Effects of ultraviolet irradiation on surface marker expression by epidermal immunocompetent cells and contact sensitization to dinitrofluorobenzene in mice

We studied the effects of ultraviolet (UV) irradiation on murine epidermal Ia-positive Langerhans cells (Ia + LC) and Thy-I-positive dendritic epidermal cells (Thy-I + dEC). We also studied contact hypersensitivity to dinitrofluorobenzene (DNFB) introduced through UV-treated epidermis. C3H/HeN mice were exposed to UVB or 8-methoxypsoralen plus UVA (PUVA). UVB and PUVA treatment led to a dramatic reduction in surface marker expression of both Ia + LC and Thy-I + dEC. High-dose UVB irradiation (360 J/m2) interfered with contact hypersensitivity to DNFB; the density of Ia + LC may thus be related to the sensitizing potential. In contrast, low-dose UVB (120 J/m2) and PUVA treatment had little effect on contact hypersensitivity despite a marked reduction in Ia + LC. The density of Thy-I + dEC appeared not to be associated with contact hypersensitivity. These results suggest that there may be a Langerhans cell density-independent mechanism for the induction of contact hypersensitivity.     

Immunohistochemistry of lymphocytes and Langerhans' cells in long-lasting allergic patch tests

A long-lasting allergic patch test is a "normal" allergic patch test that remains positive for weeks or months. An immunohistochemical study of immunocompetent cells in the skin in this rare type of patch tests was performed. Most inflammatory cells were T11 positive T-lymphocytes. The majority of these cells were of the helper/inducer phenotype (T4+), but a relative increase of T8+ cells as compared to the initial (1-2d) stages of allergic patch tests was observed. T6+ Langerhans' cells (LCs) were normal or increased in number in the epidermis, while very few dendritic cells displayed Ial antigen in the epidermis, indicating loss of Ial-staining of LCs. High to very high numbers of T6+ cells were found in the dermis. An inflammatory reaction of hair follicles with moderate numbers of T6+ cells in the peribulbar infiltrate was observed indicating that hair follicles might act as shunt pathways for allergens. A defect in down regulation of the contact hypersensitivity reaction and/or a constant antigen stimulation could be responsible for the long-lasting allergic patch tests.     

Langerhans' cells in hair follicles of the depigmenting C57Bl/Ler-vit.vit mouse. A model for human vitiligo

The C57Bl/Ler-vit.vit mouse grows a black pelage after birth. During successive hair molts, the fur loses its pigmentation. By 6 months of age, most of the fur of the animal is white. The epidermis of the ears and tail also loses its pigmentation. Histologic studies confirm that in the epidermis and hair follicles there is an absence of pigment cells identifiable by various histochemical or electron microscopic techniques. This mouse may be an excellent model in which to study the role of Langerhans' cells and the immune response in the pathogenesis of vitiligo, a study not easily done in humans. From results of prior studies, we postulated that if Langerhans' cells were involved in the destruction of melanocytes, they would be abnormal (either more or less numerous) in number during the active phase of depigmentation and normal in number after depigmentation was complete. To determine whether the Langerhans cell (Ia+/adenosine triphosphatase dendritic epidermal cell) might be involved in destruction of pigment cells, we quantified the number of Ia+ and adenosine triphosphatase dendritic cells in the hair follicles in skin from the ear, abdomen, back, and tail from male C57Bl/Ler-vit.vit mice while the fur and skin were depigmenting and after depigmentation was almost completed. We found that Langerhans' cells were normal in number during depigmentation and were most numerous after depigmentation. Previous studies indicate that Langerhans' cells in these mice are functionally defective and respond poorly to some contact allergens. From these morphologic and functional data, we conclude that Langerhans' cells probably are uninvolved in causing depigmentation in these mice. We also observed that the epithelium of hair follicles has a significantly higher (up to 1600/mm2) population density of Langerhans' cells than interfollicular skin.     

