Objective To discuss the clinical results of lateral locking plate combined with bone grafting in the treatment of distal femoral fractures in the elderly.Methods Between January 2006 and December 2013,112 senile patients with distal femoral fracture were treated at our department by lateral locking plate combined with bone grafting.They were 16 men and 96 women,aged from 67 to 85 years (average,76.1 years).By AO classification,48 cases were type 33-A1,11 type 33-A2,3 type 33-A3,7 type 33-B1,4 type 33-B2,32 type 33-C1,and 7 type 33-C2.All of them had a previous history of osteopenia.By the Singh index classification,22 cases were rated as grade Ⅰ,74 as grade Ⅱ,13 as grade Ⅲ,and 3 as grade Ⅳ.Postoperatively,the patients conformed to regular follow-up,accepted functional rehabilitation and continued medication for osteopenia.Radiographic examinations were conducted to observe fracture healing.At the last follow-up,the function of involved knee was assessed according to the criteria by Karlstrom and Olerud.Results All the 112 patients completed their follow-ups,ranging from 10 to 48 months (average,16 months).All fractures united primarily after an average of 4.2 months (range,from 3.5 to 5.0 months).According to the criteria by Karlstrom and Olerud,93 cases were rated as excellent,10 as good and 9 as fair,giving a good-to-excellent rate of 92.0％.No such complications as nonunion,malunion,plate breakage,or screw loosening were observed during the follow-ups.Conclusion Lateral locking plate combined with bone grafting is effective for the treatment of distal femoral fractures in the elderly,if followed by postoperative anti-osteopenia therapy and early functional exercises.

Objective To observe the effect of Sox9 gene overexpressian on chondrogenesis of esenchymal stem ceils (MSCs) in vitro. Methods Rabbit MSCs were obtained and purified by gradient centrifuge and adhesion culture in vitro. MSCs were transfected by recombinant Sox9 plasmid. RT-PCR and Western blot analysis were used for transfection confirmation. Chondrogenic molecules such as type I1 collagen,aggrecan and Sox9 were detected by immunohistology, RT-PCR and western blot. Cell proliferation was tested by MTT assay. Results MSCs were successfully transfected with Sox9 gene and verified by RT-PCR and Western blot analysis. The overexpression of Sox9 had no effect on the proliferation of MSCs. Transfeeted cells expressed more chondrogenic markers than the control groups. Conclusion Sox9 gene overexpression can enhance chondrogenic differentiation of MSCs.

BACKGROUND: Different isolation and culture methods for bone marrow mesenchymal stem cell (BMSCs) will result in varying cell activity and purity, which will influence repair effect of tissue engineering. OBJECTIVE: To investigate the optimized methods of isolation and culture of BMSCs in vitro and validate the efficacy of cell acquisition. DESIGN, TIME AND SETTING: In vitro cytology trial was performed at the laboratory of Department of Orthopedics, Second Hospital of Shanxi Medical University from October to December 2007. MATERIALS: Two 3-month-old New Zealand rabbits were provided by Shanxi Institute of Livestock. METHODS: Lymphocyte isolation solution was added to DMEM culture solution containing fresh bone marrow. BMSCs were obtained and purified by gradient centrifuge and adhesion culture in vitro. Non-attached cells were moved to new culture flask every 8 hours. The solution was changed firstly after 5 days of culture. Trypsinization was conducted at cell confluence of 90%. The first, third and fifth passages of cells were harvested. MAIN OUTCOME MEASURES: The morphology of BMSCs was observed with phase contrast microscope; the growth curve was drawn to determine doubling time. The percentage of the wel1 growth P2 cells were identified by CD44 staining by flow cytometry. RESULTS: Primary cultured BMSCs were oval, spindle-shaped or polygonal, and adhered to plastic surface within 48 hours, exhibiting spindle-shaped or polygonal with clear nucleus, and reached 90% confluence within 14 days. The first, third and fifth passages of BMSCs showed S-type, and were in latent phase at 1-3 days, in log phase 3 days later, and cells came into platform phase at 7-8 days. The mean doubling time was 24.22 hours. CD44-positive cells were found in the second passage, with positive rate of 93.0%. CONCLUSION: BMSCs are easily isolated and cultured in vitro with good growth and high purity by gradient centrifuge and adhesion culture with lymphocyte isolation solution.

