痘苗病毒天坛株C8L—K3L片段缺失表达载体的生物学性能研究

Construction of expression vector of recombinant vaccinia virus TianTan strain with C8L-K3L region deletion and study on biological properties of the recombinant virus

目的研究缺失C8L．K3L片段的痘苗病毒天坛株载体的生物学性能及外源基因的免疫原性。方法构建缺失C8L-K3L区域的重组痘苗病毒天坛株载体VrITT△c8-K3-gag。在4种细胞上检钡0了重组载体的增殖能力、在小鼠和家兔模型上进行毒力评价，并在BALB／c小鼠进行了免疫效果评价。结果与Vrrr相比，VTTAC8-K3-gag在CEF、BHK-21、HeLa细胞中复制能力显著下降，在Vero细胞中失去复制能力。VTYAC8-K3-gag在小鼠和家兔模型中毒力均有显著降低。VTT△C8-K3-gag免疫小鼠第4周后诱导的痘苗病毒特异结合抗体与中和抗体滴度分别达到5．6×10^3和1．0×10^4，第9周滴度弦到5．8×10^5和5．25×10^3，与VTKgpe相比均显著下降3～9倍，但E3刺激后的细胞免疫应答无显著变化。VTT△C8-K3-gag单独免疫或与DNA疫苗联合免疫诱导的HIV特异性抗体水平和细胞免疫应答均与VTKgpe无差异。结论VTrAC8-K3-gag是一个安全性较高且具有深入研究价值的HIV候选疫苗株。

Objective To study the biological properties of the recombinant vaccinia virus Tiantan strain with CSL-K3L region deletion and its immunogenicity. Methods The expression vector of recombi- nant vaccinia virus TianTan strain （ VTTACS-K3-gag ） was constructed by replacing CSL-K3L genes with HIV gag gene and GFP gene. Viral replication capacities in chicken CEF, hamster BHK-21, monkey Vero and human HeLa cell lines were detected respectively. Virulence evaluation was carried out in mice and rab- bit models, and immune effects of VTTA CS-K3-gag was evaluated in BALA/c mice model. Results The replication capacity of VTrACS-K3-gag was impaired in chicken CEF, hamster BHK-21 and human HeLa cell lines, and was completely restricted in monkey Vero cell line as compared with the parental VTT. VTT A CS-K3-gag was less virulent than VTT in mice and rabbit models. The cellular and humoral responses to HIV elicited by VTFACS-K3-gag alone or in combination with DNA vaccine were similar to that induced by VTKgpe, while binding antibody responses specific to vaccinia viruse were reduced by 3-9 times, and neu- tralizing antibody response to VTT were reduced by 4-8 times. Conclusion The study suggests that VTT A CS-K3-gag is much safer than parental VTT and is a valuable strain for further studies on HIV-vaccine candidates.