PDF(Pubmed)
Abstract：
BACKGROUND: The neuroimmune network plays a crucial role in regulating mucosal immune homeostasis within the digestive tract. Synaptosome-associated protein 25 (SNAP-25) is a presynaptic membrane-binding protein that activates ILC2s, initiating the host\'s anti-parasitic immune response.
METHODS: To investigate the effect of Moniezia benedeni (M. benedeni) infection on the distribution of SNAP-25 in the sheep\'s small intestine, the recombinant plasmid pET-28a-SNAP-25 was constructed and expressed in BL21, yielding the recombinant protein. Then, the rabbit anti-sheep SNAP-25 polyclonal antibody was prepared and immunofluorescence staining was performed with it. The expression levels of SNAP-25 in the intestines of normal and M. benedeni-infected sheep were detected by ELISA.
RESULTS: The results showed that the SNAP-25 recombinant protein was 29.3 KDa, the titer of the prepared immune serum reached 1:128,000. It was demonstrated that the rabbit anti-sheep SNAP-25 polyclonal antibody could bind to the natural protein of sheep SNAP-25 specifically. The expression levels of SNAP-25 in the sheep\'s small intestine revealed its primary presence in the muscular layer and lamina propria, particularly around nerve fibers surrounding the intestinal glands. Average expression levels in the duodenum, jejunum, and ileum were 130.32 pg/mg, 185.71 pg/mg, and 172.68 pg/mg, respectively. Under conditions of M. benedeni infection, the spatial distribution of SNAP-25-expressing nerve fibers remained consistent, but its expression level in each intestine segment was increased significantly (P < 0.05), up to 262.02 pg/mg, 276.84 pg/mg, and 326.65 pg/mg in the duodenum, jejunum, and ileum, and it was increased by 101.06%, 49.07%, and 89.16% respectively.
CONCLUSIONS: These findings suggest that M. benedeni could induce the SNAP-25 expression levels in sheep\'s intestinal nerves significantly. The results lay a foundation for further exploration of the molecular mechanism by which the gastrointestinal nerve-mucosal immune network perceives parasites in sheep.
METHODS: To investigate the effect of Moniezia benedeni (M. benedeni) infection on the distribution of SNAP-25 in the sheep\'s small intestine, the recombinant plasmid pET-28a-SNAP-25 was constructed and expressed in BL21, yielding the recombinant protein. Then, the rabbit anti-sheep SNAP-25 polyclonal antibody was prepared and immunofluorescence staining was performed with it. The expression levels of SNAP-25 in the intestines of normal and M. benedeni-infected sheep were detected by ELISA.
RESULTS: The results showed that the SNAP-25 recombinant protein was 29.3 KDa, the titer of the prepared immune serum reached 1:128,000. It was demonstrated that the rabbit anti-sheep SNAP-25 polyclonal antibody could bind to the natural protein of sheep SNAP-25 specifically. The expression levels of SNAP-25 in the sheep\'s small intestine revealed its primary presence in the muscular layer and lamina propria, particularly around nerve fibers surrounding the intestinal glands. Average expression levels in the duodenum, jejunum, and ileum were 130.32 pg/mg, 185.71 pg/mg, and 172.68 pg/mg, respectively. Under conditions of M. benedeni infection, the spatial distribution of SNAP-25-expressing nerve fibers remained consistent, but its expression level in each intestine segment was increased significantly (P < 0.05), up to 262.02 pg/mg, 276.84 pg/mg, and 326.65 pg/mg in the duodenum, jejunum, and ileum, and it was increased by 101.06%, 49.07%, and 89.16% respectively.
CONCLUSIONS: These findings suggest that M. benedeni could induce the SNAP-25 expression levels in sheep\'s intestinal nerves significantly. The results lay a foundation for further exploration of the molecular mechanism by which the gastrointestinal nerve-mucosal immune network perceives parasites in sheep.
摘要：
背景：神经免疫网络在调节消化道内的粘膜免疫稳态中起着至关重要的作用。突触体相关蛋白25（SNAP-25）是一种激活ILC2s的突触前膜结合蛋白，启动宿主的抗寄生虫免疫反应。
方法：研究Monieziabenedeni（M.benedeni）感染SNAP-25在绵羊小肠中的分布，构建重组质粒pET-28a-SNAP-25，并在BL21中表达，产生重组蛋白。然后，制备兔抗绵羊SNAP-25多克隆抗体并进行免疫荧光染色。采用酶联免疫吸附法检测SNAP-25在正常绵羊和M.benedeni感染绵羊肠道中的表达水平。
结果：结果表明，SNAP-25重组蛋白为29.3KDa，制备的免疫血清滴度达到1:128,000。结果表明，兔抗绵羊SNAP-25多克隆抗体可以特异性结合绵羊SNAP-25的天然蛋白。SNAP-25在绵羊小肠中的表达水平显示其主要存在于肌肉层和固有层，特别是围绕肠腺的神经纤维。十二指肠的平均表达水平，空肠,回肠为130.32pg/mg，185.71pg/mg,和172.68pg/mg，分别。在M.benedeni感染的情况下，表达SNAP-25的神经纤维的空间分布保持一致，但其在各肠段的表达水平显著升高(P<0.05),高达262.02pg/mg，276.84pg/mg,十二指肠中326.65pg/mg，空肠,和回肠,增加了101.06%，49.07%,和分别为89.16%。
结论：这些发现表明，贝尼得尼可以显着诱导绵羊肠神经中SNAP-25的表达水平。研究结果为进一步探索绵羊胃肠神经-黏膜免疫网络感知寄生虫的分子机制奠定了基础。
方法：研究Monieziabenedeni（M.benedeni）感染SNAP-25在绵羊小肠中的分布，构建重组质粒pET-28a-SNAP-25，并在BL21中表达，产生重组蛋白。然后，制备兔抗绵羊SNAP-25多克隆抗体并进行免疫荧光染色。采用酶联免疫吸附法检测SNAP-25在正常绵羊和M.benedeni感染绵羊肠道中的表达水平。
结果：结果表明，SNAP-25重组蛋白为29.3KDa，制备的免疫血清滴度达到1:128,000。结果表明，兔抗绵羊SNAP-25多克隆抗体可以特异性结合绵羊SNAP-25的天然蛋白。SNAP-25在绵羊小肠中的表达水平显示其主要存在于肌肉层和固有层，特别是围绕肠腺的神经纤维。十二指肠的平均表达水平，空肠,回肠为130.32pg/mg，185.71pg/mg,和172.68pg/mg，分别。在M.benedeni感染的情况下，表达SNAP-25的神经纤维的空间分布保持一致，但其在各肠段的表达水平显著升高(P<0.05),高达262.02pg/mg，276.84pg/mg,十二指肠中326.65pg/mg，空肠,和回肠,增加了101.06%，49.07%,和分别为89.16%。
结论：这些发现表明，贝尼得尼可以显着诱导绵羊肠神经中SNAP-25的表达水平。研究结果为进一步探索绵羊胃肠神经-黏膜免疫网络感知寄生虫的分子机制奠定了基础。
