AIM:To investigate the impact of various levels of glucose on endothelial progenitor cells (EPCs) proliferation, senescence, and nitric oxide (NO) secretion,and the effect of insulin under high glucose conditions.METHODS: Mononuclear cells were collected from rat bone marrow by density gradient centrifugation, cultured with medium 199, and identified to be EPCs at 7th day by flk-1 and AC133 double staining. EPCss were harvested and incubated with glucose (5, 10, 20, 40 mmol/L) or insulin (0.1, 1, 10, 100 nmol/L) under high glucose conditions for 24 h or 7 days. Proliferative capacity, senescence level and NO secretion (after 24 h of incubation) were subsequently determined.RESULTS: High glucose (40 mmol/L) markedly inhibited EPCs proliferation, accelerated EPCs senescence, and decreased NO production (all P

Objective To report the anatomical study and clinical application of the vascularized composite fibula perforating osteoseptocutaneous flap. Methods Latex injection studies were performed on 24 adult cadaveric lower limbs. The presence, prevalence, and location of the peronel artery and its perforators in the distal leg were documented. 16 vascularized fibul osteoseptocutaneous flaps were performed for reconstruction of composite soft-tissue defect of limbs. The flap areas ranged from 6 cm×4 cm to 16 cm× 8 cm. Results A mean of 5.3 perforating vessels of the peroneal artery was noted in the specimen. 40.6% of them were muscularcutanous perforators, and the other were septacutanous perforators. The perforating vessels were all located within 3.1 cm from the postlateral fibula. The average diameter was 1.1 mm at the deep fascial level, and the interval of them was 4-8 cm. There were two constant perforators which located about 15-25 cm and 4-7 cm from the lateral maleolar respectively. All of the 16 clinical cases survived uneventfully and the time to union of the graft-host junction site were 3 to 5 months. The appearance of flaps and the function of the limbs were satisfied during 10 to 36 months following up. Conclusion It is the anatomic basis that there are pedorating vessels of the peroneal constantly. The vnscularized compound fibula perforating osteoseptocutaneous flap has the advantage of flexible design. It is the safe and suitable choice in the cases when compound bone and soft tissue defects of the limbs are required for reconstruction.

Objective To explore the relationship between the interleukin-6 (IL-6) RS1800796 gene polymorphism and susceptibility of Enterovirus 71 (EV71) infection.Methods One hundred and twenty-three children with EV71 infection were selected as infection experimental group from March 2012 to December 2014 in the Central Hospital of Xiangtan,and they were divided into mild EV71 infection group (62 cases) and severe EV71 infection group (61 cases).And 52 age-and gender-matched healthy children were selected as the healthy control group.Two mL blood samples were collected from all subjects,and DNA was extracted by Beijing Optimal Boland Gene Technology LTD.The SNaPshot was used to determine the genotype for G/C polymorphism at RS1800796 position of IL-6 gene.Results The genotype frequency of IL-6 RS1800796 GG in the infection experimental group [73.2％ (90/123 cases)]was significantly higher than that in the healthy control group[48.1％ (25/52 cases)],and the difference was statistically significant (x2 =10o 215,P =0.002,OR =2.945,95 ％ CI:1.500-5.782).No significant difference was found in the distribution of genotype frequency of the IL-6 RS1800796 GG between the mild EV71 infection group and the severe EV71 infection group[71.0％ (44/62 cases)vs.75.4％ (46/61 cases),x2 =0.309,P =0.685].The G allele in IL-6 RS1800796 G/C was more frequent in the infection group (85.0％)than that in the control group (70.2％),and the difference was statistically significant (x2 =10.183,P =0.002,OR =2.399,95％ CI:1.389-4.143).No significant difference was found in allele frequency of the IL-6 RS1800796 G between the mild EV71 infection group and the severe EV71 infection group (83.1％ vs.86.9％,x2 =0.703,P =0.477).Conclusion The G allele of IL-6 RS1800796 confers susceptibility to infection of EV71.But G allele carrier will not increase the risk of severity after infection.

