BACKGROUND:Because of their immunological properties, bone marrow mesenchymal stem cells transfusion is developed as a new treatment for refractory graft-versus-host disease. OBJECTIVE:To analyze the safety and curative effect of bone marrow mesenchymal stem cells transfusion on treating different organ damages in graft-versus-host disease after al ogeneic hematopoietic stem celltransplantation. METHODS:Eight patients with malignant hematologic disease were included in this study. The patients developed severe steroid-resistant graft-versus-host disease after al ogeneic hematopoietic stem celltransplantation and received transfusion of mesenchymal stme cell(1×106 of immunosuppressive agent. RESULTS AND CONCLUSION:For the total y eight patients, six got response (two cases of complete remission, and four cases of partial remission) and two showed no remission. Four of five cutaneous damages were ameliorated and one showed no effect. For three cases of oral graft-versus-host disease, two acquired complete remission and one showed partial remission. Two cases of liver graft-versus-host disease and two cases of astro-intestinal graft-versus-host disease obtained complete remission. No response was displayed to three cases of ocular graft-versus-host disease, one case of bronchiolitis obliterans, and one case of urinary graft-versus-host disease. In the median fol ow-up of 28 months (7-62 months), three patients developed posttransplant lymphoproliferative disorders within 3 months after mesenchymal stem cells transfusion. Administration of mesenchymal stem cells is safe for treatment of severe graft-versus-host disease after al ogeneic hematopoietic stem celltransplantation. Mesenchymal stem cells transfusion may be a promising/kg) together with the primary therapy therapy for refractory cutaneous , astro-intestinal, liver and oral graft-versus-host disease but not for pulmonary, ocular and urinary graft-versus-host disease. Whether mesenchymal stem cells transfusion is associated with posttransplant lymphoproliferative disorders needs more case data.

Objective To investigate the influence of the PBSC collection yield by choosing the differ-era venous accesses in the healthy donors. Methods 118 healthy PBSC donors performing PBSC collection between January 2000 and December 2007 in our hospital were divided into four groups according to the differ-ent venous accesses. The PBSC collection yield of four groups,including mononuclear cells (MNC) count and CD34+ cells count were observed. Results In the ulnar V-ulnar V group,MNC (5.31±2.29)×108/kg,CD34+ cells (4.78±2.06)×106/kg;ulnar V- antecubital V group,MNC(5.11±2.34)×108/kg,CD34+cells(4.34±1.99)×106/kg;antecubital V- antecubital V group,MNC (5.61±1.73)±×108/kg,CD34+cells (4.60±1.42)×106/kg;ulnar V- radial V group,MNC(4.60±×1.70)×108/kg,CD34+cells (4.05±1.50)×106/kg.There was no statistical differ-ence of the PBSC collection yield between four groups (P>0.05). Conclusions Different venous accesses don't affect the PBSC collection yield in the PBSC healthy donors.

Objective To investigate the influences of cytarabine on Survivin gene expression in human leukemia K562 cell line and discuss the mechanism of drug resistance in chemotherapy. Methods The IC50 of cytarabine was chosen by MTr assay. The K562 cells were exposed to certain concentration of cytarabine for about 24 hours and 48 hours. The expression levels of Survivin gene were detected by RT-PCR and Western-blot. Results After exposed to cytarabine for 24 hours and 48 hours, the Survivin mRNA level of K562 cells was significantly elevated about 1.92 and 3.38-fold, and the protein expression level was elevated about 1.92 and 2.64-fold. Conclusion An elevated expression of Survivin was tested in K562 cells treated by cytarabine. Consequently, the elevated expression of Survivin could resist apoptosis induced by chemotherapeutic agent which probably associated with chemotherapeutic drug resistance.

Lidamycin (LDM) is a potent antitumor antibiotic. Previous studies have shown that LDM could inhibit proliferation and migration in endothelial cells. In the present report, the effect of LDM on angiogenesis of zebrafish embryo was studied. The results showed that treatment of zebrafish embryos with LDM resulted in significant inhibition of angiogenesis. Morphological observation, quantitative endogenous alkaline phosphatase (EAP) assay, alkaline phosphatase staining, and transgenic zebrafish assay were performed to evaluate vascular development defects in zebrafish. The results indicated that after the zebrafish embryos were exposed to LDM, angiogenesis defects of zebrafish embryos were observed, including pericardial edema, reduced numbers of circulating red blood cells, suppression of zebrafish vessel growth, and absences of SIV (subintestinal vein). The expression of VEGF was detected by RT-PCR assay, quantitative reverse transcriptase real-time PCR (qRT-PCR) assay and Western blotting analysis. The results revealed that LDM could inhibit the expression of VEGF protein, while the expression of mRNA was not significantly affected. The study suggests that LDM could inhibit the zebrafish embryo angiogenesis by down-regulation ofVEGF expression.