Objective To investigate the effects of sivelestat sodium on early inflammatory reaction in rats with smoke inhalation injury. Methods Forty SPF male SD rats were randomly divided into 5 groups:normal control group (A), injury group (B), smoke inhalation treated with 10 mg/kg sivelestat sodium group (C), smoke inhalation treated with 20 mg/kg sivelestat sodium group (D) and smoke inhalation treated with 30 mg/kg sivelestat sodium group (E), 8 rats for each group. After smoke inhalation injury model was established, the treatment groups were intraperitoneally injected sivelestat sodium 10 mg/kg, 20 mg/kg and 30 mg/kg separately. B group was treated with the same volume of physiological saline. After 24 hours,ELISA was used for detecting serum contents of neutrophil elastase (NE), myeloperoxidase (MPO), tumor necrosis factor-α(TNF-α) and interleukin-6 (IL-6) in five groups. Meanwhile the water content of lung tissue was measured, and the pathological changes were observed by HE staining. The thickness of alveolar septum was measured and compared between groups. Results Compared with control group, the serum levels of NE, MPO, IL-6, TNF-α, water content of the lung tissue and thickness of alveolar septum were significantly higher in other four groups (P<0.05). Compared with injury group, the serum levels of NE, MPO, IL-6, TNF-α, water content of the lung tissue and thickness of alveolar septum were significantly lower in treatment groups (P<0.05). Compared with 20 mg/kg treatment group and 30 mg/kg treatment group, the serum levels of NE, MPO, IL-6, TNF-α, water content of the lung tissue and thickness of alveolar septum were significantly lower in 10 mg/kg treatment group (P<0.05). Conclusion The result shows that sivelestat sodium can reduce the early inflammatory reaction of rats with smoke inhalation injury and attenuates the lung edema. In this experiment, the treatment effect of 10 mg/kg sivelestat sodium is better than other treatment doses.

Objective To observe the change of collagen in osteoarthritis and study the pathogenesis of osteoarthritis.Methods The right hind limb of twenty-four adult New Zealand White rabbits were immobilized with plaster cast in extension position for 10,20,30 and 40 days,respectively.Six animals without immobilization served as control.The articular cartilage of the medial femur condyle was harvested for transmission electron microscopy,in-situ hybridization of collagenⅡ,and immunohistochemistry of collagenⅠ,Ⅱ,Ⅲ.Results Transmission electron microscopy showed articular cartilage was destructed from the fine collagen fiber network of tangential zone.The fine collagen fiber network did not contain chondrocyte.Immunohistochemistry and in-situ hybridization showed that in earlier period of osteoarthritis,the collagen typeⅡand its gene expression firstly increased,then decreased with destruction of ultrastructure,and chondrocytes enhanced type Ⅱ collagen expressing and synthesizing mainly in transition zone and upper deep zone.In articular cartilage of osteoarthritis there was type Ⅲ collagen,instead of typeⅠcollagen.Conclusion In osteoarthritis,articular cartilage degenerated from tangential layer,in which collagen can not be repaired after destruction,this may contribute to the chondral degeneration.

