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Nicotinic acid receptor abnormalities in human skin cancer: implications for a role in epidermal differentiation.

Bermudez Y, Benavente CA, Meyer RG, Coyle WR, Jacobson MK, Jacobson EL - PLoS ONE (2011)

Bottom Line: Receptor transcripts are greatly over-expressed in squamous cell cancers.In normal human primary and immortalized keratinocytes, nicotinic acid receptors show plasma membrane localization and functional G(i)-mediated signaling.The results of these studies justify future genetic and pharmacological intervention studies to define possible specific role(s) of nicotinic acid receptors in human skin homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Arizona Cancer Center and Department of Pharmacology and Toxicology, College of Pharmacy, University of Arizona, Tucson, Arizona, United States of America.

ABSTRACT

Background: Chronic UV skin exposure leads to epidermal differentiation defects in humans that can be largely restored by pharmacological doses of nicotinic acid. Nicotinic acid has been identified as a ligand for the human G-protein-coupled receptors GPR109A and GPR109B that signal through G(i)-mediated inhibition of adenylyl cyclase. We have examined the expression, cellular distribution, and functionality of GPR109A/B in human skin and skin derived epidermal cells.

Results: Nicotinic acid increases epidermal differentiation in photodamaged human skin as judged by the terminal differentiation markers caspase 14 and filaggrin. Both GPR109A and GPR109B genes are transcribed in human skin and in epidermal keratinocytes, but expression in dermal fibroblasts is below limits of detection. Receptor transcripts are greatly over-expressed in squamous cell cancers. Receptor protein in normal skin is prominent from the basal through granular layers of the epidermis, with cellular localization more dispersive in the basal layer but predominantly localized at the plasma membrane in more differentiated epidermal layers. In normal human primary and immortalized keratinocytes, nicotinic acid receptors show plasma membrane localization and functional G(i)-mediated signaling. In contrast, in a squamous cell carcinoma derived cell line, receptor protein shows a more diffuse cellular localization and the receptors are nearly non-functional.

Conclusions: The results of these studies justify future genetic and pharmacological intervention studies to define possible specific role(s) of nicotinic acid receptors in human skin homeostasis.

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Related in: MedlinePlus

GPR109A and GPR109B are functional in normal human keratinocytes but are defective in malignant cells.Panel A: NHEK, HaCaT, Ras-transformed HaCaT, A-431, SCC-25, and CF3 cells were treated in the presence of 10 µM forskolin (open columns) or 10 µM forskolin and 100 µM nicotinic acid (grey columns) for 1 h followed by measurement of intracellular cAMP levels. Data are from three independent experiments and show forskolin-induced cAMP production relative to cellular protein. Panel B: The percent inhibition by nicotinic acid is shown for each cell line. For both panels A and B, Students t-test was used to compare cAMP produced without nicotinic acid to that produced with nicotinic acid, * p≤0.05.
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pone-0020487-g006: GPR109A and GPR109B are functional in normal human keratinocytes but are defective in malignant cells.Panel A: NHEK, HaCaT, Ras-transformed HaCaT, A-431, SCC-25, and CF3 cells were treated in the presence of 10 µM forskolin (open columns) or 10 µM forskolin and 100 µM nicotinic acid (grey columns) for 1 h followed by measurement of intracellular cAMP levels. Data are from three independent experiments and show forskolin-induced cAMP production relative to cellular protein. Panel B: The percent inhibition by nicotinic acid is shown for each cell line. For both panels A and B, Students t-test was used to compare cAMP produced without nicotinic acid to that produced with nicotinic acid, * p≤0.05.

Mentions: Most characterizations of the functionality of nicotinic acid receptors have been studied in cells that do not inherently express the receptors but into which the receptor genes are transfected and over-expressed. In our studies, we examined intrinsic receptor functionality by measuring the ability of nicotinic acid to inhibit forskolin-stimulated cAMP formation in the absence of heterologous expression. Under these conditions, the relative contribution of nicotinic acid sensitive cAMP formation is less than that in transfected cells. Forskolin treatment elicited cAMP formation in all of the cells studied with an accumulation of 170–200 pmol cAMP per mg protein in a 1 h incubation and the presence of 100 µM nicotinic acid inhibited the forskolin-induced cAMP production differentially in the various cell types (Fig. 6A). The relative degree of inhibition in the various cell lines by nicotinic acid is shown in Fig. 6B. Inhibition of cAMP formation by nicotinic acid through the intrinsic receptors was approximately 44% in NHEK cells, 43% in HaCaT and 39% in Ras-transformed HaCaT cells. In contrast, cAMP stimulated by forskolin was significantly reduced in the tumor-derived cells. Nicotinic acid inhibited only 23% in A-431 cells and 13% in SCC-25 cells. Forskolin treatment elicits significant cAMP accumulation in CF3 cells but nicotinic acid fails to inhibit cAMP formation, consistent with the absence of nicotinic acid receptor expression observed in dermal fibroblasts (Fig. 3A).


