Study on the Mechanism of Yangjing Zhongyu Decoction in Regulating Primordial Follicle Initiation in Model Rats with Diminished Ovarian Reserve / 中国中医药信息杂志

Objective To study the mechanism of Yangjing Zhongyu Decoction in regulating the initiation of primordial follicles in model rats with diminished ovarian reserve(DOR)based on lncRNA.Methods Three-day-old female rats were selected and their ovaries were cultured in vitro.The blank group,model group,DHEA group and Yangjing Zhongyu Decoction high-,medium-and low-dosage groups were set.The DOR model was induced by triptolide,corresponding drug containing serum was given to culture respectively.HE staining was used to observe germ cells and follicles,Western blot was used for determining the expressions of AMH,BMP15,PTEN,MST,TGF-β1,p-Smad1 protein,RT-PCR was used to detected AMH,BMP15,PTEN,MST,LTCONS-00011173,TGF-β1,Smad1 mRNA expression.Results Compared with the blank group,the number of primordial and growing follicles in the model group rats decreased(P<0.05),the expression of AMH,BMP15,TGF-β1,p-Smad1 protein in ovarian tissue decreased(P<0.05),expressions of PTEN and MST proteins increased(P<0.05),AMH,BMP15,TGF-β1,Smad1 mRNA expression decreased(P<0.05),while the expressions of PTEN,MST,and LTCONS-00011173 mRNA increased(P<0.05).Compared with the model group,the DHEA group and Yangjing Zhongyu Decoction high-and medium-dosage groups showed an increase in the number of primordial and growing follicles(P<0.05),the expressions of AMH,BMP15,TGF-β1 and p-Smad1 protein in ovarian tissue increased(P<0.05),PTEN and MST protein expressions decreased(P<0.05),AMH,BMP15,TGF-β1,Smad1 mRNA expressions increased(P<0.05),while PTEN,MST,and LTCONS-00011173 mRNA expressions decreased(P<0.05).Conclusion Yangjing Zhongyu Decoction may mediate TGF-β1/Smad1 signaling pathway through LTCONS-00011173,regulating primordial follicle initiation in DOR model rats.

Effects of genistein on myocardial fibrosis in a rat model of type 2 diabetes mellitus via TGF-β1/Smad3 signaling pathway / 中成药

AIM To explore the effects of genistein on reducing myocardial fibrosis in diabetic rats and the possible mechanism.METHODS The rat models of type 2 diabetes mellitus(T2DM)established by high-fat diet feeding and intraperitoneal streptozotocin(STZ)injection were randomly divided into the model group,the metformin group(100 mg/kg)and the low-dose and high-dose genistein groups(50,100 mg/kg),in contrast to those of the normal group given normal diet,with 10 rats in each group.After 8 weeks gavage of the corresponding drugs,the rats had detections of their weight of the body and the heart,the heart function,levels of the cardiac indices,the biomarkers of serum creatine kinase isoenzyme(CK-MB),aspartate aminotransferase(AST)and lactate dehydrogenase(LDH)activities,the extent of myocardial fibrosis,myocardial mRNA and protein expressions of transforming growth factor-β1(TGF-β1)and Smad homologue 3(Smad 3),and the myocardial distribution and expressions of Collagen Ⅰ and Collagen Ⅲ.RESULTS Compared with the model group,the genistein groups shared increased body weight,stroke output(SV)and ejection fraction(EF)(P<0.01);decreased levels of cardiac indices,left ventricular internal diameter end systole(LVIDs),CK-MB,AST and LDH activities(P<0.05,P<0.01);decreased relative area and myocardial expressions of CollagenⅠand CollagenⅢ(P<0.05,P<0.01);and decreased myocardial expressions of TGF-β1 and Smad3 mRNA and protein(P<0.05,P<0.01).Additionally,the high-dose genistein group was observed with decreased level of left ventricular internal diameter end-diastole(LVIDd)(P<0.05).CONCLUSION Genistein can protect the hearts of T2DM rat models by reducing their myocardial fibrosis via TGF-β1/Smad3 signaling pathway.

Marein alleviates myocardial fibrosis in diabetic mice / 基础医学与临床

Objective To study the effect of marein on myocardial fibrosis in diabetic mice.Methods Ten lep-tin receptor gene defective heterozygous(db/m)mice aged 5-6 weeks were selected as the control group and 30 diabetic mice with leptin receptor gene defective db/db were divided into:db/db group(db/db,n=10),metformin(Met)positive group(280 mg/kg daily,n=10)and marein drug intervention group(50 mg/kg,n=10).After continuous administration for 8 weeks,the cardiac morphological changes were observed by HE staining and Masson staining.The distribution and expression of vimentin were detected by immunohistochemis-try method.The expression of fibronectin,vimentin,and transforming growth factor-β1(TGF-β1)protein in cardiac tissue was detected by Western blot.Results Myocardial fiber hypertrophy was observed in db/db group,and myocardial structural damage was improved in metformin group and marein group.Compared with db/m group,the expression of myocardial collagen fiber in db/db group increased(P＜0.01),while the expression of myo-cardial collagen fiber in metformin group and marein group decreased(P＜0.01).Compared with the control group,the expression of vimentin in myocardial tissue of db/db group was significantly increased(P＜0.01),while the expression of vimentin in metformin group and marein group was significantly decreased(P＜0.01).The expression of fibronectin,vimentin and TGF-β1 in db/db group was significantly increased as compared with those in db/m group(P＜0.01),while the expression of fibronectin,vimentin and TGF-β1 in metformin group and marein group were significantly decreased(P＜0.01).Conclusions Marein improves myocardial fibrosis in diabetic db/db mice.

