Objective To study the effect of silencing Nek2 via RNAi on cell cycle of ovarian cancer SKOV 3 cells and the re‐lated molecular mechanism .Methods The Nek2‐siRNA was transfected into the ovarian cancer SKOV3 cells .The change of cell cy‐cle of SKOV3 cells at 48 h after transfection was examined by the flow cytometry technique ;Western blot assay was used to deter‐mine the change of level of the cell cycle related factors cyclinB1 ,CDK1 ,P27 and the phosphorylation level of the ERK1/2 after Nek2‐siRNA transfection for 48 h .Results The flow cytometry detection results showed that the proportion of the cells in G 2/M stage in the blank control group ,negative control group and RNAi group was 13 .72% ,12 .27% and 1 .56% respectively .Compared with the control group ,the number of the cells in G2/M stage in the transfected group was reduced obviously (P< 0 .05) .The Western blot detection results showed that compared with the control group ,the expression of cyclinB1 and CDK1 protein in SK‐OV3 cells was significantly reduced ,the expression of P27 was increased after silencing Nek2 and the phosphorylation level of ERK1/2 in SKOV3 cells was significantly reduced after silencing Nek2 gene(P< 0 .05) .Conclusion Silencing Nek2 gene might block the ovarian cancer cell line SKOV 3 initiating mitosis ,thus inhibit their proliferation .

Objective To assess the fixation efficiency and biocompatibility of the absorbable high strength poly L lactide (SC PLLA) screws which were designed and made by us through a new material reinforcement process, solid state compression (SC), by investigating the healing and tissue reaction of the rabbits which were given femoral osteotomy and fixed with the screws. Method The right femoral condyles of 12 New Zealand rabbits were first cut off and then fixed with a SC PLLA screw. They were sacrificed at 4, 8, 12, and 36 weeks postoperatively to exam their femurs radiologically and histologically. Results At all the sites of osteotomy was seen obvious formation of bony callus. There were no significant displacements at all the operative sides. Histologically, bony union showed at the site of osteotomy and direct connection between the bone and the implants was observed. Conclusion SC PLLA screws can provide sufficient internal fixation for the load bearing cancellous bone, and possess excellent biocompatibility.

0.05) but by IL 4 the result was significant(P0.05). Conclusion CTLA4 Ig could remarkably inhibit lymphocyte proliferation in MLC than in MLBSC; and IL 4 could be more effective in MLBSC than in MLC; the result did not show any synergetic effect of CTLA4 Ig+IL 4.

Objective To evaluate effectiveness of bone marrow derived mesenchymal stem cells(BMSCs) transduced by adenovirus mediated human bone morphogenetic protein-2 (Adv-hBMP-2) gene in the repair of diaphysic segmental bone defects of goats. Methods The tibial bone defects (2.1 cm) of 22 goats, who weighted from 18.1 kg to 29.5 kg, were established and divided into 4 groups: groupⅠ, Adv-hBMP-2 transduced BMSCs group (n=8); groupⅡ, adenovirus mediated ?-galactosidase(Adv-?gal)gene transduced BMSCs group (n=6); groupⅢ, non-transduced BMSCs group (n=6); groupⅣ, untreated group (n=2). Roentgenography, histomorphometrical analysis and biomechanical measurement were studied at various times. Results Roentgenography showed more callus in the bone defect site of group Ⅰ. At 24th week after implantation, the healing rates of group Ⅰ,Ⅱ,Ⅲ, and Ⅳ were 6/7, 1/5, 2/5, and 0/1 respectively. There was statistic significant difference of X-ray grading between groupⅠand groupⅡor Ⅲ. Histomorphometrical analysis showed much more total new trabecular bone area (TBA) of groupⅠthan that of other groups. Biomechanical strength of groupⅠwas superior to that of groupⅡand Ⅲ. Conclusion Adenovirus mediated BMP-2 gene transferred BMSCs can repair segmental bone defect in goat without osteoconductive scaffold.

[Objective]In present study an in vivo wear particles induced mouse calvarial osteolysis model was used to investigate the inhibitory effect of erythromycin over wear particles induced osteolysis.[Method]Twenty-four male eight-week-old C57BL/J6 mice were allocated into 4 groups: PMMA group receiving 30 mg PMMA particles implantation onto the calvariae,PMMA+2EM group receiving 30 mg PMMA particles implantation and EM 2mg/kg ip injection,PMMA+10EM group receiving 30 mg PMMA particles implantation and EM 10mg/kg ip injection,control group receiving sham operation.Seven days later,mice calvariae were harvested for pathology analysis.[Result]Middle suture area in the control group was 0.079 mm2?0.011 mm2,in PMMA group 0.335 mm2?0.129 mm2.In PMMA+2EM group the area was 0.094 mm2?0.019 mm2,PMMA+10EM group was 0.091 mm2?0.028 mm2.Osteoclasts number in osteolysis area in the control group were 5.3?1.0,in PMMA group 19.2?5.3,PMMA+2EM group was 6.6?1.1,PMMA+10EM group 6.1?1.9.Compared with control group,PMMA particles induced significant osteolysis(P

