Objective To investigate human immunodeficiency virus type 1 (HIV 1)specific CD8'T cell responses and related human leukocyte antigenⅠ(HLA-I)restriction sites.Methods Four Epstein-Barr virus (EBV)strains-transformed B-lymphoblastoid cell lines (BLCL)from periph eral blood monocytes (PBMC)of 4 healthy individuals were established.Using synthesized peptides in HIV-1 Gag,Vif and Nef regions as epitopes and using BLCL as antigen presenting cells (APC), the frequency of interferon-7(IFN-?)secreting cells in CD8 T cells from a HIV-infected long term nonprogressor by enzyme-linked immunospot (ELISPOT)assay in vitro were assessed.Moreover, matching analysis of HI.A-Ⅰrestriction sites were performed.Results HIV-1 epitopes were presen ted restrictively by HLA-Ⅰmolecules in specific CD8 T cell responses,and this antigen presenting had high specificity.HLA-Ⅰrestriction sitesA03,A26,C04andC08 contributed to presentepitopes in HIV-1 C-ag region,A03 and C08 to HIV-1 Nef as well.Conclusion HLA-Ⅰrestriction sites may include A03,A26,C04 and C08 in HIV-1 Gag region,and A03,C08 in Nef region.

OBJECTIVETo investigate immunodominance in CD8+ T cell responses to human immunodeficiency virus type 1 (HIV-1) epitopes.

METHODSFrequency of Interferon-gamma (IFN-gamma) secreting cells and the proliferation percentage of CD8+ T cells in PBMC from an HIV-1-infected long term nonprogressor (LTNP) were assessed after stimulation with either the 34 pools of 701 overlapping peptides covering the regions of HIV-1 Env, Pol, Gag, Vif, Nef, Tat or some single peptides, by using various assays including enzyme-linked immunospot (ELISPOT) and CFSE Carboxy-fluorescein diacetate, succinimidyl ester (CFSE) labeling and flow cytometry.

RESULTSHIV-1 Gag peptides induced the highest frequency of IFN-gamma secreting cells, followed by Nef, Tat, and Vif. Meanwhile, Env and Pol failed to induce significant responses. In the IFN-gamma ELISPOT assay, stimulation with single peptide and the corresponsive peptide pool generated analogous results. In addition, the frequencies of IFN-gamma secreting cells and the proliferation percentage of CD8+ T cells detected-ELISPOT and CFSE labeling and flow cytometry were proportional, when single peptides were used for stimulation.

CONCLUSIONCD8+ T cells can respond to some specific HIV-1 epitopes and induce immunodominant responses. As a complimentary approach to the standard of ELISPOT assay, We recommend a novel CFSE labeling and flow cytometry assay for the examination of immunodominance in studies of HIV-1 specific proliferation percentage of CD8+ T cell responses.

Adult ; CD8-Positive T-Lymphocytes ; drug effects ; immunology ; metabolism ; Enzyme-Linked Immunosorbent Assay ; methods ; Female ; HIV Infections ; immunology ; virology ; HIV-1 ; immunology ; Humans ; Immunodominant Epitopes ; immunology ; pharmacology ; Interferon-gamma ; metabolism

Objective:To evaluate the clinical efficacy of SKy bone expander system in percutaneous kyphoplasty for treat- ment of osteoporotie verterbral compression fracture.Methods:Twenty-two patients(aged 62-90 years,32 vertebrae)under- went percutaneous kyphoplasty using SKy bone expander system.The bone cement was injected into the collapsed vertebrae. The vasual analogue scale(VAS)and complications were recorded during follow up.Results:The operations were successful in all patients via unilateral or bilateral approach.The operation time ranged from 30 to 120 min.The mean volume of cement in- jected into each vertebra body was(4.8?1.1)ml,ranged from 3.1 to 6.8 ml.Extravertebral leakage of bone cement was ob- served in two vertebrae with no symptoms.All patients had their pain relieved;the VAS was 7.6?0.8 before operation,3.5?0.5 one day after operation,2.8?0.6 one week after operation,and 2.4?0.6 one month after operation,with significant difference found between preoperation and postoperation(P

Objective To report 11 cases with the tissue defects of their upper or lower limbs repaired with the anastomsed medial sural artery perforator flaps.Methods The free medial sural artery perforator flaps,with length of 8 cm to 15 cm and width of 6 cm to 14 cm were used for tissue defect reconstruction of the distal upper or lower limbs in 11 cases,including 6 females and 5 males.The flap was harvested from the ho- mo-lateral calf,confined between the posterior-medial edge of the tibia and the middle line of the calf and a- bove the distal half part of the medial sural muscle,with a same axis of this muscle.Results Ten cases survived very well,which was relatively thin,and the donor site can be acceptable.One case resulted in a complete flap necrotized and covered with a split skin graft.No obvious motor function defect was observed of the donor leg.Conclusion The anastomsed medial sural artery perforator flap is alternative donor flap for the upper or lower limb tissue defect repair,especially for the defect in the hand or foot.