Distinctive dendritic cell subsets expressing factor XIIIa, CD1a, CD1b and CD1c in mycosis fungoides and psoriasis

The papillary dermis of psoriasis and mycosis fungoides (MF) lesions is characterized by prominent collections of cells with dendritic morphology. Immunophenotypically distinct populations of cutaneous dendritic cells have been identified as CD1a+, FXIIIa-Langerhans cells (LC) and CD1a-, FXIIIa+ dermal dendritic cells (DDC). In this study, antibodies against the human GDI cluster of antigens (i.e. CD1a, CD1b and CD1c) and the DDC) marker (FXIIIa) were used to further characterize the subsets of dendritic cells in normal skin as compared to neonatal foreskin, psoriasis and MF by both immunoperoxidase and double immunofluorescence techniques. Normal skin and foreskin epidermis and dermis contained few CD1b+ or CD1c+ cells along with normal numbers of CD1a+ LC and FXIIIa+ DDC. Both MF and psoriasis were characterized by CD1a+ cells in the epidermis and dermis. FXIIIa+ cells were greatly expanded in the upper dermis of MF lesions and to a lesser degree in psoriasis as has been previously described by our group. MF contained significantly increased epidermal and dermal CD1b+ (15.7/5 high power fields [HPF] and 59.7/5 HPF respectively) and CD1c+ dendritic cells (33.8/5 HPF and 95.9/5 HPF respectively), while in psoriasis these cells were not statistically different from normal skin. Double immunofluorescence studies revealed that some (<25%) FXIIIa+ cells co-expressed CD1b and CD1c in MF>psoriasis> foreskin, while FXIIIa+ DDC never co-expressed CD1a. Thus, in contrast to normal skin in which epidermal or dermal dendritic cells rarely express CD1b and CD1c antigens, these members of the CD1 family are upregulated on both LC and DDC in benign and malignant inflammatory states. Upregulation of CD1b and CD1c on MF epidermal and dermal dendritic cells, as compared to psoriasis, foreskin and normal skin, may be useful in the immunophenotypic recognition of MF, as well as in helping to understand its immunobiology.     

Immunocompetent cells of fixed drug eruption

The positive provocation test reactions of the skin of six patients with fixed drug eruption (FDE) were studied from timed skin biopsies taken between 2 hours and 9 days after the appearance of FDE. Monoclonal antibodies to the following immunocompetent cell surface epitopes were used: T3, T4, T6, T8, T9, M1, Ia1, Drc, Leu7 and B cell. The dermal infiltrate comprised 60-80% of T lymphocytes at all the times studied. Cells with T4 and T8 epitopes were displayed in similar numbers. A transient decrease in the number of T6+ cells of the epidermis could be detected with a simultaneous and also transient increase of the T6+ cells in the dermis, which suggests a possible traffic of Langerhans' cells from the epidermis to the dermis. The epidermal Ia1+ cells showed changes similar to but less marked than the T6+ cells. The number of the dermal Ia1+ cells increased continuously. In the late biopsies these Ia1+ cells comprised up to 90% of the infiltrating cells. Except for the finding of a reduction of T6+ and Ia1+ epidermal cells, the cellular kinetics of FDE are similar to those seen in both cutaneous immunological and irritant reactions.     

A novel monoclonal antibody to a distinct subset of cutaneous dendritic cells.

A monoclonal antibody was generated by immunizing rats with Langerhans cell (LC)-enriched epidermal cells obtained from BALB/c mouse earskin after epicutaneous application of the contact sensitizer 2,4-dinitrofluorobenzene (DNFB). The antibody 4F7 detects in normal mouse skin, few dermal cells showing the morphologic, phenotypic, and functional properties of accessory dendritic cells, but lacking Birbeck granules. The capacity to stimulate allogenic T cells in the mixed leucocyte reaction resembles that of freshly isolated LCs. After DNFB application, an increased number of 4F7+ dendritic cells are found in the dermis and, in addition, some labeled dendritic cells occur in the epidermis. Some of the latter cells exhibit cytoplasmic Birbeck granules. Remarkably, there is no increase of the 4F7+ cells in the regional lymph nodes after DNFB treatment. These data suggest that the 4F7 antibody labels distinct dendritic cells of the mouse skin that are involved in the mediation of contact sensitization and probably represent immature LCs.     

