Objective To isolate and culture the rabbit mesenchymal stem cells,and explore the biological features in vitro.Methods Two milliliter of bone marrow was extracted from the tibia of juvenile rabbit under anesthesia and sterile conditions,and mesenchymal stem cells were obtained by density gradient centrifugation and inoculated to 100ml culture flask.The morphological changes,adherence rate and ultrastructure of the stem cells were observed.The surface markers and growth cycle of the mesenchymal stem cells were detected by flow cytometry,and biological features were observed by inducing the cells differentiate into osteoblasts.Results The adherence of mesenchymal stem cells started 2 hours after culturing,basically finished 20 hours later,and more than 90% of the cells were fused at the 15th day.Observed with transmission electron microscopy,the cultured cells showed typical ultrastructure of stem cells: numbers of microvillus mainly distributed on one side of the cells,irregular nuclei with indentation,and lots of cell organs.Analysis for the cell surface markers indicated positive expression of CD44 and CD90 and negative expression of CD45 in mesenchymal stem cells.Cell growth cycle assay showed that most mesenchymal stem cells(about 97%) stayed in stage G1 and G2.Three weeks after osteoblastic induction,the mesenchymal stem cells gradually produced small calcium salt crystals,the cells were positive in alkaline phosphatase staining,and Von Kossa staining showed many calcium salts in the cell clone as black reaction deposits.Conclusions The rabbit bone marrow mesenchymal stem cells isolated by density gradient centrifugation are of high purity and stable growth rate,and the cells' characters can be still maintained after serial subcultivation.Since easy to operate on isolation and proliferation,the mesenchymal stem cells could be used as the tissue engineering seed cells in repairing of intervertebral disc degeneration.

Objective To explore the effect and experience of problem-based (PBL) teaching in neurosurgical clinical training and improve the quality of the neurosurgical clinical training. Methods 600 clinical medical students of Grade 2011 in their probation in our department of neurosurgery were chosen as the subjects of study and were randomly divided into the experimental (PBL, 15 groups, 20 people/group, over 300 people) and control groups (CBL+ LBL, 15 groups, 20 people/group, over 300 people). In experimental group, PBL which is a problem-based, student-centered and teacher-directed pedagogy was adopted. In control group, CBL (case-based learning) + LBL (lecture-based learning) were adopted. After probation, two groups of students had the unified test and assessed their teaching methods by questionnaire survey and at the same time, the questionnaire survey about PBL' impact on the various aspects was done only to experimental group students. SPSS 15.0 software was used to do statistical analysis and data were processed by description analysis, t test, Mann-Whitney U test and correlation analysis. Results The scores of theoretical knowledge and general ability in PBL group [(40.57 ±3.24), (41.23 ±4.56) individually] were higher than these in control group [(37.21 ±4.05), (36.51±4.11) individually]. The differences demonstrated statistically meanings. Through the surveys, the students in experimental group showed the high satisfaction (97.0%, n=291) in PBL teaching. Most of the students considered that their abilities of team cooperation (96.7%, n=290), and self-directed learning (92.7%, n=278) were improved by PBL teaching. The students in experimental group illustrated their attitudes in satisfaction of teaching; interests of clinical training, participation of teaching and understanding of knowledge were superior to the control group. The differences demonstrated statisti-cally meanings. Conclusion PBL teaching got a good effect in neurosurgical clinical training. This method, which improved students' participation and motivation of study and contributed to the training of medical students' comprehensive abilities, is suitable for modern neurosurgical clinical training.

Programmed cell death 5(PDCD5)is a new kind of programmed cell death regulation and control gene. PDCD5′s expression is widespread,and evolution is conservative. PDCD5 has the effect of promo-ting a wide variety of tumor cells to apoptosis and inhibiting proliferation,such as lung cancer,liver cancer et al. Several laboratories′ discoveries show that PDCD5′s expression is significantly decreased when in some dis-ease cases,especially in tumor cases. PDCD5,which is related to the clinical relevance of the tumor,has a great clinical value of discovering,diagnosis and treatment of the tumor.