[Objective]To design a method of in vitro preparation and seperation for metallic wear particles around joint prosthesis and evaluate its feasibility in medical experiments.[Method]Ti-6Al-4V and Co-Gr-Mo alloys were used to make two friction jars respectirely. National inventive patent applied number 03142073.7.Lots of quadrate blocks made of the same materials are put into the jars respectively,which were then.lubri cated by man-made body fluid and vibrated on a bottle shaker.After 21 days the fluid was harvested and centrifuged to get the produced wear particles.The collected particles were studied by using element trace analysis,laser countersizer and scanning electron microscopy.The J774.A1 macrophages cultured together with these particles for 24 hours were observed under inverted phase contrast microscopy and transmission electron microscopy.[Result]A got great amounts of metallic particles with 1?m diameter coned beproduce using this method.The aver age diameter of titanium alloys(Dv90) is 1.011 and that of Co-Gr-Mo is 1.010.Particle size distribution had good consistency in different materials.Under scanning electron microscopy ,the particles had irregular shapes just like those got from revision operations.The particles taken into the J774.A1 macrophages could be seen under inverted p hase contrast microscopy and transmission electron microscopy.[Conclusion]This method is good enough to producl lots of metallic wear particles mosth like those around total joint prosthesis and can be used in further in vivo and in vitro studies about joint prosthesis loosening.

Objective To evaluate osteogenetic effectiveness of hyaluronic acid (HA) mixed with adenovirus mediated human bone morphogenetic protein-2 gene(Adv-hBMP-2) transfected bone marrow derived mesenchymal stem cells(BMSCs) of rabbits in the repair of radial shaft defects of rabbits. Methods The BMSCs were collected from bone marrow of rabbits, cultured and transfected by Adv-hBMP-2 in vitro. The radial shaft defects (1.5 cm) models of 20 rabbits (40 sides) were created and then respectively injected the cells and/or materials according to the following four groups (n=10): Ⅰ, Adv-hBMP-2 transfected BMSCs+HA; Ⅱ, Adv-hBMP-2 transfected BMSCs; Ⅲ, HA; Ⅳ, control. Results The bone defects of 6 out of 8 sides in the group Ⅰ were repaired completely and partial bone marrow cavities were revasculized. Only 3 sides were repaired completely in the group Ⅱ and no side was repaired completely in the groups Ⅲ and control. There were statistic significance between each groups in X-ray scores. In the groups ofⅠand Ⅱ, much woven callus formed in the sites of the bone defects in the 4-8th week after injection. Partial bone marrow cavities were revasculized in the 12th week after injection; while in the groups of HA and control, the sites of bone defects were mainly full of fibrous tissue and no healing happened. Both ends of the bone defects were hardened. There was significant difference statistically in the new trabecular bone area in the groups ofⅠand Ⅱ. The maximum vertical loading and elastic modulus were significant differenct in the group ⅠandⅡ. The ratios of average maximum vertical loading/normal value of radial shaft in the groups ofⅠandⅡwere 75.86% and 45.45%, respectively. Conclusion The tissue-engineered bone made by HA mixed with Adv-hBMP-2 transfected BMSCs can repair the bone defects in the radial shaft of rabbits. HA is a safe, effective carrier for tissue engineering.

Objective To repair the experimental femoral head necrosis with the implantation of tricalcium phosphate (TCP) loaded with human bone morphogenetic protein-2(hBMP-2) gene transfected bone marrow derived mesenchymal stem cells(BMSCs). Methods Established the animal models of femoral head necrosis in 22 goats by the ligation of the lateral and medial circumflex femoral arteries and delivery of liquid nitrogen into femoral head. 4 goats served as the control group without any treatment. 3 weeks after surgery, necrotic bone in the other 18 goats were removed and TCP loaded gene transfer cells were implanted into the femoral head with BMP-2 gene transfected BMSCs in right side and ?-gal gene transfected BMSCs in left side. BMP-2 concentrations in femoral head were tested by the ELISA at 1st, 2nd, 3rd week after implantation. Histological observation was done before and at 6th, 10th, 16th week after implantation. New bone volumes (NBV) were measured at 16th week after implantation by the histomorphometry method. X-ray was taken at 16th week after operation in untreated group and after treatment in BMP-2 group and ?-gal group. Results The concentration of BMP-2 in femoral head of BMP-2 group was higher than that of ?-gal group at different time point. Empty lacunae and fibrotic marrow were demonstrated before implantation. New bone was evident in the implantation field of BMP-2 group while fibrous tissues were evident in that of ?-gal group at 6th, 10th and 16th week after treatment. Histomorphology analysis indicated that the NBV in BMP-2 group was significantly higher than that in ?-gal group. Articular surface collapse were observed in the X-ray photograph of untreated group. Regular shape and normal density of femoral head in BMP-2 group compared with the irregular shape and low density of femoral head in ?-gal group could be demonstrated in the X-ray photograph 16 weeks after treatment. Conclusion Implantation of the TCP loaded with human BMP-2 gene transfected BMSCs could repair early-stage experimental femoral head necrosis.

