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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.
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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.
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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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Abstract Objective: To compare Transforming growth factor beta-1 (TGF-β1) expression in patients with and without adenomyosis. Methods: A prospective design was performed including 49 patients submitted to hysterectomy. Immunohistochemistry was performed on anatomopathological samples staged in paraffin blocks from patients with and without adenomyosis. The sample contained 28 adenomyosis cases and 21 controls. Student's t-test and multivariate logistic regression tests were used for statistical analysis. Associations were considered significant at p < 0.05. Results: We found no significant association between adenomyosis and: smoking (p = 0.75), miscarriage (p = 0.29), number of previous pregnancies (p = 0.85), curettage (p = 0.81), pelvic pain (p = 0.72) and myoma (p = 0.15). However, we did find a relationship between adenomyosis and abnormal uterine bleeding (AUB) (p = 0.02) and previous cesarean section (p = 0.02). The mean TGF-β1 intensity (mean ± SD) in the ectopic endometrium of women with adenomyosis showed no significant association (184.17 ± 9.4 vs.184.66 ± 16.08, p = 0.86) from the topic endometrium of women without adenomyosis. Conclusion: TGF-β1 expression was not increased in the ectopic endometrium of women with adenomyosis.
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@#Introduction: Epithelial-mesenchymal transition (EMT) is a process of epithelial transformation into mesenchymal cells. It is also a process that contributes to the progression of fibrosis and cancer metastasis. Transforming growth factor-beta (TGF-β), as a potent inducer of EMT, has therefore became a potential therapeutic target. However, clinical developments of TGF-β inhibitors have been un-successful due to safety risks. Hence, drug repurposing of existing safe-to-use drugs could over-come this issue. Methods: In this study, the TGF-β receptor type 1 (ALK5) was selected as the target protein. Molecular docking was performed using known ALK5 inhibitors as positive controls. Clinical drugs with similar binding affinity and amino acid interaction were selected for in vitro experimental validation. Results: ALK5 inhibitor demonstrated binding affinities ranging from -11.2 to -9.5 kcal/mol. Analysis of amino acid interaction revealed that Val219, Ala230, Lys232, and Leu340 amino acid residues are crucial for binding. Subsequent screening of clinically approved drugs against ALK5 showed top five potential drugs (ergotamine, telmisartan, saquinavir, indinavir, and nelfinavir). The selected drugs were tested in TGF-β1-induced normal human bronchial epithelial cell line, BEAS-2B. Western blot analysis showed that the drugs did not exhibit inhibitory effects on the downregulation of epithelial proteins (E-cadherin) and upregulation of mesenchymal proteins (vimentin and α-smooth muscle actin). Conclusion: Based on these experimental outcome, it is postulated that the results from molecular docking were false positives. The tested drugs in this study could serve as negative controls in future screening against ALK5 protein.
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Objective To investigate the changes of macrophage polarization during acute rejection (AR) after intestinal transplantation. Methods Six Brown Norway (BN) rats and 24 Lewis rats were divided into the sham operation group (6 Lewis rats), syngeneic transplantation group (Lewis→Lewis, 6 donors and 6 recipients) and allogeneic transplantation group (BN→Lewis, 6 donors and 6 recipients). At postoperative 7 d, the intestinal graft tissues in all groups were collected for hematoxylin-eosin (HE) staining and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay. Pathological manifestations and cell apoptosis were observed. The expression levels of serum cytokines related to M1 and M2 macrophage polarization were determined by enzyme-linked immunosorbent assay (ELISA). Surface markers of M1 and M2 macrophages of intestinal graft tissues in each group were co-localized and counted by immunofluorescence staining. Results HE staining and TUNEL assay showed that the intestinal epithelial morphology and structure were normal and no evident apoptotic bodies were found in the sham operation and syngeneic transplantation groups. At 7 d after transplantation, the epithelial villi structure of intestinal graft tissues was severely damaged, the number of crypts was decreased, the number of apoptotic bodies was increased, and inflammatory cells infiltrated into the whole intestinal wall, manifested with moderate to severe AR in the allogeneic transplantation group. ELISA revealed that the expression levels of serum cytokines related to M1 macrophage polarization, such as tumor necrosis factor (TNF)-α, interferon (IFN)-γ and interleukin (IL)-12, of the recipient rats in the allogeneic transplantation group were higher than those in the sham operation and syngeneic transplantation groups. The expression levels of serum cytokines related to M2 macrophage polarization, such as IL-10 and transforming growth factor (TGF)-β, in the syngeneic transplantation group were higher compared with those in the sham operation and allogeneic transplantation group, and the differences were statistically significant (all P<0.05). Immunofluorescence staining showed that the number of M1 macrophages in the allogeneic transplantation group was higher than those in the sham operation and syngeneic transplantation groups, and the number of M2 macrophages in the syngeneic transplantation group was higher than those in the sham operation and allogeneic transplantation groups, and the differences were statistically significant (all P<0.05). Conclusions Among the allografts with AR after intestinal transplantation, a large number of macrophages, mainly M1 macrophages secreting a large number of pro-inflammatory cytokines, infiltrate into the whole intestinal wall. Regulating the direction of macrophage polarization is a potential treatment for AR after intestinal transplantation.
