MC(1) receptors are constitutively expressed on leucocyte subpopulations with antigen presenting and cytotoxic functions

Abstract

The expression of melanocortin MC(1) receptors on human peripheral lymphocyte subsets was analysed by flow cytometry using rabbit antibodies selective for the human MC(1) receptor and a panel of monoclonal antibodies against lymphocyte differentiation markers. The MC(1) receptor was found to be constitutively expressed on monocytes/macrophages, B-lymphocytes, natural killer (NK) cells and a subset of cytotoxic T-cells. Interestingly T-helper cells appeared to be essentially devoid of MC(1) receptors. The results were confirmed by RT-PCR which indicated strong expression of MC(1) receptor mRNA in CD14(+), CD19(+) and CD56(+) cells. However, only a faint RT-PCR signal was seen in CD3(+) cells, in line with the immuno-staining results that indicated that only part of the CD3(+) cells (i.e. some of the CD8(+) cells) expressed the MC(1) receptor. The MC(1) receptors' constitutive expression on immune cells with antigen-presenting and cytotoxic functions implies important roles for the melanocortic system in the modulation of immune responses.

Fig. 1

Dual colour flow cytometric analysis of MC₁ receptor (MC₁R) expression on subsets of isolated PBMC from a healthy adult donor. Anti-CD14 mAb was used to stain monocytes/macrophages (a), anti-CD4 mAb to stain T_H cells (b), anti-CD8 mAb to stain CTLs (c), anti-CD19 mAb to stain B-cells (d) and anti-CD56 mAb to stain NK cells (e). Staining of MC₁Rs was done using the M1-Y antiserum. An irrelevant IgG1 isotype mAb and normal rabbit serum served as one of the negative controls (f). (Similar results as in F were obtained using an irrelevant IgG2a isotype mAb and normal rabbit serum; data not shown).

Fig. 2

Detection of MC₁ receptor transcripts in human peripheral blood mononuclear cells by RT-PCR. Subpopulations of PBMCs were obtained by positive selection and used for preparation of cDNA followed by RT-PCR amplification and detection on agarose gels using ethidium bromide staining. (A) From left to right lanes represent RT-PCR bands using MC₁R and β-actin selective primers for cells positively selected for CD56, CD19, CD14 and CD3. (B) RT-PCR bands obtained using an MC₁R plasmid or cDNA from cells positively selected for CD56. Indicated by arrows to the right are the mobility of standard DNA, nt 300–700. Indicated by the arrow to the left is the 686 nt size of the band amplified from the MC₁R plasmid.