Objective To investigate the vascular endothelial growth factor (VEGF) expression level by chondrocytes isolated from patients with osteoarthritis (OA) in hip or femoral neck fracture (FNF) and explore the effect of synovial fluid from OA or FNF on secretion of VEGF. Methods The cartilage tissues were collected from 12 patients with OA in hip and 8 patients with FNF. Cartilage was stained with HIM and Safranin O/Fast Green (S/F) method. The damage of cartilage was evaluated using Mankin scores.Cathepsin B which was selected for cell dedifferentiation monitoring marker and VEGF level was detected in the supernatant fluid. The synovial fluid from OA, FNF and DMEM were respectively added to the culture medium to explore their effects on regulating VEGF. Results Cartilage the Mankin scores of OA group were higher than that of FNF group. Chondrocytes gradually lost their original spherical appearance, with Cathepsin B upregulated while VEGF downregulated. The OA synovial fluid can stimulate chongdrocytes to secrete more VEGF than the one from patients with FNF. However, chondrocytes gradually produced less VEGF after passaging. Conclusion Mankin scores had good correlation with chondrocytes' VEGF production in the early stage of primary culture. Chondrocytes showed quick dedifferentiation characteristics in vitro. OA synovial fluid showed abig ger capability in stimulating chondrocytes to express more VEGF, which might indicate that OA synovial fluid participated in the pathological process of OA.

Objective To evaluate the diagnostic value of the level of plasma D-dimer, lipoproteins and carcino-embry-onic antigen(CEA) in gastrointestinal cancer.Methods The plasma D-dimer ,lipoproteins,CEA and clinicopathological data of 139 gastrointestinal cancer patients and 155 normal controls were collected and analyzed .Lipoproteins included high-density lipoprotein(HDL),low-density lipoprotein(LDL) and lipoprotein a[LP(a)].SPSS 13.0 statistical software was used to analyze the sensitivity and specificity of each examination method and to find the appropriate combination .Results The plasma D-dimer,LDL,LP( a) and CEA levels were distinctly higher in patients than those in normal controls ( P<0.001).HDL levels were significantly lower in patients than those in normal controls (P<0.001).The cutoff of D-dimer was 0.495 μg/ml , the sensitivity of D-dimer was 62.6%,and the specificity was 86.5%.The cutoff of HDL was 1.025 mmol/L, the sensitivity was 72.7%,and the specificity was 85.2%.The cutoff of LDL was 3.375 mmol/L, the sensitivity was 54%,and the specificity was 82.6%.The cutoff of LP(a) was 27.3 mg/dl, the sensitivity was 58.3%,and the speci-ficity was 87.1%.The cutoff of CEA was 2.14 ng/ml, the sensitivity was 59.7%,and the specificity was 76.8%.The sensitivity and specificity of HDL +CEA were 77.7%and 88.4%, respectively.The sensitivity and specificity of HDL +D-dimer were 70.5% and 96.1%, respectively.The sensitivity of HDL +LP(a) was 76.3%,and the specificity was 93.5%.The sensitivity of D-dimer +HDL+LP(a) was 84.2%,and the specificity was 92.3%.The sensitivity of D-dimer +HDL+CEA was 87.1%, and the specificity was 85.8%.The sensitivity and specificity of HDL +LP(a) +CEA were 85.6%and 92.3%, respectively.The sensitivity of D-dimer +HDL+CEA +LP(a) was 89.9%while the specificity was 92.3%.Conclusions Plasma D-dimer and lipoproteins can serve as tumor markers in gastrointestinal cancer .Combined detection has higher sensitivity and specificity .