Fate of melanocytes during development of the hair follicle pigmentary unit

During hair follicle morphogenesis, melanocyte precursors migrate into developing hair follicles and give rise to differentiated melanocytes that actively produce and transport pigment into the keratinocytes that form the hair shaft; however, patterns of melanocyte proliferation and differentiation during formation of the hair pigmentation unit remain to be elucidated. Using multicolor confocal microscopy and double immunofluorescence of melanogenic proteins (tyrosinase-related proteins 1 and 2, tyrosinase) and the proliferative marker Ki67, we have studied melanocyte development in C57BL/6 mouse embryonic hair follicles. Proliferating melanocyte precursors (tyrosinase-related protein-2/Ki67+ cells) are seen in the hair follicles at stages 1-2 of morphogenesis, as follicular invagination begins. In stage 3-4 hair follicles, the majority of intrafollicular melanocytes remain tyrosinase-related protein-2+ and Ki67+, whereas some located adjacent to the forming dermal papilla begin to express tyrosinase-related protein-1, an early marker of differentiation. Melanin granules appear in stage 5 hair follicles coincident with tyrosinase expression in nonproliferating tyrosinase-related protein-2+/tyrosinase-related protein-1+ melanocytes. Stage 6-8 hair follicles, those actively producing hair, show nonproliferating tyrosinase-related protein-2+ melanocytes in the bulge area, tyrosinase-related protein-2+/tyrosinase-related protein-1+ melanocytes in the outer root sheath, and tyrosinase-related protein-2+/tyrosinase-related protein-1+/tyrosinase+ melanocytes above the dermal papilla. These data suggest that melanocyte precursor cells proliferate extensively at the onset of follicle development. Progeny of these cells migrate down the developing follicle, differentiating further until reaching the area immediately above the dermal papilla, where fully differentiated nonproliferative melanin-producing melanocytes persist, contributing pigment to the growing hair shaft.     

Proliferating cells in psoriatic dermis are comprised primarily of T cells, endothelial cells, and factor XIIIa+ perivascular dendritic cells

Determination of the cell types proliferating in the dermis of patients with psoriasis should identify those cells experiencing activation or responding to growth factors in the psoriatic dermal milieu. Toward that end, sections of formalin-fixed biopsies obtained from 3H-deoxyuridine (3H-dU)-injected skin of eight psoriatic patients were immunostained, followed by autoradiography. Proliferating dermal cells exhibit silver grains from tritium emissions. The identity of the proliferating cells could then be determined by simultaneous visualization with antibodies specific for various cell types. UCHL1+ (CD45RO+) T cells (recall antigen-reactive helper T-cell subset) constituted 36.6 +/- 3.1% (mean +/- SEM, n = 6) of the proliferating dermal cells in involved skin, whereas Leu 18+ (CD45RA+) T cells (recall antigen naive T-cell subsets) comprised only 8.7 +/- 1.5% (n = 6). The Factor XIIIa+ dermal perivascular dendritic cell subset (24.9 +/- 1.5% of proliferating dermal cells, n = 6) and Factor VIII+ endothelial cells (23.0 +/- 2.3%, n = 6) represented the two other major proliferating populations in lesional psoriatic dermis. Differentiated tissue macrophages, identified by phase microscopy as melanophages or by immunostaining with antibodies to Leu M1 (CD15) or myeloid histiocyte antigen, comprised less than 5% of the proliferating population in either skin type. In addition to calculating the relative proportions of these cells to each other as percent, we also determined the density of cells, in cells/mm2 of tissue. The density of proliferating cells within these populations was increased in involved versus uninvolved skin: UCHL1+, 9.0 +/- 1.7 cells/mm2 versus 1.8 +/- 0.6 cells/mm2, p less than 0.01; Factor XIIIa+, 6.0 +/- 0.7 cells/mm2 versus 1.5 +/- 0.5 cells/mm2, p less than 0.01; Factor VIII+, 5.5 +/- 1.4 cells/mm2 versus 0.0 cells/mm2, p less than 0.05. The presence of preferential active proliferation of a T-cell subset in lesional dermis suggests that activating signals specific for this subset are contained within the psoriatic dermis in vivo. The activation of recall antigen-reactive T cells may be a driving force behind the dendritic cell and endothelial cell proliferation. Alternatively, the selective proliferation and expansion of these two constitutive cell types (Factor XIIIa+ and Factor VIII+) may result in signals that promote activation of UCHL1+ (CD45RO+) T cells.     

