Exploring the effect of Bushen Bitong recipe-containing serum on IL-1β-induced chondrocyte apoptosis based on SOX9/NF-κB/MMP-13 signaling pathway

Objective:To observe the effect and possible mechanism of action of Bushen Bitong recipe(BSBT)containing serum on IL-1β-induced chondrocyte apoptosis.Methods:Generation 3 rat chondrocytes were randomized into Control,IL-1β,IL-1β+BSBT(L),IL-1β+BSBT(M),and IL-1β+BSBT(H)groups(5%,10%and 15%BSBT-containing serum),and then 24h after intervention respectively,the cell proliferation and Apoptosis rate;Western blot detected the expression levels of Bcl-2,BAX,Caspase-3,SOX9,NF-κB p65,MMP-13 proteins in chondrocytes.ELISA detected the levels of TNF-α,IL-6,and bFGF in the supernatants of chondrocyte culture.Results:Compared with Control group,cell proliferation activity decreased,apoptosis rate increased,NF-κB p65,MMP-13 protein level and TNF-α,IL-6 level increased,and SOX9 protein level and bFGF level decreased in IL-1βgroup;compared with IL-1βgroup,different concentrations of BSBT-containing serum group,cell proliferation activity increased,and apoptosis rate decreased.NF-κB p65,MMP-13 protein level and TNF-α,IL-6 level decreased,SOX9 protein level and bFGF level increased;compared with IL-1β+BSBT(L)group,cell proliferation activity increased,apoptosis rate decreased in IL-1β+BSBT(M)and IL-1β+BSBT(H)groups,and NF-κB p65,MMP-13 protein level and TNF-αlevel decreased.13 protein levels and TNF-αand IL-6 levels decreased,and SOX9 protein levels and bFGF levels increased.Conclusion:BSBT-containing serum may promote IL-1β-induced proliferation of chondrocytes,reduce apoptosis,improve the microenvironment of chondrocytes,and promote cartilage repair through the SOX9/NF-κB/MMP-13 signaling pathway.

Effect of Bushen Yiqi Huoxue Recipe on Placental Vasculature in Pregnant Rats with Fetal Growth Restriction Induced by Passive Smoking

Interactions of vascular endothelial growth factor （VEGF） with receptors VEGFR1/Fltl and VEGFR2/Flk1, and those of angiopoietins （Ang-1, Ang-2） with receptor Tie2 play important roles in placental angiogenesis. This study investigated vascular morphology and expression of these angiogenic factors in rat placenta on the day 15, 18, 21 of gestation （D 15, D 18 and D21）. The rats were randomly assigned into 3 groups： normal group, model group [fetal growth restriction （FGR） model], and Bushen Tqi Huoxue （BYHR） recipe treatment group （BYHR group, the pregnant rats with FGR were treated with BYHR recipe）. Morphological analysis indicated that during initial villous formation, fetal nucle- ated erythrocytes （FNEs） appeared in maternal blood sinus （MBS）. Subsequently, FNEs were sur- rounded by endothelial cells to form fetal capillary （FC） and then by trophoblast cells to form villi. As pregnancy proceeded, FC density increased progressively with increasing endothelial identification staining （EIS） in normal and BYHR groups. Whereas, villous formation was suppressed, normal in- crease in FC density was impaired and EIS was weakened in model group. Quantitative PCR analysis showed that VEGF and Flkl mRNA increased over gestation in all groups, indicating that VEGF might play a pivotal role in FC growth during late gestation. VEGF mRNA was increased on D15, while de- creased on D21 in model group as compared with normal group and BYHR group. Immunohistochemi- cally, Ang-2 protein was highly expressed in FNEs, gradually disappeared as villi matured, and decreased over gestation in all groups, indicating that Ang-2 might play a pivotal role in villous formation, which was further supported by decreased Ang-2 mRNA and protein expression in model group on D 15. Ang-1 mRNA, Tie2 mRNA and Ang-1/Ang-2 ratio increased from D15 to D18 in all groups as placenta matured. Ang-1 mRNA, Tie2 mRNA and Ang-1/Ang-2 ratio were decreased on D18 in model group as compared with normal and BYHR groups, indicating delayed maturity of FGR placenta. Alterations in angiogenic factors may result in altered placental vasculature and cause placental insufficiency. BYHR recipe could balance the angiogenic factors to promote the formation and maturation of FGR placental vasculature.

