severely impacts the daily life of patients. Currently， western medicine commonly uses methods such as controlling blood sugar and blood pressure， and reducing proteinuria to treat DKD， but the efficacy is unsatisfactory， and there are many side effects. As reported， traditional Chinese medicine （TCM） treatment for DKD has many advantages， such as low cost， significant efficacy， and minimal adverse reactions. More researchers focusing on DKD are turning their attention to TCM， and progress has been made in related studies both in China and abroad. The Wnt/β-catenin signaling pathway is relatively evolutionarily conserved and plays a crucial role in normal biological development and the entire life process. Studies have demonstrated that abnormal activation of the Wnt/β-catenin signaling pathway is related to renal fibrosis， which coincides with TCM theory of "collateral diseases". By reviewing relevant literature， this article reviewed the Wnt/β-catenin signaling pathway and its role in DKD and summarized the research status of TCM monomers， single drug extracts， and TCM formulas in improving renal fibrosis and treating DKD through the improvement of glomerular mesangial cells， renal tubular epithelial cells， and podocyte injury， aiming to provide new ideas and directions for TCM treatment of DKD.
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OBJECTIVE To explore the intervention effect and related mechanism of Tongxinluo capsule on renal fibrosis in rats with diabetic nephropathy （DN）. METHODS Eight rats were selected as control group （ordinary feed）， the remaining rats were given high-glucose and high-fat diet combined with ip injection of streptozotocin （35 mg/kg） to induce DN model. Model rats were randomly divided into model group （purified water）， irbesartan group （positive control， 14.12 mg/kg） and Tongxinluo capsule group （0.3 g/kg）， including 12 rats in the model group and 11 rats for each of the other two groups. All groups were given relevant medicine or water intragastrically， once a day， for 16 consecutive weeks. After the last medication， fasting blood glucose and 24 h urinary total protein （24 h UTP） were detected. Pathological changes in renal cortex of rats in each group were observed. Serum levels of tissue-type plasminogen activator （PA） and plasminogen activator inhibitor 1 （PAI-1） were measured. mRNA expressions of transforming growth factor-β（1 TGF-β1）， type Ⅳ collagen（COL-Ⅳ）， Wnt4 and β-catenin in renal cortex of rats were detected. The protein depositions or expressions of TGF-β1， COL-Ⅳ， focal adhesion kinase （FAK）， integrin-linked kinase （ILK）， E-cadherin， PA， PAI-1， Wnt4 and β-catenin in renal cortex of rats were observed or determined. RESULTS Compared with model group， 24 h UTP of rats in Tongxinluo capsule group were all significantly reduced （P＜0.05）； pathological damage and fibrosis of renal cortex were relieved； the expression of PA in serum and renal cortex was significantly increased， while PAI-1 level was significantly reduced （P＜0.05）； the depositions of COL-Ⅳ and TGF-β1 in renal cortex were all reduced， and corresponding mRNA expression was decreased significantly （P＜0.05）； the depositions of ILK and FAK were decreased， while the deposition of E-cadherin was increased； protein and mRNA expressions of Wnt4 and β-catenin were significantly reduced （P＜0.05）. CONCLUSIONS Tongxinluo capsule can relieve pathological damage to renal tissue and renal fibrosis of DN model rats， and reduce extracellular matrix deposition. The mechanism may be related to regulation of fibrinolytic system activity， the decrease of ILK and FAK expression， and inhibition of Wnt/β-catenin signaling pathway.