Objective: To summarize the efficacy of percutaneous endoscopic transforaminal unilateral decompression for the treatment of lumbar spinal stenosis, and to investigate the incidence and possible reasons for lower extremity symptoms of non-surgical side after surgery. Methods: The clinical data of 46 patients who underwent percutaneous endoscopic transforaminal unilateral decompression for lumbar lateral recess stenosis between February 2016 and March 2018 were retrospectively analysed. There were 22 males and 24 females with an average age of 56.9 years (range, 21-90 years). The disease duration was 6 months to 12 years (mean, 4.8 years). There were 37 cases of single-segment lumbar lateral recess stenosis and 9 cases of multi-segment stenosis. All patients underwent single-segment surgery, including 2 cases of L 3, 4, 35 cases of L 4, 5, and 9 cases of L 5, S 1. For patients with multi-segment lumbar lateral recess stenosis, the single segment consistent with radicular symptom was selected for surgery. The visual analogue scale (VAS) scores of low back pain and sciatica and Oswestry disability index (ODI) scores were recorded preoperatively and at last follow-up to evaluate the relief of symptoms. Postoperative symptoms on non-surgical side of lower extremity including new or aggravated radiation pain and numbness were recorded. The surgical effectiveness was evaluated according to modified MacNab score at last follow-up. Results: All the 46 patients underwent successful operations without postoperative complications such as wound infection, permanent nerve root injury, and deep vein thrombosis. The patients were followed up 12-37 months (mean, 23.8 months). Four patients suffered from radioactive pain and numbness for the non-surgical side of lower extremities at 1-3 months postoperatively, and the symptoms were improved after conservative treatment without revision surgery. VAS scores of low back pain and sciatica and the ODI scores were significantly improved at last follow-up when compared with preoperative ones ( P<0.01). According to the criteria of modified MacNab scores, 25 cases were excellent, 17 cases were good, 3 cases were fair, and 1 case was poor. The excellent and good rate was 91.3%. Conclusion: Percutaneous endoscopic transforaminal unilateral decompression for lumbar spinal stenosis would be able to relieve pain effectively and achieve good results. Lower extremity symptoms of non-surgical side may be observed postoperatively. Although the symptoms can be relieved by conservative treatment, further investigation should be carried out for the long-term result of those cases.

Due to the insufficient supply of embryonated chicken eggs,the preparation of large quantities of inactivated influenza vaccines will require an alternative virus culture system after the emergence or reemergence of a pandemic influenza virus.The Vero cell is one of the ideal options since it was used for producing many kinds of human vaccines.However,most of the influenza viruses can not grow well in Vero cells.To develop a new influenza vaccine with Vero cells as a substrate,the virus needs to adapt to this cell substrate to maintain high growth characteristics.By serial passages in Vero cells,the B/Yunnan/2/2005va(B)strain was successfully adapted to Vero cells,with the hemagglutination titer(HAT)of the virus reaching 1:512.The high growth characteristic of this strain is stable up to 21 passages.The strain was identified by hemagglutination inhibition (HAI)test and sequencing respectively;the HA;gene sequence of the virus was cloned and analyzed.The screening and establishment of high growth B virus provides an important tool for influenza vaccine production in Vero cells.

The expression of multidrug resistant proteins in bladder cancer and clinical implication was studied. Expression of multidrug-associated protein (MRP), P-glycoprotein (P-gp), P53 and Bcl-2 proteins were detected by using immunohistochemical method in 40 specimens of bladder transitional cell carcinoma. The results showed that the positive rate of MRP, P-gp, P53 and Bcl-2 was 52.5 %, 57.5 %, 47.5 % and 62.5 % respectively. The positive rate of MRP, P-gp, P53 and Bcl-2 in the grade Ⅰ, Ⅱ and Ⅲ of tumors was 46.3 %, 38.5 %, 38.5 %, 23.1 %； 52.9 %, 39.8 %, 47.1 %, 76.4 %； 60.0 %, 80.0 %, 60.0 %, 90.0 % respectively. The positive rate of MRP, P-gp, P53 and Bcl-2 in 24 primary tumor specimens was 37.5 %, 41.7 %, 33.3 %, 45.8 % and that in 16 cases in recurrent specimens receiving chemotherapy 75.0 %, 81.3 %, 68.8 %, 87.5 % respectively. It was suggested the positive rate of MRP, P-gp, P53 and Bcl-2 was increased with the advance of tumor grade. The positive rate of four proteins in all recurrent cases was significantly increased (P<0.05). The expression of MRP, P-gp, P53 and Bcl-2 proteins might be the important factors for chemotherapy failure.