Objective To investigate the relationship of plasma concentration of NT-proBNP,copeptin and glasgow coma scale(GCS)scores,hematoma volumes in patients with acute cerebral hemorrhage.Methods 109 patients with acute cerebral hem-orrhage(the cerebral hemorrhage group)and 32 healthy individuals (the control group)admitted in our hospital from December 2011 to June 2013 were selected and detected for plasma NT-proBNP and copeptin.The levels of NT-proBNP,copeptin,glasgow co-ma scale(GCS)scores and hematoma volumes were compared between the two groups.Results The levels of plasma NT-proBNP and copeptin in the cerebral hemorrhage group were significantly higher than that in control group(P <0.05).The levels of plasma NT-proBNP and copeptin were significantly increased with the severity and the hematoma volume of the acute cerebral hemorrhage. The levels of NT-proBNP and copeptin are positively correlated with hematoma volumes(r=0.63,r=0.58,P <0.01)and negative-ly correlated with Glasgow Coma Scale(GCS)scores(r=-0.52,r=-0.46,P <0.01).Conclusion The levels of NT-proBNP and copeptin are positively correlated with hematoma volumes and negatively correlated with glasgow coma scale(GCS)scores.They are important clinical parameters to reflect the severity and hematoma volumes of the acute cerebral hemorrhage.

Objective:To prepare and screen monoclonal antibodies against Herpes simplex virus-1(HSV-1),and develop a double antibody sandwich quantitative enzyme-linked immunosorbent assay( Q-ELISA) for detection of HSV-1 particle. This method was used to control the quality of viral particle in the developing and manufacturing process of HSV-1. Methods: BALB/c mice was immunized with HSV-1 to prepare monoclonal antibodies. A double antibody sandwich Q-ELISA was developed to determine concentration of HSV-1 particle,which was based on the neutralizing monoclonal antibody 1F6 as capture antibody,and 2B1 as HRP-conjugated antibody. The performance of the reagent was evaluated,including specificity,sensitivity,precision,accuracy and linear. And the relation between the amount of virus detected by this method and the virus titer was analyzed by regression analysis method. Results: The Q-ELISA for HSV-1 particle was developed. The quantitation scope was 0. 125-2 μg/ml, the coefficient correlation was 0. 995 5, the limit of detection was 0. 125 μg/ml, the recovery was between 85. 6% and 107. 1%, the variation coefficient was lower than 10%, and the reagent does not react with other samples except HSV-1 antigen. This method has a good correlation with virus titer. Conclusion:The Q-ELISA for HSV-1 particle was successfully developed,which provide a new approach for rapid and quantitative detection of HSV-1 antigen.

AIM:To investigate the effects of celecoxib on viability , apoptosis and autophagy in acute myeloid leukemia (AML) cell lines HL-60 and HL-60A.METHODS:The HL-60 cells and HL-60A cells were cultured with vari-ous concentrations (0, 20, 40, 60, 80 and 100μmol/L) of celecoxib.The inhibitory effect of celecoxib on the cell viabil-ity was evaluated by MTT assay .Apoptosis was analyzed by Annexin-V/PI staining.Apoptosis-related and autophagy-relat-ed proteins were determined by Western blot .RESULTS:IC50 of celecoxib were 49.4 μmol/L, 32.0 μmol/L and 25.1μmol/L for HL-60 cells treated with celecoxib for 24 h, 48 h and 72 h, respectively.For HL-60A cells, the corresponding IC50 were 69.1 μmol/L, 42.5 μmol/L and 29.6 μmol/L, respectively.The results of flow cytometry analysis showed the proportions of Annexin-Ⅴ+PI-, Annexin-Ⅴ+PI+and Annexin-Ⅴ-PI+cells were increased in the HL-60 cells, and those of Annexin-Ⅴ+PI-and Annexin-Ⅴ+PI+cells were increased in the HL-60A cells treated with celecoxib for 24 h. After treated with celecoxib , the induction of apoptosis was observed , the apoptosis-related proteins cleaved caspase-3 and cleaved PARP were upregulated , the autophagy-related proteins LC3 II and P62 were both increased , and mTOR, p-mTOR, 4-EBP and p-4-EBP were not changed , indicating that celecoxib inhibited autophagy in the AML cells without the mTOR pathway involvement .CONCLUSION:Celecoxib inhibits the viability of HL-60 cells and HL-60A cells in a time-and dose-dependent manner by its effects of inducing apoptosis and necrosis .Celecoxib inhibits mTOR-independent autoph-agy in AML cells, indicating a possible way of using celecoxib for enhancing the antitumor activity of therapeutic agents to induce cytoprotective autophagy in the AML cells .