Effect of Qingre Huashi Sanjie Enema Prescription on MCP-1，TGF-β1，IL-6，and T Lymphoid Subsets in Patients with Sequelae of Pelvic Inflammatory Disease with Syndrome of Combined Dampness， Heat， and Stasis / 中国实验方剂学杂志

ObjectiveTo investigate the application value of Qingre Huashi Sanjie enema prescription in the treatment of the patients with sequelae of pelvic inflammatory disease （syndrome of combined dampness，heat，and stasis） and the effects of this prescription on inflammatory mediators and T lymphocyte subsets. MethodThe patients with sequelae of pelvic inflammatory disease （syndrome of combined dampness，heat，and stasis） treated from May 2022 to August 2023 were included in this study and randomized into two groups （79 cases）. The control group was treated with conventional Western medicine，and the observation group was treated with Qingre Huashi Sanjie enema prescription on the basis of the therapy in the control group. Both groups were treated for 12 weeks. The serum levels of monocyte chemoattractant protein-1 （MCP-1），transforming growth factor-β1 （TGF-β1），and interleukin-6 （IL-6） were measured by enzyme linked immunoserbent assay （ELISA） before and after treatment in both groups. The erythrocyte sedimentation rate （ESR） and fibrinogen （FIB） were measured by an automatic blood rheology analyzer before and after treatment in both groups. The serum levels of CD4+，CD4+/CD8+ before and after treatment in both groups were measured by flow cytometry. The traditional Chinese medicine （TCM） symptom score and the 36-item short form survey （SF-36） score were assessed before and after treatment. The uterine artery resistance index （RI），uterine artery pulsatility index （PI），and uterine artery peak systolic velocity （PSV） were measured by Doppler before and after treatment. The clinical efficacy and the occurrence of adverse reactions were compared between the two groups. ResultAfter treatment，the levels of MCP-1，TGF-β1，IL-6，ESR，and FIB decreased in both groups （P<0.01），and the decreases were larger in the observation group than in the control group （P<0.05，P<0.01）. After treatment，the serum levels of CD4+ and CD4+/CD8+ elevated in both groups （P<0.01） and the observation group had higher levels of CD4+ and CD4+/CD8+ than the control group （P<0.05，P<0.01）. The treatment in both groups decreased the TCM symptom score and TCM sign score and increased the SF-36 score （P<0.01），and the changes were more significant in the observation group than in the control group （P<0.05，P<0.01）. In addition，the treatment lowered RI and PI and elevated PSV （P<0.01），and the changes in these indicators were more significant in the observation group than in the control group （P<0.01）. The total response rate in the observation group was 93.67% （74/79），which was higher than that （79.75%，63/79） in the control group （χ2=6.645，P<0.05）. There was no significant difference in the occurrence of adverse reactions between the two groups. ConclusionFor the patients with sequelae of pelvic inflammatory disease （syndrome of combined dampness，heat，and stasis），Qingre Huashi Sanjie enema prescription can reduce inflammation，attenuate hypercoagulability，improve hemodynamics，and regulate the immune function，demonstrating a definite therapeutic effect.

The Role of TGF-β1/Smads Signaling Pathway in Anti-myocardial Fibrosis Effect of SIRT1 and Related Mechanism / 华中科技大学学报(医学版)

Objective To observe the effect of sirtuin 1(SIRT1)on rat myocardial fibrosis induced by pressure overload and the proliferation of cardiac fibroblasts induced by angiotensin Ⅱ(Ang Ⅱ),and to explore the molecular mechanisms.Methods The pressure overload-induced myocardial fibrosis was established by abdominal aorta constriction(AAC)procedure in vi-vo.After treatment with SIRT1 activator,the myocardial interstitial fibrosis and the collagen volume fraction were evaluated by Masson's trichrome staining.The protein expressions of TGF-β1/Smads were determined by immunohistochemical analy-sis.After in vitro intervention of Ang Ⅱ or Ang Ⅱ with SIRT1 activator,the fibroblasts proliferation was detected by MTT as-say.The mRNA and protein expressions of collagen Ⅰ/Ⅲ(Col1α1/3α1),SIRT1 and TGF-β1/Smads in myocardial tissue and fi-broblasts were evaluated by qRT-PCR and Western blotting.Results Compared with the sham operation group,myocardial in-terstitial fibrosis was significantly observed in the pressure overload model group,myocardial collagen volume fraction was in-creased,expressions of Col1α1/3α1 and TGF-β1/Smads were significantly increased,and SIRT1 expression was decreased.After the intervention of SRT1720,SIRT1 activator could improve the myocardial interstitial fibrosis induced by pressure overload,downregulate the expressions of Col1α1/3α1 and TGF-β1/Smads,and upregulate the expression of SIRT1.Meanwhile,correla-tion analysis showed that the protein expression of SIRT1 was negatively correlated with the expression of TGF-β1.In addition,SRT1720 also inhibited Ang Ⅱ-induced fibroblast proliferation and increased expression of Col1α1/3α1 and TGF-β1.Conclusion Activation of SIRT1 inhibits pressure overload-induced myocardial fibrosis and Ang Ⅱ-induced fibroblasts proliferation via regu-lation of the TGF-β1/Smads signaling pathway.

