Objective To investigate the effects of Xinshang Xuduan Decoction(XXD, mainly composed of Rhizoma Drynariae, Radix Dipsaci, and Eupolyphaga seu Steleophaga) on the proliferation and differentiation of rat osteoblasts and bone morphogenetic protein 2(BMP-2)expression in the osteoblasts . Methods The animal serum containing XXD was prepared by serum pharmacological method and then was mixed together with α-MEM for the cell culture. Osteoblasts were isolated from the skull bone of SD neonatal rats by collagenase digestion and were identified by alkaline phosphatase(ALP) staining and alizarin red staining. The third and fourth generations of osteoblasts were treated with XXD at the volume fraction of 5%, 10%, 20% for 24, 48 , 72 hours respectively, and then the proliferation of the cells was evaluated by Cell Counting Kit-8(CCK-8). In the other test, osteoblasts were cultured with blank control serum, and 10% serum containing XXD, Radix Dipsaci, and Rhizoma Drynariae, respectively, and then the ALP activity was examined by using ALP assay kit and the expression of BMP-2 was investigated by enzyme-linked immunosorbent assay(ELISA). Results XXD had a dose- and time-dependent effect on the proliferation of rat osteoblasts in a suitable volume fraction range from 5% to 20%, and the effect of XXD at 10% was the best. Compared with the blank control serum group, ALP activity was increased in the cells treated with 10% serum containing XXD, Radix Dipsaci, and Rhizoma Drynariae(P<0.01). On culturing day 7, the expression of BMP-2 was increased in 10% XXD group and Rhizoma Drynariae group(P<0.05). Conclusion XXD can increase the ALP activity and BMP-2 expression in the osteoblasts in vitro, so does the single herb of Rhizoma Drynariae. And their therapeutic mechanism in promoting the healing of fractures may be related with the enhancement of osteogenesis of osteoblasts.

Objective To investigate the effect and mechanism of adenovirus vector mediated SOCS 3 gene transfection in CD4+ Th cell differentiation and expression of inflammatory cytokines of mouse .Methods The CD4+ Th cells were isolated from spleen of C57bl/6 mouse and cultured .Ad‐SOCS3 were transfected into the CD4+ Th cells .PHA was used for culturing with the CD4+ Th cells .RT‐PCR were used to detect the mRNA expression ,and Western blot were used to detect the protein expression of cytokines .Results Compared with the control group ,the gene and protein expression of T‐bet ,IL‐2 ,IFN‐γ,STAT4 and IL‐12Rβ2 in the transfected group were significantly down‐regulated ,the gene and protein expression of SOCS3 ,GATA‐3 ,IL‐4 ,IL‐6 ,IL‐10 and STAT6 were significantly up‐regulated(P<0 .01) .Conclusion The results indicate that SOCS3 gene transfection can up‐regu‐late SOCS3 mRNA and protein expression in the CD4+ Th cells ,down‐regulate the JAK/STAT pathway ,inhibition of Th1 cell dif‐ferentiation ,and down regulation of inflammatory cytokine gene and protein expression ,and indirectly promote Th2 cell differentia‐tion ,and up the corresponding inflammatory cytokine gene and protein expression .

AIM:To investigate the relationship between autophagy and calcification in vascular smooth muscle cells ( VSMCs) after platelet-derived growth factor (PDGF)-BB stimulation.METHODS:Cultured VSMCs were stimulated with PDGF-BB for different time, the expression of vascular calcification-related proteins and autophagy-related proteins were detected by Western blot .The interaction be-tween Beclin1 and PI3KC3 was detected by co-immunoprecipitation.RESULTS: The expression of BMP2 and ALP showed a trend from decline to rise.ALP slumped at 12 h, and BMP2 slumped at 6 h.Moreover, the expression of Beclin-1 showed a trend from rise
to decline, and peaked at 12 h.The conversion of LC3-ⅠtoⅡincreased in a time-dependent manner , and peaked at 24 h.The ex-pression of BMP2 and ALP was increased in VSMCs incubated with PDGF-BB and autophagy inhibitor 3-MA, compared with PDGF-BB-stimulated VSMCs.Furthermore, the interaction between Beclin1 and PI3KC3 was enhanced at 6 h after PDGF-BB stimulated, peaked at 12 h, and kept in high level at 24 h.Moreover, the phosphorylation level of Beclin 1 was enhanced by PDGF-BB stimulation, and peaked at 6 h.CONCLUSION:Our findings demonstrate that PDGF-BB-induced autophagy inhibits VSMC calcification by en-hancing Beclin1 phosphorylation and interaction between Beclin 1 and PI3KC3.

