Effect of donor Langerhans cells on corneal graft rejection.

Unlike other cutaneous surfaces, the central portion of the corneal epithelium is typically devoid of Langerhans cells. The absence of Ia+ Langerhans cells in the central cornea is of more than casual interest and may explain the immunologic privilege that is characteristic of corneal allografts. The present communication summarizes previous studies that examined the role of corneal Langerhans cells in eliciting alloimmune responses and corneal graft rejection in rodents. Under normal circumstances, corneal allografts are poorly immunogenic when residing in the avascular ocular graft bed even though the graft displays large quantities of alloantigens. The afferent blockade of the immune response can be circumvented by donor-derived Langerhans cells that serve as potent immunogens for all categories of corneal allografts except grafts involving allodisparity only at class I major histocompatibility complex loci. Thus, the presence of donor-derived Langerhans cells exerts profound effects on the fate of corneal allografts.     

Antigen-presenting cells in the induction of contact hypersensitivity in mice: evidence that Langerhans cells are sufficient but not required

One explanation for the fact that certain genetically defined strains of mice prove to be resistant to effects of low dose ultraviolet B radiation on the induction of contact hypersensitivity is that ultraviolet B resistant mice possess a second pathway for antigen presentation through the skin--a pathway that is independent of epidermal Langerhans cells and beyond the reach of the damaging effects of ultraviolet B light. As a corollary, ultraviolet-B susceptible mice would be expected to be deficient in this pathway. Several experimental strategies were employed to determine whether Langerhans cells are required for the induction of contact hypersensitivity by epicutaneously applied hapten. The results reveal that tape-stripped skin supports the induction of contact hypersensitivity, whereas surgical excision of hapten-painted skin within 1 h of application fails to permit the development of contact hypersensitivity. Because the former selectively eliminates epidermal Langerhans cells while the latter deletes both Langerhans cells and dermal antigen-presenting cells, we conclude that either Langerhans cells or dermal cells are sufficient to provide antigen presentation in the induction of contact hypersensitivity. When large amounts of hapten are painted epicutaneously, or when hapten is injected subcutaneously or painted on sub-dermal tissues, contact hypersensitivity also results, indicating that induction of contact hypersensitivity does not require that antigen processing and presentation be provided by cutaneous cells. Reasons are presented for concluding that under physiologic circumstances induction of contact hypersensitivity by epicutaneous hapten application relies primarily upon the antigen-presenting capabilities of epidermal (Langerhans cells) and dermal cells.     

Langerhans cells are not damaged in contact allergic reactions in humans

Fifteen biopsies from contact allergic reactions in sensitized patients were taken at different time intervals after application of simple chemicals. Two different procedures for electron microscopy were done. No signs of damage were ever seen in Langerhans cells. The authors are therefore of the opinion that there is still no proof for a target role of Langerhans cells in contact allergic reactions in humans.     

IL-10-producing Langerhans cells and regulatory T cells are responsible for depressed contact hypersensitivity in grafted skin

Although skin grafting is a common surgical technique, the immunological state of grafted skin remains unelucidated. An experimental model has shown that the development of murine contact hypersensitivity (CHS) is depressed when mice are sensitized with a hapten through full-thickness grafted skin. We explored the immunological mechanisms underlying this hyposensitization, focusing on the fate of Langerhans cells (LCs). When FITC was applied to grafted skin, FITC-bearing LCs were capable of migrating to the draining lymph nodes. Epidermal cell suspensions isolated from the grafted skin produced a high amount of IL-10 as assessed by real-time PCR. Adoptive transfer of immune lymph node cells from the sensitized mice suppressed the CHS response of recipients in an antigen-specific manner. CD4(+)CD25(+) but not CD4(+)CD25(-) T cells purified from lymph node cells were responsible for this suppression. Finally, we detected high expression of receptor activators of nuclear factor kappa-B ligand (RANKL) in the grafted skin, and found that recombinant RANKL stimulated LCs to produce IL-10. These findings suggest that the hyposensitization of CHS through the grafted skin is not attributable merely to the reduction of LC number but that IL-10-producing LCs exert a downmodulatory effect by inducing regulatory T cells.     