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A quantitative analysis method for six principal active constituents (acubin, geniposidic acid, chlorogenic acid, pinoresinol di-O-glucopyranoside, geniposide, and pinoresinol 4-O-glucopyranoside) of crude Eucommiae Cortex (EC) and its salt-processed product extracts was developed to investigate and compare their pharmacokinetic behaviors in adenine-induced renal fibrotic rats in vivo. UHPLC-QqQ-MS/MS technology was employed. Scan was conducted in negative ion mode and quantitative determination was carried out by MRM paired ion. The established method was fully validated by specificity, linearity, precision, accuracy, stability, recovery, and matrix effect, and the results of methodological investigation met the requirements of biological sample analysis. Then, a quick, sensitive, and accurate method was successfully established, which could simultaneously measure the contents of six active constituents of crude and salt-processed EC extracts in rat plasma. After a single administration to renal fibrotic rats of crude EC and its salt-processed product extracts, the plasma concentration of each constituent at different time points was measured, the pharmacokinetic parameters were calculated and the concentration time curves were structured. The experiment was approved by the experimental animal ethics committee from Nanjing University of Chinese Medicine (No. 202103A008). The results showed that compared to the crude Eucommiae Cortex group, the tmax of aucubin, pinoresinol di-O-glucopyranoside, geniposide, and pinoresinol 4-O-glucopyranoside in the salt-processed Eucommiae Cortex group rat plasma were significantly lower than those in the crude group (P < 0.05, P < 0.01); the Cmax and AUC0-48 h of chlorogenic acid, the Cmax, AUC0-48 h and AUC0-∞ of pinoresinol di-O-glucopyranoside, and the Cmax of geniposide and pinoresinol 4-O-glucopyranoside were significantly higher than those in the crude group (P < 0.05, P < 0.01). Our investigation found that compared to crude Eucommiae Cortex, a variety of active ingredients could play a role of quick effect with higher peak blood concentration and bioavailability after oral administration of salt-processed Eucommiae Cortex, which were consistent with the traditional Chinese medicine theory of "salt-processing enhancing drug into kidney meridian", providing an experimental basis for the selection of quality control indexes and the in-depth study of processing mechanisms and metabolic rules in vivo of Eucommiae Cortex and its salt-processed product.
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ObjectiveTo investigate the role of bile acid receptor TGR5 activation in renal fibrosis induced by unilateral ischemia reperfusion injury and contralateral nephrectomy （uIRIx） model. MethodsIn vivo： C57BL/6J mice were randomly divided into Sham group， uIRIx group and uIRIx+ lithcholic acid （LCA） group with 6 mice in each group. Kidney fibrosis was induced by uIRIx model， kidney function was evaluated by blood and urine biochemical indexes， and the degree of kidney injury was evaluated by HE staining. Masson staining and immunohistochemistry were used to evaluate the degree of renal fibrosis， and Western Blotting was used to detect the expression of related index proteins of renal cortical fibrosis. Sham group and uIRIx group were set in TGR5+/+ mice and TGR5-/- mice respectively， with 6 mice in each group. The degree of renal fibrosis in each group was detected by Western Blotting. In vitro： TGF-β1 was administered to induce pro-fibrosis response in human renal tubular epithelial cell line （HK2 cells）， LCA was used for drug intervention， cytoskeleton was labeled with phalloidin-FITC staining and the expression of fibrosis related indicator protein in HK2 cells was detected by Western Blotting. ResultsIn vivo： Compared with the Sham group， plasma creatinine level （P=0.007） and urinary albumin/creatinine ratio （P=0.041） in uIRIx group were significantly increased， renal cortical protein TGR5 expression （P=0.002） was decreased， Fibronectin expression （P=0.020） and COL1A1 expression （P<0.001） were increased. At the same time， the kidney structure was damaged and collagen deposition was aggravated. LCA intervention effectively improved the kidney function and alleviated the degree of kidney injury and fibrosis. TGR5 gene knockout increased uIRIx-induced Fibronectin expression （P<0.001） and COL1A1 expression （P=0.001） compared with TGR5+/+ mice. In vitro： TGF-β1 induced morphological changes of HK2 cells， cytoskeletal depolymerization and recombination， and promoted the up-regulation of fibrosis index protein. LCA effectively inhibited the morphological changes and skeletal depolymerization induced by TGF-β1， and down-regulated the expression of fibrosis related indicator proteins. ConclusionsLCA alleviated renal fibrosis induced by uIRIx model， and knockout of TGR5 gene aggravated uIRIx induced renal fibrosis； In HK2 cells， LCA alleviated fibrogenic reaction induced by TGF-β1. This indicates that activation of TGR5 alleviates renal fibrosis induced by uIRIx.