Nicotinic acid receptor abnormalities in human skin cancer: implications for a role in epidermal differentiation.

Bermudez Y, Benavente CA, Meyer RG, Coyle WR, Jacobson MK, Jacobson EL - PLoS ONE (2011)

GPR109A and GPR109B are functional in normal human keratinocytes but are defective in malignant cells.Panel A: NHEK, HaCaT, Ras-transformed HaCaT, A-431, SCC-25, and CF3 cells were treated in the presence of 10 µM forskolin (open columns) or 10 µM forskolin and 100 µM nicotinic acid (grey columns) for 1 h followed by measurement of intracellular cAMP levels. Data are from three independent experiments and show forskolin-induced cAMP production relative to cellular protein. Panel B: The percent inhibition by nicotinic acid is shown for each cell line. For both panels A and B, Students t-test was used to compare cAMP produced without nicotinic acid to that produced with nicotinic acid, * p≤0.05.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3105069&req=5

pone-0020487-g006: GPR109A and GPR109B are functional in normal human keratinocytes but are defective in malignant cells.Panel A: NHEK, HaCaT, Ras-transformed HaCaT, A-431, SCC-25, and CF3 cells were treated in the presence of 10 µM forskolin (open columns) or 10 µM forskolin and 100 µM nicotinic acid (grey columns) for 1 h followed by measurement of intracellular cAMP levels. Data are from three independent experiments and show forskolin-induced cAMP production relative to cellular protein. Panel B: The percent inhibition by nicotinic acid is shown for each cell line. For both panels A and B, Students t-test was used to compare cAMP produced without nicotinic acid to that produced with nicotinic acid, * p≤0.05.
Mentions: Most characterizations of the functionality of nicotinic acid receptors have been studied in cells that do not inherently express the receptors but into which the receptor genes are transfected and over-expressed. In our studies, we examined intrinsic receptor functionality by measuring the ability of nicotinic acid to inhibit forskolin-stimulated cAMP formation in the absence of heterologous expression. Under these conditions, the relative contribution of nicotinic acid sensitive cAMP formation is less than that in transfected cells. Forskolin treatment elicited cAMP formation in all of the cells studied with an accumulation of 170–200 pmol cAMP per mg protein in a 1 h incubation and the presence of 100 µM nicotinic acid inhibited the forskolin-induced cAMP production differentially in the various cell types (Fig. 6A). The relative degree of inhibition in the various cell lines by nicotinic acid is shown in Fig. 6B. Inhibition of cAMP formation by nicotinic acid through the intrinsic receptors was approximately 44% in NHEK cells, 43% in HaCaT and 39% in Ras-transformed HaCaT cells. In contrast, cAMP stimulated by forskolin was significantly reduced in the tumor-derived cells. Nicotinic acid inhibited only 23% in A-431 cells and 13% in SCC-25 cells. Forskolin treatment elicits significant cAMP accumulation in CF3 cells but nicotinic acid fails to inhibit cAMP formation, consistent with the absence of nicotinic acid receptor expression observed in dermal fibroblasts (Fig. 3A).

Bottom Line: Receptor transcripts are greatly over-expressed in squamous cell cancers.In normal human primary and immortalized keratinocytes, nicotinic acid receptors show plasma membrane localization and functional G(i)-mediated signaling.The results of these studies justify future genetic and pharmacological intervention studies to define possible specific role(s) of nicotinic acid receptors in human skin homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Arizona Cancer Center and Department of Pharmacology and Toxicology, College of Pharmacy, University of Arizona, Tucson, Arizona, United States of America.

ABSTRACT

Background: Chronic UV skin exposure leads to epidermal differentiation defects in humans that can be largely restored by pharmacological doses of nicotinic acid. Nicotinic acid has been identified as a ligand for the human G-protein-coupled receptors GPR109A and GPR109B that signal through G(i)-mediated inhibition of adenylyl cyclase. We have examined the expression, cellular distribution, and functionality of GPR109A/B in human skin and skin derived epidermal cells.

Results: Nicotinic acid increases epidermal differentiation in photodamaged human skin as judged by the terminal differentiation markers caspase 14 and filaggrin. Both GPR109A and GPR109B genes are transcribed in human skin and in epidermal keratinocytes, but expression in dermal fibroblasts is below limits of detection. Receptor transcripts are greatly over-expressed in squamous cell cancers. Receptor protein in normal skin is prominent from the basal through granular layers of the epidermis, with cellular localization more dispersive in the basal layer but predominantly localized at the plasma membrane in more differentiated epidermal layers. In normal human primary and immortalized keratinocytes, nicotinic acid receptors show plasma membrane localization and functional G(i)-mediated signaling. In contrast, in a squamous cell carcinoma derived cell line, receptor protein shows a more diffuse cellular localization and the receptors are nearly non-functional.

Conclusions: The results of these studies justify future genetic and pharmacological intervention studies to define possible specific role(s) of nicotinic acid receptors in human skin homeostasis.

Show MeSH
Related in: MedlinePlus