16: Theories on the Pathogenesis of Depigmentation: Immune Hypothesis
JEAN-CLAUDE BYSTRYN
Introduction
The cause(s) of the localized, spontaneous and complete depigmentation of the skin that occurs in vitiligo and in association with some pigmented naevi and melanomas is (are) not known. The pigment loss in these condi- tions can occur in otherwise normal skin, around or within a benign or malignant pigmented lesion, or at a site distal to a pigmented lesion. These various forms of leukodermas probably result from different pathophysio- logical mechanisms as they involve the destruction of different types of pigment cells, i.e. normal or malignant melanocytes, normal or atypical naevi cells. They occur in different locations of the integument, i.e. at the site of, adjacent to, or distal to the cells that initiated the process. And they are associated with different histological abnormalities, i.e. the presence or absence of a dense cellular infiltrate. Whatever their causes, these leukoder- mas are experiments of nature that demonstrate that there are mechanisms in humans that can selectively kill pigment cells, including melanoma cells.
C H A P T E R 16
Antibodies to melanocytes in vitiligo Immune Hypothesis
The most direct and convincing evidence that vitiligo is an autoimmune disease is that specific autoantibodies to melanocyte cell surface antigens are present in the circulation of most patients with vitiligo. These antibodies were initially demonstrated by immunoprecipitation of melanocyte surface antigens and by indirect immunofluorescence (Naughton et al. 1983;
Bystryn & Naughton 1985; Bystryn & Xie 1998). Their presence has been confirmed by other techniques including complement-dependent cytotoxi- city (Norris et al. 1988; Cui et al. 1993; Yu et al. 19931, antibody-dependent cel- lular cytotoxicity (Norris et al. 1988; Yu et al. 1993), immunoblotting by live cell (Harning et al. 1991) and conventional enzyme-linked immunosorbent assay (ELISA) (Fishman et al. 1993), and passive transfer experiments (Gilhar et al. 1995). Antibodies to melanocytes are unusual in persons with nonpigmentary skin diseases.
The incidence and level of vitiligo antibodies correlates with the extent of depigmentation and the activity of the disease. They have been reported in only 50% of persons with minimal vitiligo but in 93% of those with more extensive disease (Naughton et al. 1986b). The average level of antibodies increases with increasing extent and activity of the disease (Harning et al.
1991; Cui et al. 1993). The titre of melanocyte antibodies decreases in vitiligo patients that respond to PUVA therapy (Hann et al. 19971, confirming that their level is related to disease activity.
A variety of animals develop vitiligo. All those studied have antibodies to pigment cells. These include Tervuren dogs, Siamese cats, Arabian horses (Naughton et al. 1986a), Sinclair miniswine (Cui et al. 1995) and Smyth chicken with vitiligo-like depigmentation (Austin et al. 1992). The pattern of antigens defined by pigment cell antibodies in these animals is similar to those recognized by vitiligo antibodies in humans. These observations indi- cate that humans and animals with vitiligo have similar immunological abnormalities.
Vitiligo antibodies have the functional capacity to lull pigment cells in vitro and can do so by two different mechanisms -complement-dependent cytotoxicity and antibody-dependent cellular cytotoxicity (Norris et al. 1988;
Cui et al. 1993). They also have the ability to kill melanocytes in vim, when passively administered to nude mice grafted with human skin (Gilhar et al.
1995).
Cellular immune responses to pigment cells in vitiligo
Inflammatory cells, a marker for involvement of cellular immune mecha- nisms, are usually sparse in lesions of vitiligo but not completely absent (Hann et al. 1992; A1 Badri et al. 1993a) (see Chapter 5). When present, they are most prominent at the periphery of active lesions (A1 Badri et al. 1993b;
Le Poole et al. 1996). This has variously been interpreted to mean that cellu- lar immunity is not involved in the pathogenesis of vitiligo (because the
infiltrate is sparse) or is involved (because the infiltrate is more prominent CHAPTER 1 6 where the lesions are most active). The infiltrate consists of CD3+ (T cells),
CD4+ (helper T cells), and CD8+ (cytolytic T cells) cells and macrophages.
The T cells are activated as evidenced by increased expression of class I1 HLA antigens and intercellular adhesion molecules (ICAM-I) (A1 Badri et al. 1993b1, and are closely associated with zones of melanocyte depletion (A1 Badri et al. 1993a). The characteristics of the infiltrate do not differ from those seen in other inflammatory skin diseases.