Objective To study the inhibitory effects of δ-opioid receptor activation in serumdeprivation induced apoptosis of human liver cells and the proposed protein kinase C(PKC)pathway mechanism.Methods MTT assay was used to detect the survival rate of human liver cells in vitro and Annexin V-FITC/PI double staining was used to detect the cell apoptosis rate.Flow cytometry was used to analyze cell cycle,RT PCR used to analyze the PKC mRNA and Western Blot analysis was used for detecting the protein expression of PKC and Caspase-3.Results After serum-deprivation for 48h of cultured human liver cells in vitro,significant liver cell apoptosis occurred.The apoptosis was suppressed by δ-opioid receptor activation,which manifested as a slower rate of apoptosis,decreased expression of Caspase-3and increased expression of PKC.After GF109203X was added,the inhibitory effects of DADLE decreased markedly.Conclusion Activation of δ-opioid receptor on the membrane of human liver cells has inhibitory effects on serum-deprivation induced apoptosis of liver cells.The underlying mechanism may be associated with PKC pathway activation.

Objective To investigate the changes of hypoxia-inducible factor(HIF)-1α and hexokinase-Ⅱ(HK-Ⅱ)expression in human esophageal squamous cell carcinoma and its effect in glycolysis.Methods TE13 cells and Eca109 cells were cultured under hypoxic condition(1 ％O2)for different hypoxic time(6,12,24 and 48 hours).Cells cultured under normal oxygen condition(20％O2)were set as control.The changes of HIF-1α and HK-Ⅱ expressions at protein level were detected by Western blot.HIF-1α genes were specifically silenced with RNA interference technology(RNAi),and then the changes of HIF-1α and HK-Ⅱ expression were determined by realtime PCR and Western blot.Under normal oxygen and hypoxic condition,the changes of lactic acid concentration in cell culture medium were detected by spectrophotometric method.Results Under hypoxic condition,the expression of HIF-1α and HK-Ⅱ gradually increased as hypoxic time extended(P＜0.05),reached a peak at 12h and then gradually decreased as time extended.Compared with that under normal oxygen condition,the expression of HK-Ⅱ in TE13 cells and Eca109 cells significantly increased under hypoxic condition(P＜0.05),which was more significant after 12 hours hypoxia.The result of realtime PCR indicated that under normal oxygen condition the expression of HIF-1α at RNA level in TE13/shRNA cells and Eca109/shRNA cells significantly decreased compared with TE13 cells and Eca109 cells without interference(P＜0.05).The expression of HK-Ⅱ at RNA level was consistent with the result of HIF-1α.Under normal and hypoxia condition,the expression of HK-Ⅱ at protein level in TE13/shRNA cells and Eca109/shRNA cells significantly decreased compared with TE13 cells and Eca109 cells without interference,and the difference was statistic significant(P＜0.05).The lactic acid secretion of TE13 cells and Eca109 cells under hypoxia condition(14.707 ± 3.594 and 15.062 ±3.901)was higher than that under normal oxygen condition(6.070±1.839 and 6.891±1.592,P＜0.05).The lactic acid secretion of TE13/shRNA cells and Eca109/shRNA cells significantly decreased compared with TE13 cells and Eca109 cells without interference,and the difference was statistic significant(P＜0.05).Conclusion The expressions of HIF-1α and HK-Ⅱ in human esophageal squamous cell carcinoma significantly increased under hypoxia conditions.The expression of HK-Ⅱ is closely correlated with lactic acid concentration and HIF-1α expression.HIF-1α may affect cell glycolysis through HK-Ⅱ.

Objective To evaluate the efficacy of preoperative regional intra-arterial chemotherapy (RIAC) in the treatment of resectable pancreatic head carcinoma. Methods The clinical data of 50 patients with resectable pancreatic head carcinoma who had been admitted to the Research Institute of Pancreatic Diseases of Fudan University from December 2006 to July 2007 were retrospectively analyzed. Patients were randomly divided into2 groups (n =25 in each group): patients in group A were treated with preoperative RIAC followed by regional pancreaticoduodenectomy, and patients in group B were treated with surgical procedure routinely. The lymphatic metastases in the 50 specimens of pancreatic head carcinoma were detected by histological examination with hematoxylin and eosin (HE) staining, and lymphatic micrometastases were detected by immunohistochemical method with staining of cytokeratin AE1/AE3 in 10 specimens with negative HE staining of the lymph nodes in each group. Results There was no significant difference in the incidence of complications, the length of hospital stay and the 1-, 2-year survival rates between the 2 groups (χ2 = 0.12, 2.88, P > 0.05). The incidence of positive lymph node metastasis in group A was 7.1% (52/734), which was significantly higher than 22.1% (118/532) in group B (χ2 = 60.01, P < 0.05). The incidence of lymphatic micrometastasis was 9.4% (30/319) in group A, and 9.1% (23/252) in group B, with no statistical difference between the 2 groups (χ2= 0.01, P > 0.05). Conclusions Preoperative RIAC is helpful in improving the prognosis of patients with resectable pancreatic head carcinoma by reducing the incidence of lymphatic metastasis and decreasing tumor stage.