Bone morphogenetic proteins (BMPs) are multi-functional growth factors,they are found not only inducer of new bone formation,but also have critical roles in cell growth,differentiation,migration,apoptosis,embryogenesis and organogenesis.Studies show that BMPs are closely related to tumorigenesis and metastasis,which have important practical values in tumour therapy.

Objective To introduce our experience of performing posterior or anterior renal lip incision for partial nephrectomy in the treatment of renal hilum endophytic renal cell carcinoma.Methods From Jan.2010 to Jan.2014,five female patients with renal hilum tumors were treated in our institute.The median age was 54 (51-72) years.The median tumor diameter was 4.0 (2.8-4.8) cm.The median preoperative creatinine was 53.9 (52.6-75.4) μmol/L.One of them was solitary kidney with absolute indication; three cases had basic disease with relative indication; one was with selective indication.The patients were put in supine or lateral position.After general anesthesia,we preformed partial nephrectomy by cutting posterior renal lip in 3 cases and the anterior lip in 2 cases.We clamped the renal artery,opened the renal posterior or anterior lip,then dissected the tumor beside the pseudo-capsule.After removing the tumor,we used 3-0 absorption suture to control bleeding and repair the opened collecting system.Finally,we used 2-0 absorption suture to close the renal defect.Results The median operation time was 195 (155-215) min; the median renal warm ischemia time was 35 (15-70) min; the median postoperative hospital stay was 8 (7-9) d.There was no secondary bleeding and urine leakage happened.The pathological results showed that 3 cases with clear cell carcinoma,1 with papillary carcinoma and 1 with renal medullary interstitial cell tumor.All patients showed normal kidney shape.The median postoperative creatinine was 63.0 (59.4-75.4) μmol/L.After a median follow up of 24.2 mon,all patients survive without tumor recurrence.Conclusions The short-term result of posterior or anterior renal lip incision partial nephrectomy in treating endophytic renal hilum endophytic renal cell carcinoma is safe and feasible.

Objective To investigate the expression and significance of death receptor 4(DR4) and DR5 in human craniopharyngioma.Methods The expression of DR4 and DR5 was determined by immunohistochemistry and in situ hybridization in 28 samples of craniopharyngioma and 25 samples of normal brain tissue.Results With low expression in partial normal brain tissue,DR was expressed highly in all of the craniopharyngioma samples.High DR expression in craniopharyngioma tissue differed from low DR expression in normal brain tissue(P0.05).Conclusions High DR expression is prevalent in craniopharyngioma tissue.This may contribute to the apoptosis-induced therapy of craniopharyngioma.The control of DR expression lays in protein level.This may contribute to the selective induced-apoptosis of tumor necrosis factor-related apoptosis-induced ligand.

