Objective To investigate whether 2,3,5,4'-tetrahydroxystilbene-2-O-β-glucoside(TSG)ameliorated polycystic ovary syndrome(PCOS)-like characteristics by inhibiting inflammation.Methods PCOS models were established ...Objective To investigate whether 2,3,5,4'-tetrahydroxystilbene-2-O-β-glucoside(TSG)ameliorated polycystic ovary syndrome(PCOS)-like characteristics by inhibiting inflammation.Methods PCOS models were established by injecting subcutaneously with dehydroepiandrosterone into female Sprague-Dawley rats,followed by receiving intraperitoneal injection of TSG.The granular cells(GCs)KGN were transfected with small interfering RNAs(si-NC and si-CYP19A1).The cells were preincubated with lipopolysaccharide(LPS)and then treated with or without TSG.The estrous cycle was monitored using vaginal exfoliated cells.The morphology of ovarian follicles was analyzed by H&E staining.ELISA was used to analyze estradiol(E2),testosterone(T),follicle stimulating hormone(FSH),luteinizing hormone(LH),IL-6,TNF-α,AGEs,CRP and Omentin-1 levels in serum.Immunohistochemistry was performed to analyze PCNA and CYP19A1 expressions in the GCs of ovaries.Tunel staining was executed to detect the apoptosis of GCs.Quantitative polymerase chain reaction(qPCR)and Western blot were implemented to measure the expression of CYP19A1 in the ovaries and transfected cells.qPCR was used to analyze the expression of IL-6 and TNF-αin the transfected cells treated with LPS and TSG.Results The estrous cycles were restored in TSG group.Compared with model group,the sinus follicles were reduced and corpus luteums were increased in TSG group.TSG group showed increased E2,and decreased T and LH,compared with model group.Pro-inflammatory factors(IL-6,TNF-α,CRP and AGEs)were decreased,and anti-inflammatory factor(Omentin-1)was increased in TSG group compared with those in model group.TSG could partially inhibit decrease of PNCA-positive GCs and increase of Tunel-positive GCs caused by PCOS.The CYP19A1 expression of GCs in TSG group was upregulated compared with model group.The expressions of IL-6 and TNFαin si-CYP19A1 cells were increased compared with si-NC cells.Compared with cells(si-NC and si-CYP19A1)treated without LPS,the expressions of IL-6 and TNF-αcells were increased,and the expression of CYP19A1 was downregulated in LPS-preincubated cells.Compared with cells treated with LPS,the expression of IL-6 and TNF-αwere decreased,and the expression of CYP19A1 was increased in cells treated with LPS and TSG.Compared with si-NC cells treated with LPS and TSG,the expressions of IL-6 and TNF-αcells were increased in the si-CYP19A1 cells treated with LPS and TSG.Conclusion TSG could alleviate PCOS-like characteristics by increasing the expression of CYP19A1 in GCs to inhibit inflammatory response.展开更多
Background&Objective Knee osteoarthritis(OA)is a degenerative disease,which not only induces superficial cartilage defects and full-thickness cartilage defects,but also exacerbates the microenvironment of the knee...Background&Objective Knee osteoarthritis(OA)is a degenerative disease,which not only induces superficial cartilage defects and full-thickness cartilage defects,but also exacerbates the microenvironment of the knee joint and affects the mechano-chemical responses of the organ.As a growth/repair factor,mechanical growth factor(MGF)has the function of preventing OA,promoting cartilage regeneration and repairing damaged ligaments.activating transcription factor 2(ATF-2),a transcription factor,has the property of binding to cytokines,which makes it involved in the transcriptional regulation of various pathways in response to cellular stress,inflammatory cytokine and growth factors.At present,little is known about the effect of MGF on human osteoarthritis ligament fibroblasts(OA-LFs),and whether the approach can promote OA-LFs timely response to the mechanical injury and initiate signaling pathway for cell survival.Therefore,the purpose of this study is to investigate whether MGF promotes mechanical response to ligament fibroblasts in osteoarthritis knee cavity via ATF-2.Methods OA-LFs were seeded onto six-cell BioFlex plates and suffered from 12%static mechanical stretch[60 cycles/minute(1 Hz)]for 12 hours to mimic mechanical force mediated ligament injury.Meanwhile,OA-LFs were treated with MGF before and during mechanical stretch.Intracellular reactive oxygen species(ROS)and GRP78 mRNA expression were investigated to detect the cellular stress response of OA-LFs.The scratch test was performed to detect the migration ability of cells,gelatin zymography was used to examine the effect of MGF on the activity of matrix metalloproteinase 2(MMP-2)in OA-LFs,and cell deformation was detected by phalloidin-FITC staining after stretching.Quantitative real-time polymerase chain reaction(qRT-PCR)was used to screen the messenger RNA(mRNA)expression of ATF family members after OALFs treatment with MGF.Western blotting further proved that MGF is capable to activate the p-ATF-2.Results OA delays LFs response to mechanical injury,while MGF pretreatment can promote cells timely feedback the mechanically stimuli by inducing cellular stress.MGF treatment can alleviate the decline in cell migration ability caused by mechanical injury and further promote cell migration.In addition,MGF can reduce the activity of MM P-5 and alleviate the stretch-induced deformation of OA-LFs.Furthermore,the mRNA expression of ATF-2 up-regulated in a dose-dependent manner upon MGF treatment compared with control,while the expression of ATF-5 gene was down-regulated in a dose-dependent.Protein levels showed that the expression of p-ATF-2 increased with increasing MGF concentration.Conclusions Our study shows that MGF pretreatment of OA-LFs can respond quickly to mechanical damage and accelerate the ligament injury repair by promoting cell migration,decreasing the MMP-2 activity,and remitting the cell deformation.Therefore,MGF has potential as a therapeutic for OA patients.展开更多