[Objective]To observe the effect of sciatic nerve and femoral nerve resection on heterotopic osteogenesis induced by recombined human bone morphogenetic protein-2(rhBMP-2) and discuss role of never innervation on bone regeneration and partial reasons for larger callus after bone fracture accompanying denervation.[Method]A total of 36 male ICR mice were divided into experimental group and control group at random.0.125mg rhBMP-2 /collagen composites were implanted into the right thigh muscle pouches after sciatic nerve and femoral nerve section in the experimental group and after sham operation in the control group.On the 7~(th),14~(th) and 21~(st) day after implantation,roentgenographic,biochemical,histological analyses and osteoclasts TRAP staining were performed to detect the effects of sciatic nerve and femoral nerve resection on bone growth initiated by rhBMP-2.[Result]On the 7~(th),14~(th) and 21~(st) day,wet weight of new bony tissues of the experimental group was obviously greater than that of the control group.Radiography showed range of bone formation in the experimental group was larger than that in the control,but density of new bone was lower than that of the latter.Biochemical detection showed value of AKP of the experimental group was significantly higher than that of the control on the 7~(th) day,while the content of Ca of the experimental group was higher than that of the control on the 14~(th) day and the content of Ca and P of the experimental group was lower than that of the control on the 21~(st) day.Histological observation showed area of new bone of the experimental group was greater than that of the control,but trabecular bone of the former was sparser than that of the latter.Image analysis of bony tissue showed the relative number of osteoclasts in the experimental group became higher on the 21~(st) day,while volume density and width of bone trabecula in the experimental group were lower than that of the control group.TRAP staining showed osteoclasts in the new bone of the experimental group were more activated than that of the control.[Conclusion]Nerve section results in enhancement of ability of bone regeneration by BMP in the early period,but trabecular bone becomes sparse in the middle and later period because of bone resorption by osteoclasts,which indicates nerve in nervation influences bone regemeration though direct and/or indirect way.

[Objective]To identify the expression of neurotrophic factors(NTFs)and their high affinity receptors(Trk)during the bone induction by recombined human bone morphogenetic protein-2(rhBMP-2)and discuss possible roles of NTFs on the osteoinductive activity of BMP.[Method]After a total of 36 ICR mice were divided into experimental group and control group at random,rhBMP-2/collagen sponge and collagen sponge were implanted into the right thigh muscle pouches of two groups respectively.Tissues in the implanted sites of two groups were removed on the 7~(th),14~(th) and 21~(st) day after implantation.Histological,immunohistochemical and RT-PCR analyses were performed to detect osteoinductive effect of rhBMP-2 and the expression of NTFs and Trk.[Result]RhBMP-2/collagen sponge displayed a potent ability inducing bone formation,while the expression of NTFs and Trk was observed in the course of osteoinduction by rhBMP-2.Especially on the 7~(th) day in the experimental group,NGF and TrkA positive immunostaining reached the peak in the stage of chondrogenesis,including chondroblasts,chondrocytes,hypertrophic chondrocytes and osteoblasts,then decreased in the number of positive cells and the intensity of immunostaining on the 14~(th) and 21~(st) day.BDNF was only observed in chondrocytes and cartilage matrix,and TrkB was expressed in chondroblasts and chondrocytes at the 7~(th) day after implantation.Expression of NT-3 was similar to that of NGF,while TrkC was expressed in chondroblasts and chondrocytes on day 7.Expression level of the mRNA of NTFs peaked at 7 days after the implantation,then decreased,which was nearly coincident with immunohistochemical results.[Conclusion]Many different kinds of cells are induced by rhBMP-2 express NTFs and Trk,which suggests that NTFs may play an important role in the osteogenesis initiated by BMP through direct and indirect pathways.