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Objective To study the killing effect of γδT cells on different tumor cells by inducing and expanding γδT cells in vitro.Methods Collected the peripheral venous blood of 20 healthy volunteers recruited at Xinqiao Hospital of Third Military Medical University from January to March 2018.Peripheral blood mononuclear cells (PBMCs) were induced and expanded using zoledronic acid combined with interleukin-2 (IL-2) to obtain γδT cells.The cell purity was detected on day 0,7,10,14 and 16 using flow cytometry.The killing activity was detected by CCK-8 kit.Cells cultured on the 14th day were used as effector cells,and breast cancer cell BCap-37 and liver cancer cell Bel-7402 were used as target cells,and the cell killing activity was detected at the effective target ratio (E ∶ T) of 5 ∶ 1,10 ∶ 1 and 20 ∶ 1 respectively.Results The purity of γδT cells were respectively (3.35 ± 1.32) ％,(50.76 ± 5.76) ％,(80.43 ± 4.53) ％,(90.56 ± 3.34) ％,(89.54 ± 4.42) ％ on day 0,7,10,14,16,with significant difference (F =18.431,P =0.012).The efficiency of γδT cells killing BCap-37 tumor cells were (31.3 ± 2.0) ％ (E ∶ T =5 ∶ 1),(48.7 ± 1.6) ％(E ∶T=10∶1),(71.3 ±2.4)％ (E ∶T=20 ∶ 1) respectively,with significant difference (F=16.724,P =0.016),further comparison between each two groups:10 ∶ 1 vs.5 ∶ 1 (P =0.013),20 ∶ 1 vs.5 ∶ 1 (P =0.017),20∶1 vs.10 ∶1 (P =0.011).The killing rate increased with the increase of target ratio.The efficiency of γδT cells killing Bel-7402 tumor cells were (34.5 ± 2.0) ％ (E ∶ T =5 ∶ 1),(52.4 ± 1.9) ％(E ∶ T =10 ∶ 1),(74.5 ± 1.6) ％ (E ∶ T =20 ∶ 1) respectively,with significant difference (F =18.253,P=0.013),further comparison between each two groups:10 ∶ 1 vs.5 ∶ 1 (P=0.015),20 ∶ 1 vs.5 ∶ 1 (P=0.012),20 ∶ 1 vs.10 ∶ 1 (P =0.015).The killing rate increased with the increase of target ratio.Conclusion We can obtain high purity γδT cells using zoledronic acid combined with IL-2 induced PBMCs,and the amplified γδT cells have significant killing effect on BCap-37 and Bel-7402 tumor cells.
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Objective To explore the expression and significance of CD34, CD117 on bone marrow mononuclear cells of myelodysplastic syndromes (MDS). Methods Direct immunofluorescence staining was used by means of flow cytometry. 37 patients with MDS were divided into RA/RARS/RCMD subgroup, RAEB Ⅰ/RAEB Ⅱ subgroup; favorable chromosomal subgroup, poor chromosomal subgroup; intermediate-risk Ⅰ subgroup, intermediate-risk Ⅱ subgroup, high-risk subgroup respectively according to WHO classification,cytogenetic abnormalities and international prognostic scoring system (IPSS). Results CD34 and CD117 were positive respectively in 11 of 19 patients with RMRARS/RCMD, all cases in RAEB Ⅰ/RAEB Ⅱ expressed CD34 and CD117; increased expression of CD34 and CD117 was MDS grade-related. Favorable chromosomal subgroup, 14 of 22 patients were positive for CD34, CD117, all cases in poor chromosomes expressed CD34 and CD117; there was a direct relationship between phenotytic density and poor cytogenetic risk factor. CD34 and CD117 expression was present respectively in intermediate-risk Ⅰ (9/17), intermediate-risk Ⅱ (11/11) and highrisk subgroup (9/9); the phenotypic intensity also was correlated with IPSS scores. Conclusion Detection of CD34, CD117 may be a useful tool for subtyping and predicting the prognosis of MDS.
