Objective To explore the timing of wound closure and the choice of methods in Gustilo Ⅲtype tibiofibula open fractures after emergency debridement.Methods 42 patients with Gustilo Ⅲ type tibiofibula open fractures from January 2014 to January 2015 were selected as the observation group.All patients were performed emergency fracture fixation after debridement and wound secretion culture immediately after debridement, if the training result was positive,the debridement,again until the training results for negative when square in 48 hours after closing a wound.Selected 38 patients with GustiloⅢtype tibiofibula open fractures,the prompt debridement 38 cases were fracture fixed routine treatment after closing the wound,they were set as the control group.Compared two groups of patients with wound infection and fracture healing.Results The fractures wound infection rate of the observation group was 4.76%,which was significantly lower than 15.79% of the control group,the difference was statistically significant(χ2 =10.739,P=0.000).The wound healing time and fracture healing time of the observation group were significantly better than those of the control group,there were statistically significant differences( t=9.684,7.324,all P<0.05).Conclusion GustiloⅢtype emergency open tibiofibula fracture after debridement of the wound closure time should according to the results of wound secretion culture,cultivate the results can be achieved when the negative for wound closure,according to the degree of injury of soft tissue,to choose the right means of wound closure.

Objective To observe the expression of cytotoxicity and receptors on NK cells in breast cancer,and investigate the impact of soluble MICA(sohble MHC class Ⅰ-related molecules A,sMICA) on NK cells receptors expression and cytotoxicity.Methods ELISA was used to examine the sMICA in peripheral blood.The expressions of activated receptor(NKG2D),killer inhibitory receptor (KIR)(CD158b) and NK cells were identified by flow cytometry(FCM).Cytotoxicity of NK cells to breast cancer were tested by MTT.Results sMICA was (205.36±71.27)ng/L in breast cancer patients and 81.6 % samples were detected.There were positive correlations between sMICA levels with breast cancer stages.There were no difference of NK cells percentage between breast cancer and healthy person.The cytotoxicity of NK cells and expressions of NKG2D were obviously lower in breast cancer with sMICA(+) than in healthy person,but CD158b was higher in healthy person.After cultured with sMICA,NK cells cytotoxicity decreased from(76.2±6.7)% to(48.4±4.1)% and the expression of NKG2D reduced from(92.5±7.1)% to (62.5±6.4)%,but the the expression of CD158b increased from(10.6±3.2)% to (43.6±3.4)%.IL-15 up regulated the expression of NKG2D and NK cells cytotoxicity,but decreased the expression of CD158b by co-culturation with IL-15 and sMICA in sMICA+ patients with breast cancer.Conclusion sMICA reduced the expression of NKG2D and increased the expression of Kill,which lead to the down regulation of NK cells cytotoxicity.IL-15 can reverse this effect.

Objective To construct the tranfected cell expressing the human CXCR4 gene and to identify the effect on its immigration. Methods Total RNA was isolated from peripheral blood monouclear cell (PBMC),the full-length CXCR4 gene was amplified by RT-PCR and was inserted into plasmids PBudCE4.1 which have two promtors, after the identification by digestion and sequencing,the recombinant was transfected into K562 cell by lipofectamineTM2000. After screening culture by zeocin, stable transfected K562 cell line was established, and transcription and exression of CXCR4 were checked by flow cytometry; the chemotactic activity of K562 cell transfected and untrandfected CXCR4 was analyzed by Transell plate. Results The eukaryotic expression plasmid PBudCE4.1/ CXCR4 was constructed successfully. The stable trasfected K562/CXCR4 cell lines which highly express CXCR4 was established,the chemotactic activity of K562/CXCR4 was increased significiantly than K562. ConclusionCXCR4 transfected K562 cell line was successfully established, and it can make the basis for the further research on mechanism of extramedullary infiltration in leukemia.

Objective To assess the clinical short-term to mid-term efficacy of transcatheter closure of coronary artery fistula by using patent ductus arteriosus occluder in pediatric patients. Methods During the period from Jan. 2008 to May 2013 at authors’ hospital, transcatheter closure of coronary artery fistula using patent ductus arteriosus occluder was performed in 8 pediatric patients. The clinical data, including follow-up information, were retrospectively analyzed. Results A total of 8 pediatric patients with a mean age of (4.1 ± 3.8) years were enrolled in this study. The fistula originated from the right coronary artery in five cases and from the left coronary artery in three cases. The blood flow shunted to the right atrium (n=4) or to the right ventricle (n = 4). Obstruction of the fistula was successfully accomplished in all patients. All patients were followed up for (2.2 ± 1.2) years. No procedure-related complications or cardiac ischemia occurred. Conclusion For the treatment of coronary artery fistula in pediatric patients, the use of domestic patent ductus arteriosus occluder is safe and effective with satisfactory short-term to mid-term clinical efficacy.