BACKGROUND:Recent research has shown that transplanted bone marrow mesenchymal stem cells(BMSCs) migrate to the injured regions and exert their therapeutic effects in cases of intracranial trauma, stroke, inflammation and degenerative disease.The specific mechanisms involved in their migration to lesions are still to be fully elucidated.OBJECTIVE:To explore the effects of stromal cells derived factor-1(SDF-1) and its receptor CXCR4 on the migration of transplanted BMSCs to ischemic brain lesions.DESIGN, TIME AND SETTING:The cytological in vivo study was performed at the Central Laboratory, Xinqiao Hospital, Third Military Medical University of Chinese PLA from February 2008 to February 2009.MATERIALS:Bone marrow samples were obtained from normal or primary affection non-involved bone marrow patients aged 15-40 years at the Department of Hematology, Xinqiao Hospital, Third Military Medical University of Chinese PLA.A total of 72 healthy male Sprague Dawley rats aged 3-4 months were supplied by the Experimental Animal Center, Research Institute of Surgery, Third Military Medical University of Chinese PLA.METHODS:Human BMSCs were isolated by combination of gradient centrifugation and different adherent time method.The transient middle cerebral artery occlusion(MCAO) was induced using intraluminal vascular occlusion in 54 rats, based on the method described by Nagasawa et al.The remaining 18 rats served as sham operation group, only inserted with thread for 10 mm depth.At 2, 4 and 8 days after cerebral ischemia, the expression of SDF-1 in the ischemic brain was determined by real time RT-PCR and immunohistochemistry in 9 rats from either group.The remaining 36 rat models of cerebral ischemia/reperfusion were equally and randomly assigned into a cell transplantation and solution control groups.1 mL human BMSCs(2?109/L cells) or 1 mL phosphate buffered saline were slowly infused through the caudal vein at 24 hours following reperfusion.MAIN OUTCOME MEASURES:CXCR4 mRNA and protein expression in human BMSCs was determined.SDF-1 mRNA and protein expression following ischemia/reperfusion were detected.Migration of transplanted human BMSCs into the damaged region was observed through immunohistochemistry.RESULTS:RT-PCR showed that human BMSCs were positive for CXCR4 mRNA.Immunocytochemistry revealed that CXCR4 mainly expressed in cell membrane and cytoplasm of human BMSCs.At 2, 4 and 8 days following cerebral ischemia/reperfusion, SDF-1 mRNA levels showed an increased tendency, and showed significant difference compared with the sham operation group(P

BACKGROUND: Transplanted bone marrow mesenchymal stem cells (BMSCs) migrate to the injured regions and exert their therapeutic effects. The specific mechanisms involved in their directional migration to lesions remain unclear. OBJECTIVE: To investigate the chemokine receptor CXCR4 and CX3CR1 expression of human BMSCs in hypoxia culture. DESIGN, TIME AND SETTING: The cytology in vitro study was performed at the Central Laboratory, Xingqiao Hospital, Third Military Medical University of Chinese PLA from February 2008 to February 2009. MATERIALS: Cells harvested from the iliac heparinized bone marrow were obtained by iliac crest aspiration from healthy adult volunteers, aged 15 to 40 years old, at the Department of Hematology, Xinqiao Hospital, Third Military Medical University of Chinese PLA. METHODS: Bone marrow was obtained by puncture. Human BMSCs were harvested by combination of density and gradient centrifugation and different adherent method. Cells at passage 3 were incubated in a 25 cm2 flask. When 70%-80% confluence was found, cells were incubated at 37 ℃ and saturated humidity in an incubator containing 3% O2, 5% CO2, 92% N2 for 48 hours, and those incubated under normal oxygen as controls. MAIN OUTCOME MEASURES: Morphology was examined by phase contrast microscopy. Cell surface markers were tested by flow cytometer. The CXCR4 and CX3CR1 mRNA expression were detected by real-time PCR. The CXCR4 and CX3CR1 protein expression were determined by Western blot assay. RESULTS: All of the cells had a fibroblast-like morphology cultured in vitro and reached 90% confluence at 12-14 days, with the presence of polarity arrangement and whirlpool-shape. Cells were uniformly positive for CD105 (99.38%) and CD29 (99.13%), but negative for CD14 and CD45. Exposure of BMSCs to 3% O2 increased expression of the CXCR4 mRNA and CX3CR1 mRNA, which were respectively 2.130 times and 2.361 times of normal culture; expression of the CXCR4 protein and CX3CR1 protein was respectively 1.69 times and 1.93 times of normal culture. CXCR4 and CX3CR1 mainly expressed in membrane and cytoplasm of human BMSCs. CONCLUSION: Hypoxia (3% O2) can upregulate the expression of CXCR4 and CX3CR1 in human BMSCs, which might be one of the machenisms underlying the migration of BMSCs.

