Department of Veterinary Sciences, University of Munich, Germany.
Mol Immunol. 2010 Apr;47(7-8):1619-28. doi: 10.1016/j.molimm.2010.01.011. Epub 2010 Feb 9.
Chickens provide an important model for developmental biology as well as phylogenetic studies of the immune system. In many species cytokines are important regulators of immune functions and recently we identified the chicken homologue of BAFF (B cell activating factor of the TNF family), shown to be an essential survival factor for B cells in mammals. Characterisation of BAFF function in the phylogenetically distant chicken requires in vivo studies, but despite considerable progress to date no efficient transgene or knockout technology for chickens is available. Thus the retroviral vector system RCAS (replication-competent ASLV long terminal repeat (LTR) with a Splice acceptor) was used to generate chickens stably expressing chBAFF or a soluble chBAFF neutralizing decoy receptor. Expression of RCAS based proteins was demonstrated in ovo and for the complete observation period of 2 month after hatch. In contrast to injections with recombinant proteins the prolonged RCAS based cytokine expression allowed assessment of BAFF function not only in newly hatched chicks but also in more mature birds. We demonstrate that RCAS based secreted products were biologically active, with chBAFF overexpression leading to a drastic and continuous increase in B cell numbers and neutralisation of chBAFF by expression of a soluble decoy receptor leading to severely reduced B cell numbers in transduced birds. This increase in B cell numbers was matched by increased immunoglobulin levels. Using RCASBP transduced birds we provide evidence that the RCAS system provides a fast and highly efficient gene transfer system for analysis of secreted protein function in the chicken and we show that in the chicken BAFF is necessary for B cell survival but in contrast to mammals, not only mature B cells but also immature B cells in the bursa of Fabricius are maintained by BAFF.
鸡是发育生物学和免疫系统系统发生研究的重要模型。在许多物种中，细胞因子是免疫功能的重要调节剂，最近我们鉴定了鸡 BAFF（肿瘤坏死因子家族的 B 细胞激活因子）的同源物，证明其是哺乳动物 B 细胞的必需存活因子。在系统发生上遥远的鸡中，BAFF 功能的特征需要体内研究，但尽管迄今为止取得了相当大的进展，仍然没有有效的转基因或敲除技术可用于鸡。因此，我们使用逆转录病毒载体系统 RCAS（复制能力的 ASLV 长末端重复序列（LTR）带有剪接受体）来生成稳定表达 chBAFF 或可溶性 chBAFF 中和诱饵受体的鸡。在胚胎中证明了 RCAS 基于蛋白的表达，并且在孵化后 2 个月的完整观察期内都可检测到。与重组蛋白的注射相比，RCAS 基于细胞因子的延长表达不仅允许评估新孵化小鸡中的 BAFF 功能，还允许评估更成熟鸟类中的 BAFF 功能。我们证明 RCAS 基于分泌的产物具有生物活性，过表达 chBAFF 导致 B 细胞数量急剧且持续增加，而表达可溶性诱饵受体中和 chBAFF 则导致转导鸟类中的 B 细胞数量严重减少。B 细胞数量的增加与免疫球蛋白水平的增加相匹配。使用 RCASBP 转导的鸟类，我们提供了证据表明 RCAS 系统为分析鸡中分泌蛋白的功能提供了快速而高效的基因转移系统，并且我们表明在鸡中，BAFF 是 B 细胞存活所必需的，但与哺乳动物不同，不仅成熟的 B 细胞，而且法氏囊中的未成熟 B 细胞也依赖于 BAFF 来维持。
