Induction of Epidermolysis Bullosa Acquisita in Mice by Passive Transfer of Autoantibodies from Patients (1)
Epidermolysis bullosa acquisita (EBA) is an acquired autoimmune blistering disease of the skin. Patients who are born with a defect in the gene that encodes for type VII collagen have a paucity of normal anchoring fibrils and exhibit skin fragility and skin blisters just like EBA patients.Type VII collagen is composed of three identical alpha chains, each consisting of a 145 kDa central collagenous triple-helical segment characterized by repeating Gly-X-Y amino-acid sequences, flanked by a large 145 kDa amino-terminal, non-collagenous domain (NC1), and a smaller 34 kDa carboxy-terminal non-collagenous domain (NC2). A high titer antiserum to the NC1 domain of human type VII collagen was raised by immunizing rabbits. The antibody was injected into hairless immunocompetent mice and the mice developed a skin condition that had many of the features of EBA in humans. The injection of rabbit polyclonal antibodies to the NC1 domain of mouse type VII collagen into adult nude, BALB/c, and C57BL/6 mice also induced sub-epidermal skin blisters that were reminiscent of human EBA.
The affinity-purified antibodies to the NC1 domain of type VII collagen from the sera of two EBA patients only labeled the 145 kDa NC1 domain of type VII collagen and did not label other matrix proteins including type I collagen, type IV collagen, fibronectin, laminin-1, and laminin-5. The antibodies strongly labeled the basement membrane zone (BMZ) of both human and mouse skin at dilution titers over 1:50,000. The antibody bound to the dermal side of the salt-split skin, consistent with labeling-type VII (anchoring fibril) collagen and characteristic of EBA serum.
SKH1 mice were given daily intradermal injections of affinity-purified anti-type VII collagen antibodies from EBA sera or control normal human sera at doses from 20 to 100 μg/g body weight/per day. cutaneous lesions occurred in 36 of 40 experimental mice given EBA antibodies. In contrast, none of the 10 animals that received identical doses of control IgG injections developed any cutaneous lesions. Furthermore, five animals injected with EBA IgG depleted of reactivity to the NC1 domain of type VII collagen (flow-through fractions from NC1 affinity column) did not develop blisters.
Increasing the amount of injected IgG results in an increase in the circulating titer of BMZ specific anti-type VII collagen antibody, which, in turn, was associated with increased skin lesions.
Histological examination of lesional skin from mice injected with EBA antibodies showed a full sub-epidermal blister with a clean separation between the epidermis and dermis. There was also a mild to moderate scattered dermal inflammatory infiltrate consisting mostly of mononuclear cells and a few neutrophils. Direct immunofluorescence (DIF) of perilesional and lesional skin demonstrated that all of the mice injected with EBA antibodies had human IgG deposits at the DEJ. Also, in 85% (34 out of 40) of the injected mice, murine complement component C3 deposits were detected at the DEJ. Within the dermal inflammatory infiltrate, neutrophils could be detected.
The turnover of anchoring fibril structures is thought to be very slow. Therefore, it is likely that the resident anchoring fibrils in the mice may have continued to function despite the presence of EBA autoantibodies. Rabbit antibodies to the NC1 domain of murine type VII collagen triggered an inflammatory reaction with complement activation and the recruitment of leukocytes. In this EBA murine model, blister induction required the Fc portion of the rabbit IgG and activation of terminal complement components.