Effect of Bushen Yiqi Hexue Recipe (补肾益气和血方) on Pinopodes Expression on Endometrial Surface in Embryo Implantation Dysfunctional Mice

Objective: To explore the therapeutic mechanism of Bushen Yiqi Hexue recipe (补肾益气血方, BYHR), and observe its effect on pinopodes expression on endometrial surface in mifepristone induced embryo implantation dysfunctional mice. Methods: Pregnant Kunming mice were randomly divided into the normal group, the control group and the treated group. Mice in the treated group were administered with BYHR, and those in the normal group and the control group were administered with normal saline, starting from the very first day of pregnancy (Pd1). On the Pd4, mifepristone (RU486) was subcutaneously injected into mice in the control group and the treated group. Pinopodes expression on endometrial surface at 09:30-10:00 pm on Pd4 (regarded as time point 1, T1) and at 09:30-10:00 am on Pd5 (as time point 2, T2) was determined by scanning electron microscopy. Pregnancy rate and average implanted embryos were observed on Pd7. Results: Pregnancy rate and average implanted embryos in the control group were obviously lower than those in the normal group (all P<0.01), while the two parameters were significantly higher in the treated group (P<0.05, P<0.01, respectively) than those in the control group. In the normal group, abundant developing pinopodes were distributed over the whole endometrial surface in T1, and they were altered to a great deal of fully developed pinopodes in T2. But in the control group, only a few pinopodes were expressed locally on endometrial surface in T1, showing a nonsynchronous figure of development, and they completely disappeared in T2. In the treated group, a lot of developing pinopodes were expressed like those in the normal group but somewhat lagged behind, whereas many fully developed pinopodes were expressed in T2. Conclusion: The decrease of the pinopodes in T1 and the cleaning up of them in T2 is possibly the partial mechanism of mifepristone in inducing embryo implantation dysfunction. It is indicated that BYHR could improve pinopodes expression on endometrial surface, eventually better the uterine receptivity and improve the embryo implantation.

Antitumor and synergistic effect of Chinese medicine “Bushen huayu jiedu recipe” and chemotherapy on transplanted animal hepatocarcinoma

AIM: To investigate the antitumor and synergistic effect of Chinese medicine 'Bushen huayu jiedu recipe' (recipe for invigorating the kidney, removing blood stasis and toxic substances) and chemotherapy on mice hepatocarcinoma.METHODS: Bushen huayu jiedu recipe (BSHYJDR)consisting of Chinese Cassia Bark, Psoralea, Zedoary,Rhubarb, etc. is equal to 1.5 g/mL liquid of originated herbs after being decocted, filtered, and concentrated. Kunming mice, weighing 18-22 g, were injected with 0.2 mL ascitic hepatocarcinoma H22 containing 1×107 cells/mL into armpit of the right forelimb of mice. After 24 h, the mice were weighed and randomly divided into tumor-bearing model control group, cisplatin (DDP) group, BSHYJDR high dosage group, low dosage BSHYJDR group, DDP combined with high and low dosage BSHYJDR group, 10 mice in each group. DDP group received injection intraperitoneally (ip) at the dosage of 1 mg/kg (equal to 1/10 LD50), once a day for 4 d. High and low dosage BSHYJDR groups received intragastric BSHYJDR at the dosages of 26.6 and 13.3 g/kg (20 and 10 times each of clinical adult dosage)respectively, while tumor-bearing model group received the equal volume of distilled water once a day for 10 d. On the 11th d, the mice were weighed and killed, then the tumor was dissected and weighed, the repression rate (RR) was calculated according to the mean weight of tumor (MWT).RESULTS: Compared to the model group (MWT: 1.30±0.73),DDP group (MWT: 0.41+0.09, RR: 68.46%) had a significant difference in the inhibition of hepatocarcinoma H22 (P＜0.01).High dosage BSHYJDR group (MWT: 0.69+0.29, RR:46.92%) also had a significant difference in inhibition (P＜0.05), while no difference was found in low dosage BSHYJDR group (MWT: 0.85+0.34, RR: 34.62%) (P＞0.05).When DDP was combined with high dosage BSHYJDR (MWT: 0.29±0.17, RR: 77.69%) and low dosage BSHYJDR (MWT: 0.38±0.21, RR: 70.77%) respectively, we could see improvement of the inhibition effect of DDP on transplanted hepatocarcinoma H22. DDP combined with high dosage BSHYJDR had a significant difference (P＜0.001) compared to DDP, while DDP combined with low dosage BSHYJDR only had a little improvement that is not remarkable.CONCLUSION: Chinese medicine BSHYJDR in combination with chemotherapy can inhibit transplanted hepatocarcinoma in mice.

Jianpi Bushen Recipe affects epithelial-mesenchymal transition in hepatocellular carcinoma through TGF-β and Twist

Backgroud:To explore the effects of Jianpi Bushen Recipe on the epithelial-mesenchymal transition(EMT)of nude mice with transplanted human liver cancer SK-HEP-1 cells.Methods:To establish models with transplanted liver cancer cells,nude mice were divided randomly into three groups:blank group,traditional Chinese medicine(TCM)group and control group.They were given normal saline,Jianpi Bushen Recipe and solafeni through administration by gavage for four weeks.The expression of TGF-β and twist mRNA were detected by real-time PCR.The expression of E-cadherin was detected by Western blot.Results:Compared with the blank group,the expression of transforming growth factor-P(TGF-β)mRNA and twist mRNA in the TCM group and the control group decreased significantly(P<0.05).There was no significant difference between the two groups(P>0.05).The expression of E-cadherin protein in TCM group and control group increased significantly(P<0.01).There was no significant difference between the two groups(P>0.05).Conclusion:Jianpi Bushen Recipe can inhibit the EMT of liver cancer SK-HEP-1 cells in vivo by down-regulating the expression of TGF-β/twist signaling and indirectly up-regulating E-cadherin expression in transplanted liver cancer tissue,indicating the potential of Jianpi Bushen Recipe to inhibit invasion and metastasis of liver cancer.