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AIM: To investigate the effect of ALK5 inhibitor EW-7197 on the proliferation and migration of human Tenon fibroblasts(HTFs)induced by transforming growth factor-β1(TGF-β1)and its mechanism.METHODS: The cell proliferation rate was detected by MTS assay, and the optimal concentration and time of EW-7197 were explored. Then HTFs were divided into three groups: normal control group, TGF-β1 induced group and TGF-β1+EW-7197 group. Cell migration was observed by Transwell assay. The protein expression levels of Fibronectin, α-SMA, as well as the phosphorylated Smad2, Smad3(p-Smad2, p-Smad3)were measured by Western blot.RESULTS: MTS assay showed that the proliferation rate of cells treated with 6.0 μmol/L EW-7197 for 24h was the lowest(all P<0.01). Transwell assay showed that the migrated number of HTFs in TGF-β1 induced group was 228.0±17.0/field, which was significantly more than that in normal control group(149.0±15.0/field)and TGF-β1+EW-7197 group(46.0±8.0/field; all P<0.01). Western blot showed that the protein relative expression levels of Fibronectin, α-SMA and p-Smad2, p-Smad3 of HTFs in TGF-β1 induced group were significantly higher than that in normal control group and TGF-β1+EW-7197 group(all P<0.001).CONCLUSION:EW-7197 can suppress the proliferation and migration of TGF-β1-induced HTFs through TGF-β/Smad signaling pathways.
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@#To investigate whether rare ginsenosides could alleviate idiopathic pulmonary fibrosis (IPF), C57BL/6 mice were randomly divided into control group, bleomycin (BLM)-induced IPF group, rare ginsenoside Rk1 group, rare ginsenoside Rk3 group, rare ginsenoside Rh4 group and rare ginsenoside Rg5 group.All mice except those in the control group were given bleomycin injection.The IPF model was established by BLM for 28 days.The treatment group was given ginsenoside intragastrically at the same time.After the experiment, the lung tissues of mice were collected and the pathological changes of the mice lungs were observed.The content of hydroxyproline (HYP) in mouse lung tissue was measured.The expression of IPF-related genes in mouse lung tissues was detected.In in vitro experiments, Medical Research Council cell strain-5 (MRC-5) was used to induce IPF cell model using transforming growth factor-β1 (10 ng/mL).The effects of four saponins on the expression of IPF-related genes were analyzed by MTT assay, HYP content determination and RT-qPCR.All four rare ginsenosides could effectively alleviate the pathological process such as alveolar structure destruction caused by IPF, reduce the content of HYP, and down-regulate the expression of IPF-related genes, indicating that rare ginsenosides can effectively alleviate IPF.