Objective:To explore the role of NKG2D ligand MHC-I related molecule A (MICA) in chemotherapy combined with NK cell immunotherapy in patients with advanced esophageal cancer after surgery. Methods:A total of 90 patients with esophageal cancer from Fujian Provincial Tumor Hospital were divided into three groups after surgery:40 patients of chemotherapy alone, 25 patients of chemotherapy combined with NK cell therapy with negative expression of MICA (MICA-group), and 25 patients of chemotherapy combined with NK cells therapy with positive expression of MICA (MICA+group). The efficacy was then compared. Results:Compared with the chemotherapy alone and MICA-groups, the positive rates of CD3+, CD4+T cells, NK cells, and the CD4+/CD8+ratio in peripheral blood from MICA+group were higher than those before treatment (64.2%± 6.4%vs. 51.3%± 5.6%, 39.8%± 8.2%vs. 29.5%± 3.2%, 25.3%± 2.1%vs. 16.4%±4.3%, 1.4%± 0.5%vs. 1.1%± 0.7%;P<0.05). Meanwhile, the levels of T-reg cells were lower than those before treatment (6.3%± 4.5%vs. 17.3%± 2.4%, P<0.05). No significant difference was observed between the disease control rate and response rate. Chemotherapy-induced neutropenia and peripheral neurotoxicity symptoms were significantly improved, and time to progression (TTP) and overall survival (OS) were significantly prolonged (P<0.05). No statistically significant difference was observed between the chemotherapy alone group and MICA-group (P>0.05). Conclusion:Treatment with chemotherapy and autologous NK cells on patients with advanced esophageal carcinoma and MICA positive expression can be safely transfused with only minor side effects and can effectively improve a patient's immune system, quality of life, and survival.

Objective To explore the relevance of expression of MHC class Ⅰ-related molecules A (MICA) molecule and NK cells immunotherapy in esophageal cancer patients after operation.To analyze the significance of MICA expression in NK cell immunotherapy.Methods 100 patients of esophageal cancer were divided into 3 group,surgical alone group,MICA negative with NK therapy group (MICA-group) and MICA positive with NK therapy group (MICA+ group).The immunity indicators and tumor markers including the levels of CD3+,CD4+ T cells,ratio of CD4+/CD8+, NK cells,Treg cells,the levels of Th1/Th2/Th17 cytokine,the antibody IgA,IgM,IgG and the tumor markers of CEA,SCC,CA199,CYFRA21-1 were detected before treatment and after treatment 60 days.Results The positives rates of CD3+,CD4+ T cells,NK cells and the ratio of CD4+/CD8+ in peripheral blood from MICA+ patients group were higher than those of before treatment [(68.3±7.6) ％ vs (56.2±4.1) ％,(39.8±8.2) ％ vs (30.8±4.7) ％,(22.2±4.7) ％ vs (18.7±5.5) ％,(1.49±0.30) vs (1.15±0.61),P ＜ 0.05],meanwhile the levels of Treg cells was lower than those of before treatment [(8.1± 4.0) ％ vs (13.4±4.5) ％,P ＜ 0.05].There was no statistical significant difference of positive rate of CD8+ T cells [(26.9±6.2) ％ vs (27.8±7.1) ％,P ＞ 0.05].The levels of Th1 cytokin (IL-2,IFN-γand TNF-α) increased and Th2 cytokin (IL-4,IL-6 and IL-10) decreased after treatment.The level of Th17 cytokine was not different significantly (P ＞ 0.05).The content of IgA,IgM,IgG in MICA+ group were effectively improved after treatment.The tumor markers CEA,SCC,CA199,CYFRA21-1 had no statistically change before and after treatment.Conclusion The results indicate that NK cells immunotherapy can enhance the cellular immunity and humoral immunity of MICA positive esophageal cancer patients after operation.

Objectives To assess treatment outcomes and prognosis in infants with atrial flutter (AFL).Methods Thirty-four (34) cases of infants with AFL in Hunan Children's Hospital had been analyzed for clinical features, treatment outcomes and follow-up between March, 2009 and September, 2015. Based on ECG characteristics, the patients had been divided into simple and complex AFL groups. Based on age, they had been divided into neonates and non-neonates group. The aim of this study is to compare the clinical effects of drug treatment in different types of AFL.Resultsb With digitalis alone, the cardioversion rate was 37.5%,no signiflcant difference was observed between simple and complex AFL groups (45.8% vs 12.5%,P=0.206). Combining with other drugs, the cardioversion rate was 54.5%, which showed signiflcant difference between simple and complex AFL groups (76.9% vs 22.2%,P=0.036). The overall cardioversion rate was 70.6%, which showed signiflcant difference between simple and complex AFL groups(87.5% vs 30%,P=0.003). There was no signiflcant difference in pharmaceutical cardioversion rate between neonates and non-neonates group (85.7% vs 60.0%,P=0.216). Two cases with symptoms of heart failure used synchronized cardioversion. One patient restored to sinus rhythm, and another case was still recurrent of AFL after repeated electrical cardioversion, and eventually died of cardiogenic shock. After treatment, 9 patients were still with paroxysmal AFL and atrial tachycardia episodes, including 3 cases of simple type and 6 cases of complex type who were discharged with oral digoxin and propafenone treatment at home. 24 patients were followed up (3 months to 3 years and 4 months). 16 cases restored to sinus rhythm during hospitalization had no recurrence of AFL.Conclusions The overall treatment effects of AFL in infants were good. In simple type of AFL, most of patients did not need long-term antiarrhythmic drug therapy and the prognosis was good. The prognosis of treatment with conventional drug was poor in complex AFL group, with a higher rate of recurrence of AFL.