Gentianopsis paludosa xanthone combined with probiotics inhibits colon inflammation-tumor transformation in rats by regulating TGF-β1/Smads pathway and inflammatory factors / 天津医药

Objective To investigate the mechanism of Gentianopsis paludosa xanthone(GPX)combined with probiotics in the intervention of colon inflammation-tumor transformation in rats by regulating TGF-β1/Smads pathway and inflammatory factors.Methods Ninety rats were divided into the normal group,the model group[drinking sodium dextran sulfate(DSS)for 3 days]and the intervention group by random number table method.The model group was subdivided into the inflammatory stage group,the pre-inflammatory cancer group(DMH injection for 4 weeks),the intermediate inflammatory cancer group(DMH injection for 13 weeks)and the advanced inflammatory cancer group(DMH injection for 21 weeks).The administration group was subdivided into the groups(after the first day of drinking DSS,drugs for each group were given by gavage once a day for 8 weeks)on the basis of the advanced inflammatory cancer group,including the GPX group(GPX 69.3 mg/kg),the probiotic group,the combined group(GPX+probiotics 400 mg/kg)and the thalidomide group(thalidomide 13.5 mg/kg).The disease activity index(DAI),colon length and wet mass index were compared between all groups.Characteristics of colon tumors were observed,and pathological changes of colon were observed by HE staining.The expression levels of transforming growth factor(TGF)-β1,Smad4,Smad7,interleukin(IL)-6 and tumor necrosis factor(TNF)-α were detected by Western blot assay and enzyme-linked immunosorbent assay,respectively.Results Compared with the advanced inflammatory cancer group,the administration groups showed an increase in colon length,the expression levels of TGF-β1 and Smad4 protein,a decrease in colon wall thickness,wet mass index,maximum tumor diameter,the levels of Smad7,IL-6,TNF-α,and DAI score decreased in the GPX group and the combined group(P＜0.05).The structure and morphology of intestinal mucosa were improved in the GPX group,the probiotic group and the combination group,and the structure of colonic crypt and goblet cell number were increased.Compared with the probiotic group and the GPX group,the colon wall thickness,colon wet mass index and tumor number were decreased,the protein expression levels of TGF-β1 and Smad4 were increased,and levels of IL-6 and TNF-α were decreased in the combination group(P＜0.05).Conclusion GPX combined with probiotics could inhibit the transformation of colon inflammation-tumor,and the mechanism may be related to the regulation of TGF-β1/Smads pathway and the inhibition of pro-inflammatory factors of IL-6 and TNF-α.

Expression of MMP-9 in patients with chronic rhinosinusitis and its correlation with epithelial-mesenchymal transition / 天津医药

Objective To investigate the mechanism of matrix metalloproteinase(MMP)-9 involved in epithelial mesenchymal transformation(EMT)in chronic sinusitis(CRS).Methods The expression of MMP-9 from polypoid middle turbinate tissue was detected by immunohistochemical staining qPCR and Western blot assay in 42 patients with CRS and 8 patients underwent septoplasty.Primary human nasal epithelial cells HNEpc were cultured in vitro and divided into the control group,the TGF-β1 group(5 μg/L TGF-β1 intervention)and the TGF-β1+si-MMP-9 group(transfected with si-MMP-9 and 5 μg/L TGF-β1 intervention).The expression of MMP-9 was detected by cell immunofluorescence staining.Expression levels of TGF-β1,MMP-9 and EMT-related proteins E-cadherin,vimentin and α-SMA were detected by Western blot assay.Results(1)The positive expression rate of MMP-9 was significantly higher in the nasal mucosa of CRS with nasal polyps(CRSwNP)group(54.5%,12/22)than that of the CRS without polyps(25.0%,5/20)group and the control group(12.8%,1/8).The relative expression levels of MMP-9 mRNA and protein in nasal mucosa were higher in the CRSwNP group than those in the CRSsNP group and the control group(P＜0.05).(2)Compared with the control group,the expressions levels of TGF-β1,MMP-9,vimentin and α-SMA were increased in the TGF-β1 group,while the expression of E-cadherin was decreased(P＜0.05).Compared with the TGF-β1 group,expression levels of TGF-β1,MMP-9,vimentin and α-SMA were decreased in the TGF-β1+si-MMP-9 group,and the expression of E-cadherin was increased(P＜0.05).Conclusion The expression of MMP-9 is increased in CRS patients,which may be involved in the development of CRS through the regulation of EMT.

Mechanism of Shugan Huazheng Prescription Against Liver Fibrosis Based on HIF-1α/VEGF/TGF-β1 Pathway / 中国实验方剂学杂志