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OBJECTIVE@#To investigate the effect of Bushen Chushi decoction combined with platelet-rich plasma(PRP) to treat knee osteoarthritis(KOA) in early and middle stage and its regulation on TGF-β1 and Smad-1 expression in serum.@*METHODS@#Total of 45 patients with KOA in early and middle stage from May 2020 to April 2022 were treated and divided into control group and observation group. In control group, there were 30 patients including 12 males and 18 females, aged from 43 to 69 years old with an average of(57.3±6.5) years old and disease duration ranged from 1.5 to 5.0 years with an average of(3.8±1.7) years, and there were 8 cases in gradeⅠ, 13 cases in gradeⅡ, and 9 cases in grade Ⅲ according to Kellgren-Lawrence Grade, PRP 5 ml was injected into knee joint on the first day of No1, 3 week together for 2 times. In the observation group, there were 15 cases including 7 males and 8 females, aged from 45 to 70 years old with an average of (56.7±6.2) years old and disease duration ranged from 1.8 to 5.7 years with an average of (4.0±1.8) years, there were 4 cases in gradeⅠ, 9 cases in gradeⅡand 4 cases in grade Ⅲ according to the Kellgren-Lawrence Grade, PRP 5 ml were injected into knee joints that the time and frequency were the same as those in the control group, and at the same time Bushen Chushi decoction orally were taken 1 dose per day with a total of 28 doses. All patients were treated for four weeks. Visual analogue scale(VAS) and Lequesne MG score before and after treatment were used to evaluate improvement of knee pain and joint function. The TGF-β1 and Smad-1 levels in serum were measured before and after treatment in two groups. The incidence of complications in two groups was observed.@*RESULTS@#All patients were followed up for 26 to 30 days with an average of (28.0±0.6) days. There was no significant difference in VAS and knee Lequesne MG scores between two groups before treatment(P>0.05). The scores of VAS and knee Lequesne MG on the first day after treatment in both groups were lower than those before treatment(P<0.05). The VAS and knee Lequesne MG scores in observation group were lower than those in control group(P<0.05) on the first day after treatment. The TGF-β1 level in serum after treatment were higher significantly than that before treatment in two groups(P<0.05). After treatment, TGF-β1 level in serum in observation group were lower than those in control group with statistically significant differences(P<0.05). The Smad-1 levels in serum after treatment in observation group were higher significantly than that in control group(P<0.05). The levels of Smad-1 were not statistically significant between before and after treatment(P>0.05). There was no significant difference in postopertaive complications between two groups (P>0.05).@*CONCLUSION@#The efficacy of Bushen Chushi decoction combined with PRP in treatment of early and middle KOA is better than that of PRP injection alone. The combined treatment could reduce TGF-β1 level and increase Smad-1 level in serum, which may be a mechanism to inhibit inflammation and alleviate cartilage degeneration to some extent.
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OBJECTIVES@#To investigate the effect of recombinant human fibroblast growth factor 21 (rhFGF21) on the proliferation and mineralization of cementoblasts and its mechanism.@*METHODS@#Hematoxylin eosin, immunohistochemical staining, and immunofluorescence were used to detect the expression and distribution of fibroblast growth factor 21 (FGF21) in rat periodontal tissues and cementoblasts (OCCM-30), separately. Cell Counting Kit-8 was used to detect the proliferation of OCCM-30 under treatment with rhFGF21. Alkaline phosphatase staining and Alizarin Red staining were used to detect the mineralization state of OCCM-30 after 3 and 7 days of mineralization induction. The transcription and protein expression of the osteogenic-related genes Runx2 and Osterix were detected by real-time quantitative polymerase chain reaction (PCR) and Western blot analysis. The expression levels of genes of transforming growth factor β (TGFβ)/bone morphogenetic protein (BMP) signaling pathway in OCCM-30 were detected through PCR array analysis.@*RESULTS@#FGF21 was expressed in rat periodontal tissues and OCCM-30. Although rhFGF21 had no significant effect on the proliferation of OCCM-30, treatment with 50 ng/mL rhFGF21 could promote the mineralization of OCCM-30 cells after 7 days of mineralization induction. The transcriptional levels of Runx2 and Osterix increased significantly at 3 days of mineralization induction and decreased at 5 days of mineralization induction. Western blot analysis showed that the protein expression levels of Runx2 and Osterix increased during mineralization induction. rhFGF21 up-regulated Bmpr1b protein expression in cells.@*CONCLUSIONS@#rhFGF21 can promote the mineralization ability of OCCM-30. This effect is related to the activation of the TGFβ/BMP signaling pathway.
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Humanos , Ratas , Animales , Cemento Dental , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Diferenciación Celular , Proteínas Morfogenéticas Óseas/metabolismo , Factor de Crecimiento Transformador beta/farmacologíaRESUMEN
