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ObjectiveTo study the effect of Bushen Huoxuetang on the apoptosis and the expression of B-cell lymphoma （Bcl-2）-associated X protein （Bax）/ Bcl-2 and cleaved cysteine-containing aspartate proteolytic enzyme-3 （cleaved Caspase-3） in the nude mouse model of bone metastasis of breast cancer， and explore the mechanism of Bushen Huoxuetang in inhibiting bone destruction. MethodThirty BALB/c female nude mice were randomly assigned into blank group （n=6） and model group （n=24）. The suspension of 4T1 breast cancer cells was injected into the tibia of mouse right lower limb to establish model of bone metastasis of breast cancer. The successfully modeled nude mice were randomly assigned into model group， Bushen Huoxuetang group， zoledronic acid group， and combined drug group， with 6 mice in each group. Bushen Huoxuetang was administrated at a dose of 36.67 g·kg-1， once a day， and zoledronic acid was administrated by subcutaneous injection at a dose of 100 μg·kg-1， twice a week. The combined drug group was administrated with the same doses of Bushen Huoxuetang group by gavage and zoledronic acid by subcutaneous injection. The mice in the blank group and the model group were administrated with the same volume of distilled water by gavage for 14 days. On the next day at the end of drug administration， the mice were sacrificed by cervical dislocation. The general situation and weight changes of the mice were examined. The right lower limb was collected， and X-ray scanning and hematoxylin-eosin （HE） staining methods were used for observation of pathological changes in the bone. The terminal deoxynucleotidyl transferase dUTP nick-end labeling （TUNEL） was employed to detect the apoptosis of bone tissue in nude mice， and Western blot to determine the expression of Bax/Bcl-2 and cleaved Caspase-3 in the bone tissue. ResultCompared with the blank group， the modeling reduced the body weight （P<0.01） and increased the right lower limb weight of the nude mice （P<0.01）. Compared with the model group， Bushen Huoxuetang， zoledronic acid， and their combination increased the body weight （P<0.01） and decreased the right lower limb weight （P<0.01）. Compared with the blank group， the other groups showed obvious tumor cell atypia， deep nuclear staining， and clear bone metastasis， and the model group showed obvious osteolytic damage in right lower limb and loss of proximal tibia and knee joint. Compared with the model group， Bushen Huoxuetang， zoledronic acid， and their combination reduced the osteolytic lesions in the right lower limb and recovered part of the bone structure， demonstrating an inhibitory effect on bone destruction. The TUNEL assay showed that the model group had lower apoptosis rate of bone metastatic tumor cells than the blank group， Bushen Huoxuetang group， zoledronic acid group， and combined drug group （P<0.01）. Compared with the blank group， the modeling down-regulated the expression of Bax and cleaved Caspase-3 （P<0.01） and up-regulated the expression of Bcl-2 （P<0.01）. Compared with the model group， Bushen Huoxuetang， zoledronic acid， and their combination up-regulated the expression of Bax （P<0.01） and cleaved Caspase-3 （P<0.05， P<0.01） and down-regulated the expression of Bcl-2 （P<0.05， P<0.01）. ConclusionBushen Huoxuetang may inhibit bone destruction in the nude mouse model of bone metastasis of breast cancer by up-regulating the expression of Bax， down-regulating the expression of Bcl-2， activating cleaved Caspase-3， and further inducing apoptosis.