ObjectiveTo observe the therapeutic effect of Shugan Huazheng prescription on hepatic fibrosis model rats induced by carbon tetrachloride （CCl4） and explore whether it plays its role through hypoxia-induced factor-1α/vascular endothelial growth factor/transforming growth factor-β1 （HIF-1α/VEGF/TGF-β1） pathway. MethodA total of 54 male SPF SD rats were randomly divided into six groups： blank group， model group， colchicine group （0.2 mg·kg-1）， and high-， medium-， and low-dose groups （29.52， 14.76， and 7.38 g·kg-1） of Shugan Huazheng prescription， with nine rats in each group. The molding was conducted three times a week for eight weeks. Administration began the day after the first injection， and the drug intervention was once a day for eight weeks. On the day after the last administration， the rats were deprived of food and water， and they were killed the next day， during which the physiological status of each group of rats was dynamically monitored. The pathological changes in the liver were observed by hematoxylin-eosin （HE） staining， and the content of hydroxyproline （HYP） and angiotensin Ⅱ （AngⅡ） in liver tissue were detected by enzyme-related immunosorbent assay （ELISA）. Real-time fluorescent quantitative PCR （Real-time PCR） was used to determine the mRNA expression levels of HIF-1α， VEGF， and TGF-β1 in liver tissue， and immunohistochemical method （IHC） and Western blot were used to detect the protein expression levels of HIF-1α， VEGF， and TGF-β1 in liver tissue. ResultCompared with the blank group， the overall condition of rats in the model group decreased significantly. The proliferation of connective tissue and the increase in adipose cells between hepatocytes were obvious. The content of HYP and Ang was increased. The mRNA and protein expressions of HIF-1α， VEGF， and TGF-β1 were increased to varying degrees （P<0.05）. Compared with the model group， the proliferation of connective tissue and inflammatory cell infiltration in the liver tissue of colchicine and Shugan Huazheng prescription groups were reduced. The content of HYP and Ang was decreased. The mRNA and protein expression levels of HIF-1α， VEGF， and TGF-β1 were decreased， and the colchicine group and high-dose group of Shugan Huazheng prescription were the most significant （P<0.05）. ConclusionShugan Huazheng prescription has an obvious therapeutic effect on CCl4-induced hepatic fibrosis model rats. Its therapeutic mechanism may be related to the regulation of the HIF-1α/VEGF/TGF-β1 signaling pathway and the improvement of hepatic hypoxia， vascular remodeling， and the syndrome of Qi deficiency and blood stasis in hepatic fibrosis.

Intervention effect and mechanism of breviscapine on hepatic fibrosis in rats / 中国药房

OBJECTIVE To investigate the intervention effect and potential mechanism of breviscapine on hepatic fibrosis （HF） in rats based on the transforming growth factor-β（1 TGF-β1）/Smad2/extracellular signal-regulated protein kinase 1（ERK1） and Kelch-like epichlorohydrin-associated protein 1（Keap1）/nuclear factor-erythroid 2-related factor 2（Nrf2）/heme oxygenase-1（HO-1） pathways. METHODS Totally 60 rats were randomly divided into normal control group， model group， breviscapine low-dose， medium-dose and high-dose groups （5.4， 10.8， 21.6 mg/kg）， and colchicine group （positive control， 0.45 mg/kg）， with 10 rats in each group， half male and half female. Except for the normal control group， HF model of the other groups was induced by carbon tetrachloride. Subsequently， each drug group was given corresponding medicine by gavage once a day for 28 days. The liver appearance of rats in each group was observed and their liver coefficients were calculated. The levels of alanineaminotransferase （ALT） and aspartate aminotransferase （AST）in serum， those of ALT， AST， superoxide dismutase （SOD），malondialdehyde （MDA） and glutathione peroxidase （GSH- Px） in liver tissue were detected. The liver tissue inflammatory and fibrotic changes were observed. The protein and mRNA expressions of TGF-β1， Smad2， ERK1， Nrf2， Keap1 and HO-in liver tissue were detected. RESULTS Compared with the normal control group， the model group showed large areas of white nodular lesions in the liver， obvious inflammatory cell infiltration and collagen fiber deposition. The body weight， the levels of SOD and GSH-Px in liver tissue， the protein and mRNA expressions of Nrf2 and HO-1 were significantly lowered in the model group （P＜0.05）； the liver coefficient， the percentage of Masson staining positive area， ALT and AST levels of serum and liver tissue， MDA level of liver tissue， the protein and mRNA expressions of TGF-β1， Smad2， ERK1 and Keap1 were significantly increased （P＜0.05）. Compared with the model group， the liver lesions of rats in each drug group were improved， and the above quantitative indexes were generally reversed （P＜0.05）. CONCLUSIONS Breviscapine has a good intervention effect on HF rats， which may be related to inhibiting TGF-β1/Smad2/ERK1 pathway for anti-fibrosis and regulating Keap1/Nrf2/HO-1 pathway to inhibit oxidative stress.

Effect Xuefu Zhuyu decoction on endothelial-to-mesenchymal transition of pulmonary artery endothelial cells and its mechanism / 中国药理学通报

Aim To investigate the effect of Xuefu Zhuyu decoction on transforming growth factor-β1(TGF-β1 ) -induced endothelial-to-mesenchymal transition (EndMT) of pulmonary microvascular endothelial cells ( PMVEC), and further analyze the mechanism related to the TGF-β1/Smad signaling pathway. Method To construct an EndMT cell model, PMVEC was treated with TGF-β1 (5 μg · L

Effect of Qianyang Yuyin Granules on Mesenchymal Transdifferentiation of Renal Tubular Epithelial Cells Induced by TGF-β1 / 中国实验方剂学杂志