A Meta-analysis on the Treatment of Insulin Resistance in Polycystic Ovary Syndrome by Bushen Huatan Recipe

Objective:To systematically evaluate the efficacy and safety of Bushen Huatan recipe compared with metformin in the treatment of insulin resistance in polycystic ovary syndrome(PCOS).Methods Researches were reviewed by searching the China National Knowledge Infrastructure(CNKI),VIP network,WanFang database,SinoMed,Cochrane Library,Web of Science,PubMed and Embase up to December 8,2019.We retrieved the RCTs of Bushen Huatan recipe and metformin in the treatment of insulin resistance in polycystic ovary syndrome and applied Revman5.3 software for meta analysis.Results A total of 5 RCTs were included,with a total of 285 PCOS patients.The results showed:①There was no significant difference between the two groups in LH[WMD=-2.68,95%CI(-6.21,0.84),P=0.14]、FSH(WMD=0.36,95%CI(-0.59,1.32),P=0.45]、LH/FSH(WMD=-0.29,95%CI(-1.06,0.48),P=0.46)、fasting blood glucose(FPG)[WMD=0.05,95%CI(-0.18,0.28),P=0.67]、fasting insulin(FINS)[WMD=-0.91,95%CI(-15.28,13.46),P=0.9].②TCM syndrome score:Bushen Huatan recipe group[WMD=-3.10,95%CI(-5.39,-0.81),P=0.008]was better than metformin group;③Total clinical effective rate:Bushen Huatan recipe group[RR=1.50,95%CI(1.18,1.92),P=0.001]was better than metformin group.Conclusion Bushen Huatan recipe has the same effect as metformin in improving the sex hormones and glucose metabolism of patients with PCOS complicated with insulin resistance,and can effectively improve the clinical symptoms of patients,reduce TCM syndrome score,and improve the total therapeutic efficiency.

Efficacy of Jianpi Bushen Huatan Recipe in the treatment of obese polycystic ovarian syndrome:A meta-analysis

Objective:To evaluate the effects of Jianpi Bushen Huatan Recipe on sex hormone levels、blood lipid and blood glucose levels in the treatment of obese poly-cystic ovarian syndrome(PCOS).Methods:Data bases including the CNKI、Vip、Wanfang were searched.Studies about randomized controlled trials(RCT)of the effectiveness of Jianpi Bushen Huatan Recipe Combined with Metformin in the Treatment of obese PCOS were included.Rev Man 5.3 software was used for meta analysis.Results:A total of 11 studies were enrolled.Meta-analysis showed that Jianpi Bushen Huatan Recipe Combined with Metformin significantly improved the clinical total effective rate of patien ts(RR=1.32,95%CI(1.20,1.45),P<0.00001);reduced the body mass index(BMI)(SMD=-0.92,95%CI(-1.13,-0.72),P<0.00001),luteinizing hormone(LH)(SMD=-1.00,95%CI(-1.37,-0.62),P<0.00001),LH/FSH(SMD=-1.03,95%CI(-1.34,-0.71),P<0.00001),insulin resistant index(HOMA-IR)(SMD=-0.59,95%CI(-0.79,-0.40),P<0.00001),fasting insulin(FIN)(SMD=-0.77,95%CI(-1.19,-0.35),P=0.0003),total cholesterol(TC)(SMD=-0.63,95%CI(-0.88,-0.38),P<0.00001),triglycerides(TG)(SMD=-0.60,95%CI(-0.83,-0.36),P<0.00001),highdensity lipoprotein(HDL)(SMD=-0.41,95%CI(-0.51,-0.32),P<0.00001).However,there was no statistically significant in waist-to-hip ratio(WHR),follicle stimulating hormone(FSH),fasting blood sugar(FBS)and low density lipoprotein(LDL).Conclusions:Jianpi Bushen Huatan Recipe Combined with Metformin in the treatment of obese PCOS has an advantage in improving the sex hormone level,blood lipid and blood glucose level,and is superior to metformin alone.

A meta-analysis on the improvement of endometrial receptivity of PCOS by Bushen recipe combined with Clomiphene

