THE ROLE OF FRAGMENTED FIBRONECTIN IN THE MESANGIAL IGA-DEPOSITION AND PROLIFERATION IN IGA NEPHROPATHY.

免疫球蛋白A （IgA）在肾小球系膜基质中的沉积是诊断IgA肾病的先决条件，循环中含有IgA的复合物参与了这一过程。这些复合物的组成已被广泛研究，但碎片化纤维连接蛋白尚未报道。由于碎片化纤维连接蛋白本身参与细胞外基质的组装，我们研究了患者血清中是否存在碎片化纤维连接蛋白并能够结合IgA。方法将IgA肾病患者血清应用于肝素亲和柱，用Western-blot技术对流出液进行抗人纤维连接蛋白单克隆抗体的分析。用组织蛋白酶D消化天然纤维连接蛋白，得到与血清纤维连接蛋白相似的片段，用纯化的IgA1和组织蛋白酶D消化的纤维连接蛋白的混合物，用jacalin-琼脂糖亲和纯化，模拟纤维连接蛋白结合IgA的能力。发现约43 kDa的片段仅在抗体FN 1-1的患者中发生反应，该抗体对含有二硫键的纤维连接蛋白的羧基端区域具有特异性。通过组织蛋白酶d消化天然分子产生类似大小的片段，并在体外与IgA1结合。本研究表明，约43 kDa的纤维连接蛋白羧基末端片段能够结合IgA1，并且在患者血清中存在类似大小的片段。由于已知羧基末端二硫键在将外源性纤维连接蛋白组装到细胞外基质中是至关重要的，因此在患者中发现的IgA1与片段的亲和力可能具有致病潜力，参与IgA肾病的细胞外扩张和IgA沉积。

BackgroundsImmunoglobulin A (IgA) deposition in the glomerular mesangial matrix is a prerequisite for the diagnosis of IgA nephropathy, and circulating IgA-containing complex has been implicated in this process. Composition of these complexes has been extensively studied, but fragmented fibronectin has not been reported. Since fragmented fibronectin by itself is known to be involved in the assembly of extracellular matrix, we investigated whether fragmented fibronectin exists in patient's serum and is capable of binding IgA.MethodsSerum from patients with IgA nephropathy was applied to heparin-affinity column and the effluent was analyzed by Western-blot technique with a set of anti-human fibronectin monoclonal antibodies. Native fibronectin was digested with cathepsin D to obtain fragment similar to that of serum fibronectin, and the capacity of fibronectin to bind IgA was modeled with a mixture ofpurified IgA1 and cathepsin D-digested fibronectin using affinity purification on jacalin-agarose.FindingsA fragment at about 43 kDa reacted only in the patients with antibody FN 1-1, which is specific to the carboxyl-terminal region of fibronectin containing a disulfide bond. A similar-sized fragment was generated by cathepsin D-digestion of the native molecule and was bound to IgA1 in vitro.InterpretationThis study demonstrates that about a 43 kDa carboxylterminal fragment of fibronectin was able to bind IgA1 and a fragment of similar size was present in patients'sera. Since the carboxyl-terminal disulfide bond is known to be critical in assembling exogenous fibronectin into the extracellular matrix, the affinity of IgA1 to a fragment found in patients may have pathogenic potential to participate in extracellular expansion and IgA-deposition in IgA nephropathy.