ObjectiveBy observing the effect of Qianyang Yuyin granules on the phenotype of renal tubule epithelial cells， the intervention of Qianyang Yuyin granule on renal interstitial fibrosis was investigated. MethodThe renal tubular epithelial cells （HK-2） were treated with different concentrations of transforming growth factor （TGF）-β1 （5， 10， 15， 20， 25 μg·L-1） for 24 hours， and cell morphology and growth state were observed with an inverted phase contrast microscope. The 20 μg·L-1 was selected as the most appropriate concentration of TGF-β1 according to Western blot results for subsequent experiments. HK-2 cells were divided into six groups： blank group， TGF-β1 group （concentration of 20 μg·L-1）， low， medium， and high dose Qianyang Yuyin granule groups （concentration of 0.5， 1， 2 g·L-1）， and valsartan group （1 × 10-5 mol·L-1）. The cell activity was measured by cell proliferation and cell counting kit-8 （CCK-8）. The cell migration ability was detected by scratch test. The Transwell method was used to detect the invasiveness of cells. Western blot was used to detect levels of fibronectin （FN）， E-cadherin， α-smooth muscle activator （α-SMA）， Vimentin， collagen type Ⅰ（Col Ⅰ）， collagen type Ⅳ（Col Ⅳ）， and other related proteins. ResultTGF-β1 stimulating epithelial-mesenchymal transition （EMT） in renal tubular epithelial cells was time- and concentration-dependent. Compared with the blank group， higher concentration in the TGF-β1 group indicates longer intervention time and more obvious long spindle change of cells， and the migration and invasion ability of the cells was significantly enhanced. The protein expression level of FN， α-SMA， Vimentin， Col Ⅰ， and Col Ⅳ increased significantly （P<0.05， P<0.01）， while the expression level of E-cadherin protein decreased （P<0.05）. Compared with the TGF-β1 group， Qianyang Yuyin granule groups could maintain normal cell morphology， and the migration and invasion ability of the cells was inhibited. The protein expression level of FN， α-SMA， Vimentin， Col Ⅰ， and Col Ⅳ decreased （P<0.05， P<0.01）， and the expression of E-cadherin protein was significantly restored （P<0.05）. ConclusionQianyang Yuyin granule can reverse TGF-β1-induced interstitial transformation of renal tubular epithelial cells by reducing the phenotypic expression of mesenchymal cells and increasing the phenotypic expression of epithelial cells.

Suppression of NLRP3 inflammasome by ivermectin ameliorates bleomycin-induced pulmonary fibrosis / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Ivermectin is a US Food and Drug Administration (FDA)-approved antiparasitic agent with antiviral and anti-inflammatory properties. Although recent studies reported the possible anti-inflammatory activity of ivermectin in respiratory injuries, its potential therapeutic effect on pulmonary fibrosis (PF) has not been investigated. This study aimed to explore the ability of ivermectin (0.6 mg/kg) to alleviate bleomycin-induced biochemical derangements and histological changes in an experimental PF rat model. This can provide the means to validate the clinical utility of ivermectin as a treatment option for idiopathic PF. The results showed that ivermectin mitigated the bleomycin-evoked pulmonary injury, as manifested by the reduced infiltration of inflammatory cells, as well as decreased the inflammation and fibrosis scores. Intriguingly, ivermectin decreased collagen fiber deposition and suppressed transforming growth factor-‍β1 (TGF-‍β1) and fibronectin protein expression, highlighting its anti-fibrotic activity. This study revealed for the first time that ivermectin can suppress the nucleotide-binding oligomerization domain (NOD)‍-like receptor family pyrin domain-containing protein 3 (NLRP3) inflammasome, as manifested by the reduced gene expression of NLRP3 and the apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), with a subsequent decline in the interleukin‍-‍1β (IL‍-‍1β) level. In addition, ivermectin inhibited the expression of intracellular nuclear factor-‍κB (NF‍-‍κB) and hypoxia‑inducible factor‑1α (HIF‍-‍1α) proteins along with lowering the oxidative stress and apoptotic markers. Altogether, this study revealed that ivermectin could ameliorate pulmonary inflammation and fibrosis induced by bleomycin. These beneficial effects were mediated, at least partly, via the downregulation of TGF-‍β1 and fibronectin, as well as the suppression of NLRP3 inflammasome through modulating the expression of HIF‑1α and NF-‍κB.

Effect of TRB3 on the Function of Gastric Cancer Cells by Regulating TGF-β1/Smad3/PAI-1 Axis / 医学研究杂志

Objective To study the effect of TRB3 on gastric cancer cells function by regulating TGF-β1/Smad3/PAI-1 axis.Methods The expression of TRB3 protein in gastric cancer cell lines(AGS,HGC-27,NCI-N87,SNU5 and MKN45)and normal human gastric mucosal cells(GES-1)was detected by Western blot.Knockdown of TRB3 in gastric cancer cell AGS,CCK-8,fluores-cenceactivated cell sorter(FACS),Transwell assay,scratch assay and Western blot were used to detect cell proliferation,apoptosis,in-vasion,migration and the changes of TGF-β1/Smad3/PAI-1 signal pathway proteins.On the basis of knockdown of TRB3,TGF-β1 treated gastric cancer cell AGS,and the above experiments were repeated again.Results Compared with GES-1 cells,the expression of TRB3 protein increased in gastric cancer cell lines.the proliferation ability and the viability of cells(52％±12％,P＜0.001)were decreased;migration ability and cell healing rate(24％±6％,P＜0.001)were decreased;the invasive ability and the number of inva-sive cells(107±23,P＜0.001)were decreased;TGF-β1/Smad3/PAI-1 pathway was inhibited,and the expression of TGF-β1,p-SMAD3 and PAI-1 were decreased;the apoptosis and the proportion of apoptotic cells(17％±3％,P＜0.001)were increased.On the basis of knockdown of TRB3,TGF-β1 was used to treated gastric cancer cells AGS,the cell viability(73％±10％,P＜0.01)was increased;the migration ability and cell healing rate(63％±8％,P＜0.01)were increased;the number of invasive cells(252±20,P＜0.01)was increased;cell apoptosis and the percentage of apoptosis(12％±2％,P＜0.01)was reduced.Conclusion TRB3 pro-motes the development of gastric cancer by activating of TGF-β1/Smad3/PAI-1 signaling pathway.