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Diabetic nephropathy （DN） is one of the common chronic kidney diseases （CKD） worldwide and a major cause of end-stage renal disease （ESRD）， seriously threatening and affecting the life and health of the global population. Currently， the pathogenesis of DN is considered to be closely related to factors such as glucose metabolism disorders， abnormal lipid metabolism， oxidative stress， activation of inflammatory factors， autophagy， and cell apoptosis in the continuous high-glucose environment of the body. Renal fibrosis is an important pathological feature and ultimate pathological outcome of DN. Timely intervention in renal fibrosis is of significant clinical and practical importance for the prevention and treatment of DN. Due to the limitations of western medicine in treating DN， traditional Chinese medicine （TCM） intervention in the process of renal fibrosis in DN has been widely used as a routine and potential treatment method due to its multi-component， multi-effect， and multi-target effects， effectively delaying the progression of the disease. It has been found that the Notch signaling pathway plays an important role in the development and maintenance of homeostasis in the body， and abnormal activation of the Notch signaling pathway is associated with DN. Activation of this signaling pathway plays a key role in the process of renal fibrosis. This article reviewed the regulatory mechanism of the Notch signaling pathway in renal fibrosis in DN， focusing on the relationship between targeting Notch signaling pathway by Chinese medicinal monomers and prescriptions and renal fibrosis in DN in order to provide a theoretical basis for the development of new drugs， basic research， and clinical application of TCM in the prevention and treatment of DN.
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Renal fibrosis， the final pathological outcome of end-stage chronic kidney diseases， is associated with inflammation， oxidative stress， epithelial-mesenchymal transdifferentiation （EMT）， and extracellular matrix deposition. It belongs to the categories of edema， ischuria， anuria and vomiting， and consumptive disease in traditional Chinese medicine （TCM）， with the key pathogenesis of Qi deficiency and blood stasis and the primary treatment principle of replenishing Qi and activating blood. Astragali Radix-Salviae Miltiorrhizae Radix et Rhizoma mainly contains astragalosides， polysaccharides， calycosin， salvianolic acid， and tanshinone， with the effect of tonifying Qi and activating blood. Studies have shown that this herb pair and its active components can delay the progress of renal fibrosis by regulating multiple signaling pathways. With consideration to the pathogenesis of Qi deficiency and blood stasis， this article reviews the research progress in the mitigation of renal fibrosis by Astragali Radix-Salviae Miltiorrhizae Radix et Rhizoma from the aspects of protecting glomerular filtration barrier， inhibiting EMT and mesangial cell proliferation， improving renal hemodynamics， and protecting renal function. Furthermore， the mechanisms were summarized. Specifically， Astragali Radix-Salviae Miltiorrhizae Radix et Rhizoma and its effective components can improve mitochondrial function and fatty acid metabolism， alleviate endoplasmic reticulum stress and autophagy disorders， and inhibit immune inflammation and oxidative stress by regulating nuclear factor E2-related factor 2 （Nrf2）/PTEN-induced kinase 1 （Pink1）， Nrf2/antioxidant response element （ARE）， tumor necrosis factor-α （TNF-α）/nuclear transcription factor-κB （NF-κB）， miR-21/Smad7/transforming growth factor beta （TGF-β）， Wnt/β-catenin， long non-coding RNA-taurine up-regulated gene 1 （lncRNA-TUG1）/tumor necrosis factor receptor-associated factor 5 （TRAF5）， Ras-related C3 botulinum toxin substrate 1 （Rac1）/cell division cycle protein 42 （CDC42）， Ras homolog （Rho）/Rho-associated coiled-coil containing protein kinase （ROCK）， phosphatidylinositol-3-kinase （PI3K）/protein kinase B （Akt）， Janus kinase （JAK）/signal transducer and activator of transcription （STAT）， peroxisome proliferator-activated receptor α （PPARα）/peroxisome proliferator-activated receptor γ coactivator l alpha （PGC-1α）， and p38 mitogen-activated protein kinase （p38 MAPK）. This review aims to provide references for the relevant research， give play to the role of Astragali Radix-Salviae Miltiorrhizae Radix et Rhizoma， and provide guidance for the clinical treatment of renal fibrosis.