文摘Objective To investigate whether 2,3,5,4'-tetrahydroxystilbene-2-O-β-glucoside(TSG)ameliorated polycystic ovary syndrome(PCOS)-like characteristics by inhibiting inflammation.Methods PCOS models were established by injecting subcutaneously with dehydroepiandrosterone into female Sprague-Dawley rats,followed by receiving intraperitoneal injection of TSG.The granular cells(GCs)KGN were transfected with small interfering RNAs(si-NC and si-CYP19A1).The cells were preincubated with lipopolysaccharide(LPS)and then treated with or without TSG.The estrous cycle was monitored using vaginal exfoliated cells.The morphology of ovarian follicles was analyzed by H&E staining.ELISA was used to analyze estradiol(E2),testosterone(T),follicle stimulating hormone(FSH),luteinizing hormone(LH),IL-6,TNF-α,AGEs,CRP and Omentin-1 levels in serum.Immunohistochemistry was performed to analyze PCNA and CYP19A1 expressions in the GCs of ovaries.Tunel staining was executed to detect the apoptosis of GCs.Quantitative polymerase chain reaction(qPCR)and Western blot were implemented to measure the expression of CYP19A1 in the ovaries and transfected cells.qPCR was used to analyze the expression of IL-6 and TNF-αin the transfected cells treated with LPS and TSG.Results The estrous cycles were restored in TSG group.Compared with model group,the sinus follicles were reduced and corpus luteums were increased in TSG group.TSG group showed increased E2,and decreased T and LH,compared with model group.Pro-inflammatory factors(IL-6,TNF-α,CRP and AGEs)were decreased,and anti-inflammatory factor(Omentin-1)was increased in TSG group compared with those in model group.TSG could partially inhibit decrease of PNCA-positive GCs and increase of Tunel-positive GCs caused by PCOS.The CYP19A1 expression of GCs in TSG group was upregulated compared with model group.The expressions of IL-6 and TNFαin si-CYP19A1 cells were increased compared with si-NC cells.Compared with cells(si-NC and si-CYP19A1)treated without LPS,the expressions of IL-6 and TNF-αcells were increased,and the expression of CYP19A1 was downregulated in LPS-preincubated cells.Compared with cells treated with LPS,the expression of IL-6 and TNF-αwere decreased,and the expression of CYP19A1 was increased in cells treated with LPS and TSG.Compared with si-NC cells treated with LPS and TSG,the expressions of IL-6 and TNF-αcells were increased in the si-CYP19A1 cells treated with LPS and TSG.Conclusion TSG could alleviate PCOS-like characteristics by increasing the expression of CYP19A1 in GCs to inhibit inflammatory response.
基金supported by the National Natural Science Foundation of China ( 11532004,31270990, 31600762)Innovation and Attracting Talents Program for College and University( “111”Project) ( B06023)
文摘Background&Objective Knee osteoarthritis(OA)is a degenerative disease,which not only induces superficial cartilage defects and full-thickness cartilage defects,but also exacerbates the microenvironment of the knee joint and affects the mechano-chemical responses of the organ.As a growth/repair factor,mechanical growth factor(MGF)has the function of preventing OA,promoting cartilage regeneration and repairing damaged ligaments.activating transcription factor 2(ATF-2),a transcription factor,has the property of binding to cytokines,which makes it involved in the transcriptional regulation of various pathways in response to cellular stress,inflammatory cytokine and growth factors.At present,little is known about the effect of MGF on human osteoarthritis ligament fibroblasts(OA-LFs),and whether the approach can promote OA-LFs timely response to the mechanical injury and initiate signaling pathway for cell survival.Therefore,the purpose of this study is to investigate whether MGF promotes mechanical response to ligament fibroblasts in osteoarthritis knee cavity via ATF-2.Methods OA-LFs were seeded onto six-cell BioFlex plates and suffered from 12%static mechanical stretch[60 cycles/minute(1 Hz)]for 12 hours to mimic mechanical force mediated ligament injury.Meanwhile,OA-LFs were treated with MGF before and during mechanical stretch.Intracellular reactive oxygen species(ROS)and GRP78 mRNA expression were investigated to detect the cellular stress response of OA-LFs.The scratch test was performed to detect the migration ability of cells,gelatin zymography was used to examine the effect of MGF on the activity of matrix metalloproteinase 2(MMP-2)in OA-LFs,and cell deformation was detected by phalloidin-FITC staining after stretching.Quantitative real-time polymerase chain reaction(qRT-PCR)was used to screen the messenger RNA(mRNA)expression of ATF family members after OALFs treatment with MGF.Western blotting further proved that MGF is capable to activate the p-ATF-2.Results OA delays LFs response to mechanical injury,while MGF pretreatment can promote cells timely feedback the mechanically stimuli by inducing cellular stress.MGF treatment can alleviate the decline in cell migration ability caused by mechanical injury and further promote cell migration.In addition,MGF can reduce the activity of MM P-5 and alleviate the stretch-induced deformation of OA-LFs.Furthermore,the mRNA expression of ATF-2 up-regulated in a dose-dependent manner upon MGF treatment compared with control,while the expression of ATF-5 gene was down-regulated in a dose-dependent.Protein levels showed that the expression of p-ATF-2 increased with increasing MGF concentration.Conclusions Our study shows that MGF pretreatment of OA-LFs can respond quickly to mechanical damage and accelerate the ligament injury repair by promoting cell migration,decreasing the MMP-2 activity,and remitting the cell deformation.Therefore,MGF has potential as a therapeutic for OA patients.