The expression of multidrug resistant proteins in bladder cancer and clinical implication was studied. Expression of multidrug-associated protein (MRP), P-glycoprotein (P-gp), P53 and Bcl-2 proteins were detected by using immunohistochemical method in 40 specimens of bladder transitional cell carcinoma. The results showed that the positive rate of MRP, P-gp, P53 and Bcl-2 was 52.5 %, 57.5 %, 47.5 % and 62.5 % respectively. The positive rate of MRP, P-gp, P53 and Bcl-2 in the grade Ⅰ, Ⅱ and Ⅲ of tumors was 46.3 %, 38.5 %, 38.5 %, 23.1 %； 52.9 %, 39.8 %, 47.1 %, 76.4 %； 60.0 %, 80.0 %, 60.0 %, 90.0 % respectively. The positive rate of MRP, P-gp, P53 and Bcl-2 in 24 primary tumor specimens was 37.5 %, 41.7 %, 33.3 %, 45.8 % and that in 16 cases in recurrent specimens receiving chemotherapy 75.0 %, 81.3 %, 68.8 %, 87.5 % respectively. It was suggested the positive rate of MRP, P-gp, P53 and Bcl-2 was increased with the advance of tumor grade. The positive rate of four proteins in all recurrent cases was significantly increased (P<0.05). The expression of MRP, P-gp, P53 and Bcl-2 proteins might be the important factors for chemotherapy failure.

Objective:To investigate the prevalence of enterovirus 71 (EV71) and the genetic characteristics of VP1 region in common hand, foot and mouth disease (HFMD) cases in Shenzhen from 2016 to 2022.Methods:Throat swabs from mild HFMD in Shenzhen sentinel hospitals were collected from 2016 to 2022. A total of 38 EV71-positive samples were screened from these throat swabs and were sequenced. Then, the VP1 sequence of these EV71-positive samples were analyzed for their phylogenetic evolution by bioimformatics software DNAStar and MEGA 6.Results:From 2016 to 2022, the number of EV71 infections among HFMD patients in Shenzhen sentinel hospitals decreased from 136 in 2016 to 0 in 2022. The mumber of EV71 infections in 2018 and 2019 decreased by 96.3%(257/267) compared to that in 2016 and 2017. From 2020 to 2022, the number of EV71 infections decreased to 0. During this period, the EV71 vaccination rate among HFMD patients increased from 6.4% to 39.6%; Evolutionary analysis showed that the nucleotide homology and amino acid homology between 38 EV71 sample strains in Shenzhen from 2016 to 2022 were 91.8%-99.9% and 98.3%-100.0%, all belonging to the C4a subgenotype; Among them, 26 strains wene local epidemic strains, and 11 strains were imported from other provinces, with a close genetic relationship with epidemic strains in Hainan, Yunnan, Sichuan, Tianjin, Henan, Jilin, and other places. One strain from 2017 had the closest genetic relationship with the US epidemic strain OP207969-USA-2017. Further comparing the EV71 epidemic strains in Shenzhen from 2016 to 2022 and EV71 severe strains, it was found that the EV71 strains in Shenzhen carried four amino acid mutation sites related to severe condition, named R22H, K43R, I249V and T289A.Conclusions:The EV71 epidemic strains in Shenzhen from 2016 to 2022 all belong to the C4a subgenotype, and the number of EV71 infection shows a downward trend with the increase of vaccine coverage rate. At the same time, the distribution of EV71 virus strains in Shenzhen shows a significant decrease in local strains and a predominance of imported strains. There are a total of four amino acid mutation sites associated with severe cases in the EV71 sample strains in Shenzhen from 2016 to 2022. Among them, 22R and 289T are located at the N and C ends of VP1, which are related to EV71 adsorption and targeting cells. The 43R site is associated with binding ability to Annexin2 protein, which enhances cell binding ability.