BACKGROUND:The main component of cartilage, type Ⅱ col agen gene expression in chondrocyte is positively correlated with SOX9 concentration in a dose-dependent manner.
OBJECTIVE:To observe the variation of SOX9 and type Ⅱ col agen mRNA content at different periods in the differentiation process (osteogenic, chondrogenic, adipogenic induction) of mesenchymal stem cel s, and to explore the correlation of SOX9 expression and type Ⅱ col agen.
METHODS:Bone marrow mesenchymal stem cel s were isolated from 4-week-old Kunming mice, and cultured in vitro to passage 3. The cel phenotype was identified with flow cytometry. Cel s were divided into three groups and subjected to three kinds of induction conditions favorable for adipogenic, chondrogenic and osteogenic differentiation, and each group was observed at three time points. In addition, the non-induced cel s were used as a control group. The total RNA of cel s was extracted at 3, 7, 14 days after induction, and SOX9 and type Ⅱ col agen mRNA was quantified with reverse transcription-polymerase chain reaction. The induced cel s were stained by immunofluorescence to observe the differentiation and perform statistical analysis.
RESULTS AND CONCLUSION:Passage 3 bone marrow mesenchymal stem cel s grew wel , and cel phenotype was confirmed as stem cel s by flow cytometry. The staining results showed that, the cel s differentiated into chondrocytes, adipocytes and osteoblasts. The SOX9 mRNA levels in the induced cel s were the highest in chondrogenic differentiation group, then in osteogenic differentiation group, and the lowest in adipogenic differentiation group. Type Ⅱ col agen mRNA levels in the induced cel s were the highest in chondrogenic differentiation group, then in adipogenic differentiation group, and the lowest in osteogenic differentiation group. SOX9 expression in chondrogenic differentiation group increased at 3 and 7 days, and then decreased at 14 days. While type Ⅱ col agen expression increased at 3, 7, 14 days. SOX9 mRNA levels increased as the osteogenic differentiation, while type Ⅱ col agen expression gradual y decreased. There was no significant difference in the SOX9 mRNA expression between adipogenic differentiation group and control group (P>0.05), while type Ⅱ col agen expression was not regularly changed. Experimental findings suggest that, critical effect of SOX9 in chondrogenic differentiation is better than that in osteogenic and adipogenic differentiation. SOX9 is associated with type Ⅱcol agen, which may alter along with the SOX9 in the early chondrogenic differentiation;SOX9 may play a fine-tuning role in the process of chondrogenic and osteogenic differentiation.

_ Objective_ To evaluate the efficacy of unilateral subtotal adrenalectomy in the treatment of bilateral adrenal solitary neoplasma causing Cushing's syndrome and to elaborate the therapeutic principle. Methods From 2007 to 2013, a total of ten patients were diagnosed with Cushing's syndrome caused by bilateral solitary adrenal neoplasma. We compared patients'clinical symptoms, hormone profiles, biochemical and metabolic parameters, and imaging data before and after the surgery. Five of them chose the optimal neoplasma based on the lateralization ratio of adrenal venous sampling result and the other 5 patients chose the optimal neoplasma based on the diameter of the mass reflected by the computed tomography result and were then operated. Results After the unilateral subtotal adrenalectomy,the24-hour urinary free cortisol decreased significantly(P<0.05)and the midnight serum cortisol level also significantly reduced(P<0. 01). Plasma adrenocorticotropic hormone level increased significantly(P<0. 01). Nine patients of them did not need contralateral adrenalectomy and one patient received contralateral adrelectomy because of the remnant of Cushingnoid symptoms. Conclusion Unilateral subtotal adrenalectomy is an effective and safe way to treat Cushing's syndrome caused by bilateral solitary neoplasma.

Objective To investigate the value of prospectively electrocardiogram (ECG)-triggered high-pitch coronary computed tomography angiography at 70 kV and 30 mL contrast medium with Stellar detector dual-source CT.Methods 60 patients with the BMI<24.9 kg/m2 were randomized into two groups:Group A (30 cases)with the tube voltage of 70 kV and 30 mL contrast medi-um.Group B (30 cases)with the tube voltage of 100 kV and 50 mL contrast medium.All the patients were performed with a Stellar detector dual-source CT using a prospectively ECG triggered high-pitch spiral technique.The images of Group A were reconstructed with SAFIRE technique,while Group B were with FBP technique.The objective image quality includes CT attenuation,SNR, CNR.And radiation dose was also calculated.The subjective was evaluated by 4-scale point.Independent sample t test was used.Re-sults There was no difference for subjective image quality between the groups,A:(3.38±0.942),B:(3.50±0.682),(t=-0.562, P >0.05).The CT attenuation of the 70 kV group were higher than that of the 100 kV group for all the segments(P <0.01).The SNR and the CNR have no statistically significant difference between the groups (P >0.05).Compared with the 100 kV group,the radiation dose of the 70 kV group was reduced by 76.5% (A:0.19±0.023 mSv,B:0.81±0.101 mSv,P <0.01).Conclusion Using 70 kV with 30 mL contrast medium in Stellar detector dual-source CT coronary angiography for the patients with a normal BMI could obtain qualified diagnostic image with low radiation dose and contrast medium.