Objective: To systematically evaluate the efficacy and safety of Bushen Recipe combined with clomiphene compared with clomiphene in improving endometrial receptivity of polycystic ovary syndrome. Methods: The time limit is from the establishment of the database to January 2, 2020. A randomized controlled trial was conducted to compare the effects of Bushen Recipe combined with clomiphene and clomiphene on the endometrial receptivity of polycystic ovary syndrome (PCOS). Meta analysis was performed with RevMan5.3 software. Results: A total of 13 RCTS were included, with a total of 797 PCOS patients. The results of this meta-analysis show: there were statistically significant differences in the improvement of endometrial thickness [WMD=1.62,95%CI(1.22, 2.01), P <0.00001], endometrial type[RR=1.51,95%CI(1.28, 1.78), P <0.00001], uterine artery pulse index(PI) [WMD=-0.18,95%CI(-0.27, -0.09) , P <0.0001], uterine artery resistance index(RI)[WMD=-0.08,95%CI(-0.11, -0.05) , P <0.00001], pregnancy rate [RR=2.18,95%CI(1.55, 3.05), P <0.00001], TCM syndrome score[WMD=-6.97,95%CI(-8.97, -4.97) , P <0.00001], effective rate [RR=1.25, 95%CI (1.13, 1.37), P < 0.00001]. Conclusion: Compared with clomiphene alone, Bushen Recipe combined with clomiphene can effectively increase the thickness of endometrium and improve the type of endometrium, promote the blood perfusion of endometrium, improve the pregnancy rate, improve the clinical symptoms, reduce the TCM syndrome score, improve the total efficiency of treatment, with high clinical application value.

Effect of Jiangtang Bushen Recipe (降糖补肾方) in Intervention Treatment of Patients with Impaired Glucose Tolerance

Objective:To evaluate the intervention effect of diet, exercise and Jiangtang Bushen Recipe (JBR, 降糖补肾方), a Chinese herbal recipe, in preventing the progress of patients with impaired glucose tolerance ( IGT ) to diabetes mellitus (DM) type 2. Methods: Fifty-one IGT patients with their diagnosis conformed to the diagnosis standard of WHO, 1999, were randomly divided into the control group (n = 26) and the TCM group (n=25). Patients in the control group attended to the educational course for DM and received dietotherapy and kinetotherapy, and to those in the TCM group, under these treatments, JBR was given additionally. Oral glucose tolerance test (OGTT), body weight index (BWI), levels of blood lipids and fasting insulin of all the patients were examined after 3 months, 6 months and 12 months of treatment. The total observation time was 1 year. Results:Except the 6 cases out of the 51 patients (11.7%), on whom the observa-tion discontinued, in the control group, as compared with before treatment, levels of fasting insulin and fasting blood glucose after treatment were not changed significantly ( P > 0. 05 ) , also insignificant difference was shown in levels of total cholesterol (TC) and triglyceride (TG), thoughthe two indexes lowered slightly after treatment (P>0. 05), but significant difference was shown in comparison of OGTT/2h, blood glucose and BWI (P<0. 05). While in the TCM group, fasting blood glucose was changed insignificantly (P> 0. 05) , but there was significant difference in comparison of fasting insulin, TC, BWI, OGTT/2h and plasma glucose levels (P<0. 01) respectively before and after treatment. At the end of the stud-y, the cumulative cases with conversion to diabetes were 3 (13. 6%) in the control group, and 1 (4. 3%) in the TCM group, x2 test showed insignificant difference in comparison of diabetes conversion rate between the two groups (P>0. 05), however, the TCM group showed a better year conversion rate of normal glucose tolerance than that in the control group (x2 = 8. 31, P<0. 01). Conclusion: TCM intervention is possibly effective in delaying the conversion of IGT to DM type 2, and plays integrative effeciency in impelling IGT patients to health. The favorable education and treatment of DM controlling, including dieto- and kineto-therapy may also be advantageous in IGT intervention, but could not be effective in blocking the advance of IGT.Original article on CJITWM (Chin) 2004 ;24 (4): 317

Effect of bushen tiaopi recipe on sex hormones, immune function and oxidative stress in patients with climacteric syndrome

Objective: To explore the effect of Bushen Tiaopi Recipe on sex hormones, immune function and oxidative stress in patients with climacteric syndrome and guide clinical medication. Methods: A total of 140 cases of climacteric syndrome patients who were treated in our hospital from May 2016 to May 2017 were selected according to the lottery method, 70 cases in each group. Control group was given Li Fumin treatment, and the observation group was given Li Fumin combined with Bushen Tiaopi Recipe. The levels of sex hormones, T lymphocyte subsets, leptin, adiponectin and oxidative stress were measured before and after treatment in both groups. Result: There was no significant difference in the levels of sex hormones, T lymphocyte subsets, leptin, adiponectin and oxidative stress before treatment between the two groups. After 12 weeks of drug treatment, the level of sex hormone P increased and the levels of LH and FSH decreased in both groups. And the research group P was (21.89±2.29) nmol/L higher than the control group, and the LH and FSH were (4.28±0.43) IU/L and (4.91±0.50) IU/L were lower than the control group. The LP level was decreased and the APN level increased in both groups. Moreover LP level in the research group was (15.34±1.52) μg/L lower than that in the control group, and the APN level was (8.39±0.84) mg/L higher than that in the control group. The SOD level was increased and MDA level decreased in both groups. And SOD level in the research group was (33.27±3.29) U/L, which was higher than that in the control group, and the MDA level was (12.47±1.25) nmol/mL, which was lower than that in the control group. CD4+and CD4+/CD8+levels were significantly elevated in both groups, and CD8+levels were significantly decreased. In the research group, CD4+and CD4+/CD8+were (28.87±2.69)% and (1.41±0.14), respectively, higher than the control group, and the number of CD8+cells was (21.05±2.23)% lower than the control group. Conclusion:Bushen Tiaopi can significantly improve sex hormone levels in climacteric syndrome patients, improve immune function, relieve oxidative stress, and improve treatment effects.