AIM: To investigate the effect of the intravitreal injection of vascular endothelial growth factor-A165(VEGF-A165)on the scleral remodeling of guinea pigs with form-deprivation myopia(FDM).METHODS: A total of 120 tricolor guinea pigs, aged three weeks, were randomly divided into 6 groups, with 20 in each group. The blank group did not undergo any intervention. In the FDM group, only the FDM model was established. In the phosphate buffer saline(PBS)group, 2.5 μL of PBS was injected into the vitreous cavity before establishing the FDM model. In the 1ng group, 5ng group, and 10ng group, VEGF-A165 was injected into the vitreous cavity at concentrations of 1, 5 and 10ng, respectively, before the establishment of the FDM model. The FDM model was established by covering the right eyes of guinea pigs with translucent balloons for 14d. The diopter and axial length of the right eyes were measured before and after covering. After 14d, the content of dopamine(DA)in retina was measured by high performance liquid chromatography. Additionally, the mRNA and protein expression levels of matrix metalloproteinase-2(MMP-2), tissue inhibitor of matrix metalloproteinase-2(TIMP-2), transforming growth factor(TGF)-β1, TGF-β2 and α-smooth muscle actin(α-SMA)in sclera were detected by reverse transcription polymerase chain reaction(RT-PCR)and Western blot.RESULTS: Before covering, there were no significant differences in the diopter and axial length of the right eyes of guinea pigs in all groups(P>0.05). After 14d of modeling, when compared with the blank group, FDM group showed an increase in the degree of myopia in the right eye, a prolongation of the axial length, a decrease in the content of DA in the retina, and an increase in the expression of MMP-2, TGF-β2 and α-SMA in the sclera. Conversely, the expression of TIMP-2 and TGF-β1 were decreased(P<0.01). However, in comparison to the FDM group, the degree of myopia in the 1ng, 5ng, and 10ng groups of guinea pigs decreased, the growth trend of axial length slowed, the content of DA in the retina increased, and the expression of MMP-2, TGF-β2 and α-SMA in the sclera decreased. Furthermore, the expression of TIMP-2 and TGF-β1 in the sclera increased(P<0.01). As the concentration of intravitreal injection of VEGF-A165 increased, the degree of myopia in the right eye of guinea pigs gradually increased, and the axial length gradually prolonged. The content of DA in the retina gradually decreased, the expression of MMP-2, TGF-β2, and α-SMA in the sclera gradually increased, while the expression of TIMP-2 and TGF-β1 decreased gradually.CONCLUSION: Intravitreal injection of VEGF-A165 can increase the content of DA in the retina of FDM guinea pigs, affect the expression of MMP-2, TIMP-2, TGF-β1, TGF-β2 and α-SMA in the sclera, and inhibit scleral remodeling of guinea pigs. Notably, the VEGF-A165 at the concentration of 1ng showed the most significant efficacy.
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AIM: To investigate the mechanism of pyrrolidine dithiocarbamate(PDTC)on transforming growth factor-beta 2(TGF-β2)-induced epithelial-mesenchymal transition(EMT)in human lens epithelial cells(LECs).METHODS: LECs were treated with various doses of PDTC chemicals following TGF-β2 caused EMT on these cells. Cell proliferation and lateral migration were discovered using the CCK-8 and cell scratch test. The markers of EMT, including E-cadherin, α-SMA and nuclear factor-κB(NF-κB)signaling pathway-related expression, were tested by Western Blot as well as the changes in the expression of the apoptosis-related proteins BAX, BCL-2, Caspase-3, and Cyclin D1.RESULTS: The proliferation and migration viability of cells in the TGF-β2 treated group was increased compared to the group without TGF-β2, and the expression of α-SMA increased whereas the E-cadherin expression decreased. With the effect of TGF-β2, NF-κB p65 and phosphorylated NF-κB p65 expression increased, the concentration of TGF-β2 that had the greatest capacity for proliferation and migration was 10 ng/mL(P<0.05). Mechanism study of PDTC-induced EMT reversal and apoptosis showed that cell viability and migratory capability were both significantly reduced after PDTC intervention; PDTC prevents IκB phosphorylation, thus inhibiting NF-κB nuclear translocation. Protein associated to the NF-κB signaling pathway, and protein expression of NF-κB/IκBα/p-IκBα/Iκκ-α/p-Iκκ-α was decreased(P<0.05), PDTC increased the expression of the pro-apoptotic protein BAX/Caspase-3, expression of the inhibitor of apoptosis protein BCL-2 and the cell cycle protein Cyclin D1 was reduced. The expression of NF-κB/IκB mRNA was reduced, expression of the apoptosis-related mRNA BAX increased, while BCL-2 reduced.CONCLUSION: The EMT in LECs cells induced by TGF-β2 can be significantly reversed by PDTC, which may be related to the decreased expression of NF-κB p65/IκB/Iκκ-α and activation of apoptosis-related protein. PDTC can reverse EMT by inhibiting NF-κB signaling pathway and induce apoptosis of abnormally proliferated cells, which will provide new potential therapeutic agents for posterior capsular opacification(PCO)treatment.
