Knospe V, Gregerson D S, Donoso L A
Department of Ophthalmology, University of Minnesota, Minneapolis 55455.
Autoimmunity. 1989;4(3):153-69. doi: 10.3109/08916938909003046.
The factors which lead to selection of dominant antigenic sites concentrated in discreet regions of proteins and polypeptides are important to the development of antigen-specific immunotherapies for autoimmune diseases and for vaccine design. In this study, the main immunogenic regions of the immunopathogenic autoantigen, retinal S-antigen, have been identified by examination of the specificity of antibody responses of different species. Using cyanogen bromide and synthetic peptides in western blots and the ELISA, the specificities of antisera from rabbits, guinea pigs, rats and 19 inbred strains of mice were tested. All animals produced high titers of antibody to S-antigen with the exception of PL/J mice. Antibodies which bound epitopes contained in peptide CB46, a 46 amino acid-containing peptide located at the C-terminus of S-antigen, were dominant in all species and strains tested. The epitopes in CB46 were multiple, overlapping, and concentrated in a stretch of approximately 30 residues. Two overlapping synthetic peptides from that region substantially competed the anti-CB46 response of all animals. Antibodies which recognized peptide CB47, a 47 residue peptide from the N-terminus, comprised the next most common group. This epitope was similar in all mice and overlapped the epitope defined by rat antibodies. All anti-CB47 antibodies mapped to an 11 residue region of CB47. Eleven strains of mice did not respond to CB47 after one immunization with S-antigen; however, multiple immunizations readily converted all animals so tested to CB47 responders. Rabbits and guinea pigs exhibited very weak responses to CB47 following one immunization; multiple immunizations increased the response minimally. Rats produced a strong antibody response to peptide CB123, which contains the known uveitogenic sites, while very little activity to CB123 was raised in rabbits and guinea pigs. Only 3 murine strains, LP, LP.R3, and B10.R3-71, responded with antibodies to CB123 and the epitope was mapped to a 30 residue region which in rats also contains two distinct pathogenic sites and an antibody epitope. Only rats and rabbits made antibody to the CB35 peptide; the epitopes were contained within an 18 residue sequence. The results show that a main immunogenic region is located in S-antigen near the C-terminus and is independent of species or MHC. Less dominant, species and strain-dependent immunogenic regions were found in three other areas, i.e. peptides CB47, CB123 and CB35.
导致优势抗原表位集中在蛋白质和多肽离散区域的因素，对于自身免疫性疾病的抗原特异性免疫疗法及疫苗设计的发展至关重要。在本研究中，通过检测不同物种抗体反应的特异性，已确定了免疫致病性自身抗原视网膜S抗原的主要免疫原性区域。利用溴化氰和合成肽进行蛋白质印迹法及酶联免疫吸附测定，检测了来自兔、豚鼠、大鼠及19个近交系小鼠的抗血清的特异性。除PL/J小鼠外，所有动物均产生了高滴度的抗S抗原抗体。与位于S抗原C末端的含46个氨基酸的肽CB46中所含表位结合的抗体，在所有测试的物种和品系中均占主导地位。CB46中的表位是多个、重叠的，且集中在一段约30个残基的区域。来自该区域的两个重叠合成肽在很大程度上能竞争所有动物的抗CB46反应。识别来自N末端的含47个残基的肽CB47的抗体，构成了下一个最常见的组。该表位在所有小鼠中相似，且与大鼠抗体所界定的表位重叠。所有抗CB47抗体均定位在CB47的一个11个残基的区域。用S抗原免疫一次后，11个小鼠品系对CB47无反应；然而，多次免疫能使所有经测试的动物轻易转变为对CB47有反应的动物。兔和豚鼠在一次免疫后对CB47的反应非常弱；多次免疫仅使反应略有增加。大鼠对含有已知葡萄膜炎致病位点的肽CB123产生强烈抗体反应，而兔和豚鼠对CB123产生的活性则非常低。只有3个小鼠品系，即LP、LP.R3和B10.R3 - 71，对CB123产生抗体反应，且该表位定位在一个30个残基的区域，在大鼠中该区域也包含两个不同的致病位点和一个抗体表位。只有大鼠和兔产生了针对CB35肽的抗体；表位包含在一个18个残基的序列中。结果表明，一个主要免疫原性区域位于S抗原靠近C末端的位置且与物种或主要组织相容性复合体无关。在其他三个区域，即肽CB47、CB123和CB35中，发现了不太占主导地位、依赖物种和品系的免疫原性区域。
