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Objective @#To investigate the role and mechanism of bone formation caused by the ratio of advanced platelet-rich fibrin (A-PRF) and β-tricalcium phosphate (β-TCP) in rabbit femur defect model, which provides a new idea for clinical treatment of bone defect.@*Methods @#Twenty-four New Zealand white rabbits were divided into model group, 1∶1 complex group (A-PRF∶β-TCP=1∶1), 2∶1 complex group (A-PRF∶β- TCP=2∶1) and 4∶1 complex group (A-PRF∶β- TCP=4∶1）, with 6 rabbits in each group. Femoral defect models were constructed in each group. In the composite group, the bone defect was filled with composite material, while in the model group, no material was filled. After 8 weeks, the animals were euthanized and specimens were collected. Bone mineral density (BMD), bone volume fraction (BV/TV), trabecular thickness (Tb.Th), trabecular separation (Tb.SP) and trabecular number (Tb.N) in femoral defect tissue were measured by micro-CT and photographed. Hematoxylin - eosin staining was used to detect the pathological changes of new bone tissue. The morphological changes of the new bone tissue were observed by scanning electron microscopy. Determination of phospho-mitogen activated protein kinase p38 (p-p38MAPK), CCAAT/enhancer binding protein homologous protein (CHOP) and phospho-cysteine aspartic protease-3 (p-Caspase3) in newborn femur by ELISA. The mRNA expressions of osteoprotegerin (OPG), bone morphogenetic protein-2 (BMP-2), receptor activator of nuclear factor kappa-B ligand (RANKL) and p38MAPK were detected by real-time quantitative PCR. The expression of OPG, BMP-2, RANKL, p-p38MAPK and p-Caspase3 protein in the new bone tissue was observed by immunohistochemistry. @*Results @#In the model group, bone formation in the femoral defect area was slow and osteogenic quality was poor. Compared with the model group, the bone formation and neocapillaries of femoral defect area in the complex group was good, BMD, BV.TV, Tb.Th, Tb.N were increased, and Tb.Sp were decreased, the expressions of p-p38MAPK, CHOP and p-Caspase3 were decreased, and the mRNA and protein expressions of OPG and BMP-2 were increased. The mRNA expression of RANKL and p38MAPK was decreased. Apoptosis in new bone tissue of each group showed the lowest apoptosis rate in samples of the 2∶1 complex group (P<0.05); A-PRF: β-TCP=2∶1 ratio has the best osteogenic effect. @*Conclusion@#The complex composed of A-PRF and β-TCP can promote the expression of OPG, inhibit the expression of RANKL and phosphorylation of p38MAPK, reduce the apoptosis of new bone tissue cells, and promote osteogenic differentiation.
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AIM: To explore the effect of inhibiting proliferation and inducing apoptosis in human primary gastric tumor by the Agkis-trodon halys venom anti-tumor component (AHVAC-). METHODS: Human primary gastric cancer cells were isolated by trypsin digestion, serum-free culture, and purified by differential adherence method, and cells were identified by immunohistochemistry. Cell proliferation and toxicity assay (CCK-8) was used to detect the inhibition rate of AHVAC- in different concentrations of primary gastric cancer cells. Immunohistochemistry was used to verify the apoptosis of human primary gastric cancer cells induced by AHVAC- and the morphological changes were observed by Hematoxylin-Eosin staining (HE staining). AHVAC--induced primary gastric cancer cell transformation rate was detected by flow cytometry Annexin V/PI double staining.RESULTS: Seven human primary gastric cancer cells were successfully isolated and purified, and 11 cases failed. Immunohistochemical identifications of carcinoembryonic antigen (CEA) and broad-spectrum keratin protein (AE1/AE3) were positive for both antibodies. AHVAC- inhibited the proliferation of human primary gastric cancer cells and showed a dose-dependent effect (P<0.01). Immunohistochemistry showed that the expression level of cysteine aspartic protease-3 (Caspase-3) up-regulated with the increase of AHVAC- concentration. HE staining showed that with the increase of AHVAC- concentration, the cell gap increased, nuclear pyknosis, and apoptosis cells increased. Flow cytometry showed that the apoptosis rate of human primary gastric cancer cells up-regulated with the increase of AHVAC- concentration (P<0.05). CONCLUSION: AHVAC- can inhibit the proliferation and induce apoptosis in human primary gastric cancer cells in a dose-dependent manner.
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OBJECTIVE:To study the effect of grape seed procyanidins on protein expressions of cysteine aspartic protease 3 (Caspase-3), tumor necrosis factor-related receptors (TRAF6) in renal tissue of rats with renal ischemia-reperfusion injury (RIRI),and explore the protective mechanism of procyanidins on RIRI. METHODS:50 rats were randomly divided into sham op-eration group,model group,Shenfukang capsules group(positive control,600 mg/kg),procyanidins low-dose,high-dose groups (100,150 mg/kg),10 in each group. All rats were intragastrically administrated once a day,for 7 d. After administration,rats in other groups except for sham operation group were established the RIRI model.After the modeling successed of 24 h,levels of creati-nine(Cr),urea nitrogen(BUN)in serum,and protein expressions of Caspase-3,TRAF6 in renal tissue of rats in each group were detected,and apoptotic rate of renal tubular epithelial cells was determined. RESULTS:Compared with sham operation group,lev-els of Cr,BUN in serum in model group were significantly increased(P<0.05);protein expressions of Caspase-3,TRAF6 in re-nal tissue were obviously enhanced (P<0.05);apoptotic rate of renal tubular epithelial cells was obviously increased (P<0.05). Compared with model group,levels of Cr,BUN in serum in each administration group were obviously decreased(P<0.05);pro-tein expressions of Caspase-3,TRAF6 in renal tissue were obviously weakened(P<0.05);apoptotic rate of renal tubular epithelial cells was obviously decreased(P<0.05);and procyanidins high-dose group showed superior effect to low-dose group and Shenfu-kang granules group (P<0.05). CONCLUSIONS:Grape seed procyanidins can relieve the RIRI of rats,which may be achieved by reducing protein expressions of Caspase-3 and TRAF6 to inhibit the apoptotic rate of renal tubular epithelial cells.