Ontogeny of Ia-positive and Thy-1-positive leukocytes of murine epidermis

Murine epidermis harbors 2 populations of dendritic leukocytes: Langerhans cells (LC) and Thy-1-positive dendritic epidermal cells (Thy-1 +DEC). In the adult mouse these cell types are morphologically distinct and display highly characteristic phenotypes. LC bear Ia-antigens and a group of markers typical for mononuclear phagocytes: Fc- and C3bi-receptors, macrophage-specific antigen F4/80, and membrane ATPase. Thy-1 +DEC, in contrast, lack these markers but express high levels of Thy-1 and asialo-GM1 (asGM1) antigen. Since LC and Thy-1 +DEC share a common origin from the bone marrow we expected to gain insight into their relationship by studying their ontogenetic development. Epidermal sheets from fetal and newborn C3H/He and C57B1/6 mice obtained at defined ages from day 17 of gestation up to day 30 of postnatal life were monitored for the emergence of the above-mentioned markers for LC and Thy-1 +DEC. In double-labeling experiments LC markers were first detected by visualizing the monoclonal antibodies by a sensitive triple-layer rhodamine-immunofluorescence technique; in a second step, after appropriate blocking procedures, Thy-1 and asGM1 antigens were demonstrated by direct and indirect immunofluorescence. We found that in fetal epidermis, only few cells expressed either Thy-1 or Ia (4 and 1 cells/mm2, respectively, on day 18 of gestation). The bulk of Thy-1 +DEC and Ia +EC appeared only after birth. Adult proportions of Thy-1 +DEC and Ia +EC were reached at around 1 month of postnatal life. In contrast, all the other LC markers were expressed on a substantial number of fetal dendritic cells (280 cells/mm2 on day 18 of gestation), indicating the presence of phenotypically immature Ia-negative LC in fetal epidermis. By day 4 of postnatal life all F4/80 +EC and ATPase +EC (i.e., LC) had acquired Ia-antigens. Surprisingly, LC also bore asGM1 antigens, which in the adult epidermis are strictly confined to Thy-1 +DEC, up to day 5 of postnatal life. Thus, LC in fetal and early newborn epidermis are not yet fully differentiated. As they differentiate, they acquire Ia antigens and lose asGM1 antigens. In contrast, a phenotypically immature Thy-1 +DEC population could not be traced with the markers used. Thy-1 +DEC appear to be characterized by a stable phenotype (Thy-1+/asGM1+) throughout their lifetime.     

