Liver fibrosis is a dynamic wound-healing response characterized by the agglutination of the extracellular matrix (ECM). Si-Wu-Tang (SWT), a traditional Chinese medicine (TCM) formula, is known for treating gynecological diseases and liver fibrosis. Our previous studies demonstrated that long non-coding RNA H19 (H19) was markedly upregulated in fibrotic livers while its deficiency markedly reversed fibrogenesis. However, the mechanisms by which SWT influences H19 remain unclear. Thus, we established a bile duct ligation (BDL)-induced liver fibrosis model to evaluate the hepatoprotective effects of SWT on various cells in the liver. Our results showed that SWT markedly improved ECM deposition and bile duct reactions in the liver. Notably, SWT relieved liver fibrosis by regulating the transcription of genes involved in the cytoskeleton remodeling, primarily in hepatic stellate cells (HSCs), and influencing cytoskeleton-related angiogenesis and hepatocellular injury. This modulation collectively led to reduced ECM deposition. Through extensive bioinformatics analyses, we determined that H19 acted as a miRNA sponge and mainly inhibited miR-200, miR-211, and let7b, thereby regulating the above cellular regulatory pathways. Meanwhile, SWT reversed H19-related miRNAs and signaling pathways, diminishing ECM deposition and liver fibrosis. However, these protective effects of SWT were diminished with the overexpression of H19 in vivo. In conclusion, our study elucidates the underlying mechanisms of SWT from the perspective of H19-related signal networks and proposes a potential SWT-based therapeutic strategy for the treatment of liver fibrosis.
Humans ; RNA, Long Noncoding/genetics* ; Liver Cirrhosis/genetics* ; Liver/metabolism* ; Hepatic Stellate Cells/pathology* ; MicroRNAs/metabolism* ; Extracellular Matrix/metabolism* ; Drugs, Chinese Herbal

ObjectiveTo investigate the protective effect and regulatory mechanism of berberine （BBR） against the senescence of ovarian granulosa cells. MethodA cell senescence model in the human ovarian granulosa-like tumor （KGN） cell line was induced by H₂O₂. A control group， a model group， and high-dose （1 μmol·L^-1） and low-dose （0.5 μmol·L^-1） BBR groups were set up. The cells in the model group and the BBR groups were incubated with 10 μmol·L^-1 H₂O₂ for 40 min. The effect of BBR on KGN cell proliferation was detected by cell counting kit-8 （CCK-8） assay. The effect of BBR on the senescence of KGN cells was detected by β-galactosidase staining. The effects of BBR on the apoptosis and ROS content of KGN cells were detected by flow cytometry. The effects of BBR on the mRNA expression of B-cell lymphoma-2 （Bcl-2）/Bcl-2-associated X protein （Bax）， cysteinyl aspartate-specific protease-3 （Caspase-3）， forkhead transcription factor O1 （FoxO1）， and catalase （CAT） was detected by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. Western blot was used to detect the effects of BBR on protein expression of silent information regulator1 （SIRT1）， superoxide dismutase 2 （SOD2）， c-Jun N-terminal kinase （JNK）， FoxO1， autophagy-associated protein microtubule-associated protein light chain 3Ⅱ （LC3BⅡ）， mammalian ortholog of yeast Atg6 （Beclin-1）， and ubiquitin-binding protein p62. ResultAfter H₂O₂ induction for 40 min， the cell proliferation rate of the model group decreased compared with that of the control group （P<0.01）， and the cell proliferation rates of the BBR groups increased compared with that of the model group （P<0.05）. The results of β-galactosidase staining showed that the cells of the model group showed significant senescence compared with those of the control group （P<0.01）， and the cellular senescence in the BBR groups was reduced compared with that of the model group （P<0.01）. As revealed by flow cytometry， compared with the control group， the model group showed increased apoptosis rate （P<0.01）， and compared with the model group， BBR groups showed decreased apoptosis rates （P<0.05）. Meanwhile， the ROS content in the model group increased compared with that in the control group （P<0.01）， and compared with the model group， the BBR groups showed reduced cellular ROS content （P<0.01）. The Real-time PCR results showed that compared with the control group， the model group showed decreased mRNA expression of CAT and Bcl-2/Bax in KGN cells and increased mRNA expression of Caspase-3 and FoxO1 （P<0.05）， and compared with the model group， the BBR groups showed increased mRNA expression of CAT and Bcl-2/Bax （P<0.05） and reduced mRNA expression of Caspase-3 and FoxO1 in KGN cells （P<0.05）. As revealed by Western blot results， SIRT1， SOD2， and p62 protein levels decreased in the model group compared with those in the control group （P<0.01）， and JNK FoxO1， LC3BⅡ， and Beclin-1 protein levels increased （P<0.05）. After BBR intervention， SIRT1， SOD2， and p62 protein levels increased （P<0.01）， and JNK， FoxO1， LC3BⅡ， and Beclin-1 protein levels decreased compared with those in the model group （P<0.05）. ConclusionBBR has an inhibitory effect on ovarian granulosa cell senescence， and the mechanism is related to the inhibition of apoptosis and autophagy mediated by the SIRT1/FoxO1 pathway.

