Safety and effectiveness of tumor-ablative chemotherapy combined with low intensity modified conditioning regimen for 30 patients with hematologic malignancies receiving allogeneic hematopoietic stem cell transplantation / 中国实验血液学杂志

This study was aimed to investigate the safety and effectiveness of tumor-ablative Chemotherapy combined with low intensity conditioning regiment BUCy/TBICy for patients with hematologic malignancies receiving allogeneic hematopoietic stem cell transplantation (allo-HSCT). The clinical data of 30 patients with hematologic malignancies received above-mentioned therapeutic method from January 2012 to January 2013 was analyzed retrospectively, and the engraftment, GVHD, infection, conditioning-related toxicity, relapse and survival rates were evaluated. All the patients signed the informed consent before transplantation. The median follow-up duration was 20.5 (16.3-27.3) months. The results indicated that all the patients had been engrafted successfully. One year overall survival (OS) and disease-free survival (DFS) rates were 93.3% and 83.3% respectively. No conditioning-related toxicity occurred. The incidences of II-IV grade aGVHD was 37.9%, among which incidence of III-IV grade aGVHD was 3.4%; incidence of extensive cGVHD was 13.8%. So far, 1 case relapsed, 1 case displayed graft rejection, and poor function of graft occurred in 1 case, death occurred in 2 cases(6.7%). It is concluded that tumor-ablative chemotherapy combined with low intensity-modified BUCy/TBICy is safe and effective in allogeneic hematopoietic stem cell transplantation for hematologic malignancies, and it is useful to reduce relapse of hematologic malignancies after transplantation.

Conditioning regimen containing fludarabine instead of cyclophosphamide for haploidentical hematopoietic stem cell transplantation / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the feasibility and safety of conditioning regimen containing fludarabine (Flud) for haploidentical hematopoietic stem cell transplantation (HSCT).</p><p><b>METHODS</b>Preparative regimen containing Flud 40 mgxm(-2)xd(-1) on day -7 to -3 in place of cyclophosphamide (CTX) for haploidentical HSCT was given to 35 patients with hematologic malignancies (4 standard risk, 16 high risk, 15 relapse with no remission). All donors received rhG-CSF followed by HSC harvest. One patient received peripheral blood HSCT (PBSCT), one bone marrow transplantation (BMT), and the others BM combination with PBSCT. The regimen-associated side effect, engraftment, incidence of graft-versus-host disease (GVHD) and disease-free survival (DFS) probabilities were observed.</p><p><b>RESULTS</b>All patients achieved sustained, full donor-type engraftment. Thirty-four patients obtained primary durable engraftment, and 1 who rejected graft from his mother obtained successful durable engraftment after the second graft from his father. The cumulative incidence of grade III-IV acute GVHD and chronic GVHD was 12.1% and 31.7%, respectively. With a follow-up duration of 8-25 months, 6 patients were dead, in which 3 died of relapse, 2 of acute GVHD, 1 of fungal infection, none died of regimen-associated side effect. The other 29 patients remained alive and DFS probability was 79.7%.</p><p><b>CONCLUSION</b>Flud based conditioning regimens for haploidentical HSCT is safe and feasible, which reduces regimen-associated side effect, with no increasing the rate of relapse and infection, and decreases the incidence of aGVHD.</p>

Experimental study of human umbilical cord blood derived stromal cells transfected with recombinant adenoviral vector co-expressing VCAM-1 and GFP / 中国实验血液学杂志

This study was aimed to investigate the effect of vcam-1 gene-modified human umbilical cord blood derived stromal cells (CBDSCs) on hematopoietic regulation so as to establish the experimental foundation for further study. The target gene vcam-1 was cloned into the shuttle plasmid with the report gene GFP. The recombinant shuttle plasmid was transformed into BJ5183 bacteria to recombine with backbone vector pAdeasy-l, and the recombinant adenoviral vector ad-vcam-1-gfp was confirmed after transfection with CBDSCs. The results indicated that two fragments of about 9 kb and 2 kb were obtained after digestion of recombinant plasmid pAdTrack-vcam-1 with NotIand XhoI, and single fragment of 600 bp was obtained after amplification with PCR; two fragments of about 31 kb and 4 kb were obtained after digestion of recombinant plasmid pad-vcam-1-gfp with PacI, which suggested a successful homologous recombination. The expression of vcam-1 gene in ad-vcam-1-gfp transfected CBDSCs could be detected by immunocytochemistry, RT-PCR and fluorescent microscopy. It is concluded that the recombinant adenoviral vector ad-vcam-1-gfp has been constructed successfully, and the expression of vcam-1 is up-regulated in CBDSCs transfected by gene ad-vcam-1-gfp.