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ObjectiveTo investigate the protective effect and underlying mechanism of Gandou Fumu decoction （GDFMT） on renal fibrosis in a mouse model of Wilson's disease. MethodSixty adult male toxic milk （TX） mice were randomly divided into a model group， high-， medium-， and low-dose GDFMT groups， and a positive control （penicillamine） group， and another 12 wild-type mice were assigned to the normal group. The high-， medium-， and low-dose GDFMT groups were administered GDFMT at 13.92， 6.96， 3.48 g·kg-1， respectively， and the positive control group received penicillamine at 0.1 g·kg-1， while the model and normal groups were given an equal volume of 0.9% saline solution by gavage once a day for 4 consecutive weeks. Enzyme-linked immunosorbent assay （ELISA） was used to measure the levels of blood urea nitrogen （BUN）， creatinine （CRE）， type Ⅲ procollagen （PC-Ⅲ）， and type Ⅳ collagen （C-Ⅳ） in the serum. Histological changes in the mouse kidneys were examined by hematoxylin-eosin （HE） and Masson's trichrome staining. Immunofluorescence was used to assess the protein expression of leptin， Janus kinase 2 （JAK2）， and signal transducer and activator of transcription （STAT） in renal cells. Real-time polymerase chain reaction （Real-time PCR） was performed to analyze the mRNA expression levels of leptin， leptin receptor（OB-R）， JAK2， and STAT. Western blot was used to detect the expression of transforming growth factor-β1 （TGF-β1） and monocyte chemoattractant protein-1 （MCP-1）. ResultCompared with the normal group， the model mice exhibited a significant increase in BUN， CRE， PC-Ⅲ， and C-Ⅳ levels （P<0.01）. Compared with the model group， the high- and medium-dose GDFMT groups and the penicillamine groups showed significant decreases in these parameters （P<0.05， P<0.01）， with the high-dose GDFMT group demonstrating the most significant reduction （P<0.01）. The histological examination of renal tissue revealed fibrosis in the model group， while the fibrotic damage was mitigated to varying degrees after drug intervention， with improvement in fibrosis. Immunofluorescence results showed that leptin， JAK2， and STAT3 protein expression levels were significantly upregulated in the renal fibrosis of the model group. After GDFMT intervention， the fluorescence intensity decreased， with the high-dose GDFMT group showing the lowest intensity. Real-time PCR results demonstrated that leptin， OB-R， JAK2， and STAT3 mRNA expression levels were significantly elevated in the model group compared with those in the normal group， while the high- and medium-dose GDFMT groups and the penicillamine group showed significant reductions in their expression levels （P<0.05， P<0.01）. Western blot analysis revealed that TGF-β1 and MCP-1 expression levels were significantly increased in the model group （P<0.01）， and the high- and medium-dose GDFMT groups exhibited significant reductions in their expression levels （P<0.01）. ConclusionGDFMT can alleviate renal fibrosis damage in TX mice， and its mechanism of action may be related to the regulation of leptin and the JAK/STAT signaling pathway.