Effect of Combination of Bushen Jianpi Recipe (补肾健脾方) and Erythropoietin on Serum Tumor Necrosis Factor a in Patients with Anemia

Objective: To study the effect of treatment of renal anemia by combination of Bushen Jianpi Recipe (补贤健脾方, BJR, a Chinese experienced recipe for supplementing Shen and supporting Pi) and low dosage erythropoietin (EPO), and the influence of treatment on change of serum tumor necrosis factor α (TNF-α) so as to explore the possible mechanism of integrative Chinese and Western medicine (ICWM) in treating renal anemia. Methods: Patients with renal anemia were randomly divided into two groups, the ICWM group and the control group, supporting treatment such as dialysis, supplementing of ferrous, foliac acid and vitamin B12, was given to both groups. Additionally, to the ICWM group, 50 IU/kg of EPO subcutaneous injection for twice every week, and oral intake of BJR, one dose per day taken in two parts, were given, and to the control group, EPO alone, 50IU/kg by subcutaneous injecting, 3 times perweek, was given. The therapeutic course for two groups was 3 months. Blood levels of hemoglobin (Hb), hematocrit (Hct), TNF-α were measured before treatment and the therapeutic effect was observed. Results: After treatment, the levels of Hb and Hct increased significantly in both groups (P<0. 01) , comparison between the two groups in Hb and Hct after treatment for 3 months showed significant difference (P<0. 05). The serum level of TNF-α was markedly higher than normal range in both groups before treatment, it significantly lowered after treatment in the ICWM group (P< 0.05), but unchanged in the control group. Conclusion: Combination of BJR and EPO could significantly inhibit the production of TNF-α, this may be an important factor for ICWM in effectively improving sensitivity to EPO.Original article on CJITWM (Chin) 2004;24 (2): 106

Effective ingredients, potential targets and mechanism in cancers treatment of Bushen Jiedu Sanjie recipe

Objective: To analyze the active compounds, potential drug targets and corresponding therapy cancer of Bushen Jiedu Sanjie Recipe (BSJDSJR) based on network pharmacology and bioinformatics technology. Methods: The network databases including Cancer HSP, TCMSP, TCMID, TCM-PTD, TCM Database@Taiwan and DrugBank were applied to screen the active compounds, potential drug targets and corresponding cancers of BSJDSJR. Cytoscape3.3 software was used to construct the network between active compounds of Chinese medicine and the corresponding targets. Then, the enrichment of biological processes and KEGG pathways of BSJDSJR were analyzed using DAVID database. Results: According to Oral bioavailability (OB)≥30% and drug like index (DL)≥0.18, 129 active compounds in BSJDSJR were screened out and they targeted to 301 proteins. These targets were closely associated with the occurrence of various cancers, such as bladder cancer, pancreatic cancer, non-small cell lung cancer and colorectal cancer. Further investigation showed that, there were lots of active compounds in BSJDSJR are closely connected with the COX-2/β-catenin signaling pathway, STAT3 signaling pathway and MAPK/ERK signaling pathway. Conclusions: Based on the network pharmacology, the study disclosed the active chemical compounds, potential targets and possible action cancers of BSJDSJR.

Mechanism of treating recurrent abortion with “Peiyuan Bushen Antai formula” based on network pharmacology and molecular docking

Objective:To study the effective compound action target signal pathway of Peiyuan bushenan taifang(PYBSATF)has achieved good clinical efficacy in the treatment of recurrent spontaneous abortion(RSA),but its mechanism has not been clarified because of its complex components.In this study,network pharmacology is applied to study the effective compounds,targets and signal pathways of PYBSATF in the treatment of RSA,so as to reveal its pharmacological mechanism of action in the treatment of recurrent spontaneous abortion.Methods:The Traditional Chinese Medicine Systems Pharmacology database(TCMSP)and CNKI are used to obtain the main compounds and drug action targets of PYBSATF.Genecards and the Online Mendelian Inheritance of Man databases(OMIM)are searched to collect the known genes related to RSA,so as to construct compound-target network and screen out the common target proteins and main active compounds.We also use string database to construct a visual protein-protein interaction network(PPI).Cluster Profiler and R software were used to analyze the common targets of drugs and diseases for GO function analysis and KEGG pathway enrichment analysis.Finally,the compound and the protein sequences were conducted according to the node parameters,so that the core protein and core compounds are used to perform molecular docking.Results:186 potential active components and 65 predicted action targets of PYBSATF were screened out.At the same time,1658 genes were also screened out to be closely related to RSA,among which 65genes overlaped with PYBSATF targets and were considered to be related to way of treatment.PPI network showed that VEGFA,IL6,EGFR,MAPK8 and ESR1were the core targets of PYBSATF for the treatment of RSA.GO and KEGG enrichment analysis obtained 93 biological processes of cells(P<0.01)and 87 signaling pathways(P<0.01).PYBSATF played a pharmacological role through a variety of pathways including anti-inflammatory,anti-apoptosis,promoting proliferation and angiogenesis.Molecular docking showed that most active components and key targets of PYBSATF had strong efficiency.Conclusion:Through the study of network pharmacology,it predicted that PYBSATF might treat RSA through multiple targets and multiple signal pathways.Significantly,the predictive targets may be potential targets for treatment of RSA.