Connexin 43 expression and interacellular communicating function in acute leukemia bone marrow stroma cells / 中国实验血液学杂志

This study was purposed to investigate the connexin 43 (Cx43) expression level in acute leukemia bone marrow stromal cells (ABMSCs) and normal bone marrow stromal cells (NBMSCs), and to explore the difference in communicating functions between these cells. The Cx43 expression levels of ABMSCs and NBMSCs were detected by using immunohistochemistry and computer gray scale assay, and the difference of gap junction intercellular communication (GJIC) was examined through dry transfer technique. The results showed that expression level of Cx43 in ABMSCs was lower than that in NBMSCs and its function of GJIC in ABMSCs was also weaker than that in NBMSCs. It is concluded that cell-cell communication function is lowered in ABMSCs.

Hypoxia responsive element regulated herpes simplex virus-thymidine kinase system enhances killing effect of gancyclovir on Ewing's sarcoma cell line under hypoxic condition / 中华儿科杂志

<p><b>OBJECTIVE</b>To find out a possible approach to improve the effectiveness of radiotherapy and chemotherapy for Ewing's sarcoma by constructing a eukaryotic expression vector expressing herpes simplex virus-thymidine kinase (HSV-TK) regulated by hypoxia responsive element (HRE) under hypoxia and to evaluate the effects of this HRE regulated HSV-TK system on killing effect of gancyclovir (GCV) on Ewing's sarcoma cell line SK-ES under hypoxic condition.</p><p><b>METHODS</b>The HRE was synthesized according to the literature and cloned into the enhancer site of pIRES(2)-EGFP vector to obtain the pHRE recombinant plasmid. The HSV-TK was amplified by PCR and cloned into the multiple clone site of pIRES(2)-EGFP and pHRE to obtain pTK and pHRE-TK recombinant plasmid. The human Ewing's sarcoma cell line SK-ES was transfected by pTK or pHRE-TK recombinant plasmid with liposome and then was exposed to normoxic (21% oxygen) or hypoxic (3% oxygen) condition. The expression of enhanced green fluorescent protein (EGFP) was monitored by fluorescent microscopy. The sensitivity of human Ewing's sarcoma cell line SK-ES transfected with pTK or pHRE-TK recombinant plasmid to the anti-tumour drug GCV was determined with the method of tetrazolium (MTT) after treating with GCV for five days.</p><p><b>RESULTS</b>(1) The result of sequencing showed that the recombinant plasmid pHRE contained HRE, and that the recombinant plasmid pTK and pHRE-TK contained HSV-TK gene in the sense direction. (2) Comparison of fluorescent optical density (FOD) showed that (1) the EGFP FOD value of pHRE and pHRE-TK group cells exposed to hypoxia was significantly higher than those exposed to normoxia (P < 0.01); (2) when the cells were exposed to hypoxia, the EGFP FOD value of pHRE and pHRE-TK group cells was significantly higher than that of pTK and empty vector group (P < 0.01); (3) there was no significant difference among the four groups of cells when they were exposed to normoxia (P > 0.05). (3) Comparison of the sensitivity of four groups of cells to GCV showed that (1) the cells in pHRE-TK and pTK groups were much more sensitive to GCV than the cells in pHRE group under hypoxia condition (P < 0.01), the higher the GCV concentration, the greater the difference; (2) the cells of pHRE-TK group were more sensitive to GCV than those in pTK group under hypoxic condition (P < 0.01), but was almost equally sensitive under normoxic condition (P > 0.05); (3) the pHRE-TK group cells had higher sensitivity to GCV under hypoxia than normoxia (P < 0.01) while the pTK group cells had almost the same sensitivity to GCV under hypoxia and normoxia (P > 0.05).</p><p><b>CONCLUSION</b>(1) The eukaryotic expression vector expressing herpes simplex virus-thymidine kinase (HSV-TK) regulated by hypoxia responsive element (HRE) under hypoxia was constructed successfully. (2) HRE could up-regulate expression of EGFP by SK-ES cells under hypoxia condition. (3) HRE could enhance the killing effect of HSV-TK/GCV system on human Ewing's sarcoma cell line SK-ES under hypoxic condition.</p>