Objective To construct and identify lentiviral vectors carrying the connexin43 (CX43) gene. Methods Plasmids containing the CX43 gene and lentiviral vectors were digested using EcoRI/XbaI restriction enzymes, and the target gene fragments were cloned into the lentival vectors to result in CX43 recombinant lentiviral vectors (pHBLV-CMVIE-IRES-ZsGreen-CX43). CX43 recombinant lentiviral vectors were identified by restriction enzyme digestion and electrophoresis, and sequencing was carried on only for correct vectors after identification. The successfully-constructed CX43 recombinant lentiviral vectors and packaging plasmids were mixed and contransfected into 293FT cell for packaging and producting virus. Then the virus was collecting, concentrated and titrated in 293FT cells. Finally, the expression of CX43 gene was assessed by real-time polymerase chain reaction(PCR). Results The restriction enzyme digestion and electrophoresis and sequencing results proved that CX43 recombinant lentiviral vectors were constructed correctly. Lentiviral concentrated virus suspension titer was 1× 108/ml. The up-regulation expression of CX43 was detected correctly by real-time PCR in 293FT cells. Conclusions Stable lentiviral vectors expressing CX43 gene is constructed successfully.

Objective To investigate the gene expression of connexin43(Cx43) and its effect on gap junction intercellular communication(GJIC) of acute leukemia bone marrow stromal cells(ALBMSCs).Methods After ALBMSCs were transfected by recombinant adenovirus Ad-Cx43-GFP,the expression of report gene GFP and the transfection efficiency were monitored by fluorescent microscopy.RT-PCR,Western blot and immunocytochemical method were used to detect the mRNA and protein expressions of Cx43.Dye transfer procedure was performed to examine the GJIC function.Results After transfected by Ad-Cx43-GFP for 24 h,the expression of GFP in ALBMSCs was detected by fluorescent microscopy and the transfection efficiency was(82.7?2.16)%;The mRNA and protein expressions of Cx43 in ALBMSCs transfected by Ad-Cx43-GFP were higher than those not transfected by Ad-Cx43-GFP(P

Objective: To investigate the safety and efficacy of haploidentical hematopoietic stem cell transplantation with different intensity conditioning regimen in the treatment of childhood aplastic anemia (AA) . Methods: Thirty-seven AA patients who underwent haploidentical transplantation in BaYi Children's Hospital Affiliated to PLA Army General Hospital from January 2013 to January 2017 were enrolled. According to the dosage of conditioning regimen, 34 patients excluding 3 other conditioning regimens were divided into high-dosage group (regimen 2, 22 cases) and low-dosage group (regimen 3, 12 cases). The data of Engraftment, graft-vs-host disease (GVHD), hematopoietic reconstitution, relapse, infection, overall survival (OS) were analyzed. The comparison between the two groups was tested by χ² test. Results: A total of 35 of 37 patients achieved primary engraftment; 2 cases died of regimen-related toxicity and severe infection before the infusing of the grafts. The activation rate of CMV and EBV was 60% (21/35) . Post-transplant lymphocyte disease (PTLD) of lung occurred in one case. The cumulative incidences of acute GVHD grade Ⅰ-Ⅳ and chronic GVHD were 29% (10/35) and 34% (12/35) respectively and the incidence of extensive chronic GVHD was 6% (2/35) . The median follow-up time was 18.8 (2.9-44.1) months, the OS was 92% (34/37) .All survived patients were no longer dependent on blood transfusion and none of them had recurrence. Comparing the rates of overall survival(86%(19/22) vs.100%(12/12)) and rates of chronic GVHD(40%(8/20) vs. 17%(2/12)) in regimen 2 and regimen 3 group, there were no significant difference (χ²=1.742, 1.841, all P>0.05) . Significant difference was found at the incidence of Ⅰ-Ⅳ acute GVHD (10% (2/20) vs. 50% (6/12) ,χ²=6.200, P=0.013). Conclusions Haploidentical hematopoietic stem cell transplantation is effective and safe. It is suitable for patients who are not eligible for matched donor transplantation. Application of reduced dose preconditioning in haploid transplantation is worth exploring.