VEGFR2 mediated RAS/MAPK signaling pathway to explore the mechanism of Bushen Antai Granule in the treatment of recurrent abortion

Objective:To investigate the effect of Bushen Antai Granule on the mRNA and protein expression of Ras protein/mitogen activated protein kinase mediated by vascular endothelial growth factor receptor 2 with small interference RNA interference.Methods:Method to construct the placenta microvascular endothelial cells,and the preparation of kidney fetus granule drug-containing serum,select the best drug-containing serum concentration,it can be divided into normal group,the serum siRNA-NC normal serum group,drug serum,siRNA normal serum group,siRNA drug serum group,using real-time fluorescent quantitative PCR,Western blotting,immunofluorescence test respectively the RAS/MAPK mRNA and protein expression.Results:Results there was no significant difference in Ras and MAPK mRNA and protein expression between the normal group and the negative control group(P>0.05).The mRNA and protein expressions of Ras and MAPK in the drug serum group were significantly higher than those in the normal serum group(P<0.01).Ras and MAPK mRNA and protein expression were significantly decreased in siRNA1 normal serum group compared with normal serum group(P<0.01).Ras,MAPK mRNA and protein expression in siRNA1 drug serum group were significantly different from that in siRNA1 normal serum group(P<0.01).Conclusion:Conclusion The therapeutic effect of Bushen Antai Granule on recurrent abortion may be realized by upregulation of RAS/MAPK mRNA and protein expression.

补肾活血方治疗肾虚血瘀型卵巢储备功能下降患者的临床疗效观察

目的观察补肾活血方对肾虚血瘀型卵巢储备功能下降(DOR)患者的治疗效果。方法选取北京妇产医院中医科2021年1月至2022年12月就诊的180例诊断为DOR的肾虚血瘀型患者作为研究对象,采用计算机随机分组法分为雌二醇片/雌二醇地屈孕酮片组(西药组)、补肾活血方组(中药组)和补肾活血方联合雌二醇片/雌二醇地屈孕酮片组(联合组),各60例。观察三组治疗前后卵泡刺激素(FSH)、黄体生成素(LH)、雌二醇(E_(2))、抗米勒管激素(AMH)、窦卵泡数(AFC)、卵巢体积、卵巢动脉和子宫动脉阻力指数(RI)、子宫内膜厚度的变化,比较三组中医证候评分及疗效。结果11例患者(4例来自西药组,3例来自中药组,4例来自联合组)退出研究。6个月后,三组FSH和LH均显著降低,E_(2)和AMH升高(P<0.05);西药组和联合组较中药组FSH降低更显著,E_(2)升高更显著(P<0.01)。治疗后,中药组和联合组卵巢体积增加(P<0.05),AFC显著增加(P<0.01),卵巢动脉RI显著降低(P<0.01);西药组卵巢体积增加不显著(P>0.05),AFC显著增加(P<0.05),卵巢动脉RI显著降低(P<0.05);三组子宫动脉RI均无显著改善(P>0.05)。三组子宫内膜厚度均显著增加(P<0.05),西药组和联合组较中药组增加更显著(P<0.01)。与治疗前相比,三组治疗后中医证候评分均显著降低(P<0.05),且中药组和联合组中医证候评分降低的程度大于西药组(P<0.05)。西药组总有效率为71.43%,中药组为82.46%,联合组为85.96%;联合组的疗效优于西药组(P<0.05)。结论补肾活血方能够通过平衡激素水平、促进排卵和修复子宫内膜对DOR患者产生良好的治疗效果,联合应用雌二醇片/雌二醇地屈孕酮片可能是治疗DOR的一种行之有效的治疗策略。

补肾健脾活血方调控Apaf-1/CHOP-Caspase-9/Bcl-2通路抑制成骨细胞氧化应激

目的探讨补肾健脾活血方对减轻大鼠成骨细胞氧化应激损伤和调控线粒体介导的细胞凋亡的机制。方法提取大鼠原代成骨细胞,通过CCK8实验检测补肾健脾活血方干预下对成骨细胞增殖活性的影响、碱性磷酸酶(alkaline phosphatase,ALP)染色检测成骨细胞ALP活性、茜素红染色检测成骨细胞的矿化情况、实时定量PCR检测Apaf-1和CHOP基因的mRNA表达、蛋白质印迹检测Caspase-9和Bcl-2蛋白表达量。结果CKK8实验和碱性磷酸酶染色表明氧化应激损伤的大鼠成骨细胞增殖活性显著降低,成骨分化受抑制,同时Apaf-1和CHOP的mRNA表达以及Caspase-9的蛋白表达明显增加(P≤0.01),Bcl-2蛋白表达则显著减少(P≤0.01);经补肾健脾活血方干预后,成骨细胞增殖活性得到明显的增强、成骨分化增加,Apaf-1和CHOP的mRNA及Caspase-9的蛋白表达均显著减少(P<0.05),而Bcl-2的蛋白表达则显著升高(P≤0.0001)。结论补肾健脾活血方可以促进成骨细胞增殖和成骨分化,显著降低Apaf-1和CHOP的mRNA及Caspase-9的蛋白表达,升高Bcl-2的蛋白表达量。补肾健脾活血方在减轻成骨细胞的氧化应激损伤和调控线粒体介导的细胞凋亡中起着重要作用。