Objective:To investigate the cytotoxic effects of CTL cell induced by DCs loaded with exosomes derived from hepatoma Huh-7 cells(T-exo).Methods: Exosomes derived from hepatoma Huh-7 cells were isolated and purified by combination of ultrafiltration centrifugation and sucrose density gradient centrifugation.Morphology of exosomes was observed under transmission electron microscopy and the expression of CD 9,CD63,HSP70 and AFP was detected by Western blot.DCs were induced with peripheral blood monocytes isolated from healthy donors.Flow cytometry was used to analysis surface markers of the DCs loaded with T-exo.WST-1 light absorption measurement was adopted to evaluate the T cell proliferation ability.Annexin-V/PI Flow cytometry were respectively used to examined cytotoxicity against the tumor cells.Results:Exosomes isolated and extracted from culture supernatant of Huh-7 cells presented as circular or elliptical vesicle with bilayer membrane , unequal in size , and with diameter of 50 to 100 nm.Western blot showed that the T-exo expressed CD9,CD63,HSP70 and AFP molecules.DCs loaded with T-exo caused significantly higher T cell pro-liferation and cytotoxic effect against AFP positive Huh-7 cells as compare to gainst AFP negative SMMC 7721 cells and un-loaded control group ( P<0.05 ).Conclusion: T-exosome loaded-DC can promote proliferation and induce significant cytotoxic effect of CTL against Huh-7 cells.

Objective:To explore the association of microsatellite polymorphism of MICA gene with susceptibility to esophageal cancer.Methods: PCR-STR microsatellite genotyped technique was used to detect the polymorphism of MICA in Exon 5 in 103 cases of esophageal cancer and 84 cases of normal controls.Constructed of eukaryotic expression vector in esophageal carcinoma with high frequency of occurrence of the MICA allele.NK cells killing effect to 293T cells after alleles MICA transfected were assayed by LDH and the effect on target was 20∶1.ELISA was used to test supernatants sMICA of 293T cell after transfected.Results: Identified five allelic genes in MICA Exon 5 with esophageal cancer.Each allele and its frequency respectively were:MICA-A4(9.71%),MICA-A5(22.3%),MICA-A5.1(40.8%),MICA-A6(15.5%),MICA-A9(11.7%).MICA-A5.1 showed significant difference comparison with the control group.After 293T cell line was transfected MICA allele,MICA-A5.1 group was less sensitive to NK cytotoxicity compared to other groups[(30.4±6.3)%,P<0.05].The secretion of soluble MICA increased(135.7±6.2)pg/ml.Conclusion: Esophageal cancer was relevent with the MICA-A5.1 polymorphism of MICA Exon 5 alleles.Its risk is higher than other alleles.

Objective To screen serum biomarkers in patients with hepatocelhlar carcinoma by using surface-enhanced laser desorption and ionization time-of-flight mass spectrometry(SELDI-TOF-MS)technique and to evaluate its clinical implication in the patients whose alpha-fetoprotein were negative in the sera. Methods Proteomic spectra were generated by mass spectroscopy in 112 cases, including 57 cases AFP-negative hepatocellular carcinoma,and 55 cases of healthy control. The consequence was analyzed and the characteristic preteomic peaks was selected by using Biomarker Wizard. Results Seven low expressed potential biomarker were indentified with the mass-to-charge ratio of 4.2×103, 4.1×103, 6.7×103, 5.7×103, 6.5×103, 6.9×103, 5.8×103. The sensitivity for diagnosing hepatic cancer were 88.23 % and specificity was 92.31%. These peaks were not correlated with age, sex, tumor mass size, pathology grading and cirrhosis in hepatocellular carcinoma. Conclusion SELDI-TOF-MS offers a unique platform for the proteomic detection of hepatocellular carcinoma.It also offers an auxiliary diagnosis method to the patients whose alpha-fetoprotein are negative in the serum.