Objective To summarize the clinical efficacy of free KISS lateral femoral circumflex artery perforator flap in repairing defects of limbs.Methods Twelve cases were suffered wound surface defects on hands and feet.And the defects were repaired by flap construction from October,2010 to May,2013,based on the characteristics of the defects combined with anatomical features of the free lateral femoral circumflex artery perforator flap.Length of flap was adopted as the width for direct suture in the flap donor.Results Postoperative flap and donor area preliminarily healed.There was no vascular crisis.Twelve cases received 6-18 months followed-up (averaged of 12 months).The skin flap was good in color and texture.The dorsal flap was a bit bloated.Linear scar was remained in distal flap donor area.The quadriceps muscle power level 5,knee flexion,extension 10°-180°.Quadriceps strength,knee flexion and stretch activities were all normal.The flaps recovered protective sense.Four cases had tendon adhesion after hand tendon transplantation.The finger function was well recovered after release.At the last followup,the functions of the upper limbs were evaluated according to the trial evaluation standard of the Hand Surgery Association of Chinese Medical Association:6 cases were excellent,1 case was good,and 1 case was qualified.Conclusion The design of the lobulated tissue flap of the lateral femoral circumflex artery descending branch is flexible.Large area of the surface defect can be repaired.The flap donor area is directly sutured.It is an ideal method to repair the wound tissue defect.

Objective To observe the PML protein expression of hepatocellar carcinoma tissue and cells lines and As 2 O3 regulate its expression .Methods Immunohistochemistry was used to examine the PML protein expression of hepatocellar carcinoma tissue . Western blot analysis were used to observe PML protein expression of hepatocellar carcinoma tissue of 12 cases ,5 hepatocellar car-cinoma cell lines ,such as HuH7 ,HepG2 ,Hep3B ,SMMC-7721 ,MHC97H .Western blot analysis was used to detected the PML pro-tein expression of these hepatocellar carcinoma cell lines after 72-96 h treated with 0 .25 μg/mL of As2 O3 .Results Immunohisch-enmical staining showed that the PML protein was expressed in both cytoplasm and nucleus ,did not well-distributed in hepatocellar carcinoma cells .There was no significant differences of PML protein expressed among differently differentiated stages of hepatocel-lar carcinoma cells .Western blot analysis found that hepatocellar carcinoma tissues of 12 cases with hepatocellar carcinoma ex-pressed PML protein ,and there was significant difference of PML protein expressed among 12 cases suffer with hepatocellar carci-noma .hepatocellar carcinoma cell lines ,such as HuH7 ,HepG2 ,Hep3B ,SMMC-7721 and MHC97H all expressed PML protein ,and there was little difference of PML protein expressed among hepatocellar carcinoma cell lines .The PML protein expression of HuH7 ,HepG2 ,Hep3B ,SMMC-7721 and MHC97H cell after 72-96 h treated with 0 .25 μg/mL of As2O3 significant decreased . Conclusion Hepatocellar carcinoma tissue and cells may express PML protein ,and As2 O3 may regulate this protein expression as well .PML protein may be the target molecule of As2 O3 treating HCC .

Objective This study aimed to investigate whether platelet-derived growth factor CC could enhance resolution ,or-ganization and recanalization of venous thrombus .Methods The experimental models of deep vein thrombosis in rats were induced as previously described and modified .The rats were randomly divided into four groups according to the dose of PDGF-CC(Group A :0 ng/100 μL ;Group B :100 ng/100 μL ;Group C :200 ng/100 μL and Group D :500 ng/100 μL) .The samples were obtained at 7 days after operation and the expressions of protein VEGF were detected by Western blot analysis ,respectively .Immunohistochemi-cal staining was used to observe the types of the positive cells expressing vWF in thrombus sections .HE staining and Masson stai-ning were used to analyze the area of resolution and organization in venous thrombi .The capillary number was analyzed quantitative-ly by high-power microscope .The area of thrombus and collagen fiber in each section were measured and analyzed with image analy-sis software .Results Animal models of venous thrombogenesis were performed successfully .The data of Western blot analysis demonstrated that the expressions of VEGF protein were significantly increased in group C and D than group B and A ,which were no statistical significance between group C and group D at day 7 after operation .The positive cells of expressing vWF mostly located in the thrombus border and venous wall and the types of the positive cells by immunohistochemical staining were ECs .The resolu-tion rates ,organization rates and recanalization capillary numbers were significantly higher in group C and D than those in group B and A ,and those in group B were higher than those in group A .Conclusion PDGF-CC could enhance resolution ,organization and recanalization of vein thrombi effectively through therapeutic angiogenesis pathway ,which provides a novel strategy for gene thera-py of venous thromboembolism disease .