Cellular kinetics of delayed hypersensitivity test reactions to topical glucocorticosteriods

Summary The phenotypes of the infiltrating cells in 13 patients with delayed hypersensitivity to topical glucocorticosteriods (GCS) were studied from sequential biopsies of positive epicutaneous test reactions by using the avidin-biotin-complex (ABC) technique. Monoclonal antibodies were used to identify the cells with the following phenotypes: T3, T4/T4a, T6, T8, T9, T11, M1, Ia1 (HLA-DR), interleukin-2 receptor/T26a, and dendritic reticular cell. The cellular kinetics of GCS hypersensitivity reactions were compared with delayed hypersensitivity reactions caused by allergens not related to GCS. In both GCS and non-GCS reactions the epidermal dendritic T6 + cells were more numerous than dendritic Ia1 + cells. There was a decrease in the number of both cell types during these reactions; in GCS reactions the decrease in the number of T6 + cells was seen later than in non-GCS reactions. Ia1 + keratinocytes were seen at sites near dermal infiltrates. Compared with the non-GCS delayed hypersensitivity reaction, there were fewer pan T (T11 +/T3+) in the GCS reaction. The relative numbers of M1 + monocytes and the T4/T8 ratio were substantially lower in the latter; these findings can be explained as a GCS effect which modulates the delayed type hypersensitivity reaction.     

Distribution of CD1a-positive cells in psoriatic skin during the evolution of the lesions

Normal skin and psoriatic lesions from 35 patients were investigated immunohistochemically with regard to the CD1a+ cell population (Langerhans' cells and indeterminate cells) in the epidermis as well as in the dermal infiltrate. In the normal-appearing skin, we found the regularly typical pattern of CD1a+ dendritic cells in suprabasal position, but in lesional skin of chronic psoriasis the CD1a+ cells were scattered in the acanthotic epidermis. In initial lesions, CD1a+ cells represent up to 50-60% of the infiltrating cells of the dermal compartment, in several cases being preferentially localized in the upper part of the papillar dermis close up to the epidermal CD1a+ cells in basal position, whereas in chronic psoriasis they represent less than 10%. These results suggest that in psoriasis vulgaris, CD1a+ cells actively migrate between the epidermis and the dermal vessels.     

A model system using tape stripping for characterization of Langerhans cell-precursors in vivo

Little is known about the immigration of bone marrow-derived progenitors of Langerhans cells (LC) into the epidermis. We developed an in vivo system based on the tape stripping method that allowed us to study the immigration of LC into the epidermis after intradermal injection of bone marrow-derived dendritic cells (DC). Tape stripping induced a mechanical disruption of the epidermal barrier that led to skin inflammation and subsequent emigration of LC and dermal DC from the skin. Emigrating LC and dermal DC were observed in lymphatic vessels, and the numbers of LC and dermal DC in the draining lymph node increased. Up to 500 times more injected precursors migrated into tape-stripped epidermis as compared with unstripped epidermis. Newly immigrated cells were slender with one or two dendrites and acquired a more dendritic morphology after 2-4 days. They were both MHC II-positive and negative and they did not express Langerin/CD207, nor macrophage-mannose receptor/CD206 and Fc-epsilon receptor I. In contrast, all cells that had entered the epidermis expressed CD11c and CCR6, suggesting that they were LC. We conclude that this experimental system may serve as a valuable tool for the further characterization of LC-precursors and the conditions necessary for LC-immigration into the epidermis.     

Changes in expression of Ia, Thy-1, and Ly-5 antigens in epidermal cells during delayed contact sensitivity reactions in mice: flow cytometric analysis