New progress of mechanism of action of miRNA-21 in diabetic kidney disease and Chinese medicine intervention / 天津医药

Diabetic kidney disease is one of the complications of diabetes,which can progress to end-stage renal disease.In recent years,it has been found that miRNAs have become a research hotspot,with miRNA-21 regulating transforming growth factor β1(TGF-β1)/Smads,phosphoinositide 3-kinase(PI3K)/protein kinase B(AKT),Wnt/β-catenin and other signaling pathways to promote the progress of diabetic kidney disease.Studies have showed that traditional Chinese medicine has a regulatory effect on the expression of miRNA-21 and can target miRNA-21 to regulate TGF-β1/Smads,phosphatase and tensin homolog/PI3K/AKT/mammalian target of rapamycin(mTOR),peroxisome proliferator activated receptors and other signal transduction pathways to trigger signal cascade reactions,which intervene in pathological processes such as fibrosis,inflammation,oxidative stress and autophagy.In this article,the role of miRNA-21 in diabetic kidney disease and the intervention of traditional Chinese medicine were summarized,in order to provide some reference for the treatment of diabetic kidney disease and the development of new drugs.

Ethyl Acetate Extract of Qigesan Intervenes in Migration and Invasion of Esophageal Cancer Cells via Inhibiting TGF-β1 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo explore the mechanism of Qigesan （QGS） in intervening in the migration and invasion of esophageal carcinoma TE-1 cells. MethodMicroarray technology was used to screen differentially expressed genes （DEGs） in the normal group and the QGS group， and the ontological functions and signaling pathways of DEGs were analyzed. The thiazolyl tetrazolium （MTT） assay was used to detect the effect of QGS on the viability of TE-1 cells. In the subsequent experiments for verification， a blank group， a transforming growth factor-β1 （TGF-β1） group， a TGF-β1 + QGS group， and a TGF-β1 + SB431542 group were set up. The cell morphology in each experimental group was observed by microscopy. The migration and invasion abilities of cells were detected by wound healing assay， and the mRNA expression levels of E-Cadherin， vimentin， Smad2， and Smad7 were detected by Real-time quantitative polymerase chain reaction （Real-time PCR）. The protein expression of E-Cadherin， vimentin， p-Smad2/3， Smad2/3， and Smad7 was detected by Western blot. ResultThere were 1 487 DEGs between the QGS group and the blank group， including 1 080 down-regulated ones （accounting for 72.63%） and 407 up-regulated ones. The down-regulated genes were mainly involved in biological processes such as cytoskeletal protein binding， ATP binding， adenylate nucleotide binding， and adenylate ribonucleotide binding， and the involved Kyoto Encyclopedia of Genes and Genomes （KEGG） pathways included TGF-β signaling pathway， cell cycle， extracellular matrix-receptor interaction protein， tumor pathways， and oocyte meiosis. The up-regulated genes were mainly involved in RNA binding， DNA binding， transcriptional regulator activity， transcriptional activator activity， and nucleotide binding， and the KEGG pathways involved mainly included mitogen-activated protein kinase （MAPK） signaling pathway， bladder cancer， renal cell carcinoma， cancer pathways， and p53 signaling pathway. Compared with the blank group， the inhibition rate of cell viability of TE-1 cells increased after QGS （20， 30， 40， 60， 80 mg·L-1） intervention for 12， 24， 36， 48， 60 h （P<0.05）， and the inhibition rate was time- and dose-dependent. Compared with the blank group， the TGF-β1 group showed lengthened cells with fibroblast phenotype. Compared with the TGF-β1 group， the TGF-β1 + QGS group showed shortened cells with normal morphology and epithelial phenotype. The cell morphology in the TGF-β1 + SB431542 group was similar to that of the TGF-β1 + QGS group. Compared with the blank group， the TGF-β1 group showed potentiated ability of cell migration and invasion （P<0.05）. Compared with the TGF-β1 group， the TGF-β1 + QGS group and the TGF-β1 + SB431542 group showed inhibited and weakened migration and invasion abilities of cells （P<0.05）. However， there was no significant difference in migration and invasion abilities between the TGF-β1 + QGS group and the TGF-β1 + SB431542 group. The mRNA expression levels of vimentin and Smad2 in the TGF-β1 group were higher （P<0.05）， and the mRNA expression levels of E-Cadherin and Smad7 were lower （P<0.05） than those in the blank group. Compared with the TGF-β1 group， the TGF-β1 + QGS group and the TGF-β1+ SB431542 group exhibited decreased expression levels of vimentin and Smad2 mRNA （P<0.05）， and elevated expression levels of E-Cadherin and Smad7 mRNA （P<0.05）. Compared with the blank group， the TGF-β1 group showed up-regulated protein expression levels of vimentin， p-Smad2/3， and Smad2/3 （P<0.05）， and reduced protein expression levels of E-Cadherin and Smad7 （P<0.05）. Compared with the TGF-β1 group， the TGF-β1 + QGS group and the TGF-β1 + SB431542 group displayed decreased protein expression levels of vimentin， p-Smad2/3， and Smad2/3 （P<0.05）， and increased protein expression levels of E-Cadherin and Smad7 （P<0.05）. ConclusionThe ethyl acetate extract of QGS inhibits the epithelial-mesenchymal transition （EMT） of TE-1 cells through the TGF-β1 pathway to reduce the migration and invasion of TE-1 cells.