Enriched epidermal Langerhans cells are potent antigen-presenting cells for T cells

Human epidermis was separated from dermis by means of a suction blister device and dissociated with trypsin. The epidermal cell (Ec) suspensions contained 2-6% Langerhans cells (Lc). Using a new rosette technique for enrichment of Lc, suspensions were obtained that contained 50-92% viable Lc. Ec, enriched Lc, or peripheral blood monocytes (Mo) were cocultured with or without antigens (Candida albicans, herpes simplex virus) and autologous T lymphocytes from sensitized donors. Strong proliferative T-cell responses were obtained provided Ec, enriched Lc, or Mo were also present. Furthermore, Lc were more effective than similar numbers of Mo in inducing T-cell responses to the antigens tested, and Lc did not require the presence of significant numbers of keratinocytes to exert this function.     

Langerhans cells in normal human skin capillaries

Cells with Langerhans granules are reported for the first time in normal human skin capillaries.     

Dermal dendritic cells,but not Langerhans cells,are critical in murine single epicutaneous sensitization

A murine repeated protein‐patch model has been established to study epicutaneous sensitization in atopic dermatitis. This model has shown a predominant Th2 and a weak Th1 response in both BALB/c and C57BL/6 mice. However, Th responses induced in the repeated model are not consistent with the generally accepted theory that BALB/c and C57BL/6 mice are Th2 and Th1 prone and are representatives of human atopy and non‐atopy, respectively. In this study, a single protein‐patch model was established, which showed in addition to the Th2 response, a remarkable Th1 response in C57BL/6 mice, but not in BALB/c mice. Moreover, using muLangerin‐DTR mice, we demonstrated that dermal dendritic cells, but not Langerhans cells, are critical in single epicutaneous sensitization in both strains of mice.     

Biology of Langerhans cells: analysis by experiments to deplete Langerhans cells from human skin

In vivo studies have demonstrated that various treatments of skin, e.g., UV irradiation, topical corticosteroids, and tape-stripping, will temporarily deplete the epidermis of Langerhans cells (LC). Whether this loss represents simply a loss of cell surface markers unique to LC, or actual depletion of cells, is unknown. By design, normal human skin transplanted to the congenitally athymic (nude) mouse is a system devoid of circulating precursors for human LC. Because LC have been shown to be of bone marrow origin, any depletion of these cells in this system should be permanent. Treatments to deplete LC from human skin grafts on nude mice after grafting included: (a) large doses of UV radiation (400 mJ/cm2 every 48 h, X 3), (b) potent high-dose topical corticosteroids (2.5 mg betamethasone valerate/cm2 every day, X 5), (c) tape-stripping (X 20). Treatments before grafting included: (a) treating donor skin with 900 R of gamma irradiation, (b) complement fixing monoclonal antibody to Ia-like antigens of LC, followed by fresh complement, (c) monoclonal antibody conjugated to toxins. Quantitation of the number of LC was analyzed on control and treated epidermal sheets using immunodiagnostic reagents, anti-HLA-DR, and surface ectoenzymes , ATPase. Results show that both UV irradiation and topical corticosteroids reduce the number of LC by these analyses. However, within 3 weeks, recovery to pretreatment levels has occurred. X-irradiation and tape-stripping were without effect. Despite evidence that the monoclonal antibody, complement, and toxic systems were delivered to the LC within the epidermis, there is no evidence that these treatments resulted in a decrease in LC. It appears that LC are currently either long-lived or replaced locally from a proliferative pool and that certain cell membrane determinants of human LC are somewhat differentially sensitive to UV radiation and corticosteroids.     

Immunohistochemical study of Langerhans cells in skin tumors

The behavior of Langerhans cells in skin tumors was investigated by immunohistochemical techniques using OKT6. OKT6-positive cells were numerous in squamous cell carcinomas, seborrheic keratoses and keratoacanthomas. They were rare in basal cell carcinomas, Bowen's disease, eccrine poromas, extramammary Paget's disease and warts. In solar keratosis, the number of OKT6-positive cells was almost equal or slightly larger compared to the normal epidermis. These results indicate that the density of Langerhans cells may not correlate with the degree of malignancy but, to a certain extent, with the nature of the membrane of tumor cells occurring e.g. during keratinization.     