AIM:To construct AFT024-SCF cell line and HPC-Lhx2 cell line for confirming the biological function of AFT024-SCF.METHODS:The HPC-Lhx2 cell line, AFT024-SCF cell line and AFT024-GFP cell line were constructed by retro-viral infection.The expression of stem cell factor(SCF) in AFT024-SCF cells was detected by real-time PCR and Western blot.SCF in the supernatant of AFT024-SCF was detected with ELISA.The supernatant of AFT024-SCF and AFT024-GFP were collected and then diluted (1:10) with basic IMDM medium.So we made 4 culture medium:AFT024-SCF medium was used for experiment group, AFT024-GFP medium was used for endogenous negative control, IM-DM basic medium was used for exogenous negative control, and IMDM basic medium with SCF was used for positive con-trol.SCF-dependent HPC-Lhx2 cell line was cultured in these 4 different medium for 72 h.According to MTT method and colony forming experiment, the biological function of AFT024-SCF was confirmed by the proliferation ability of SCF-depend-ent HPC-Lhx2 cell line.RESULTS:SCF was highly expressed in AFT024-SCF cells.After cultured for 72 h, neither IM-DM basic medium nor GFP-AFT024 medium support HPC-Lhx2 cell line proliferation.However, AFT024-SCF medium supported HPC-Lhx2 cell line expansion as well as the positive control medium.CONCLUSION:AFT024-SCF cells ex-press SCF successfully and recombinant SCF can be replaced by the supernatant of AFT024-SCF culture medium for expan-ding HPC-Lhx2 cell line in vitro.

Objective To investigate that the relationship of serum concentration of copeptin ,procalcitonin(PCT )and early diagnosis ,prognosis in patients with cerebral hemorrhage‐associated pulmonary infection .Methods One hundred and twenty pa‐tients with acute cerebral hemorrhage ,acute cerebral hemorrhage‐associated pulmonary infection and 60 healthy individuals (the control group) admitted in our hospital from January 2012 to December 2013 were selected and detected for serum copeptin and procalcitonin .The differences of serum copeptin ,procalcitonin levels were compared in controls ,in patients with cerebral hemor‐rhage and cerebral hemorrhage‐associated pulmonary infection and their correlation was analyzed .Results The levels of serum copeptin in the cerebral hemorrhage‐associated pulmonary infection were significantly higher than that in control group and the cer‐ebral hemorrhage (P<0 .05) .The levels of serum procalcitonin in control group and the cerebral hemorrhage were significantly lower than that in the cerebral hemorrhage‐associated pulmonary infection ,the levels of serum C‐reactive protein ,copeptin ,procalci‐tonin and the APACHEⅡ scores of the patients with survival group were significantly lower than those with non‐survival group (P<0 .05) .Conclusion The levels of serum copeptin and procalcitonin are correlated intimately with cerebral hemorrhage‐associat‐ed pulmonary infection .They are important clinical parameters to reflect the early diagnosis and prognosis of cerebral hemorrhage‐associated pulmonary infection .

Objective To evaluate the value of neck blood vessel colored doppler ultrasound (NBVCDU) and magnetic resonance angiography (MRA) to extracranial carotid artery stenosis in patients with transient ischemic attack (TIA).Methods After implementing NBVCDU and MRA examinations at the same time,45 TIA patients with at least one examination showing arteriostenosis in extracranial section were chosen to carry out cerebral digital subtraction angiography( DSA ),then the stenosis rate was calculated by American symptomatic carotid endarterectomy trial (NASCET) method.Results Regarding DSA as the gold standard,for 45 TIA patients that having 180 arteriostenosis in extracranial section, sensitivity,specificity,accuracy of NBVCDU examination was 93.51% ,95.15% ,94.44%, Kappa = 0.735; sensitivity,specificity,accuracy of MRA was 92.21% ,94.17% ,93.33% , Kappa =0.681; sensitivity,specificity,accuracy of NBVCDU combined with MPA was 97.40% ,99.03% ,98.33%, Kappa = 0.872.Conclusions The sensitivity and accuracy of arteriostenosis in extracranial section by NBVCDU examination is higher than that by MRA, and it is suitable in the crowd primary examination.NBVCDU combined with MRA has shown good consistence with DSA for diagnosing arteriostenosis in extracranial section,but can't replace DSA comlpetely.