Lentiviral gp34/OX40L Gene Transfer into Dendritic Cells Facilitates Alloreactive CD4<Superscript>+</Superscript> T-Cell Response In Vitro

Gene-modified dendritic cells (DCs) are promising targets for cancer immunotherapy. In this study, we demonstrated that lentiviral transduction of DCs with the gp34/OX40L gene, one of the costimulatory molecules, facilitates alloreactive CD4+ T-cell response in vitro. We achieved a 20% to 40% efficiency of gp34/OX40L gene transfer into DCs by the lentiviral vector, and lentiviral gp34/OX40L gene transfer did not alter the surface phenotype of either immature or mature DCs, suggesting that expression of gp34/OX40L did not induce the maturation of immature DCs and that gp34/OX40L-transduced DCs could fully differentiate into mature DCs. gp34/OX40L gene transfer facilitated an allogeneic CD4+ T-cell response in vitro by mature DCs but not by immature DCs. Dose escalation of the transgene induced an increasing amount of gp34/OX40L expression, leading to an increasing level of up-regulation of the allogeneic CD4+ T-cell response. The addition of anti-gp34 monoclonal antibody totally abrogated this up-regulation. These results suggest that this facilitation of allogeneic CD4+ T-cell response is specifically dependent on gp34/OX40L expressed on transduced DCs. Taken together, our findings show that gp34/ OX40L plays an important role in allogeneic CD4+ T-cell activation by DCs and that lentiviral gp34/OX40L gene transfer into DCs may be a useful strategy for cancer immunotherapy.     

胰岛素样生长因子-1基因修饰骨髓基质细胞对大鼠脑缺血的神经保护作用

目的：探讨胰岛素样生长因子-1（IGF-1）基因修饰的骨髓基质细胞（MSCs）对大鼠脑缺血的神经保护作用及其机制。方法健康雄性Wistar大鼠48只分为假手术组、缺血模型组、 MSCs组、MSCs-IGF-1组,每组12只。除假手术组外,其余组采用改良线栓法建立左侧大脑中动脉缺血再灌注模型。再灌注24 h后MSCs组、MSCs-IGF-1组分别采用MSCs、IGF-1修饰的MSCs 干预治疗。每组于再灌注后3 d和14 d行神经功能缺损评分,采用免疫组化法测 BrdU阳细胞数量, TUNEL法测大鼠神经细胞凋亡数量,分光光度计测脑组织超氧化物歧化酶（SOD）活性和丙二醛（MDA）含量。结果再灌注14 d后MSCs组与MSCs-IGF-1组的神经功能缺损评分较缺血模型组明显降低（ P＜00．5）,而MSCs-IGF-1组神经功能缺损评分低于MSCs组（P＜0．05）。再灌注后3 d及14 d,假手术组与缺血模型组未见明显BrdU阳性细胞,MSCs-IGF-1组BrdU阳性细胞数较MSCs组明显增多（P＜0．05）。再灌注后3 d及14 d,MSCs组与MSCs-IGF-1组梗死半球神经细胞凋亡数目较缺血模型组相比明显减少（P＜0．05）,MSCs-IGF-1组比MSCs组凋亡细胞明显减少（P＜0．05）。在灌注后3 d及14 d,与缺血模型组相比,MSCs组脑组织中SOD活性明显升高（P＜0．05）,MDA含量明显下降（P＜0．05）,且MSCs-IGF-1组SOD活性较MSCs组更高（P＜0．05）,MDA含量更低（ P＜0．05）。结论 IGF-1基因修饰骨髓基质细胞治疗干预比单纯骨髓基质细胞干预对脑缺血具有更好的神经保护作用,其机制可能与增强骨髓基质细胞的抗氧化应激能力、减少神经细胞凋亡有关。     

转基因髓核细胞移植对兔腰椎间盘退变的影响

目的：椎间盘退变是引起腰背痛的主要原因。将腺相关病毒（adeno-associated virus,AAV）介导的结缔组织生长因子（connective tissue growth factor,CTGF）和基质金属蛋白酶组织抑制因子1(tissue inhibitor of metalloproteinases 1,TIMP1) 双基因体外联合转染兔椎间盘髓核细胞后,再移植于退变椎间盘内,观察椎间高度、II型胶原和蛋白多糖合成的变化,以探讨体外延缓或逆转椎间盘细胞退变的方法。方法 20只新西兰大白兔应用CT引导下的微创穿刺法建立兔椎间盘退变模型。以rAAV2-CTGF-IRES-TIMP1体外转染培养的兔髓核细胞,再将其显微注射于退变椎间盘内作为细胞移植组;退变椎间盘内注入磷酸盐缓冲液（PBS）作为退变对照组;正常椎间盘作为空白对照组。分别于6、10、14周行X线和MRI检查观察椎间高度的变化及椎间盘信号的改变,Western-blot鉴定细胞因子CTGF和TIMP1在椎间盘内的表达,35S整合法测定兔髓核组织蛋白多糖合成率,RT-PCR检测II型胶原及蛋白多糖mRNA的表达。结果 MRI证实CT引导下经皮微创穿刺2周后椎间盘开始退变。与退变对照组相比,MRI显示细胞移植组椎间盘退变明显减轻,转基因兔髓核细胞移植可减缓椎间高度(DHI)下降的发生率,转基因细胞移植组较退变对照组相比,Ⅱ型胶原含量和蛋白多糖的生物合成明显增加,差别有统计学意义（P﹤0.05）。结论 经皮CT 引导下穿刺可成功建立兔椎间盘退变模型。转双基因CTGF和TIMP1 髓核细胞移植可有助于椎间高度的维持,促进椎间盘内蛋白多糖和II型胶原的生物合成,明显延缓椎间盘退变。