D.-H. Kalden

The Role of α‐MSH as a Modulator of Cutaneous Inflammation

Abstract: Among various neuropeptides such as substance P, calcitonin gene‐related peptide and others, α‐melanocyte‐stimulating hormone (α‐MSH) was found to be produced in the skin. Moreover, melanocortin receptor 1 (MC‐1R), which is specific for α‐MSH and ACTH, is expressed in the skin on keratinocytes, dendritic cells, macrophages and endothelial cells. In monocytes, macrophages and dendritic cells α‐MSH inhibits the production and activity of immunoregulatory and proinflammatory cytokines such as IL‐2, IFN‐γ, TNF‐α, and IL‐1. It downregulates the expression of costimulatory molecules such as CD86 and CD40 and induces the production of suppressor factors such as the cytokine synthesis inhibitory factor IL‐10. On endothelial cells α‐MSH is capable of downregulating the LPS‐induced expression of adhesion molecules such as vascular cell adhesion molecule (VCAM) and E‐selectin. Moreover, the LPS‐induced activation of transcription factors such as NFκB is downregulated by α‐MSH. In a mouse model i.v. or topical application of α‐MSH was found to inhibit the induction phase as well as the effector phase of contact hypersensitivity (CHS) reactions and to induce hapten‐specific tolerance. These findings indicate that the production of immunosuppressing neuropeptides such as α‐MSH by epidermal cells may play an essential role during the pathogenesis of immune and inflammatory reactions in the skin.

Mechanisms of the Antiinflammatory Effects of α-MSH: Role of Transcription Factor NF-κB and Adhesion Molecule Expression

ABSTRACT: The recruitment of leukocytes from the circulation to inflamed tissue is regulated by the expression of adhesion molecules on both leukocytes and endothelial cells. The proopiomelanocortin‐derived peptide α‐melanocyte stimulating hormone (α‐MSH) is known to modulate inflammation. Thus, we investigated the influence of α‐MSH on the LPS‐induced expression of the adhesion molecules E‐selectin and VCAM‐1 on endothelial cells. Human microvascular endothelial cells (HMEC‐1) were treated with LPS (100 ng/ml) alone or in the presence of α‐MSH (10−8 to 10−16 M). RT‐PCR analysis showed that α‐MSH significantly reduced LPS‐induced expression of VCAM‐1 and E‐selectin. Since many adhesion molecules contain regulatory NF‐κB sites in their promoter region, the role of α‐MSH in the activation of the transcription factor NF‐αB was also investigated. α‐MSH significantly downregulated the LPS‐mediated activation of NF‐κB, in a dose‐dependent manner. These findings indicate that modulation of the transcription factor NF‐κB is a crucial molecular event, one that seems to be responsible for the antiinflammatory effects of α‐MSH.

Human Peripheral Blood‐Derived Dendritic Cells Express Functional Melanocortin Receptor MC‐1R

ABSTRACT: The neuropeptide, α‐melanocyte‐stimulating hormone (α‐MSH) is well known for its immunomodulating capabilities. α‐MSH antagonizes the activity of numerous proinflammatory mediators; for example, Interleukin‐1 (IL‐1), IL‐6, tumor necrosis factor α (TNFα), and bacterial endotoxin. In vivoα‐MSH has been shown to suppress a contact hypersensitivity reaction in mice, and to induce hapten‐specific tolerance. Since antigen presenting cells (APC) represent key elements for tolerance induction, the effect of α‐MSH, and the expression of its receptor‐melanocortin receptor‐1 (MC‐1R), on human peripheral blood‐derived monocytes and dendritic cells (DC), was investigated. Semiquantitative RT‐PCR demonstrated that monocytes and DC express MC‐1R, but none of the other members of the MC‐receptor family. Moreover, the extent of MC‐1R expression correlated with the state of activation of these cells. Since the major ligand of MC‐1R is α‐MSH the question of whether α‐MSH affects the function of monocyte derived DC was further investigated. We found that the expression of the costimulatory molecules CD 86 and CD 40 was downregulated on DC in the presence of α‐MSH. Thus, α‐MSH may exert its immunosuppressive effects by altering the function of APC.

Molecular Basis of the α‐MSH/IL‐1 Antagonism

ABSTRACT: The neuropeptide α‐melanocyte stimulating hormone (α‐MSH) is recognized as a potent mediator of immune and inflammatory reactions. Accordingly, α‐MSH in vitro, as well as in vivo, antagonizes the proinflammatory activities of cytokines such as interleukin‐1 (IL‐1), IL‐6, and tumor necrosis factor a (TNFα). Since the molecular basis of these antiinflammatory effects is not well known, the influence of α‐MSH on IL‐1β‐induced chemokine production and transcription factor activation was investigated in human keratinocytes. α‐MSH, in a dose‐dependent manner, after 48 h, significantly reduced the IL‐1b mediated secretion of the C‐X‐C chemokines IL‐8 and Groα. This was confirmed by semiquantitative RT‐PCR, which revealed a marked downregulation in IL‐8 and Groα mRNA expression. Furthermore, we determined the effect of α‐MSH on the IL‐1β‐induced activation of the nuclear factor κB (NFκB)‐a major transcription factor for chemokine genes. Electrophoretic mobility‐shift‐assays showed that α‐MSH, in a dose range from 10−6 to 10−12 M, significantly downregulated the IL‐1b‐induced activation of NFκB 10 minutes after stimulation. Therefore, NFκB inactivation by α‐MSH appears to be a crucial event, one that is responsible for the downregulation of cytokine gene transcription.