Ia antigen-bearing (Ia+) Langerhans cells have attained an important position as immunocompetent cells in the epidermis. Recently there have been successive reports on other new possible candidates for immunocompetent cells in the epidermis, i.e., Ia+ keratinocytes and dendritic Thy-1 antigen-bearing (Thy-1+) epidermal cells which also express Ly-5 antigen and asialo-GM1. Based on our previous findings that in allergic contact sensitivity reactions, keratinocytes express Ia antigen 3-9 days postchallenge, in this report, we have attempted to define more clearly the dynamic changes of Ia+ keratinocytes and dendritic Thy-1+ epidermal cells by enumeration of the precise percentages of Ia+, Thy-1+, and Ly-5 antigen-bearing (Ly-5+) cells in epidermal cells at various times of the challenge phase in allergic contact sensitivity reactions by use of a fluorescence-activated cell sorter. By 24 h postchallenge, the percentages of Ia+, Thy-1+, and Ly-5+ cells showed hardly any change. There were approximately 2% Ia+ cells, 50% Thy-1+ cells which consist of 2 populations (i.e., 45% weakly Thy-1 antigen-positive cells and 4% strongly Thy-1 antigen-positive cells), and 3.5% Ly-5+ cells. From 48 h postchallenge, however, the percentage of Ia+ cells and that of Thy-1+ cells began to increase and reached a plateau, with approximately 20% Ia+ cells and 70% Thy-1+ cells, respectively, at 120 h postchallenge. The change of the percentages of Ly-5+ cells appears to correspond to that of strongly Thy-1 antigen-positive cells. Only at 48 h postchallenge, Ly-5+ cells and strongly Thy-1 antigen-positive cells showed a small increase in number, comprising approximately 10% of the epidermal cells. These data suggest that among Thy-1+ epidermal cells, strongly Thy-1 antigen-positive cells correspond to dendritic Thy-1+ epidermal cells, and in contact sensitivity reactions in mice, dendritic Thy-1+ epidermal cells show only a minor dynamic change in contrast to Ia+ cells, in which more than 15% of keratinocytes express Ia antigen from 48 h postchallenge.     

Alterations in cutaneous immune reactivity to dinitrofluorobenzene in graying C57BL/vi.vi mice

The fur of the C57BL/vi.vi mouse is black at 6 weeks of age. By 6 months of age the animals are white and there are no identifiable pigment cells within the epidermis or hair bulbs. Human subjects with vitiligo exhibit loss of epidermal pigment cells. The loss of pigment cells in human subjects with vitiligo has been associated with loss of cutaneous immune reactivity to contact allergens. Therefore, studies were performed to determine whether loss of pigment cells in these depigmenting mice also was associated with loss of the cutaneous immune response. The number of Ia-positive (Ia +) Langerhans cells (LC)/mm2 on the back and the ear, the sites of sensitization and challenge with dinitrofluorobenzene (DNFB), was quantified before, during, and after depigmentation. We observed that there were fewer LC/mm2 on the back and the ear before and after pigment loss in the graying mice than in the normal control C57BL/6 mice. The young pigmented C57BL/vi.vi mice were capable of developing moderate contact hypersensitivity; the older depigmented mice did not sensitize to DNFB. We conclude that the depigmented mice, like human subjects with vitiligo, have a loss of contact hypersensitivity associated with a loss of pigment cells within the epidermis. In the mouse, loss of melanocytes is associated with a decrease in the population density of Ia + cells.     

A comparative study of allergic and primary irritant contact dermatitis with dinitrochlorobenzene (DNCB) in dogs.

Attempts were made to induce allergic contact dermatitis in dogs, a species generally considered poorly responsive to experimental allergic contact dermatitis. Yound Beagles were sensitized to 2,4 dinitrochlorobenzene (DNCB) by multiple intradermal injections. Two weeks after sensitization, these dogs were challenged topically with 0.1% DNCB by a standard closed-patch technique. Sensitization evidenced by various degrees of reaction following challenge was established in all of 14 pups used, while 7 nonsensitized control pups did not react to challenge. Primary irritant contact dermatitis was induced in the skin of nonsensitized Beagle pups by 1%, 5%, and 10% solutions of DNCB. In allergic contact dermatitis the sites of challenge were grossly indurated, erythematous, and edematous. Histologically at these sites there was an infiltration of mononuclear cells which reached maximum intensity at 3 to 4 days. Accumulations of lymphoid cells were marked around sweat galnds and hair follicles. Penetration of leukocytes into these cutaneous adnexa was associated with degenerative processes in their cellular structures. Mononuclear cell infiltration into the epidermis was mild. Spongiosis was observed in the epidermis, but vesicle formation was rare. In primary irritant contact dermatitis gross lesions were characterized by severe erythema, edema, and gangreen of the skin. Microscopically, the main lesions were necrosis of the epidermal cells, separation of the epidermis from the dermis, dermal edema, and massive infiltration of the dermis with polymorphonuclear cells.     