Objective To study the effects of modified Danggui Beimu Kushen Pills on the expressions of TLR2, TLR4, TLR6, TRAF6 and MyD88 in tumor tissues on MFC gastric cancer bearing mice; To discuss relevant mechanism of action. Methods MFC gastric cancer bearing mice were employed to perform anti-tumor experiment in vivo in this study. A total of eligible 48 mice were randomly divided into model group, DDP positive control group, modified Danggui Beimu Kushen Pills high-dose and low-dose groups, modified Danggui Beimu Kushen Pills high-dose and low-dose combined with DDP groups. The treatment was conducted once a day, and lasted for 14 continuous days. After the last administration of gavage orally treatment, all mice were anaesthetized and killed by cervical dislocation method to obtain tumor tissue completely for further HE staining measure and detection of TLR2, TLR4, TLR6, TRAF6 and MyD88 in tumor tissue with the method of RT-qPCR and immunohistochemistry. Meanwhile, the tumor growth was observed and the general conditions of mice were recorded. Results The model group was rich in tumor cells; the sizes of cells were different; the volume was large; the nucleus was deeply stained and the heterotypic shape was obvious, and the small focal necrosis was seen. The number of tumor cells in each administration group was reduced; the arrangement was loose; the cell volume was significantly reduced, and the nuclear pyknosis was reduced. Cell necrosis significantly increased; the number of interstitial blood vessels decreased; collagen fibers increased, especially in modified Danggui Beimu Kushen Pills high-dose and low-dose combined with DDP groups. Compared with the model group, the expressions of TLR2, TLR4, TLR6, TRAF6 and MyD88 mRNA and protein decreased in each administration group. TLR2, TLR4, TLR6, TRAF6 and MyD88 were lighter and weakly positive expressed in modified Danggui Beimu Kushen Pillshigh-dose and low-dose combined with DDP groups, the protein changes were more obvious Compared with DDP positive control group, modified Danggui Beimu Kushen Pills high-dose and low-dose groups. Conclusion Modified Danggui Beimu Kushen Pills can down-regulate TLR2, TLR4, TLR6, TRAF6 and MyD88 expression of tumor tissue in MFC gastric cancer bearing mice at both mRNA and protein levels to play anti-tumor pharmacology action.

Objective To explore the relationship between interleukin(IL)-13 RS20541 gene polymorphism and susceptibility of enterovirus 71 (EV71) infection in children's hand-foot-mouth disease.Methods Blood samples were collected from 123 children with EV71 infection from the Central Hospital of Xiangtan (experimental group),and they were divided into mild EV71 infection group (n =62) and severe EV71 infection group (n =61) according to their severity.And 52 healthy children without EV71 infection were selected as the controls,with age and sex matched.Two mL blood samples stored in the-80 ℃ freezer,were collected from all subjects,and DNA was extracted by Beijing ubiolab genetic technology company limited.The SNaPshot was used to determine genotype for G/A polymorphism at RS20541 position of IL-13 gene.SPSS 18.0 software was used to analyze the data.Results IL-13 RS20541 loci had 3 genotypes:AA,GA,GG;the frequency of AA,GA,GG in the experimental group was 4.07％,44.71％,51.22％,which was significantly lower than that in the healthy control group (the frequency of AA,GA,GG were 11.54％,32.69％,55.77％),there was no statistically significant difference in genotypes (x2 =4.676,P ＞ 0.05);there was no statistically significant difference in allele frequency (the frequency of A,G in experimental group was 26.42 ％,73.58 ％,and that of the healthy control group was 27.88 ％,72.12 ％;x2 =0.080,P ＞ 0.05).EV71 infection caused by mild group,severe group and healthy controls genotype frequencies between the 3 groups [(AA + GA) were 53.22％,44.26％,44.23％;GG were 46.78％,55.74％,55.77％;x2 =1.294,P ＞ 0.05] and allele frequency (A were 30.65％,22.13％,27.88％;G were 69.35％,77.87％,72.12％;x2 =2.349,P ＞ 0.05) among the mild group,severe group and healthy control group had no statistical significance.Conclusion There is no correlation between the IL-13RS20541 gene polymorphism and EV 71 infection in children with the hand-foot-mouth disease.