Objective To study the effects of serum of the Zhuartgguqiang jin tablets on the proliferation and osteogenic differentiation of bone marrow stromal stem cells (BMSCs) of aged rats.Methods The BMSCs were obtained from male SD rats of 18 months.The cell surface markers CD34,CD31,CD29 were detected by flow cytometry.The proliferation of BMSCs treated by different concentration of medicated serum (2.5％,5％,10％) at 24,48,72hours was detected by CCK-8 method.The BMSCs were divided into 4 groups,which were osteogenic induction group,the Chinese medicated osteogenic induction group,Chinese medicine group,the blank group.After the BMSCs were cultured for 7 days,alkaline phosphatase (ALP) vitality was detected.After the BMSCs were cultured for 14 days,the BMSCs were stained by alizarin red,the mRNA of ALP and osteocalcin(OC) was detected.Results The expressionof CD29 was positive,and the expression of CD31 and CD34 were negative.2.5％,5％ of the medicated serum could promote cell proliferation.The ALP vitality of osteogenic induction group[202.76 ± 15.44(U/gprot)],the Chinese medicated osteogenic induction group [240.48 ± 18.55 (U/gprot)],Chinese medicine group [178.87 ± 17.29 (U/gprot)]were significantly higher than that of blank group [111.24 ± 20.71 (U/gprot)] (t =22.50,7.985,3.535,all P ＜0.01).The ALP activity of Chinese medicated osteogenic induction group was higher than that of osteogenic induction group (t =3.103,P ＜ 0.05).The ALP gene relative OD value of osteogenic induction group (0.40 ± 0.20) and Chinese medicated osteogenic induction group (0.60 ± 0.06) were higher than the blank group (0.09 ± 0.03) (t =2.372,9.547,all P ＜0.01).The ALP gene expression of Chinese medicated osteogenic induction group was obviously higher than that of the osteogenic induction group(P ＜0.05).The OC gene relative OD value of osteogenic induction group(0.58 ± 0.09) and Chinese medicated osteogenic induction group(0.76 ± 0.11) were higher than the blank group(0.41 ± 0.02) (P ＜ 0.05,P ＜ 0.01).The OC gene expression of Chinese medicated osteogenic inductiongroup was obviously higher than that of the osteogenic induction group(t =2.673,P ＜ 0.05).Conclusion Medicated serum of Zhuangguqiangjin tablets can promote the proliferation and osteogenic differentiation of BMSCs of aged rats,which may be the mechanism of prevention senile osteoporosis.

Objective To evaluate the role of conventional protein kinase Cγ (cPKCγ)/growthassociated protein-43 (GAP-43) signaling pathway in ketamine-induced apoptosis in hippocampal neurons of developing rats in an in vitro experiment.Methods Primarily cultured hippocampal neurons were seeded in culture plates at a density of 1×10.6 cells/ml and divided into 2 groups (n=10 each) using a random number table:control group (C group) and ketamine group (K group).Group C received no treatment.Ketamine was added with the final concentration of 300 μmol/L in group K.At 12 h of culture or incubation,the apoptosis in hippocampal neurons was detected by flow cytometry.The apoptotic rate was calculated.The expression of cPKCγ,GAP-43 and phosphorylated GAP-43 in hippocampal neurons was measured by Western blot.Results Compared with group C,the apoptotic rates of hippocampal neurons were significantly increased,and the expression of cPKCγ,GAP-43 and phosphorylated GAP-43 was down-regulated in group K (P<0.01).Conclusion The mechanism by which ketamine induces apoptosis in hippocampal neurons of developing rats may be related to inhibition of cPKCγ/GAP-43 signaling pathway activation in an in vitro experiment.