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Objective To examine the frequency and the course of cytomegalovims infection after allogeneic hematopoietic stem cell transplantation, and correlation of transplant factors with Cytomegalovirus (CMV) infection and viral load. Methods Using real-time polymerase chain reaction, we detected the copies of CMV-DNA in blood samples of the 62 patients after allo-HSCT. Furthermore, we studied the relationship between transplant factors and CMV infection. Results Among the total, 23 cases were contracted with CMV infection, 4 cases developed to CMV disease. 22 cases were cured and 1 case died. Course of CMV infection influenced the viral load significantly. Donor type, stem cell source, use of ATG, Ⅱ-Ⅳ grade aGVHD, use of glucocorticoid, complicating with other infection and use of cellular filter significantly influenced CMV infection. However, in multivariate analysis, none of them was the independent risk factors. Conclusion Real-time polymerase chain reaction may be used to early diagnose of the CMV infection and to guide treatment. Many factors influenced CMV infection. Early diagnosis and treatment could decrease the morbidity and mortality of CMV infection.
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BACKGROUND:ABO-incompatibility between donor and recipient is not a barrier for Successful allogeneic hematopoietic stem cell transplantation even though it is well established that major ABO incompatibility may lead to prolonged destruction of donor-derived erythrocytos and prolonged transfusioil requirements.OBJECTIVE:To explore the effect of ABO.incompatible on clinical characteristics in allogeneic-hematopoietic stem cell transplantation.DESIGN:A retrospective observation.SETTING:Department of Hematology.the Affiliated Drum Tower Hospital of Nanjing University Medical School.PARTICIPANTS:Fourteen patients(11 males and 3 feiliales,aged 15-60 years old)with malignant hematologic diseases who received ABO-incompatible allogeneic hematopoietic stem cell transplantation in the Affiliated Drum Tower Hospital of Nanjing University Medical School from May 2002 to September 2007 Were recruited for this study.Of the 14 patients,7 were human leukocyte antigen(HLA).matched,and the other 7 were HLA-half-matched.Controls were 11 patients who received ABO-compatibility bone marrow transplantation during the same period.Written informed consents for receiving allogeneic hematopoietic stem cell transplantation were obtained from each reciplent.The donors were sibling sister,sibling brother.son and mother,and they all agreed to provide marrow for transplantation.T1lis experiment was given an approval by the Ethics Committees of the hospital.METHODS:Regimen conditioning:HLA-matched transplantation regimen conditioning consisted of busulfan(Bu)and cyclophosphamide(Cy).HLA-half-matched transplantation regimen conditioning adopted GIAC program from Beijing People's Hospital.The GIAC program consisted of 4 parts:G:granulocyte colony-stimulating factors used for donors;I:stronger immunosuppressive regimen conditioning used for recipients;A: antihuman thymocyte globulin added:C: combined transplantation of bone marrow and peripheral blood;Perfusion of hematopoietic stem cells:The marrow from ABO-incompatible donor depleted erythrocytes by hydroxyethyl starch sedimentation.MAIN OUTCOME MEASURES:Adverse reaction.complication and hematologic recovery after ABO-incompatibility stem cell transplantation.RESULTS:One out of fourteen recipients developed pure red cell aplasia(PRCA)and dropped out of final analysis.Hematologic recovery:The median time of erythrocyte recovery after ABO-incompatible stem cell transplantation was delayed compared with ABO compatible stem cell transplantation (t=2.352.P＜0.05).There were no significant difieFences in the recovery of neutrophils and platelets between ABO-incompatible group and ABO-compatible group(P＞0.05).The median time of recovery of the erythrocyte and the blood type switching was delayed in HLA-mis-matched allogeneic hematopoietic stem cell transplantation compared with HLA-matched allogeneic hematopoietic stem cell transplantation,but without significant difference(P＞0.05).Complications:During the stem cell transfusion following transplantation.none of 14 Patieats had hemolytic complications or delayed haemolysis.CONCLUSION:There was no evidence of ABO-incompatibility between donor and recipient is a barrier for successful allogeneic hematopoietic stem cell transplantation.