This study was aimed to investigate the effects of Si-Wu mixture on the secretion of E2 and the expression of CYP19al gene in granulosa cells after cell injury.Ovarian granulosa cells of SD rats were treated with cisplatin (CDDP).And the expression levels of E2 and CYP19a1 were determined by different testing methods.The results showed that the level of E2 induced by radioimmunoassay of the Si-Wu group was significantly higher than that of the CDDP group.Meanwhile,the group which added TGF-β3 protein pathway blocker was lower than others.The results of immunohistochemistry and western blotting showed that the expression level of CYP 19a1 of the Si-Wu group was higher than that of the CDDP group.In the western blotting,the group which added blocker was significantly lower than the non-blocking group.It was concluded that the pharmacological serum of Si-Wu mixture can enhance the level of E2 in CDDP cells through TGF-β3 protein pathway.And the effect is accomplished by the intervention of CYP 19a1.

ObjectiveTo study the effects of tanshinoneⅡA on proliferation of cervical squamous cancer Siha cells; To discuss its possible molecular mechanism.Methods Cervical squamous cancer Siha cells were treated with different doses of tanshinoneⅡA. The effects of tanshinoneⅡA on proliferation of Siha cells were measured by MTT assay and flow cytometry analysis. The effects of tanshinoneⅡA on expression levels of phospho-extracellular regulate kinase (p-ERK) and Cyclin D in Siha cells were measured by Western blot.Results 1×10-5, 5×10-6, 1×10-6, 5×10-7 mol/L tanshinoneⅡA significantly inhibited Siha cell proliferation and such effect could be enhanced by ERα antagonist MPP and attenuated by ERβ antagonist PHTPP. 1×10-5, 5×10-6, 1×10-6 tanshinoneⅡA could significantly decrease the proliferation index of Siha cells. 1×10-5, 5×10-6, 1×10-6, 5×10-7 mol/L tanshinoneⅡA could significantly reduce the protein expression levels of p-ERK and Cyclin D of Siha cells.ConclusionTanshinoneⅡA can inhibit cervical squamous cancer Siha cell proliferation and such effect is realized via estrogen receptor pathway. TanshinoneⅡA plays anti-proliferation roles by reducing the expression levels of p-ERK and Cyclin D.

This study was aimed to observe the influence of β-sitosterol (BSS) on estrogen receptor (ER) positive the human breast cancer cell line T47D and to study its mechanisms. ER antagonist ICI182 780 was employed to observe the influence on the proliferation. Proliferations of T47D cells influenced by different concentrations of BSS were analyzed by MTT assay. Cell cycle analyses were examined by flow cytometry. The protein expression of cyclin D1 was measured by western blot analysis and cyclin D1 mRNA was quantified by real-time PCR assay. The results showed that BSS in high dose exhibited significant inhibitory effects that were partly antagonized by ICI182 780 and decreased the proliferative index on T47D cells. However, BSS in low dose obviously promoted the proliferation that was completely inhibited by ICI182 780 and increased the proliferative index on T47D cells. The mRNA and protein levels of cyclin D1 were increased in low-dose BSS. The effect was blocked by ICI182 780. It was concluded that BSS in low concentration had phytoestrogenic effect by up-regulating the expression of cyclin D1 via ER pathway.

This study was aimed to investigate effects of Er-Xian (EX) decoction in follicular granulosa cell apoptosis in chemotherapy-induced premature ovarian failure (POF) of rats. SD rats were randomly divided into the normal group, model group, positive group and EX decoction group. Intraperitoneal injection of cisplatin was used in the establishment of chemotherapy-induced POF rat model. Intragastric administration of corresponding medication was give to each group for four weeks. The general conditions of rats were observed. The serum contents of E2, FSH, LH and Pro were determined. The ovarian tissue morphology was observed. Granulosa cell apoptosis was analyzed by TUNEL. The expressions of BAX, Bcl-2 and VEGF protein were detected by immunohistochemistry. The results showed that in the model group, the body weight decreased with disordered estrous cycle. After treatment, the weight gained, and the estrous cycle recovered. Compared with the normal group, in the model group, E2 decreased, FSH and LH levels increased. There was no significant difference in the content of progesterone among different groups. Detection of TUNEL showed that the positive expression in the model group was increased compared with the normal group (P < 0.05). The positive expression of EX decoction group was reduced compared to the positive group (P < 0.05). Compared with the normal group, the expression of BAX was obviously increased and Bcl-2 was decreased in the model group. After EX decoction treatment, the expression of BAX was decreased and the Bcl-2 was increased compared to the model group. The expression of VEGF protein in the model group was obviously less than that in the normal group. It was concluded that cisplatin can induce follicular granulosa cell apoptosis, and cause premature ovarian function recession. EX decoction can adjust the content of different hormones in serum, alter expression of apoptosis related protein to reduce the damage of cisplatin on ovarian follicular development, in order to promote follicular development, improve and enhance the ovarian function.