Non-coding RNA-mediated TGF-β1/Smads Signaling Pathway and Liver Fibrosis， and Intervention by Active Components of Chinese Medicinal： A Review / 中国实验方剂学杂志

Liver fibrosis is a wound healing response that occurs in the setting of chronic liver injury and is caused by imbalance in the synthesis and degradation of extracellular matrix （ECM）. If left untreated， it can progress to liver cirrhosis and hepatocellular carcinoma. The activation of hepatic stellate cell （HSC） is now well established as a central driver of liver fibrosis. The activated HSC will transform into myofibroblasts that produce ECM protein. Transforming growth factor-β1 （TGF-β1） can induce the activation of hepatic stellate cell （HSC）， and TGF-β1/Smads signaling pathway is one of the important pathways to promote liver fibrosis. Non-coding RNA （ncRNA） does not encode proteins during the transcription but plays an important regulatory role in the post-transcriptional process of genes. Accumulating evidence shows that the occurrence of liver fibrosis is closely related to the abnormal expression of ncRNA which participates in the activation of HSC by regulating TGF-β1 signal transduction and then affects the process of liver fibrosis. MiRNA-mediated TGF-β1/Smads signaling pathway can not only promote liver fibrosis but also play a role in anti-fibrosis. Long non-coding RNA （lncRNA） not only promotes the development of liver fibrosis by binding to target genes but also enhances TGF-β1 signal transduction by acting as competitive endogenous RNA. circular RNA （circRNA） acts as a ''sponge'' to regulate TGF-β1/Smads pathway， thereby inhibiting HSC activation and exerting the anti-liver fibrosis effect. Chinese medicinal plays an essential part in the prevention and treatment of liver fibrosis， and the active components can inhibit TGF-β1/Smads pathway by regulating the expression of miRNA， thus alleviating liver fibrosis. This article reviews the role and mechanism of miRNA-， lncRNA- and circRNA-mediated TGF-β1/Smads signaling pathway in liver fibrosis and summarizes the anti-liver fibrosis effect of active components of Chinese medicinals by regulating miRNA-mediated TGF-β1/Smads signaling pathway， which can serve as a reference for clinical treatment of liver fibrosis and the development of new drugs.

Cytokine receptor-like factor 1 (CRLF1) promotes cardiac fibrosis via ERK1/2 signaling pathway / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Cardiac fibrosis is a cause of morbidity and mortality in people with heart disease. Anti-fibrosis treatment is a significant therapy for heart disease, but there is still no thorough understanding of fibrotic mechanisms. This study was carried out to ascertain the functions of cytokine receptor-like factor 1 (CRLF1) in cardiac fibrosis and clarify its regulatory mechanisms. We found that CRLF1 was expressed predominantly in cardiac fibroblasts. Its expression was up-regulated not only in a mouse heart fibrotic model induced by myocardial infarction, but also in mouse and human cardiac fibroblasts provoked by transforming growth factor-‍β1 (TGF‍-‍β1). Gain- and loss-of-function experiments of CRLF1 were carried out in neonatal mice cardiac fibroblasts (NMCFs) with or without TGF-‍β1 stimulation. CRLF1 overexpression increased cell viability, collagen production, cell proliferation capacity, and myofibroblast transformation of NMCFs with or without TGF‍-‍β1 stimulation, while silencing of CRLF1 had the opposite effects. An inhibitor of the extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathway and different inhibitors of TGF-‍β1 signaling cascades, comprising mothers against decapentaplegic homolog (SMAD)‍-dependent and SMAD-independent pathways, were applied to investigate the mechanisms involved. CRLF1 exerted its functions by activating the ERK1/2 signaling pathway. Furthermore, the SMAD-dependent pathway, not the SMAD-independent pathway, was responsible for CRLF1 up-regulation in NMCFs treated with TGF-‍β1. In summary, activation of the TGF-‍β1/SMAD signaling pathway in cardiac fibrosis increased CRLF1 expression. CRLF1 then aggravated cardiac fibrosis by activating the ERK1/2 signaling pathway. CRLF1 could become a novel potential target for intervention and remedy of cardiac fibrosis.

ZLY18 protects against angiotensin II-induced cardiac fibrosis via inhibiting TGF-β1/Smads pathway / 中国药理学通报