Epidermal Langerhans cells in myelodysplastic syndromes are abnormal

The myelodysplastic syndromes (MDS) represent clonal disorders of the hematopoietic stem cell that are associated with quantitative and qualitative disturbances of the peripheral blood cells and a high risk for the transition to overt leukemia. As epidermal Langerhans cells (LC) are bone-marrow-derived cells, we were interested to see whether they are altered in patients with MDS. Epidermal sheets were prepared from biopsies taken from the thighs of nine patients with MDS and five control persons and processed for immunoperoxidase staining of CD1a antigens. The density and morphology of CD1a+ cells (i.e., LC) was evaluated by visual assessment as well as automatic image analysis. The density of LC was reduced in seven of nine patients (range, 30-75% of normal), whereas the morphology of LC appeared to be altered in all MDS patients in that the LC displayed large and bizarre cell bodies with only a few and often abnormally long dendrites. The HLA-DR expression by LC was not altered, as shown by double immunofluorescence staining of CD1a and HLA-DR antigens. Ultrastructurally, LC again appeared enlarged and often presented with bizarre nuclei, yet displayed no other abnormalities. Our findings suggest that LC are abnormal in MDS and might even indicate a more wide-spread involvement of the dendritic cell lineage in this syndrome.     

Langerhans cells but not monocytes are capable of antigen presentation in vitro in corticosteroid contact hypersensitivity

Corticosteroids suppress delayed-type hypersensitivity (DTH) reactions in vivo and impair lymphoid cell functions in vitro. In contact hypersensitivity (CHS) to corticosteroids, however, the corticosteroids are capable of inducing DTH responses in vivo. The present study examined the capacity of corticosteroids to induce in vitro proliferation of T lymphocytes from patients with CHS to corticosteroids. With peripheral blood mononuclear adherent cells as antigen-presenting cells (APC) and hydrocortisone-17-butyrate (H-17-B) as hapten, no proliferation responses were detected of T lymphocytes from patients with CHS to H-17-B. However, when epidermal Langerhans cells (LC) were used as APC, weak proliferation responses were observed.     

Migration of Langerhans cells from carcinogen-treated sheep skin.

To define the mechanism(s) of carcinogen depletion of Langerhans cells (LC) from skin, the migration of LC from the skin to the regional lymph node was examined in carcinogen-treated, antigen-treated, and control sheep. This was assessed by cannulation of afferent lymphatic vessels that drain the treated areas of skin or the efferent lymphatic draining the regional lymph node. Cells draining from test or control skin were continuously collected and enumerated by indirect immunofluorescence and flow cytometry using specific anti-CD1 monoclonal antibodies. There was a marked increase in the rate of LC migration in the 8 h following the application of the contact sensitizing antigen trinitrochlorobenzene (TNCB). The chemical carcinogen 7,12-dimethylbenz(a)anthracene (DMBA) triggered a tenfold-greater migration of LC compared with TNCB--with the peak response at 5 d. After DMBA treatment LC were also detected in the efferent lymph of the regional lymph node. It is concluded that the depletion of LC from carcinogen-treated skin is due to the increased LC migration and not carcinogen-induced cell death.     

Further evidence for the self-reproducing capacity of Langerhans cells in human skin

The limited number of Langerhans cells (LC) in the epidermis is one of the main reasons for the technical difficulties in resolving the question of LC kinetics. In the present paper, we describe a method to evaluate the LC replication potential in epidermis. The procedure is based on the specific incorporation of bromodeoxyuridine (BrdU), a thymidine analogue, into the DNA during the S-phase of the cell cycle. Mice, bearing human skin grafts, were injected s.c. every 6 h for up to 17 days with BrdU. At different times, the incorporated BrdU as well as the human epidermal LC were revealed on skin sections using anti-BrdU and OKT-6 monoclonal antibodies, respectively. After 6 h, 4.9% of the LC were labeled with BrdU. Then, the number of OKT-6(+) BrdU(+) cells increased in a linear manner and achieved 34% at 120 h, 67% at 240 h, and 94% at 400 h during the course of continuous labeling procedures. Based on this result we calculated a total cell cycle time of 392 h (16.3 days) and 12 h for the S-phase for human epidermal LC. Applying this technique, we were able to show also that 48 h after local treatment with 12-O-tetradecanoylphorbol-13-acetate or after stripping, the number of BrdU-labeled LC was considerably increased. Furthermore, after i.p. injection of colchicine in the nude mouse, human epidermal LC undergoing mitosis were evidenced by electron microscopy in the graft. From these results we conclude that the LC are actively cycling--therewith a self-reproducing cell population in human epidermis.