Objective Salidroside is a major active component of integripetal rhodiola herbal medicine, which has a significant activity against hypoxia and ischemia.This study was to investigate the effects of side chain carbon losssalidroside analogues (N04) on the expressions of HIF-1α-related factors in the hypoxia-injured EAhy926 human vascular endothelial cells.Methods EAhy926 human umbilical vein endothelial cells in the logarithmic growth phase were randomly divided into a normal control, a hypoxia model control, a salidroside, a high-dose N04, a medium-dose N04, and a low-dose N04 group.The hypoxia model was established by depriving the culture medium of sugar and serum and culturing the EAhy926 cells in an environment of 95%N2+5%CO2 for 2 hours, followed by intervention with salidroside at 1×10-6 mol/L and N04 at 1×10-6, 1×10-7, and 1×10-8 mol/L, respectively.Then, the activity of the cells was detected by MTT assay, their LDH activity examined by spectrophotometry, the mRNA expressions of HIF-1α and VEGF measured by RT-PCR, the protein expressions of HIF-1α, VEGF and pVHL determined by Western blot, and the activity of eNOS measured by ELISA.Results Compared with the normal control group, the hypoxia model cells showed significantly reduced activity (0.51±0.05 vs 0.27±0.02, P<0.01), an elevated LDH level ([6.65±1.43] vs [78.82±2.33] U/L, P<0.01), and decreased eNOS activity ([1.56±0.23] vs [1.16±0.20] U/100 mL, P<0.01).In comparison with the hypoxia model group, the cells treated with high-, medium-, and low-dose N04 exhibited remarkably increased activity (0.27±0.02 vs 0.0.42±0.05, 0.40±0.03 and 0.37±0.04, P<0.01), a reduced LDH level ([78.82±2.33] vs [53.05±3.90], [58.42±4.45] and [62.73±3.63] U/L, P<0.01), and increased eNOS activity ([1.16±0.20] vs [3.01±0.47], [2.60±0.26] and [2.32±0.29] U/100 mL, P<0.01).The activity of eNOS was also increased in the salidroside group ([2.32±0.29] U/100 mL, P<0.01).The cell activity in the high-and medium-dose N04 groups was markedly higher than that in the salidroside group (P<0.05), and so was the eNOS activity in the high-dose N04 group and the LDH level in the medium-and low-dose N04 groups (P<0.05).In comparison with the normal control group, the expressions of HIF-1α mRNA, HIF-1α protein and VEGF protein were significantly up-regulated in the hypoxia model group (P<0.01) while that of the pVHL protein markedly down-regulated (P<0.01).Compared with the hypoxia model group, the expressions of HIF-1α mRNA, HIF-1α protein and VEGF protein were remarkably reduced (P<0.05), while that of the pVHL protein markedly elevated (P<0.05).Both the expressions of VEGF mRNA and HIF-1α protein were significantly lower in the medium-and low-dose N04 groups than in the salidroside group (P<0.05).Conclusion N04 can protect vascular endothelial cells against hypoxia-induced injury by regulating the expression of HIF-1α-related factors and eNOS.

Objective To explore the effectiveness of our self-designed 3D printing relay template in the treatment of malunion of irregular intra-articular ankle fractures.Methods From October 2013 to October 2015,our self-designed 3D printing relay template was used in 14 patients with severe malunion of intra-articular ankle fracture to assist their intra-articular osteotomy.They were 10 males and 4 females,with an average age of 40.5 years (from 34 to 59 years).Thin slice CT scan was preformed preoperatively for all the ankles involved.The computer fictitious tibial and fibular models for location and osteotomy navigation templates were acquired from the data of CT scan after processing,design and reconstruction by software.After the navigation templates which fit the size of solid ankle were printed by a 3D printer,they were sterilized and reserved.The guide plates were used in turn during operation.The irregular malunited bone blocks in the posterior malleolus were completely resected along the osteotomy template.After fracture reduction,the bone blocks were stabilized with internal fixation.Fracture union time was recorded.Scores of visual analogue scale (VAS),American Orthopaedic Foot and Ankle Society (AOFAS) and the short form-36 (SF-36),ankle ROM and complications were also recorded at the final follow-ups.Results The 14 patients obtained an average follow-up of 28.6 months (from 14 to 35 months).Bony union was achieved after an average of 3.7months (from 3 to 6 months).At the final follow-ups,on average,VAS score (2.2 ± O.8),AOFAS ankle and hindfoot score (81.1 ± 6.8),SF-36 score (82.5 ± 5.2),dorsal extension (27.5 ° ± 2.5°),and plantar flexion (24.2° ±6.3°) were significantly improved compared with the preoperative values (6.3 ± 1.8,43.1 ± 12.0,37.6 ± 9.9,14.1° ± 3.8° and 14.4° ± 3.9°,respectively) (P ＜ 0.05).No implant failure,soft tissue complications or post-traumatic arthritis occurred during the follow-up.Conclusion Since our self-designed 3D printing relay templates for osteotomy can accurately position the fracture lines of the distal tibia via limited incision and invasion,they help a lot in precise osteotomy and anatomical reduction of malunited intra-articular ankle fracture,preventing incidence of traumatic arthritis.