Neufing Petra J, Clancy Robert M, Jackson Michael W, Tran Hai Bac, Buyon Jill P, Gordon Tom P
Department of Immunology, Allergy and Arthritis, Flinders University, Adelaide, South Australia, Australia.
Arthritis Rheum. 2005 Dec;52(12):3934-42. doi: 10.1002/art.21486.
Opsonization of apoptotic cells by autoantibodies bound to surface membrane-translocated La/SSB antigens may initiate tissue damage in the setting of congenital heart block. By injecting pregnant mice with human anti-La antibodies, we previously demonstrated the formation of IgG-apoptotic cell complexes in the developing mouse fetus; however, the binding of anti-La antibodies to human-specific epitopes could not be addressed. Accordingly, the objective of the current study was to delineate the epitope specificity of human La antibodies that are exposed on the surface of apoptotic cells.
We used fluorescence microscopy and flow cytometry to assess the binding of human anti-La antibodies affinity purified against immunodominant epitopes of La to human cells undergoing spontaneous apoptosis, in a murine xenograft model in vivo and in cultured human fetal cardiocytes rendered apoptotic in vitro, respectively.
Anti-La antibodies bound to immunodominant epitopes of La within the NH(2)-terminus and the RNA recognition motif (RRM) region of apoptotic human cells, in both xenografts and fetal cardiocytes. In contrast, human antibodies affinity purified against the COOH-terminal La epitope did not bind apoptotic cells in either model. This defines the topology of redistributed La during apoptosis, with surface exposure of the NH(2)-terminus and RRM regions. The potential importance of anti-La NH(2)-terminal and anti-La RRM specificity was confirmed by detection of this reactivity in mothers of children with congenital heart block.
These findings provide insight into both the molecular modification of the La autoantigen during apoptosis and the specificity of antibodies capable of binding to surface-exposed La. Subsequent formation of surface immune complexes may lead to tissue injury in patients with autoimmune diseases such as congenital heart block.
与表面膜转位的La/SSB抗原结合的自身抗体对凋亡细胞的调理作用可能在先天性心脏传导阻滞的情况下引发组织损伤。通过给怀孕小鼠注射人抗La抗体，我们之前在发育中的小鼠胎儿中证明了IgG-凋亡细胞复合物的形成；然而，抗La抗体与人特异性表位的结合尚未得到研究。因此，本研究的目的是确定凋亡细胞表面暴露的人La抗体的表位特异性。
我们分别在体内小鼠异种移植模型和体外诱导凋亡的培养人胎儿心肌细胞中，使用荧光显微镜和流式细胞术评估针对La免疫显性表位亲和纯化的人抗La抗体与自发凋亡的人细胞的结合情况。
在异种移植和胎儿心肌细胞中，抗La抗体均与凋亡人细胞NH(2)末端和RNA识别基序（RRM）区域内的La免疫显性表位结合。相比之下，针对COOH末端La表位亲和纯化的人抗体在两种模型中均未与凋亡细胞结合。这确定了凋亡过程中重新分布的La的拓扑结构，即NH(2)末端和RRM区域暴露于表面。先天性心脏传导阻滞患儿母亲中这种反应性的检测证实了抗La NH(2)末端和抗La RRM特异性的潜在重要性。
这些发现为凋亡过程中La自身抗原的分子修饰以及能够结合表面暴露的La的抗体的特异性提供了见解。随后表面免疫复合物的形成可能导致自身免疫性疾病（如先天性心脏传导阻滞）患者的组织损伤。