Unfortunately, there are few direct studies of the ability of cellular mechanisms specifically to kill pigment cells in vitiligo. An early study by Mitchell et al. (1980) found that lymphocytes from vitiligo patients were unreactive to melanoma cells, though it is unclear whether the melanoma cells used were pigmented. In Smyth chickens with vitiligo-like amelanosis, the infiltrate follows the appearance of morphologically abnormal melanocytes, but actual disappearance of melanocytes does not occur if the infiltrate is prevented by neonatal bursectomy (Boissy et al. 1984). This sug- gests that the infiltrate does not initiate injury to melanocytes but aggra- vates it. Whether the same is true in humans is not known.
While these observations suggest that cellular mechanisms may play a role in the pathogenesis of vitiligo, the evidence is much weaker than that for antibody responses.
h m u n e Hypothesis
Vitiligo antigens
The pigment cell antigens defined by antibodies in patients with vitiligo called 'vitiligo antigens' play a critical role in the pathogenesis of this disease as they may be both the cause and the target of immune responses that damage melanocytes.
Vitiligo antibodies are predominantly directed to melanocyte antigens of 4045, 75 and 90 kDa (referred to as VIT40, VIT75, and VIT90, respec- tively) and occasionally to other antigens of 35 and 150 kDa (Cui et al. 1992).
All these antigens are expressed on the cell surface. The most interesting is VIT90, because it is selectively expressed on pigment cells, particularly melanocytes. By contrast VIT40 and VIT75 are common tissue antigens expressed on both pigment and unrelated cells, and expressed more strongly on melanoma than on melanocytes. Little is known about the iden- tity of these antigens, apart from VIT40 which shares a cross-reacting epitope with, or is tightly bound to, class I HLA. VIT90 is unrelated to cur- rently known pigment cells antigens of similar size, including gp100, p97, and SlOO (Cui et al. 1995). VIT75 may be related to tyrosinase or tyrosinase- related protein-1 (TRP-l), as all three molecules have similar molecular weights. However, the available evidence suggests this is unlikely. While antibodies to tyrosinase were initially reported in 61% of 26 patients with vitiligo (Song et al. 19941, subsequent studies have not confirmed this association. Some investigators have detected tyrosinase antibodies in only a minority (Baharav et al. 1996), 11% (Kemp et al. 1997), or none (Xie &
131
CHAPTER 16
Immune Hypothesis Bystryn 1996) of patients with active vitiligo in three subsequent studies.
Thus, antibodies to tyrosinase do not seem to be present often enough in vitiligo to be vitiligo antibodies.
TRP-1 has been suggested to be a vitiligo antigen as passive administra- tion of antibody to this molecule into mice causes loss of pigmentation (Hara et al. 1995). However, immunodepletion experiments show that VIT75 and gp75 are immunologically distinct (Cui et al. 1995). The protein, gp75, is a cytoplasmic antigen while VIT75 is on the cell surface. Antibodies to gp75 have not been found in patients with vitiligo (Song et al. 1994), and the depig- mentation caused by anti-gp75 is restricted to regrowing hairs (Hara et al.
1995) whereas pigment cells in hair follicles are usually spared in vitiligo.
Many molecules on pigment cells have a molecular weight of approximately 75 kDa. Several of these have partial sequence homology with tyrosinase. At this stage VIT75 does not appear to be tyrosinase or TRP-1, but a distinct antigen that may have some homology with these molecules.
That some vitiligo antigens are normal tissue antigens also expressed by nonpigment cells does not exclude their playing a role in the pathogenesis of vitiligo. Melanocytes are unusually susceptible to be damaged by immune mechanisms (Norris et al. 1988). The LD50 of melanocytes to perox- ide-mediated injury, a major mechanism by which immune cells damage their target, is 10 times greater than that of keratinocytes and 100 times greater than that of fibroblasts. Thus, immune reactions to normal antigens expressed by melanocytes could damage these cells while sparing more resistant unrelated cells expressing the same antigens, resulting in a selec- tive destruction of melanocytes.
The antigenic phenotype of melanocytes derived from hair follicles and from overlying epidermis in the same individual differ from each other (Tobin & Bystryn 1996). Hair follicle melanocytes express more of the anti- gens recognized by antibodies associated with alopecia areata and less of those recognized by vitiligo antibodies, while the reverse is true for epider- mal melanocytes. These differences may account for epidermal but not hair follicle melanocytes being preferentially destroyed in vitiligo, while the reverse is true in alopecia areata; and for the hair follicle serving as the source for melanocytes in lesions of vitiligo that repigment (Cui et al. 1991).