Antigen-presenting cells in essential fatty acid-deficient murine epidermis: keratinocytes bearing class II (Ia) antigens may potentiate the accessory cell function of Langerhans cells.

Essential fatty acid deficiency (EFAD) is a useful model for studying the role of (n-6) fatty acid metabolism in normal physiology. Because cutaneous manifestations are among the earliest signs of EFAD and because abnormalities in the distribution and function of tissue macrophages have been documented in EFAD rodents, we studied the distribution and function of Class II MHC (Ia) antigen-bearing cells in EFAD C57B1/6 mouse epidermis. Immunofluorescence studies revealed 1.9-9.6 (mean +/- SEM = 5.2 +/- 2.6) times more class II MHC (Ia) antigen-bearing epidermal cells in suspensions prepared from EFAD as compared to normal skin. Analysis of epidermal sheets demonstrated similar numbers of dendritic Ia+ and NLDC145+ cells in EFAD and normal epidermis, however. This discrepancy occurred because some keratinocytes in EFAD epidermal sheets expressed class II MHC (Ia) antigens, whereas keratinocytes in normal mouse epidermis did not. Two-color flow cytometry confirmed that all Ia+ cells in normal epidermis are Langerhans (Ia+ NLDC145+) cells, whereas Ia+ cells in EFAD epidermis are comprised of Langerhans cells and a subpopulation of keratinocytes (Ia+ NLDC145-). Similar levels of Ia antigens were expressed on EFAD and normal Langerhans cells. EFAD and normal epidermal cells were also compared in several in vitro assays of accessory cell function. Epidermal cells prepared from EFAD C57B1/6 mice present the protein antigen DNP-Ova to primed helper T cells more effectively than epidermal cells prepared from normal animals. EFAD epidermal cells are also more potent stimulators of T cells in primary and secondary allogeneic mixed lymphocyte-epidermal cell reactions than normal epidermal cells. The functional differences between EFAD and normal epidermal cells do not appear to result from increased cytokine release or decreased prostaglandin production by EFAD epidermal cells. In view of these findings and the observation that the antigen-presenting cell activity of EFAD epidermal cells correlates with the number of Ia+ keratinocytes in epidermal cell preparations, Ia+ keratinocytes (in the presence of Langerhans cells) may potentiate cutaneous immune responses in vitro and perhaps in vivo as well. These results also suggest that (n-6) fatty acids or metabolites of (n-6) fatty acids are involved in regulating the expression of class II MHC (Ia) antigens by keratinocytes in vivo.     

The functional maturation of Langerhans cells in CTL induction during tissue culture

In the previous report, we investigated the effects of tissue culture on the APC function of murine epidermal Langerhans cells (LC) in the induction of allo-CTL in vitro, and found that (1) cultured ear LC expressed increased amounts of Ia antigens on their cell surface, and (2) they induced extremely enhanced levels of CTL over those produced by freshly prepared ear LC and also (3) cultured tail LC were proved to be able to induce CTL for the first time. It seemed that tissue culture decreased the functional heterogeneity among the murine ear LC and tail LC in addition to that among Ia+ APC in spleen and in epidermis. In this study, we investigated the culture conditions that increase the APC function of LC. Our data indicate that LC cultured with dermal components exhibited more enhanced APC function than LC cultured in single cell suspension with only epidermal cells. Recent studies indicate that IL-1 and GM-CSF, which keratinocytes release, are essential for freshly prepared LC to mature into highly efficient APC that resemble splenic dendritic cells. We found dermal factors are more important than epidermal ones for LC to mature in tissue culture.     