Circulating DNA is cell-free DNA existing in plasma or serum. It has already been verified that circulating DNA of cancer patients is derived from tumor cells. Therefore, it is of great value to detect the changes in the quantity and quality of the circulating DNA in cancer patients for early diagnosis and prognosis. The advantages of the detection of circulating DNA such as micro-trauma, convenient access to samples, possibility of continuous and dynamic monitoring, make it a promising tumor mark. This review recapitulates the application of circulating DNA analysis in hepatocellular carcinoma patients for diagnosis and prognosis.

Objective To explore the clinical curative effect on non-small cell lung cancer (NSCLC) patients by cinobufacini injection combined with first-line chemotherapy.Methods Eighty patients with NSCLC from January 2013 to January 2015 in our hospital were selected as the research objects.Then they were divided into the observation group (n =40)and the control group (n =40)by random number tables.The patients in control group accepted docetaxel and cisplatin combination chemotherapy regimens (TP).While the observation group accepted cinobufacini injection on the basis of the control group.Then the local control, adverse reactions and prognosis of the two groups were compared.Results The local control of observation group was 77.5%,while the control group was 62.5%,the local control of observation group was obviously higher than that of the control group (χ2 =5.36,P =0.03).Leucopenia incidence of the observation group was 27.5%,the control group was 50.0%,and the incidence of the observation group was obviously lower than that of the control group (χ2 =4.27,P =0.04).There was no statistically significant difference between the two groups in diarrhea,stomachache,vomiting,tinnitus (17.5% vs.27.5%,χ2 =1.15,P =0.28;25.0% vs.45.0%,χ2 =3.52,P =0.06;5.0% vs.7.5%,χ2 =0.34,P =0.56;7.5% vs.10.0%,χ2 =0.16,P =0.69).There was statistically significant difference between the two groups in median survival time (97 d vs.45 d,HR =8.934,χ2 =9.928,P <0.05).Conclusion The cinobufacini injection combined with docetaxel can effectively reduce the incidence of myelosuppression,and improve survival and local control with high safety,and the clinical effect is remarkable and can improve the prognosis of patients.

Objective To evaluate of the clinical effect of triple arthrodesis and bone grafting combined with plat screw internal fixation for treatment of necrosis of talus. Methods The clinical data of 24 patients with necrosis of talus who were treated by triple arthrodesis and bone grafting combined with plat screw internal fixation from April 2011 to January 2015 in Xiangya Second Hospital of Central South Univer-sity were retrospectively analyzed. And the clinical effect were measured by AOFAS Ankle Hind-foot Scale System. Results The 24 patients were followed up for 26 months averagely (12~36 months). According to AOFAS Ankle Hind-foot Scale System, there were 4 cases of mod-erate and 20 cases of bad before treatment, and there were 18 cases of excellent, 4 cases of good, and 2 cases of moderate after the treat-ment. Conclusion It can achieve good clinical results in treatment of necrosis of talus through triple arthrodesis and bone grafting combined with plat screw internal fixation.

Acrocephalosyndactylia ; pathology ; Fetus ; pathology ; Humans

Objective To evaluate the clinical value of the oral contrast-enhanced three-dimensional spiral CT cholangiography to display better the intrahepatic and extrahepatic bile ducts. Methods Before spiral CT scanning, the preparation was similar to the conventional X-ray oral cholecystography. The collected parameters included 3-5 mm slice, 1 or 1.5 pitch, 120 kV, 220-250 mAs, and 1 mm reconstruction image. Volume rendering was used to construct the 3D bile duct images. Results In the 20 patients without serious biliary obstruction, oral three-dimensional CT cholangiography displayed the grade Ⅰ branch of hepatic ducts clearly and the displaying rate was 100%. The displaying of the grade Ⅱ branch of hepatic ducts was not good enough and the rate was 60%. The grade Ⅲ branch of hepatic ducts could not be displayed. Conclusion As the substitute and supplement of invasive ERCP, spiral CT 3D cholangiography is safe, simple, painless, and non-invasive. Furthermore, spiral CT 3D cholangiography can display the space structure of bile duct tree more clearly than ERCP. This technique is of great clinical value.