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Objective:Rat model of RA complicated with pulmonary fibrosis were constructed to observe the degree of improvement of pulmonary fibrosis in RA rats by JAK2 inhibitor CEP33779 and the possible mechanisms.Methods:①The RA models were constructed by subcutaneous injection of 0.2 ml (1 mg/ml) of bovine type Ⅱ collagen into the tail of the rats on the day of modeling development (d0); intratracheal injection of 100 μl bleomycin (2.5 mg/kg) was used to induce pulmonary fibrosis model at d13. In vivo study: model rats were randomly divided into the normal group, pulmonary fibrosis group, pulmonary fibrosis CEP treatment group, RA complicated with pulmonary fibrosis group, and RA complicated pulmonary fibrosis CEP treatment group. Rats in the treatment group was given CEP (10 ml/kg) qd by gavage from d14 to week 4. The right hind foot of the rats was measured for joints swelling and the arthritis index score were measured, lung compliance (Cst) and lung specific gravity were measured. In addition, the pathological changes of the left lung were observed by HE and Masson staining, and the extracellular matrix level of the right lung was measured by protein immunoblotting (WB). ② In vitro study: TGF-β 1 (10 ng/ml) was applied to stimulate human embryonic lung fibroblasts (HFL1) for 24 h and 48h, and p-JAK2 expression was detected by immunofluorescence. After HFL1 inoculation of culture plates, the control group, TGF-β 1 stimulation group, TGF-β 1+ LY2109761 (TGFβ-R1/2 inhibitor group, 0.5 mmol/L and 2 mmol/L) group, TGF-β 1+CEP (0.1 mmol/L and 1.0 mmol/L) group were co-incubated for 48 h, and the expression levels of TGFβ-R2, α-SMA, JAK2, and Col 1 were measured by WB. Comparisons between multiple groups were made by Tukey′s test, and comparisons between the two groups were analyzed by independent t-test. Results:① In vivo study, compared with the control group (1.45±0.04), joint swelling was increased at d13 [(2.54±0.16) in RA+PF+Vehicle group, t=16.02, P<0.001], and the mean arthritis index score and toe volume were decreased 3 days after CEP treatment(d16) [(2.89±0.11), t=5.78, P<0.001; (1.92±0.13), t=6.85, P<0.001]. For rats with pulmonary fibrosis, all had different degrees of lung enlargement, increased lung specific gravity, decreased Cst, and increased lung inflammation and fibrosis[(0.96±0.06), t=19.76, P<0.001; (0.26±0.09), t=17.64, P<0.001; (3.63±1.51), t=6.00, P<0.001; (1.75±0.71), t=5.84, P<0.001]. After CEP gavage, rats that had RA complicated with pulmonary fibrosis had reduced lung swelling, decreased lung specific gravity, increased Cst [(0.82±0.05), t=5.76, P<0.001; (0.43±0.18), t=2.31, P=0.038], and the scores of pulmonary fibrosis and inflammation [(3.00±1.00); (1.56±0.52)] all showed a trend of decrease, but did not reach statistical difference CEP inhibited the expression of TGF-β 1, TGFβ-R2, α-SMA, Fn and JAK2 in lung tissue of pulmonary fibrosis rats, and the differences among the five groups were statistically significant ( F=9.02, P=0.017; F=4.86, P=0.048; F=6.57, P=0.032; F=11.26, P=0.010; F=13.32, P=0.007). ② In vitro study, TGF-β 1 stimulated HFL1 showed stronger phosphorylated JAK2 (p-JAK2) fluorescent signal WB showed a significant increase in the expression of TGFβ-R2, α-SMA, JAK2 and Col1, and after LY and CEP intervention, the above proteins were reduced in a concentration-dependent manner, with statistically significant differences among all five groups ( F=337.30, P<0.001; F=20.61, P<0.001; F=100.60, P<0.001; F=180.90, P<0.001). Conclusion:JAK2 inhibitors can ameliorate RA-related pulmonary fibrosis, and the mechanism may be through interfering with the "crosstalk" between JAK2 and TGF-β 1 signaling pathway.
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Objective:To study the early predictive values of serum thrombospondin-1(TSP-1)and transforming growth factor-β1(TGF-β1) for bronchopulmonary dysplasia(BPD)in preterm infants.Methods:From September 2020 to April 2022, preterm infants with gestational age<32 weeks and ≥28 weeks as well as birth weight<1 500 g admitted to neonatal intensive care unit within 2 hours after birth were enrolled in the study.The dynamic changes of serum TSP-1 and TGF-β1 levels in preterm infants were observed on 1st, 7th, 14th, and 28th day after birth.Preterm infants were divided into BPD group and non-BPD group according to the diagnostic criteria of BPD.Receiver operating characteristic(ROC) curve and area under curve(AUC)was used to analyze the predictive value of serum TSP-1 and TGF-β1 for preterm infants with BPD.Results:According to the diagnostic criteria of BPD, 38 cases were in the BPD group and 52 cases in the non-BPD group.There was no significant difference in gestational age, birth weight and gender between the two groups( P>0.05). The levels of TSP-1 and TGF-β1 in the serum of BPD group were gradually increased, which were significantly higher than those of non-BPD group on the 1st, 7th, 14th, and 28th day( P<0.001). ROC results showed that AUC of TSP-1, TGF-β1 and their combination for predicting BPD were 0.889(95% CI 0.819~0.959), 0.826(95% CI 0.743~0.910), and 0.923(95% CI 0.870~0.976), respectively.The sensitivity were 86.80%, 86.70%, 89.50%, and the specificity were 86.50%, 73.10%, 80.80%, respectively.Cutoff values of TSP-1 and TGF-β1 for predicting BPD were 44.50 μg/L and 6.13 μg/L, respectively. Conclusion:Combined detection of serum TSP-1 and TGF-β1 on the first day after birth has an early predictive value for BPD in preterm infants.