补肾安胎方治疗肾虚型早期先兆流产患者的临床疗效及对其Th1/Th2相关因子、Th17/Treg细胞平衡的影响

目的探讨补肾安胎方治疗肾虚型早期先兆流产临床疗效及对Th1/Th2相关因子、Th17/Treg细胞平衡的影响。方法选取2021年1月—2022年12月期间南京市妇幼保健院收治的126例肾虚型早期先兆流产患者为研究对象,按随机数字表法分为对照组和观察组,每组各63例。对照组给予口服地屈孕酮治疗,观察组在对照组基础上增加补肾安胎方治疗。连续治疗14 d后,观察比较两组患者临床疗效及治疗前后中医证候积分、性激素[人绒毛膜促性腺激素(Human chorionic gonadotropin,β-HCG)、孕酮(Progesterone,P)、雌二醇(Estradiol,E_(2))、卵泡刺激素(Follicle stimulating hormone,FSH)]、Th1/Th2相关因子[白细胞介素-2(Interleukin-2,IL-2)、白细胞介素-4(Interleukin-4,IL-4)、白细胞介素-10(Interleukin-10,IL-10)、干扰素-γ(Interferonγ,INF-γ)]、Th17/Treg水平。结果治疗后观察组临床总有效率92.06%(58/63)明显高于对照组79.37%(50/63),差异有统计学意义(P<0.05)。治疗后两组患者阴道流血时间、出血量、小腹疼痛或坠胀、腰酸痛、头晕耳鸣、夜尿频数、两膝酸软积分及总积分均较治疗前降低,差异有统计学意义(P<0.05);且观察组阴道流血时间、出血量、小腹疼痛或坠胀、腰酸痛、头晕耳鸣、夜尿频数、两膝酸软积分及总积分均明显低于对照组,差异有统计学意义(P<0.05)。治疗后两组患者性激素β-HCG、P、E_(2)、FSH水平均较治疗前升高,差异有统计学意义(P<0.05);且观察组性激素β-HCG、P、E_(2)、FSH水平均明显高于对照组,差异有统计学意义(P<0.05)。治疗后两组患者血清IL-4、IL-10水平较治疗前升高,IL-2、INF-γ水平较治疗前降低,差异有统计学意义(P<0.05);且观察组IL-4、IL-10水平高于对照组,IL-2、INF-γ水平低于对照组,差异有统计学意义(P<0.05)。治疗后两组患者血清Th17及Th17/Treg水平较治疗前降低,差异有统计学意义(P<0.05);且观察组血清Th17及Th17/Treg水平较明显低于对照组,差异有统计学意义(P<0.05)。两组患者Treg水平治疗前后比较,差异无统计学意义(P>0.05)。结论在地屈孕酮治疗基础上结合补肾安胎方治疗,可显著缓解肾虚型早期先兆流产患者临床症状,并可改善性激素水平及免疫状态。

补肾降浊方在糖尿病肾病中的作用及其机制研究

目的 探究补肾降浊方在糖尿病肾病中的作用及其机制研究。方法 复制糖尿病肾病大鼠模型,使用11.04 g/(kg·d)补肾降浊方治疗。根据不同方法将大鼠随机分为Control组、DN组、DN+补肾降浊方组,每组6只。通过生化分析仪测定尿样中的尿白蛋白水平、血尿素氮和血肌酐含量,HE染色检测肾组织病理损伤,Masson染色检测肾组织纤维化情况,实时荧光定量聚合酶链反应检测SIRT1 mRNA,Western blotting检测MAPK/NF-κB信号通路相关蛋白的表达。结果 DN组血尿素氮、血清肌酐、尿白蛋白水平较Control组升高(P <0.05),补肾降浊方组较DN组降低(P <0.05)。DN组SIRT1 mRNA相对表达量较Control组降低(P <0.05),DN+补肾降浊方组较DN组升高(P <0.05)。Control组肾小球和肾小管形态完整,而DN组肾小管基底膜增厚,肾小球肥大,表明DN组大鼠肾功能受到严重损害。与DN组比较,DN+补肾降浊方组大鼠肾小管基底膜增厚和肾小球肥大情况明显减少。与Control组比较,DN组肾小球和肾小管腔中胶原蛋白的沉积量明显增加。与DN组比较,DN+补肾降浊方组大鼠肾小球和肾小管腔中胶原蛋白的沉积量明显减少。DN组SIRT1蛋白较Control组降低(P <0.05),p-p38/p38、p-NF-κB p65/NF-κB p65较Control组升高(P <0.05);DN+补肾降浊方组SIRT1蛋白较DN组升高(P <0.05),pp38/p38、p-NF-κB p65/NF-κB p65较DN组降低(P <0.05)。结论 补肾降浊方促进SIRT1的表达,抑制MAPK/NF-κB信号通路,改善糖尿病肾病肾组织损伤。