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Renal fibrosis is the main pathological foundation of chronic kidney diseases progressing to end-stage renal diseases. With complex pathogenic factors and prolonged disease course， it threatens the quality of life of patients and brings about heavy financial burden to medical care. In the instance of intestinal flora disturbance， the internal homeostasis is broken， resulting in various "imbalances". The "combination of state and target" endows the syndrome differentiation-based treatment of renal fibrosis with new connotation from the perspective of intestinal flora reconstruction and microbial diversity restoration. In addition， traditional Chinese medicine （TCM）-targeted intervention of intestinal microecology has unique advantages under the principle of "treating different diseases with the same method"， which can guide the diagnosis and treatment of renal fibrosis. To be specific， TCM emphasizes macroscopic regulation of state and microscopic targeting. In view of the inflammatory response， accumulation of endotoxin， and excessive deposition of extracellular matrix （ECM） in the process of renal fibrosis， the strategies for treating this disease have been developed， such as alleviating dampness，removing turbid toxin， and relieving deficiency and stasis. Famous prescriptions in ancient books or compound Chinese medicine prescriptions， classical formulas， Chinese medicine monomers， or active components of Chinese medicine target intestinal microecology. Therefore， from the perspective of common pathogenic factors of renal diseases （renal fibrosis） or pathological product-intestinal microecological imbalance， this article combines TCM basic theory with modern medical pathogenesis， and summarizes the research on TCM intervention of renal fibrosis by regulating intestinal microecology and the scientific connotation of renal fibrosis， which is expected to provide ideas and methods for the product development and related preparations and in-depth molecular biological research.
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OBJECTIVE@#To observe the effect of Shenbing Decoction Ⅲ for improving renal function and pathology in rats with 5/6 nephrectomy and analyze its therapeutic mechanism for renal fibrosis in chronic kidney disease using network pharmacology combined with molecular docking.@*METHODS@#Forty male SD rats were randomized into two groups to receive two-staged 5/6 nephrectomy (n=30) or sham operation (n=10), and 2 weeks after the final operation, serum creatinine level of the rats was measured. The rats with nephrectomy were further randomized into Shenbing Decoction Ⅲ group, losartan group and model group for daily treatment with the corresponding drugs via gavage starting at 1 week after 5/6 nephrectomy. After 16 weeks of treatment, serum creatinine and urea nitrogen levels of the rats were measured, and HE staining and Western blotting were used to examine the changes in renal pathology and fibrosis-related factors. Network pharmacology combined with molecular docking study was performed to explore the therapeutic mechanism Shenbing Decoction Ⅲ against renal fibrosis in chronic kidney disease, and Western blotting was used to verify the expressions of the core targets.@*RESULTS@#Compared with those in the model group, the rats receiving 5/6 nephrectomy and Shenbing Decoction Ⅲ treatment showed significantly reduced serum creatinine and urea nitrogen levels, lessened renal pathologies, and improvement of the changes in epithelial mesenchymal transition-related proteins. Network pharmacological analysis showed that the main active ingredients of Shenbing Decoction Ⅲ were acacetin, apigenin, eupatilin, quercetin, kaempferol and luteolin, and the key targets included STAT3, SRC, CTNNB1, PIK3R1 and AKT1. Molecular docking study revealed that the active ingredients of Shenbing Decoction Ⅲ had good binding activity to the key targets. Western blotting showed that in rats with 5/6 nephrectomy, treatment with Shenbing Decoction Ⅲ obviously restored the protein expression of STAT3, PI3K, and AKT in renal tissue.@*CONCLUSION@#Shenbing Decoction Ⅲ can reduce renal injury induced by 5/6 nephrectomy in rats, and its therapeutic effects are mediated possibly by its main pharmacologically active ingredients that alleviate renal fibrosis via modulating multiple targets including STAT3, PIK3R1, and AKT1.