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Objective:To investigate the effects of alum ice nanoemulsion on VEGF and TGF-β1 in hypertrophic scar based on Notch signaling pathway.Methods:Totally 144 SD rats were divided into blank control group, model group, triamcinolone acetonide group and alum ice nanoemulsion low-, medium- and high-dose groups according to random number table method, with 24 rats in each group. Except for the blank control group, the rats in other groups were prepared with deep Ⅱ ° burn models. 24 hours after the successful modeling, the model group was given the same amount of normal saline, the rats in alum ice nanoemulsion low-, medium- and high-dose groups were given 8.15, 6.30 and 32.60 mg/ml alum ice nanoemulsion respectively, and the triamcinolone acetonide group was given triamcinolone acetonide twice a day, 0.2 ml each time, for 35 consecutive days. At 14, 21, 28 and 35 d, the collagen fiber surface density was calculated by VG staining. The protein expressions of vascular endothelial growth factor (VEGF), transforming growth factor-β1 (TGF-β1), Notch1 and Jagged1 were detected by Western Blot. The expressions of Notch1 mRNA and Jagged1 mRNA were detected by RT-PCR.Results:Compared with model group, triamcinolone acetonide and different doses of alum ice nanoemulsion groups could decrease collagen fiber surface density, protein expressions of VEGF, TGF-β1, Notch1, Jagged1 and mRNA expressions of Notch1, Jagged1 in different degrees ( P<0.05). Compared with the triamcinolone acetonide group, the collagen fiber surface density, protein expressions of VEGF, TGF-β1, Notch1 and Jagged1 and mRNA expressions of Notch1, Jagged1 in the alum ice nanoemulsion medium-dosage group decreased ( P<0.05). Conclusion:Alum ice nanoemulsion can inhibit hypertrophic scar formation, and its mechanism is related to down-regulating Notch signal pathway related molecules Notch1, Jagged1 protein and mRNA levels, and then down-regulating VEGF and TGF-β1 protein expressions.
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Objective:To investigate the protective effect and possible mechanism of Tangshenbao on renal damage in diabetic nephropathy (DN) rats.Methods:Totally 36 SPF male SD rats were randomly divided into normal group ( n=6) and model group ( n=30). The DN rat model was prepared by single high-dose intraperitoneal injection of STZ. According to the random number table method, the rats were divided into model group, irbesartan group and Tangshenbao low-, medium- and high-dosage groups, with 6 rats in each group. Drug intervention lasted for 8 weeks. The general condition and body weight of rats in each group were recorded. The blood glucose, kidney index, 24 h urine protein (24 h UTP), SCr and BUN levels were detected. The pathological morphology of renal tissue was observed by PAS staining and transmission electron microscopy. The mRNA and protein expressions of Ets-1, TGF-β1, Smad2 and Smad3 in renal tissue were detected by real-time fluorescence quantitative PCR and Western blot. Results:Compared with model group, the body weight of Tangshenbao low, medium and high dose groups and irbesartan group significantly increased ( P<0.01). The kidney index decreased ( P<0.05 or P<0.01). The contents of 24 hUTP, BUN and SCr significantly decreased ( P<0.05 or P<0.01). Glomerular volume was significantly reduced ( P<0.05 or P<0.01), the mRNA expressions of Ets-1 (1.59 ± 0.06, 1.47 ± 0.04, 1.31 ± 0.03, 1.39 ± 0.03 vs. 1.64 ± 0.04), TGF-β1 (1.65 ± 0.05, 1.59 ± 0.03, 1.38 ± 0.05, 1.49 ± 0.04 vs. 1.77 ± 0.08), Smad2 (1.48 ± 0.05,1.39 ± 0.05, 1.22 ± 0.03, 1.31 ± 0.04 vs. 1.54 ± 0.05), Smad3 (1.57 ± 0.04, 1.48 ± 0.03, 1.28 ± 0.03, 1.39 ± 0.02 vs. 1.64 ± 0.05) in renal tissue of rats significantly decreased ( P<0.05 or P<0.01), the protein expressions of Ets-1 (1.33 ± 0.32, 1.16 ± 0.38, 0.77 ± 0.06, 0.84 ± 0.06 vs. 1.97 ± 0.43), TGF-β1 ( 1.35 ± 0.14, 1.24 ± 0.22, 0.94 ± 0.13, 1.07 ± 0.06 vs. 1.63 ± 0.20), Smad2 (1.24 ± 0.26, 1.14 ± 0.31, 0.77 ± 0.28, 0.85 ± 0.19 vs. 1.72 ± 0.34) and Smad3 (1.29 ± 0.14, 1.19 ± 0.21, 0.85 ± 0.39, 0.90 ± 0.37 vs. 1.76 ± 0.21) decreased ( P<0.05 or P<0.01). Conclusion:Tangshenbao can improve renal damage in DN rats, and its mechanism may be related to the inhibition of Ets-1 expression and TGF-β1/Smads signaling pathway.