Aim To explore the effects of tanshinone IIA on cell proliferation via G protein-coupled estrogen receptor inductive and regulative pathway in typical es-trogen receptor and G protein-coupled estrogen receptor positive T47D breast cancer cells. Methods The pro-liferation rate of T47 D cells influenced by tanshinone IIA was analyzed by MTT assay. G protein-coupled es-trogen receptor agonist G1 and GPER antagonist G15 were employed as tools. GPER SiRNA was applied to build GPER gene silence T47D cells. GPER expres-sion influenced by tanshinone IIA was measured by Western blot. Results The proliferation rates of T47D cells treated with 1 × 10 -5 mol·L-1 - 1 × 10 -7 mol· L-1 of tanshinone IIA were decreased significantly. Such effects could be attenuated by G1 or enhanced by G15 . Growth of GPER SiRNA transfected T47 D cells were significantly inhibited by 1 × 10 -5 mol·L-1 - 1 × 10 -7 mol·L-1 of tanshinone IIA treating. Result of Western blot showed that tanshinone IIA at 1 × 10 -5 mol· L-1 and 1 × 10 -6 mol · L-1 could induce de-crease of GPER protein expression in T47D cells. Conclusions Tanshinone IIA shows inhibitory effects on proliferation rate of T47 D breast cancer cells via GPER pathway. Tanshinone IIA could perform regula-tive function on GPER expression level in target cells.

Aim To isolate and characterize the human circulating fibrocytes from human peripheral blood and explore the effects of curcumin on human circulating fi-brocytes.Methods The cells were isolated and puri-fied by density gradient centrifugation,and identified by flow cytometry and immunocytochemistry .Then , CCK-8 and flow cytometry were used to study the effect of curcumin on the proliferation as well as COL I ex-pression of human circulating fibrocytes,respectively. Results After being isolated the cells expressed CD34,CD45 and COLⅠ,among which 79.7% were both CD45 and collagen I positive,typical of human circulating fibrocytes.Curcumin could exert regulatory effects on proliferation of human circulating fibrocytes. Exposure of the cells to curcumin for as short as 24 hours promoted their growth,while prolonged treatment (72 h ) significantly inhibited cell propagation and downregulated the COLⅠ levels,best manifested at a concentration as high as 20 μmol · L-1 .Conclusion The proliferation of cells and COLⅠexpressions can be effectively inhibited by curcumin with the prolonged action period and high concentrations.

Postmenopausal osteoporosis is one of the commonest systemic bone metabolism diseases among menopausal women, mainly caused by lowering internal estrogen. Although Hormone Replacement Therapy (HRT) is an effective method in clinical practice for years, it shows side-effect in increasing gynecological carcinoma. It has already been proved by clinical tests that multiple traditional Chinese medicine formulas and their monomer ingredients and phytoestrogen-like active constituents contained in traditional Chinese medicines are effective on treating osteoporosis with relatively less side-effects comparing with HRT. They show protective and therapeutic effects by acting on estrogen receptors of targeted tissues and targeted cells and then affecting expressions of bone metabolism-related regulatory proteins and factors in downstream signal conduct paths. Recent studies on estrogen related receptor (ERR) provide new possibilities and pathways for mechanism of traditional Chinese medicine and their active constituents in osteoporosis.
Chemistry, Pharmaceutical ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; therapeutic use ; Humans ; Medicine, Chinese Traditional ; methods ; Osteoporosis, Postmenopausal ; drug therapy ; Receptors, Estrogen ; metabolism

OBJECTIVEResearch on the phytoestrogenic effect and its possible mechanism of formononetin.

METHODTo evaluate the estrogenic effect and mechanisms of formononetin through the test of its influence on proliferation and ER subtype expression of T47D cells.

RESULTThe proliferation rates of T47D cells treated with 1 x 10(-7) -1 x 10(-6) mol x L(-1) formononetin were not increased. On the influence of ICI182, 780, the proliferation rates of T47D cells treated with 1 x 10(-7) 1 x 10(-6) mol x L(-1) formononetin were decreased. Formonenetin could induce the augment of ERalpha expression significantly of T47D.

CONCLUSIONFormonenetin has phytoestrogenic effect Formonenetin can not accelerate ER(+) T47D cell proliferation. But the expression level of ERalpha subtype in T47D cells change significantly with certain concentrations of formonenetin.

Breast Neoplasms ; drug therapy ; genetics ; metabolism ; physiopathology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Female ; Humans ; Isoflavones ; pharmacology ; Phytoestrogens ; pharmacology ; Receptors, Estrogen ; genetics ; metabolism