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ObjectiveTo predict the underlying mechanism of Bushen Huoxuetang in treating osteoporosis related to endocrine therapy in breast cancer by network pharmacology and to verify the results through in vitro cell model. MethodThe main effective components and targets of Bushen Huoxuetang were screened out through network pharmacology， and the targets of osteoporosis related to endocrine therapy in breast cancer were further obtained. The intersected targets were analyzed by Gene Ontology （GO） functional annotation and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment. Kaplan Meier plotter was used to analyze the survival of crucial targets. Finally， the inhibitory activity against cell proliferation was evaluated by in vitro methye thiazolye telrazlium（MTT） assay. The key targets and pathways were verified by Western blot， and the mRNA expression of the key targets was evaluated by real-time polymerase chain reaction（Real-time PCR）. ResultA total of 716 active components and 249 key targets of Bushen Huoxuetang were obtained from network pharmacology. There were 135 common targets， among which protein kinase B（Akt）1 and hypoxia-inducible factor-1α （HIF-1α） were two key targets. Additionally， 531 biological processes， 62 cellular components， 162 molecular functions， and 145 signaling pathways including breast cancer and endocrine resistance were involved. The key targets were effectively enriched in phosphatidylinositol 3-kinases（PI3K）/Akt and HIF-1 signaling pathways. According to the MTT assay， the cell proliferation rate and cell motility of MCF-7 and T47D cells in the luminal A cell line were reduced by Bushen Huoxuetang treatment （22.5， 45， 90 g·L-1， and 45， 90， 180 g·L-1） for 48 h as compared with the blank group. As revealed by Western blot， MCF-7 cells were treated with Bushen Huoxuetang （0， 15， 60 g·L-1） for 48 h， and the relative expression of p-PI3K， PI3K， p-Akt， Akt， and HIF-1α was decreased in a dose-dependent manner as compared with the blank group （P<0.05， P<0.01）. Real-time PCR was used to detect the mRNA expression of the key target HIF-1α. The results showed that the mRNA expression of HIF-1α in MCF-7 cells was decreased with the increase in the dose （P<0.01）， and the change was in a concentration-dependent manner. ConclusionThe mechanism of Bushen Huoxuetang in the treatment of osteoporosis related to endocrine therapy in breast cancer may be related to the key targets including Akt1 and HIF-1α through the PI3K/Akt/HIF-1α signaling pathway.
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Objective:To discuss the clinical efficacy of modified Bushen Huoxuetang combined with autologous bone grafting and locking compression plate （LCP） in treating nonunion of long bone fractures， and the effect on microcirculation， osteogenic differentiation factor and bone metabolism index. Method:A total of 70 patients were randomly divided into control group and observation group by random number table， with 35 cases in each group. Patients in both groups received LCP. Patients in control group got Dieda Shenggu granule， 10 g/time， 1 time/day. Patients in observation group got Bushen Huoxuetang， 1 dose/day. The course of treatment lasted for 3 months， and 3-month follow-up data were recorded. On a weekly basis， the main symptoms， such as pain， tenderness， longitudinal percussion pain and swelling were checked， and the time of disappearing of main symptoms and signs were compared. On a weekly basis， a X-ray examination was performed for callus formation and fracture line， and the fracture healing time was recorded. Before and after treatment， Fugl-Meyer （FMA） was scored， and levels of fibrinogen （FIB）， whole blood viscosity （BV） （high shear， low shear）， plasma viscosity （PV）， platelet aggregation rate （PAR）， <italic>D</italic>-Dimer （<italic>D</italic>-D）， bone morphogenetic protein-2 （BMP-2）， BMP-7， insulin-like growth factor-1 （IGF-1）， vascular endothelial growth factor （VEGF）， transforming growth factor-<italic>β</italic><sub>1</sub> （TGF-<italic>β</italic><sub>1</sub>）， osteocalcin （BGP）， osteoprotegerin （OPG）， procollagen type Ⅰ N-terminal propeptideserum amino pro peptide （PINP）， serum type 1 collagen cross-linked C-terminal peptide （S-CTX） and serum tartrate resistant acid phosphatase （TRACP） of type I procollagen were detected， and the safety was evaluated. Result:Disappearance time of symptoms and signs and fracture healing time in observation group were all lower than those in control group （<italic>P</italic><0.01）. At the third month after treatment， and during the three-month follow-up， scores of callus and FMA （upper and lower limbs） in observation group were all higher than those in control group （<italic>P</italic><0.01）. Levels of <italic>D</italic>-D， FIB， PAR， BV and PV （high-cut and low-cut）， BMP-2， BMP-7， IGF-1， VEGF， TGF-<italic>β</italic><sub>1</sub>， S-CTX and TRACP were all lower than those in control group （<italic>P</italic><0.01）， whereas levels of BGP， OPG and PINP were higher than those in control group （<italic>P</italic><0.01）. The curative effect of fracture healing was better than that of control group （<italic>Z</italic>=1.977， <italic>P</italic><0.05）. And the limb function recovery was superior to that in control group （<italic>Z</italic>=1.970， <italic>P</italic><0.05）. Conclusion:Based on autogenous bone and LCP， modified Bushen Huoxuetang can promote the fracture healing， shorten the course of disease， and promote the recovery of limb function， with a good clinical efficacy. It can improve microcirculation， promote the expression of osteogenic differentiation factor， regulate bone metabolism， and play a role in promoting fracture healing， with a safety in clinical use.