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Background Radiation-induced liver damage is a major complication for primary liver cancer and other upper abdominal tumors during radiation therapy. The early biological effects of radiation-induced liver damage at different doses of radiation and its mechanisms of action have not yet been elucidated. Objective To establish X-ray-induced radioactive mouse liver damage model and explore the level of oxidative stress and its correlation with nuclear factor-κB (NF-κB) and transforming growth factor-β1 (TGF-β1). Methods A total of 24 male C57BL/6J mice aged 6 weeks were randomly divided into 4 groups (control, 0.8 Gy, 1.6 Gy, and 4 Gy), with 6 mice in each group. X-rays irradiated the whole body of mice singly in each dose group. At 24 h after radiation, histopathological changes in mouse liver were evaluated; peripheral blood cell count, serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels, as well as liver tissue superoxide dismutase (SOD) activity, malondialdehyde (MDA) level, reduced glutathione (GSH) level, and 8-hydroxy-2′-deoxyguanosine (8-OHdG) level were measured; real-time fluorescence quantitative PCR was used to detect liver tissue NF-κB p65 and TGF-β1 mRNA expression levels; the correlations of oxidative stress indicators with NF-κB p65 and TGF-β1 mRNA expression levels were analyzed by Pearson correlation. Results Compared with the control group, at 24 h after different doses of X-ray radiation, early injury-related histopathological changes were observed in liver, and the serum levels of AST and ALT were significantly increased in the 4 Gy group (P<0.05); the numbers of peripheral blood leukocytes and lymphocytes were decreased in the radiation exposure groups (P<0.05), showing a decreasing trend with increasing radiation doses; the levels of liver oxidative stress indicators (MDA, SOD, and GSH) in exposed mice were significantly increased (P<0.05), showing an increasing trend with increasing radiation doses. The liver 8-OHdG were significantly increased in the 1.6 Gy and 4 Gy groups compared with the control and the 0.8 Gy groups, respectively (P<0.05). The NF-κB p65 and TGF-β1 mRNA expression levels in the liver of mice were significantly increased in the 1.6 Gy and 4 Gy groups compared with the control group (P<0.05). The TGF-β1 mRNA expression level also exhibited an increasing trend with increasing radiation doses. The results of correlation analysis showed that the levels of MDA, SOD, GSH, and 8-OHdG in liver tissues were significantly and positively correlated with the expression levels of NF-κB p65 and TGF-β1 mRNA (P<0.05). Conclusion X-rays of various doses can affect the degree of liver injury, peripheral blood cell count, serum levels of AST and ALT, and liver oxidative stress levels in mice. The level of oxidative stress induced by X-ray is positively correlated with NF-κB and TGF-β1 in liver tissues, and it may participate in the process of radiation-induced liver injury.
