Xing Jing, Xiao Yue'e, Tang Xiaoqian, Sheng Xiuzhen, Zhan Wenbin
Laboratory of Pathology and Immunology of Aquatic Animals, KLMME, Ocean University of China, Qingdao 266003, PR China.
Laboratory of Pathology and Immunology of Aquatic Animals, KLMME, Ocean University of China, Qingdao 266003, PR China; Laboratory for Marine Fisheries Science and Food Production Processes, Qingdao National Laboratory for Marine Science and Technology, No. 1 Wenhai Road, Aoshanwei Town, Qingdao, PR China.
Fish Shellfish Immunol. 2017 Jul;66:78-85. doi: 10.1016/j.fsi.2017.05.017. Epub 2017 May 5.
In acquired immunity, T lymphocytes regulate the immune responses of B lymphocytes, including the IgM B lymphocyte counts and antibody production. In this paper, Cyclosporine A (CsA) and Rapamycin (RaPa) were used, and their inhibition on T lymphocytes and immune responses of B lymphocytes in flounder (Paralichthys olivaceus) were investigated. Flounder was injected with Keyhole Limpet Hemocyanin (KLH), a mixture of KLH and CsA (KLH + CsA), or a mixture of KLH and RaPa (KLH + RaPa). Then, the proportions of T and IgM B lymphocytes (PT and PB) in peripheral blood leukocytes (PBL) were analysed by flow cytometry (FCM), total antibodies (TA) and KLH specific antibodies (KA) in serum were measured by ELISA, and expression of 9 immune-related genes in the spleen and kidneys were determined using q-PCR. On the other hand, the PBL culture was treated with Concanavalin A (ConA), a mixture of ConA and CsA, and a mixture of ConA and RaPa. Then the PT and PB were measured, and the cell proliferation was examined using the MTT method. The results showed that the PT peaked on the 5 day in the KLH group, KLH + CsA group and KLH + RaPa group. The maximum inhibition rates (MIR) of CsA and RaPa were 27.44% ± 0.50% and 21.37% ± 2.06%, respectively. The PB peaked at the 5 week, and the MIR of CsA and RaPa were 44.51% ± 1.36% and 33.3% ± 0.65%, respectively. The KA and TA peaked at the 5 week. The MIR of CsA and RaPa on TA were 40.31% ± 1.59% and 32.96% ± 2.21%, respectively, and were 27.77% ± 2.02% and 23.41% ± 1.08% for KA, respectively. Nine immune-related genes had significantly lower expression in the KLH + CsA group and KLH + RaPa group compared to the KLH group. The proliferation of the PBL culture was inhibited by CsA or RaPa, and the inhibition rate of CsA and RaPa for PT was 18.14% ± 1.08% and 17.88% ± 1.02%, respectively, and the inhibition rates for PB were 3.03% ± 0.57% and 2.95% ± 0.53%, respectively. The results demonstrated that T lymphocytes counts were inhibited by CsA and RaPa, followed by suppression of IgM B lymphocytes and antibody production, which suggests that T lymphocytes regulate the immune response of B lymphocytes in flounder.
在获得性免疫中，T淋巴细胞调节B淋巴细胞的免疫反应，包括IgM B淋巴细胞计数和抗体产生。本文使用了环孢素A（CsA）和雷帕霉素（RaPa），研究了它们对牙鲆（Paralichthys olivaceus）T淋巴细胞以及B淋巴细胞免疫反应的抑制作用。给牙鲆注射钥孔戚血蓝蛋白（KLH）、KLH与CsA的混合物（KLH + CsA）或KLH与RaPa的混合物（KLH + RaPa）。然后，通过流式细胞术（FCM）分析外周血白细胞（PBL）中T和IgM B淋巴细胞的比例（PT和PB），用酶联免疫吸附测定（ELISA）法检测血清中的总抗体（TA）和KLH特异性抗体（KA），并使用定量聚合酶链反应（q-PCR）测定脾脏和肾脏中9个免疫相关基因的表达。另一方面，用刀豆蛋白A（ConA）、ConA与CsA的混合物以及ConA与RaPa的混合物处理PBL培养物。然后测定PT和PB，并使用MTT法检测细胞增殖情况。结果显示，KLH组、KLH + CsA组和KLH + RaPa组中PT在第5天达到峰值。CsA和RaPa的最大抑制率（MIR）分别为27.44% ± 0.50%和2l.37% ± 2.06%。PB在第5周达到峰值，CsA和RaPa的MIR分别为44.51% ± 1.36%和33.3% ± 0.65%。KA和TA在第5周达到峰值。CsA和RaPa对TA的MIR分别为40.31% ± 1.59%和32.96% ± 2.21%，对KA的MIR分别为27.77% ± 2.02%和23.41% ± 1.08%。与KLH组相比，KLH + CsA组和KLH + RaPa组中9个免疫相关基因的表达显著降低。CsA或RaPa抑制了PBL培养物的增殖，CsA和RaPa对PT的抑制率分别为18.14% ± 1.08%和17.88% ± 1.02%，对PB的抑制率分别为3.03% ± 0.57%和2.95% ± 0.53%。结果表明，CsA和RaPa抑制了T淋巴细胞计数，随后抑制了IgM B淋巴细胞和抗体产生，这表明T淋巴细胞调节牙鲆中B淋巴细胞的免疫反应。