α-Melanocyte-Stimulating Hormone as a Mediator of Tolerance Induction

There is accumulating evidence for a strong interaction between components of the nervous system and the immune system. Accordingly, specific receptors for neuropeptides were found to be expressed on immunocompetent cells and several neuropeptides were recognized as potent regulators of immune and inflammatory reactions. Among various neuropeptides such as substance P, calcitonin gene-related peptide and others α-melanocyte-stimulating hormone (α-MSH) was found to be produced in the skin. Moreover, melanocortin receptor 1 which is specific for α-MSH and ACTH is expressed in the skin on keratinocytes, dendritic cells, macrophages and endothelial cells. In monocytes, macrophages and dendritic cells α-MSH inhibits the production and activity of immunoregulatory and proinflammatory cytokines such as IL-2, IFNγ and IL-1. It downregulates the expression of costimulatory molecules such as CD86 and CD40 and induces the production of suppressor factors such as the cytokine synthesis inhibitory factor IL-10. On endothelial cells α-MSH is capable of downregulating the LPS-induced expression of adhesion molecules such as vascular cellular adhesion molecules and E-selectin. Moreover, the LPS-induced activation of transcription factors such as NFκB is downregulated by α-MSH. In a mouse model intravenous or topical application of α-MSH was found to inhibit the induction as well as the effector phase of a contact hypersensitivity reaction and to induce hapten-specific tolerance. Moreover, there is evidence that the N-terminal tripeptide of α-MSH is sufficient for its in vitro and in vivo immunomodulatory effects. These findings indicate that the production of immunosuppressing neuropeptides such as α-MSH by epidermal cells may play an essential role during the pathogenesis of immune and inflammatory reactions in the skin.

Expression of Functional Melanocortin Receptors and Proopiomelanocortin Peptides by Human Dermal Microvascular Endothelial Cells

ABSTRACT: Human dermal microvascular endothelial cells (HDMEC) are capable of mediating leukocyte‐endothelial interactions by the expression of cellular adhesion molecules and the release of proinflammatory cytokines and chemokines during cutaneous inflammation. Recent studies support the important role for proopiomelanocortin (POMC) peptides, such as α‐melanocyte stimulating hormone (α‐MSH), as immunomodulators in the cutaneous immune system. The purpose of the studies described here was to determine whether HDMEC serves as both target and source for POMC peptides. RT‐PCR and Northern blot studies demonstrated the constitutive expression of mRNA for the adrenocorticotropin (ACTH) and α‐MSH‐specific melanocortin receptor 1 (MC‐1R) in HDMEC, and the microvascular endothelial cell line HMEC‐1 that could be upregulated by stimulation with IL‐1β and α‐MSH. HDMEC responded to stimulation by α‐MSH with a dose‐ and time‐dependent synthesis and release of the CXC chemokines, IL‐8 and GROα. Likewise, α‐MSH augmented HDMEC chemokine release induced by TNF or IL‐1. HDMEC were found to constitutively express POMC and prohormone convertase 1 (PC‐1); the latter being required to generate ACTH from the POMC prohormone. POMC and PC‐1 mRNA expression are increased as a result of stimulation with UVB and UVA1 radiation, IL‐1, and α‐MSH. In addition, UV‐radiation is capable of inducing the release of HDMEC, ACTH, and α‐MSH in a time‐ and dose‐dependent fashion. Thus, these data provide evidence that HDMEC are capable of expressing functional MC‐1R, POMC, and PC‐1 mRNA; and of releasing POMC peptides with UV light, IL‐1, and α‐MSH as regulatory factors. The expression and regulation of these peptides may be of importance, not only for the autocrine or paracrine regulation of physiologic functions of dermal endothelial cells, but also for the regulation of certain microvascular‐mediated cutaneous or systemic inflammatory responses.

Expression of Proopiomelanocortin Peptides and Prohormone Convertases by Human Dermal Microvascular Endothelial Cells

Proopiomelanocortin (POMC) peptides such as α -melanocyte stimulating hormone ( α -MSH) and adrenocorticotropin (ACTH) are expressed in the epidermal and dermal compartments of the skin. In epidermal cells, such as keratinocytes or melanocytes, the transcription and release of POMC-peptides in vivo and in vitro increases as a result of noxious stimuli such as UV-light or infection, either directly, or possibly by the induction of secondary mediators such as IL-1. POMC-peptides, especially α -MSH, are considered to be potent immunomodulators with antipyretic and immunosuppressive properties. The expression and regulation of cellular adhesion molecules and cytokines by dermal microvascular endothelial cells is crucial in mediating leukocyte-endothelial cell interaction and transmigration into the extravascular tissue during cutaneous inflammation. 1,2 In this paper we address the hypothesis that human dermal microvascular endothelial cells (HDMEC) are capable of expressing both the POMC and prohormone convertases (PC) that are required to generate POMC peptides.