表达hCGβ的重组乳酸杆菌经阴道免疫小鼠后的体液免疫应答

Mucosall immunization via vagina with Lactobacillus expressing hCGβinduces an anti-hCGβmucosal hCGβmucosal and systemic immune response

目的用表达hCGβ的重组乳酸杆菌活疫苗经阴道黏膜途径免疫不同品系小鼠产生抗hCGβ免疫应答及机制。方法用108、109、1010个重组乳酸杆菌Lb hCGβ经阴道黏膜免疫6～8周龄雌性BALB／c和C257BL/6小鼠。3周后用相同剂量加强免疫一次。间接ELISA检测初次免疫后2～8周血清和阴道灌洗液中抗hCGβIgG和IgA抗体滴度。分离免疫小鼠脾、子宫、阴道黏膜的淋巴细胞,流式细胞仪分析其中B和T细胞的增殖情况,ELISPOT分析分泌抗hCGβIgG和IgA抗体分泌细胞的克隆数。结果对不同品系小鼠黏膜接种109、1010 Lb hCGβ可诱导相近的免疫应答;而108诱导的抗体滴度较低。在加强免疫后,109和1010 Lb hCGβ诱导的抗血清中和hCGβ抗原的能力>100ng／ml。Lb hCGβ阴道黏膜免疫后T和B细胞增殖;脾脏、子宫及阴道内均存在抗hCGβIgG及IgA抗体分泌细胞。结论重组乳酸杆菌活疫苗Lb hCGβ可经阴道黏膜免疫诱导有效的免疫应答,产生的应答与免疫剂量呈正相关。黏膜局部及全身T和B细胞克隆增殖;子宫及阴道内抗hCGβIgG和IgA分泌细胞可能是Lb hCGβ阴道黏膜免疫诱导体液免疫的主要来源。

Objective To study anti-hCGβ antibody response and its mechanism after mucosal immnunization via vagina by Lactobacillus expressing hCGβ. Methods The female BALB/c and C57BU6 mice were immunized via vagina by 10^8, 10^9, 10^10 Lb hCGβ, a recombinant Lactobacillus expressing hCGβ and then boosted with the same dose 3 weeks later. An indirect ELISA was used to determine anti-hCGβ IgG and IgA antibodies in vaginal lavage and serum, which were got at the end of 2-8 weeks after the primary immunization. The lymphocytes of spleen, uterus and vagina were separated and the proliferation of B and T ceils was analyzed by flow cytometry. The count of antibody secreting cells of anti-hCGβ IgG or IgA was evaluated by ELISPOT. Results The mucosal immunization of 10^9, 10^10 Lb hCGβ induced resemble response while 10^8 only induce lower response. The antiserum of 10^9 ＆ 10^10 Lb hCGβ immunization after booster could neutralize more than 100 ng/ml hCG antigen both in BALB/c and C57BL/6 mice. Proliferation of T and B cells was obtained after vaginal inoculation with Lb hCGβ. The IgG or IgA antibody secreting cells were appeared in spleen, uterus and vagina. Conclusion Lb hCGβ immunization via vagina can induce effective response, which is positively related with the immunization dose. The presence of lgG and IgA antibody secreting cells in uterus and vagina might be one of the mechanisms of Lb hCGβ inducing anti-hCGβ humoral immunity in vagina.