Antimitochondrial Autoantibodies in Pemphigus Vulgaris

Abstract

A loss of epidermal cohesion in pemphigus vulgaris (PV) results from autoantibody action on keratinocytes (KCs) activating the signaling kinases and executioner caspases that damage KCs, causing their shrinkage, detachment from neighboring cells, and rounding up (apoptolysis). In this study, we found that PV antibody binding leads to activation of epidermal growth factor receptor kinase, Src, p38 MAPK, and JNK in KCs with time pattern variations from patient to patient. Both extrinsic and intrinsic apoptotic pathways were also activated. Although Fas ligand neutralizing antibody could inhibit the former pathway, the mechanism of activation of the latter remained unknown. PV antibodies increased cytochrome c release, suggesting damage to mitochondria. The immunoblotting experiments revealed penetration of PVIgG into the subcellular mitochondrial fraction. The antimitochondrial antibodies from different PV patients recognized distinct combinations of antigens with apparent molecular sizes of 25, 30, 35, 57, 60, and 100 kDa. Antimitochondrial antibodies were pathogenic because their absorption abolished the ability of PVIgG to cause keratinocyte detachment both in vitro and in vivo. The downstream signaling of antimitochondrial antibodies involved JNK and late p38 MAPK activation, whereas the signaling of anti-desmoglein 3 (Dsg3) antibody involved JNK and biphasic p38 MAPK activation. Using KCs grown from Dsg3؊/؊ mice, we determined that Dsg3 did not serve as a surrogate antigen allowing antimitochondrial antibodies to enter KCs. The PVIgG-induced activation of epidermal growth factor receptor and Src was affected neither in Dsg3KCs nor due to absorption of antimitochondrial antibodies. These results demonstrated that apoptolysis in PV is a complex process initiated by at least three classes of autoantibodies directed against desmosomal, mitochondrial, and other keratinocyte self-antigens. These autoantibodies synergize with the proapoptotic serum and tissue factors to trigger both extrinsic and intrinsic pathways of cell death and break the epidermal cohesion, leading to blisters. Further elucidation of the primary signaling events downstream of PV autoantigens will be crucial for the development of a more successful therapy for PV patients.Pemphigus vulgaris (PV) 3 is an IgG autoantibody-mediated disease of skin and mucosa caused by a loss of epidermal cohesion and manifested by progressive blistering and non-healing erosions. The mechanism of detachment of keratinocytes (KCs) in PV, termed acantholysis, is a subject of intensive research. Elucidation of the pathophysiologic mechanism of acantholysis should facilitate development of novel pharmacologic approaches to prevent and treat blistering without systemic corticosteroids, a mainstay of treatment of PV patients. Acantholysis can be blocked both by inhibitors of signaling kinases, such as p38 MAPK (3), mammalian target of rapamycin (4), Src, and epidermal growth factor receptor (EGFR) kinase (2, 4 -7) and by other tyrosine kinases, phospholipase C, calmodulin...

Mechanisms of Mitochondrial Damage in Keratinocytes by Pemphigus Vulgaris Antibodies

“…We became interested in MtAbs because we sought to elucidate the mechanism of intrinsic apoptosis of KCs in PV originally demonstrated by us (17) and confirmed by others (18,19). The direct evidence that MtAbs are critical to disease pathology, rather than a bystander phenomena in PV, was provided by the studies demonstrating that PV IgGs enter KCs and specifically bind to a number of mitochondrial proteins, which is associated with the mitochondrial damage manifested by cytochrome c release (20). Most importantly, adsorption of MtAbs abolished the ability of the IgG fraction of PV serum (PVIgG) to cause keratinocyte detachment (i.e.…”

“…One-day-old pups delivered by the BALB/c mice purchased from the Jackson Laboratory were used to investigate the effect of mitochondria-protecting drugs on the extent of epidermal acantholysis induced by passive transfer of 1 mg/g of body weight of PVIgG. The pups were injected subcutaneously with PVIgG with or without test drugs and examined 24 h later for the extent of epidermal acantholysis, as detailed elsewhere (20). Briefly, the euthanized animals were snap frozen in liquid nitrogen, cross-sectioned at the umbilicus level, embedded into the OCT compound (Miles Scientific, Naperville, IL), and stained by hematoxylin and eosin.…”

Desmosomal Cadherins and Signaling: Lessons from Autoimmune Disease

“…The involvement of p38MAPK is well established, which may require its downstream kinase MAPKAP kinase 2 (MK2) (Berkowitz et al, 2005(Berkowitz et al, , 2006Mao et al, 2013) and epidermal growth factor receptor (EGF-R) and/or Src signaling (Bektas et al, 2013;Chernyavsky et al, 2007;Frusic-Zlotkin et al, 2006;Tsang et al, 2012b). Other molecules implicated in pemphigus are Akt/mTOR and FAK (Gil et al, 2012;Pretel et al, 2009) as well as JNK (Marchenko et al, 2010), CDK-2 (Lanza et al, 2008), PERP (Nguyen et al, 2009) and factors associated with apoptotic signaling such as caspases 3, 6, 8, and 9 as well as Bcl/Bax and others (Li et al, 2009;Marchenko et al, 2010). However, for most signaling pathways the precise mechanisms by which they reduce Dsg-mediated binding and desmosomal integrity are not well understood.…”

Desmosome assembly, homeostasis, and desmosomal disease

“…[135][136][137] Classically, pemphigus can be classified into two major groups, ie, pemphigus vulgaris (PV), which is characterized by suprabasal acantholysis and by autoantibodies against Dsg3 or both Dsg3 and Dsg1, and pemphigus foliaceus, which is characterized by superficial acantholysis in the granular cell layer of the epidermis and autoantibodies against Dsg1. Additional desmosomal autoantigens include Dscs, such Dsc3, 138,139 as well as Pg, 140 Dsp, 141 and Pkp3. 142 Insights into pathomechanisms underlying the generation of acantholysis in PV after autoantibodies bind to desmosomal and nondesmosomal antigens on the keratinocyte surface have lead to two conflicting, yet complementary hypotheses.…”