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Masculino , Animais , Ratos , Ratos Sprague-Dawley , Simulação de Acoplamento Molecular , Farmacologia em Rede , Creatinina , Insuficiência Renal Crônica/tratamento farmacológico , Fibrose , UreiaRESUMO
This article aimed to explore the theoretical connotation and mechanism of clearing damp-heat method in the treatment of chronic kidney disease (CKD), provide theoretical support for clearing damp-heat method in the treatment of chronic kidney disease, and further explain the modern scientific connotation of "damp-heat impairing kidney". Modern Traditional Chinese Medicine (TCM) believes that damp-heat is an important pathogenesis of kidney damage. Clearing damp-heat method plays a key role in inhibiting CKD immune inflammatory response, improving oxidative stress and antagonizing renal fibrosis. The mechanism is mainly related to the regulation of TNF-α level, blocking NF-κB signaling pathway, inhibiting inflammatory cytokines, antagonizing TGF-β1 secretion and other pathways.
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Objective:To investigate the role of immunoglobulin-like domain-containing receptor 2 (Ildr2) in renal fibrosis induced by ischemia-reperfusion.Methods:Ildr2 knockout mice (KO group) were constructed using CRISPR/Cas9 technology, and wild-type mice were as the control group (WT group). The unilateral renal ischemia-reperfusion (UIR) model (UIR group) was constructed by clamping the left renal pedicle, and was divided into KO-UIR group and WT-UIR group after modeling. Sham operation mice (sham group) were not treated with ischemia. Serum creatinine was measured by creatinine oxidase method. Blood urea nitrogen was detected by the diacetyloxime colorimetric method. The urinary albumin level was measured by enzyme-linked immunosorbent assay, and urinary albumin/creatinine ratio was calculated. HE, PAS and MASSON staining were used to detect the infiltration of inflammatory cells and the degree of fibrosis in renal tissues. The mRNA expression levels of Ildr2, kidney injury-associated molecules neutrophil gelatinase-associated lipocalin ( NGAL) and kidney injury molecule-1 ( KIM-1), fibrosis markers typeⅠcollagen α 1 ( Col1α1), fibronectin 1 ( Fn1), α-smooth muscle actin ( α-SMA) and connective tissue growth factor ( CTGF), as well as inflammation-related molecules macrophage marker F4/80 and monocyte chemoattractant protein-1 ( MCP-1) were detected by real time quantitative PCR (qRT-PCR). The protein levels of Ildr2, α-SMA and Col1α1 were detected by immunofluorescence and Western blotting. Results:(1) qRT-PCR and Western blotting showed that the expression levels of Ildr2 mRNA and protein in UIR group were significantly lower than those in sham group (both P<0.05). (2) There were no significant differences in body weight, serum creatinine, blood urea nitrogen, total cholesterol, low density lipoprotein, high density lipoprotein and triglyceride between KO group and WT group (all P>0.05). qRT-PCR results showed that there were no significant differences in the mRNA expression levels of NGAL, KIM-1, α-SMA, Col1α1, CTGF, Fn1, MCP-1 and F4/80 between KO group and WT group (all P>0.05). Histological staining showed no abnormal inflammatory cell infiltration and interstitial fibrosis between KO group and WT group. (3) Compared with the WT-UIR group, serum creatinine and blood urea nitrogen in the KO-UIR group were significantly higher (both P<0.05). qRT-PCR results showed that the mRNA expression levels of NGAL, F4/80, MCP-1, Col1α1, α-SMA, and CTGF in the KO-UIR group were significantly higher than those in the WT-UIR group (all P<0.05). Immunofluorescence and Western blotting results also showed that the protein expression levels of Col1α1 and α-SMA in the KO-UIR group were significantly higher than those in the WT-UIR group (all P<0.05). Histological staining showed that, compare with WT-UIR group, KO-UIR group had more severe inflammatory infiltration and more collagen fiber deposition. Conclusion:Ildr2 knockout does not cause phenotypic changes in mice under normal physiological conditions. Ildr2 plays a regulatory role in UIR injury, and Ildr2 deletion aggravates the degree of renal fibrosis induced by UIR.