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目的筛选可拮抗原发性胆汁性肝硬化(PBC)中PDC-E2_(165-174)特异性CTL功能的模拟肽。方法以自身抗原表位PDC-E2_(165-174)(LLAEIETDKA)为原型肽,采用丙氨酸突变扫描法,通过亲和力分析及其对PDC-E2_(165-174)特异性CTL细胞毒性和分泌细胞因子等功能的抑制效应,从中筛选拮抗性模拟肽。结果8条模拟肽中,5号位突变的模拟肽(I5A)显示与HLA-A*0201分子高亲和力,5名PBC患者PDc-E2 特异性CTL对其负载的T2细胞的杀伤率均小于10％,并且I5A也未能诱导PBC患者PDC-E2特异性CTL 产生IFN-γ。在原型肽的存在下,I5A可抑制PDC-E2_(165-174)CTL细胞毒性和分泌IFN-γ,随着模拟肽浓度增加,抑制效应越强,在10 μmol／L浓度时可以明显抑制PDC-E2_(165-174)CTL的细胞毒活性和分泌IFN-γ的能力。结论模拟肽I5A(LLAEAETDKA)可能是PDC-E2_(165-174)特异性CTL的一个拮抗性多肽,其抑制效应并不是其能竞争结合靶细胞上的HLA-A*0201分子,而是对TCR的一种拮抗作用,为将来设计以TCR为基础的特异性免疫治疗提供实验依据。
Objective To screen mimotope peptides that can antagonize PDC-E2 (165-174) specific CTL function in primary biliary cirrhosis (PBC). Methods Using the autoantigen epitope PDC-E2_ (165-174) (LLAEIETDKA) as the prototype peptide, by alanine mutation scanning method and its affinity to PDC-E2 (165-174) specific CTL cytotoxicity and secretion Cytokines and other functions of the inhibitory effect, screening of antagonistic peptidomimetics. Results The mimic peptide (I5A) mutated at position 5 showed high affinity to HLA-A * 0201 molecule. The killing rate of PDc-E2-specific CTL on T2 cells loaded by 5 PDBCs was less than 10 %, And I5A also failed to induce PDC-E2-specific CTLs to produce IFN-γ in PBC patients. In the presence of the prototype peptide, I5A inhibited the cytotoxicity of PDC-E2 (165-174) CTL cells and secreted IFN-γ. With the increase of the mimetic peptide concentration, the stronger the inhibitory effect, the inhibitory effect of PDC at the concentration of 10 μmol / L -E2_ (165-174) cytotoxic activity of CTLs and the ability to secrete IFN-γ. Conclusion Peptide I5A (LLAEAETDKA) may be an antagonistic polypeptide of PDC-E2 (165-174) -specific CTL and its inhibitory effect is not that it can compete for binding to HLA-A * 0201 on target cells but rather to TCR It provides an experimental basis for the future design of TCR-based immunotherapy.