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Aim To investigate the therapeutic effect of Jiangtang Wan (JTW) on mioe with diabetic kidney disease (DKD) and to explore its potential moiecuiar mechanism on ameliorating renal injury and fibrosis. Methods C57BL/6 mice were randomly divided into four groups: the control group, the model group, JTW group (1250 mg • kg
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OBJECTIVE@#Huangqi Decoction (HQD), a classical traditional Chinese medicine formula, has been used as a valid treatment for alleviating liver fibrosis; however, the underlying molecular mechanism is still unknown. Although our previous studies showed that microRNA-663a (miR-663a) suppresses the proliferation and activation of hepatic stellate cells (HSCs) and the transforming growth factor-β/small mothers against decapentaplegic (TGF-β/Smad) pathway, whether long noncoding RNAs (lncRNAs) are involved in HSC activation via the miR-663a/TGF-β/Smad signaling pathway has not yet reported. The present study aimed to investigate the roles of lncRNA lnc-C18orf26-1 in the activation of HSCs and the mechanism by which HQD inhibits hepatic fibrosis.@*METHODS@#The expression levels of lnc-C18orf26-1, miR-663a and related genes were measured by quantitative reverse transcription-polymerase chain reaction. HSCs were transfected with the miR-663a mimic or inhibitor and lnc-C18orf26-1 small interfering RNAs. The water-soluble tetrazolium salt-1 assay was used to assess the proliferation rate of HSCs. Changes in lncRNA expression were evaluated in miR-663a-overexpressing HSCs by using microarray to identify miR-663a-regulated lncRNAs. RNA hybrid was used to predict the potential miR-663a binding sites on lncRNAs. Luciferase reporter assays further confirmed the interaction between miR-663a and the lncRNA. The expression levels of collagen α-2(I) chain (COL1A2), α-smooth muscle actin (α-SMA) and TGF-β/Smad signaling pathway-related proteins were determined using Western blotting.@*RESULTS@#Lnc-C18orf26-1 was upregulated in TGF-β1-activated HSCs and competitively bound to miR-663a. Knockdown of lnc-C18orf26-1 inhibited HSC proliferation and activation, downregulated TGF-β1-stimulated α-SMA and COL1A2 expression, and inhibited the TGF-β1/Smad signaling pathway. HQD suppressed the proliferation and activation of HSCs. HQD increased miR-663a expression and decreased lnc-C18orf26-1 expression in HSCs. Further studies showed that HQD inhibited the expression of COL1A2, α-SMA, TGF-β1, TGF-β type I receptor (TGF-βRI) and phosphorylated Smad2 (p-Smad2) in HSCs, and these effects were reversed by miR-663a inhibitor treatment.@*CONCLUSION@#Our study identified lnc-C18orf26-1 and miR-663a as promising therapeutic targets for hepatic fibrosis. HQD inhibits HSC proliferation and activation at least partially by regulating the lnc-C18orf26-1/miR-663a/TGF-β1/TGF-βRI/p-Smad2 axis.
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Humanos , Factor de Crecimiento Transformador beta/farmacología , Factor de Crecimiento Transformador beta1/metabolismo , ARN Largo no Codificante/farmacología , Medicamentos Herbarios Chinos/farmacología , MicroARNs/genética , Células Estrelladas Hepáticas/patología , Cirrosis Hepática/metabolismo , Proliferación Celular , Factores de Crecimiento Transformadores/farmacologíaRESUMEN
OBJECTIVE@#To study effects of Shenmai Injection on hypertensive heart failure and its mechanism for inhibiting myocardial fibrosis.@*METHODS@#Salt-sensitive (Dahl/SS) rats were fed with normal diet (0.3% NaCl) and the high-salt diet (8% NaCl) to observe the changes in blood pressure and heart function, as the control group and the model group. Salt-insensitive rats (SS-13BN) were fed with the high-salt diet (8% NaCl) as the negative control group. After modeling, the model rats were randomly divided into heart failure (HF) group, Shenmai Injection (SMI) group and pirfenidone (PFD) group by a random number table, with 6 rats in each group. They were given sterilized water, SMI and pirfenidone, respectively. Blood pressure, cardiac function, fibrosis and related molecular expression were detected by sphygmomanometer, echocardiogram, enzyme linked immunosorbent assay (ELISA), hematoxylin-eosin staining, Masson staining, immunofluorescence and qPCR analysis.@*RESULTS@#After high-salt feeding, compared with the control and negative control group, in the model group the blood pressure increased significantly, the left ventricular ejection fraction (LVEF) and left ventricular fraction shortening (LVFS) were significantly reduced, and the serum NT-proBNP concentration increased significantly (all P<0.05); furthermore, the arrangement of myocardial cells was disordered, the edema was severe, and the degree of myocardial fibrosis was also significantly increased (P<0.05); the protein and mRNA expressions of collagen type I (Col I) were up-regulated (P<0.05), and the mRNA expressions of transforming growth factor β 1 (TGF- β 1), Smad2 and Smad3 were significantly up-regulated (P<0.05). Compared with HF group, after intervention of Shenmai Injection, LVEF and LVFS increased, myocardial morphology was improved, collagen volume fraction decreased significantly (P<0.05), and the mRNA expressions of Col I, TGF- β 1, Smad2 and Smad3, as well as Col I protein expression, were all significantly down-regulated (all P<0.05).@*CONCLUSION@#Myocardial fibrosis is the main pathological manifestation of hypertensive heart failure, and Shenmai Injection could inhibit myocardial fibrosis and effectively improve heart failure by regulating TGF-β 1/Smad signaling pathway.