Langerhans cell migration into ultraviolet light-induced squamous skin tumors is unrelated to anti-tumor immunity.

There has been much speculation as to the role of Langerhans cells (LC) in the induction of anti-tumor immunity. Whereas there is considerable circumstantial evidence that disruptions in the density and function of these cells during the early stages of ultraviolet (UV) light- and chemical carcinogen-induced carcinogenesis may be important for enabling developing neoplasms to escape immune destruction, the role of the large number of these cells found infiltrating developed skin tumors is less clear. To investigate this we have compared the LC density infiltrating transplanted non-immunogenic and immunogenic UV-induced murine tumors as well as LC in the epidermis overlying the tumors. Whereas two non-immunogenic tumor lines attracted large numbers of Ia+ dendritic cells, an immunogenic tumor line did not. Similar results were obtained whether the tumors were transplanted into syngeneic immunocompetent or athymic immunodeficient mice. Hence, there was no relationship between tumor immunogenicity or host immunocompetence and Ia+ dendritic cell density. Furthermore, there was no correlation with the pattern of T-cell infiltration of the tumors or CD4/CD8 cell ratio. Our results also indicate that whereas UV light decreased Ia+ cell density, both in the epidermis and the tumors, it did not inhibit the tumors from attracting Ia+ dendritic cells. Thus, the Ia+ dendritic cells infiltrating skin tumors are unlikely to indicate a host immune response to the tumor, but are more likely to be attracted by tumor-derived cytokines.     

Distribution of major histocompatibility antigens in normal skin

The distribution of major histocompatibility antigens HLA-A,B,C (HLA), beta 2-microglobulin (beta 2m), and Ia-like antigens (HLA-DR; Ia) in normal skin was studied in frozen tissue sections by a four-step immunoperoxidase method and an avidin-biotin method employing monoclonal antibodies. HLA and beta 2m were present on the basal and spinous keratinocytes of the epidermis, on the outer root sheath epithelium in the infundibulum of the hair follicle, and on the excretory sebaceous duct epithelium. Ia-positive dendritic cells were found in the epidermis and hair follicles, but they were more frequent in the infundibulum and isthmus of the hair follicle than in its inferior portion or in the epidermis. In the straight eccrine duct, HLA and beta 2m-positivity was most striking in its lower portion. In the superficial duct, there was a less intense staining using the four-step procedure, but when an avidin-biotin method was used, the difference was less apparent. In contrast, the acrosyringial epithelium was markedly Ia-positive with decreasing intensity of staining as the duct penetrated the dermis. No HLA or Ia antigens were identified in eccrine glands and apocrine glands. Eccrine glands were slightly beta 2m-positive. HLA and beta 2m were uniformly present in non-dilated and dilated intradermal apocrine ducts.     

Distributional changes of Langerhans cells in human skin during irritant contact dermatitis

We used light and electron microscopic immunocytochemistry to study distributional changes in the human Langerhans cell (LC) system during the first 14 days of a mild irritancy caused by sodium lauryl sulphate (SLS). A marked initial decrease in epidermal LC was noted possibly resulting from migration from the epidermis to the dermis and from irreversible cell damage. Several studies have previously found an unchanged number of LC in SLS-induced contact irritant dermatitis, but these studies may not have taken into account the fact that SLS is effectively absorbed from the test chamber. Unless certain precautions are taken the SLS concentration rapidly falls to topical levels that have no effect on the LC system. Simultaneously with the decrease in the epidermis we observed an increase in dermal CD1a+ cells, confirming an often reported finding. There is, however, no consensus as to the identity of these cells, and several authors have reported that such cells lack LC granules and thus these cells have often been classed as indeterminate cells. We found that, during irritant contact dermatitis, provided an adequate number of sections were scrutinized in the electron microscope, all dermal CD1+ cells contained Birbeck granules.