基于Akt/mTOR信号通路探讨补肾活血方对去卵巢大鼠软骨细胞自噬的影响

目的:研究补肾活血方是否可以通过调控Akt/mTOR信号通路促进去卵巢大鼠软骨细胞自噬反应来保护关节软骨。方法:选取30只SPF级12周龄雌性SD大鼠,体重(247.0±7.0)g,先随机选择6只为空白对照组,剩余大鼠采用卵巢切除术结合右膝关节腔注射碘乙酸钠(monosodium iodoacetate,MIA)建立绝经后膝骨关节炎模型,随机分为模型组、BSHXR-L组、BSHXR-M组、BSHXR-H组,每组6只。运用肉眼观察评分、番红O-固绿染色、免疫组化等方法明确补肾活血方对大鼠关节软骨损伤的保护作用,Western-blot检测自噬相关蛋白的表达,qPCR检测Akt、mTOR及下游自噬基因的相对表达。结果:造模后BSHXR(L、M、H)各组均可减轻软骨组织形态学损伤,免疫组化示Collagen-Ⅱ、Aggrecan表达逐渐升高,基质金属蛋白酶-13(matrix metallopriteinase-13,MMP-13)表达逐渐降低,其中BSHXR-M组和BSHXR-H组与模型组相比差异均有统计学意义(P<0.05)。Western-blot结果示BSHXR(L、M、H)各组自噬通路蛋白p-Akt/Akt、p-mTOR/mTOR受到抑制,下游蛋白Beclin-1、LC3Ⅱ表达逐渐升高,而p62逐渐降低,呈剂量效应。qPCR结果示BSHXR(L、M、H)各组均可促进Beclin-1、LC3ⅡmRNA的相对表达,抑制p62、Akt、mTOR mRNA的相对表达,与模型组相比差异有统计学意义(P<0.05)。结论:补肾活血方可以通过抑制Akt/mTOR信号通路增强软骨细胞自噬反应,进而发挥保护软骨的作用。

基于大鼠肝星状细胞氧化应激探讨化浊解毒补肾方治疗绝经后女性肝纤维化机制

目的通过研究化浊解毒补肾方含药血清对过氧化氢(H_(2)O_(2))诱导大鼠肝星状细胞(hepatic stellate cells,HSCs)氧化应激的影响,进一步探讨该方治疗绝经后女性肝纤维化的机制。方法制备化浊解毒补肾方含药血清和正常血清。将肝星状细胞随机分为空白组、模型组、空白血清组、氟维司群(ICI 182780,以下简记为ICI)组、中药组、中药+ICI组、雌二醇(estradiol,E2)组、E2+ICI组,随后按照分组处理细胞。采用流式细胞术检测细胞内活性氧(reactive oxygen species,ROS)水平;ELISA法检测细胞中丙二醛(malondialdehyde,MDA)、超氧化物歧化酶(superoxide dismutase,SOD)水平;免疫荧光染色法检测细胞中α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)表达;Western Blot法检测胞浆细胞色素C(cytochrome C)、环氧化酶-Ⅰ(cyclooxygenase-Ⅰ,COX-Ⅰ)、α-SMA、Ⅰ型胶原蛋白(collagenⅠ)表达。结果与空白组比较,模型组细胞中ROS、MDA水平和α-SMA、collagenⅠ蛋白表达以及α-SMA平均荧光强度均明显升高(P<0.0001,P<0.001),SOD水平、cytochrome C、COX-Ⅰ蛋白表达均明显降低(P<0.0001);与模型组比较,中药组和E2组ROS、MDA水平和α-SMA、collagenⅠ蛋白表达以及α-SMA平均荧光强度均有明显降低(P<0.0001,P<0.001,P<0.01,P<0.05),SOD水平、cytochrome C、COX-Ⅰ蛋白表达均有明显升高(P<0.0001);与中药组和E2组比较,在各自加入ICI处理后,对应的ROS、MDA水平和α-SMA、collagenⅠ蛋白表达以及α-SMA平均荧光强度均有明显升高(P<0.0001,P<0.001,P<0.05),SOD水平和cytochrome C、COX-Ⅰ蛋白表达均有明显降低(P<0.001,P<0.05,P<0.01)。结论化浊解毒补肾方含药血清可通过雌激素受体降低ROS、MDA水平和α-SMA、collagenⅠ蛋白表达以及α-SMA平均荧光强度,提高SOD水平和cytochrome C、COX-Ⅰ蛋白表达,从而提高抗氧化能力,减轻氧化应激反应,最终起到防治绝经后女性肝纤维化的作用。