Ex vivo pathogenicity of anti-laminin γ1 autoantibodies.

Autoimmunity against laminins has been described in several autoimmune diseases (including mucous membrane pemphigoid, anti-laminin γ1 pemphigoid, and connective tissue diseases), in pregnancy loss, and in infections such as Chagas disease. Except for anti-laminin-332 mucous membrane pemphigoid, adequate evidence has been lacking for the tissue injury potential of laminin-specific antibodies and the pathogenic epitopes. We evaluated the pathogenic potential of antibodies targeting laminin γ1, a major constituent of basement membranes and the main antigen in anti-laminin γ1 pemphigoid. Rabbit antibodies were generated against fragments of the N-terminus and C-terminus of murine laminin γ1, and their ability to disrupt ligand interactions and/or to activate complement and granulocytes was assessed using previously established ex vivo assays. Our findings document a pathogenic potential of antibodies targeting the laminin γ1 N-terminus. These antibodies interfere with the binding of nidogen to laminin and can activate granulocytes and the complement cascade. We detected antibodies with different degrees of reactivity with laminin γ1 N-terminus in patients with anti-laminin γ1 pemphigoid, cutaneous lupus erythematosus, and scleroderma. Our results provide mechanistic insights into the tissue damage associated with laminin autoimmunity and could facilitate development of appropriate diagnostic tools and therapeutic strategies.

Structural elucidation of full-length nidogen and the laminin-nidogen complex in solution.

Nidogen-1 is a key basement membrane protein that is required for many biological activities. It is one of the central elements in organizing basal laminae including those in the skin, muscle, and the nervous system. The self-assembling extracellular matrix that also incorporates fibulins, fibronectin and integrins is clamped together by networks formed between nidogen, perlecan, laminin and collagen IV. To date, the full-length version of nidogen-1 has not been studied in detail in terms of its solution conformation and shape because of its susceptibility to proteolysis. In the current study, we have expressed and purified full-length nidogen-1 and have investigated its solution behavior using size-exclusion chromatography (SEC), dynamic light scattering (DLS) and small angle X-ray scattering (SAXS). The ab initio shape reconstruction of the complex between nidogen-1 and the laminin γ-1 short arm confirms that the interaction is mediated solely by the C-terminal domains: the rest of the domains of both proteins do not participate in complex formation.