Anna Wankowicz-Kalinska

Alpha-type-1 polarized dendritic cells: A novel immunization tool with optimized CTL-inducing activity

Using the principle of functional polarization of dendritic cells (DCs), we have developed a novel protocol to generate human DCs combining the three features critical for the induction of type-1 immunity: (a) fully mature status; (b) responsiveness to secondary lymphoid organ chemokines; and (c) high interleukin-12p70 (IL-12p70)-producing ability. We show that IFN-α and polyinosinic:polycytidylic acid (p-I:C) synergize with the “classical” type-1-polarizing cytokine cocktail [tumor necrosis factor α (TNFα)/IL-1β/IFNγ], allowing for serum-free generation of fully mature type-1-polarized DCs (DC1). Such “α-type-1-polarized DC(s)” (αDC1) show high migratory responses to the CCR7 ligand, 6C-kine but produce much higher levels of IL-12p70 as compared to TNFα/IL-1β/IL-6/prostaglandin E2 (PGE2)-matured DCs (sDC), the current “gold standard” in DC-based cancer vaccination. A single round of in vitro sensitization with αDC1 (versus sDCs) induces up to 40-fold higher numbers of long-lived CTLs against melanoma-associated antigens: MART-1, gp100, and tyrosinase. Serum-free generation of αDC1 allows, for the first time, the clinical application of DCs that combine the key three features important for their efficacy as anticancer vaccines.

Local immune response in skin of generalized vitiligo patients. Destruction of melanocytes is associated with the prominent presence of CLA+ T cells at the perilesional site.

In situ immune infiltrates in lesional, perilesional, and nonlesional skin biopsies from patients with vitiligo were analyzed by immunohistochemistry and compared with immune infiltrates found in the skin of normal healthy donors and relevant disease controls. An increased influx of activated skin-homing T cells and macrophages were seen in the perilesional biopsies. The overall percentages of cutaneous leukocyte-associated antigen-positive (CLA+) T cells were similar to those found in normal healthy donors. This is compatible with the similar expression of E-selectin. Most strikingly, however, the CLA+ T cells in perilesional skin were mainly clustered in the vicinity of disappearing melanocytes, and 60% to 66% of these interacting T cells expressed perforin and granzyme-B. The perforin+/granzyme-B+ cells were not seen in locations different from that of disappearing melanocytes. Interestingly, the majority of the infiltrating T cells were HLA-DR/CD8+. Another hallmark of the present study is the focal expression of intercellular adhesion molecule (ICAM)-1 and HLA-DR in the epidermis at the site of interaction between the immune infiltrates and the disappearing melanocytes. The data presented in this study are consistent with a major role for skin-homing T cells in the death of melanocytes seen in vitiligo.

Immunopolarization of CD4 + and CD8 + T Cells to Type-1–Like is Associated with Melanocyte Loss in Human Vitiligo

Vitiligo is an autoimmune condition characterized by loss of epidermal melanocytes. High frequencies of melanocyte-reactive cytotoxic T cells in the peripheral blood of vitiligo patients and the observed correlation between perilesional T-cell infiltration and melanocyte loss in situ suggest the important role of cellular autoimmunity in the pathogenesis of this disease. We isolated T cells from both perilesional and nonlesional skin biopsies obtained from five vitiligo patients, then cloned and analyzed their profile of cytokine production after short-term, nonspecific expansion in vitro. Perilesional T-cell clones (TCC) derived from patients with vitiligo exhibited a predominant Type-1–like cytokine secretion profile, whereas the degree of Type-1 polarization in uninvolved skin-derived TCC correlated with the process of microscopically observed melanocyte destruction in situ. Detailed analysis of broad spectrum of cytokines produced by perilesional- and nonlesional-derived CD4+ and CD8+ TCC confirmed polarization toward Type-1–like in both CD4 and CD8 compartments, which paralleled depigmentation process observed locally in the skin. Furthermore, CD8+ TCC derived from two patients also were analyzed for reactivity against autologous melanocytes. The antimelanocyte cytotoxic reactivity was observed among CD8+ TCC isolated from perilesional biopsies of two patients with vitiligo. Finally, in two of five patients, tetramer analysis revealed presence of high frequencies of Mart-1–specific CD8 T cells in T-cell lines derived from perilesional skin. Altogether our data support the role of cellular mechanisms playing a significant part in the destruction of melanocytes in human autoimmune vitiligo.

A symbiotic concept of autoimmunity and tumour immunity: lessons from vitiligo

Vitiligo is a skin disease in which melanocytes (MCs) are eradicated from lesional epidermis, resulting in disfiguring loss of pigment. MCs are destroyed by MC-reactive T cells, as well as other non-immune and immune components. Similarities exist between the autoimmunity observed in vitiligo and the tumour immunity observed in melanoma immuno-surveillance. An analysis of these mechanisms might lead to the development of new therapies for both vitiligo and melanoma.

Autoimmune melanocyte destruction in vitiligo

V itiligo is a disorder of pigmentation characterized by the presence of milk-white skin macules. The term vitiligo may have evolved from either the Latin word vitium, meaning fault, or vitelius meaning spotted calf (Kovacs, 1998). Pigment-producing cells (melanocytes) are absent from vitiligo lesions (Le Poole et al, 1993b); their loss represents a key event in the pathogenesis of the disease. Vitiligo lesions can change in size and shape over time and can develop at any age, but in approximately half of all cases the disease onset is before the age of twenty (Lerner, 1959). Clinical presentations include (a) segmental vitiligo, characterized by lesions that occur in a dermatomal, asymmetric distribution (of limited clinical significance); (b) focal vitiligo, characterized by a limited number of small lesions; (c) generalized vitiligo, the most common type of vitiligo, where lesions occur with bilateral, symmetrical distribution; and (d) universal vitiligo, indicating complete or almost complete depigmentation (Mosher et al, 1993). The incidence of this disease varies greatly between populations (from 0.14% to 8.8%), but the worldwide incidence is considered to be between 1% and 2% (Mosher et al, 1993). As well as the enhanced ultraviolet (UV) sensitivity of depigmented spots, vitiligo also is a cosmetically disabling disorder that often leads to psychologic stress (Porter et al, 1979). In addition, due to the occurrence of similar hypopigmented skin lesions in the early phase of leprosy, vitiligo is a true social stigma in countries in which leprosy is endemic. This problem is probably best emphasized by the terminology used in southern India, where vitiligo is known as ven kushtam, meaning “white leprosy” (Mosher et al, 1993). There are three main hypotheses for the pathogenesis of vitiligo: self destruction, neural, and autoimmune (Kovacs, 1998). In a number of recent studies, strong evidence in favor of the autoimmune hypothesis has been obtained. This review will discuss the relevant new evidence for autoimmune melanocyte destruction in vitiligo.