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Objective::To explore the clinical efficacy of modified Bushen Huoxuetang on kidney deficiency and blood stasis type early unexplained recurrent abortion and its effect on intestinal flora. Method::Totally 90 patients with kidney deficiency and blood stasis type early unexplained recurrent abortion were selected from March 2017 to October 2018.According to the random number table, they were divided into control group and observation group, with 45 cases in each group. The control group was given Bushen Huoxue capsule, while the observation group was given modified Bushen Huoxuetang. After treatment, the clinical efficacy, traditional Chinese medicine (TCM) syndrome scores and adverse reactions of two groups were compared, and the changes of serum inflammatory factors, coagulation function and intestinal flora were detected before and after treatment. Result::After treatment, the total effective rate of the observation group was 91.11%, which was higher than 77.78%of the control group (P<0.05). The scores of TCM syndromes in observation group were significantly lower than that in control group (P<0.05). The levels of C-reactive protein (CRP), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and interleukin-8 (IL-8) in the observation group were significantly lower than those in control group (P<0.05). The thromboplastin time (APTT), thrombin time (TT) and prothrombin time (PT) in observation group were significantly higher than those in control group, while fibrinogen (FIB) was significantly lower than that in control group (P<0.05). The numbers of enterococcus, yeast and Enterobacter in observation group were significantly lower than those of control group, while the numbers of bifidobacteria and Lactobacillus in observation group were significantly higher than those of control group (P<0.05). The incidence of adverse reactions in observation group was 11.11%, which was lower than 28.89%of the control group (P<0.05). Conclusion::Modified Bushen Huoxuetang has a good clinical efficacy in treating kidney deficiency and blood stasis type early unexplained recurrent abortion, and can reduce the TCM syndrome score. Its mechanism may be related to the reduction of inflammation, and improvement of coagulation function and intestinal flora, with a good safety.
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Objective: To observe the effect of Jiawei Bushen Huoxuetang on osteoporotic vertebral compression fractures (OVCF), bone density and bone metabolism. Method: One hundred and sixteen patients with OVCF operation were randomly divided into control group (58 cases) and observation group (58 cases) by random number table. Patients in control group got Alendronate sodium tablets, 70 mg/time, 1 time/week. Menatetrenone soft capsules, 15 mg/time, 3 times/days after meals. Calcium carbonate D3 chewable tablets (Ⅱ), 1 tablet/time, 2 times/days. Based on the treatment in control group, patients in observation group received additional Jiawei Bushen Huoxue decoction, 1 dose/day. The course of treatment was 24 weeks in both groups. The treatment of traditional Chinese medicine(TCM) fumigation was used in combination with the TCM for 2 weeks. At the 4th, 8th, 12th and 24th weeks after treatment, pain degree of waist and back were evaluated by pain visual analogue scale (VAS), and waist dysfunction was evaluated by Oswestry disability index (ODI). Before and after treatment, anterior vertebral height (AVBH), Cobb, femoral neck, lumbar spine bone mineral density and Chinese osteoporosis quality of life scale were evaluated. Before and after treatment, levels of carboxy terminal propeptide of type I collagen (CICP), C-terminal cross-linking peptide of type I collagen (CTX-I), tartrate resistant acid phosphatase (TRACP), bone alkaline phosphatase (BALP) and bone glaprotein(BGP) were detected. Result:By rank sum test, the clinical efficacy in observation was better than that in control group (Z=2.026, Pth, 8th, 12th and 24th weeks after treatment, scores of lumbago and back pain VAS and ODI were lower than those in control group (PPPPPConclusion:Jiawei Bushen Huoxuetang can enhance bone mineral density, regulate bone metabolism, reduce back pain, promote healing of fracture, and ameliorate osteoporosis, with obvious clinical efficacy.