Aim To explore the effect of ZLY18 on angiotensin II-induced cardiac fibrosis and the underlying mechanism. Methods Ang II was used to induce cardiac fibrosis in vitro and in vivo. Cardiac fibroblasts were divided into blank control group, model group and medicine group. The medicine group was subdivided into ZLY18(L)group, ZLY18(M)group and ZLY18(H)group. Compound ZLY18 was given 1, 2, 5 μmol·L-1 respectively. C57BL/6 mice were randomly divided into control group, model group and medicine group. The medicine group were subdivided into ZLY18(L)group, ZLY18(M)group and ZLY18(H)group. Compound ZLY18 was given 10,20 and 50 mg·kg-1 respectively. Both the model group and the medicine group were given with Ang II to induce cardiac fibrosis. The changes of protein levels were detected by Western blot and immunofluorescence. The changes of cardiac function indexes in C57BL/6 mice were detected by small animal echocardiography. The morphology, cell arrangement and collagen fibers of cardiac fibroblasts were observed by tissue section staining and other methods. Results The model of Ang II-induced myocardial fibrosis was successfully established at the cell and animal levels, and ZLY18 treatment improved the elevated fibrosis-related protein caused by Ang II and abnormal cardiac function in mice. Moreover, ZLY18 was able to inhibit the increased phosphorylation of TGF-1 and Smad3 caused by Ang II and increased Smad2/3 nuclear entry, suggesting that the antifibrotic effect of ZLY18 might be related to the activation of TGF-1/Smads signaling pathway. Conclusions ZLY18 has a protective effect on Ang II-induced cardiac fibrosis. ZLY18 may inhibit TGF-β/Smads signaling pathway activation to exert anti-fibrotic effects.

Effects of betulinic acid on epithelial-mesenchymal transition of human gastric cancer MKN-45 cells / 中国药理学通报

Aim To observe the effect of betulinic acid (BA) on the migration and invasion of human gastric cancer MKN-45 cells induced by transforming growth factor-pi (TGF-β1), and to explore the effect of BA on epithelial-mesenchymal transition (EMT) and the potential mechanism. Methods The MKN-45 cells were cultivated in vitro, and the effects of different concentrations of BA on the proliferation of MKN-45 cells at 24, 48 and 72 h were detected using CCK-8 method. The effects of BA (5, 10, 20 jjunol • L) and TGF-01 inhibitor LY2109761 (10

Mechanism of Dahuang Zhechongwan in Treating Myocardial Fibrosis of Rats via TGF-β1/Smads/miR29 Pathway / 中国实验方剂学杂志

ObjectiveTraditional Chinese medicine， namely Dahuang Zhechongwan （DHZCW） was used to treat myocardial fibrosis in model rats， observe its effect on myocardial fibrosis in rats， and explore its action mechanism. MethodThirty-six SPF male Kunming rats were divided into blank group， model group， low-， medium-， high-dose groups of DHZCW （0.056， 0.084， 0.168 g·kg-1）， captopril group （10 mg·kg-1）， with six rats in each group. Except for the blank group， the other groups were intraperitoneally injected isoproterenol solution of 5 mg·kg-1 for 15 consecutive days to replicate the myocardial fibrosis model. At the beginning of modeling， the rats in each group took drugs， and they were sacrificed 28 days after administration. Serum and heart tissue were collected for the corresponding detection. Hematoxylin-eosin （HE） staining and Masson staining were used to observe tissue inflammation， cellular degeneration， necrosis， and fibrosis. The contents of hydroxyproline （HYP）， tumor necrosis factor-α （TNF-α）， interleukin-1β （IL-1β）， interleukin-6 （IL-6）， hyaluronic acid （HA）， laminin （LN）， type-Ⅲ procollagen （PC Ⅲ） in serum of rats and rats were determined by enzyme-related immunosorbent assay （ELISA）. The expression levels of key pathway proteins transforming growth factor-β1 （TGF-β1）， α-smooth muscle actin （α-SMA）， Smad2， Smad3， and Smad7 were detected by Western blot. The expression levels of key pathway genes TGF-β1， α-SMA， Smad2， Smad3， Smad7， miR-29a-5p， miR-29b-2-5p， and miR-29c-5p were detected by Real-time quantitative polymerase chain reaction （Real-time PCR）. ResultCompared with the blank group， the pathological changes of fibrosis in the model group were obvious， the contents of serum HYP， TNF-α， IL-1β， IL-6， HA， LN， and PCⅢ were increased （P<0.01）， the protein expression levels of TGF-β1， α-SMA， Smad2， and Smad3 were increased； the protein expression level of Smad7 was decreased （P<0.01）. The mRNA expression levels of TGF-β1， α-SMA， Smad2， and Smad3 were increased （P<0.05， P<0.01）， while those of Smad7， miR-29a-5p， miR-29b-2-5p， and miR-29c-5p were decreased （P<0.01）. Compared with the model group， after 28 days of administration， serum HYP， TNF-α， IL-1β， IL-6， HA， LN， and PCⅢ in high-， medium-， and low-dose groups of DHZCW and captopril groups were decreased （P<0.01）. Except for the low-dose group， the protein contents of TGF-β1， α-SMA， Smad2， and Smad3 were decreased， while the protein content of Smad7 was increased （P<0.01）. The mRNA expression levels of TGF-β1， Smad2， α-SMA， and Smad3 in high-dose group of DHZCW were decreased （P<0.05，P<0.01）， while those of Smad7， miR-29a-5p， miR-29b-2-5p， and miR-29c-5p were increased （P<0.05）. The mRNA expressions of TGF-β1， Smad2， and Smad3 in the medium-dose group of DHZCW were decreased （P<0.05， P<0.01）， while mRNA expression of Smad7 was increased （P<0.01）. The mRNA levels of TGF-β1 and Smad2 in the low-dose group of DHZCW were decreased （P<0.01）. ConclusionDHZCW can improve myocardial fibrosis in rats， and its action mechanism may be related to the regulation of the TGF-β1/Smads/miR-29 pathway. In addition， there is dose dependence in the range of 0.056-0.168 g·kg-1， and the effect of the high-dose group is more stable.