A murine model for the development of melanocytic nevi and their progression to melanoma

Abstract

Acquired melanocytic nevi are commonly found in sun exposed and unexposed human skin, but the potential for their transformation into invasive melanoma is not clear. Therefore, a mouse model of nevus initiation and progression was developed in C3H/HeN mice using a modified chemical carcinogenesis protocol. Nevi develop due to DNA damage initiated by dimethylbenz(a) anthracene (DMBA) followed by chronic promotion with 12‐O‐tetradecanoyl‐phorbol‐13‐acetate (TPA). Dysplastic pigmented skin lesions appeared in 7–9 wk with 100% penetrance. Nests of melanocytic cells appeared in a subset of skin draining lymph nodes (dLN) by 25 wk, but not in age matched controls. Immunohistochemistry, real‐time PCR, and flow cytometric analyses confirmed their melanocytic origin. Transformed cells were present in a subset of nevi and dLNs, which exhibited anchorage‐independent growth, tumor development, and metastasis in nude mice. Approximately 50% of the cell lines contained H‐Ras mutations and lost tumor suppressor p16Ink4a expression. While most studies of melanoma focus on tumor progression in transgenic mouse models where the mutations are present from birth, our model permits investigation of acquired mutations at the earliest stages of nevus initiation and promotion of nevus cell transformation. This robust nevus/melanoma model may prove useful for identifying genetic loci associated with nevus formation, novel oncogenic pathways, tumor targets for immune‐prevention, screening therapeutics, and elucidating mechanisms of immune surveillance and immune evasion. © 2015 The Authors. Molecular Carcinogenesis, published by Wiley Periodicals, Inc.

IL-23 Inhibits Melanoma Development by Augmenting DNA Repair and Modulating T Cell Subpopulations

“…After a 5 day rest, they were painted with 100μg DMBA in 100μl acetone and then treated twice weekly with topical 12.5μg TPA (20nmol) (16). Before isolation of nevi or LNs, mice were rested for 5 weeks and then sacrificed (16). …”

“…Nevus biopsies or lymph nodes were processed for hematoxylin and eosin (H&E) staining (16). Corresponding sections were also melanin bleached using 0.25% potassium permanganate and 5% oxalic acid solutions and were processed for H&E or fluorescent staining (16).…”

“…Corresponding sections were also melanin bleached using 0.25% potassium permanganate and 5% oxalic acid solutions and were processed for H&E or fluorescent staining (16). Images were captured with an Olympus DP70 digital camera and further analyzed using ImagePro Plus software v6.0 (Media Cybernetics, Inc., Silver Springs, MD.…”

“…Lesional tissues and adjacent skin from carcinogen treated mice or age matched untreated skin were collected for histological examination or were subjected to digestion and processing into single cell suspensions for flow cytometric staining (16). …”

“…Total RNA was extracted from individual samples using Trizol reagent (Invitrogen, Carlsbad, CA) as described elsewhere (16). Primers for Gapdh , Mut-Hras , p16 INK4a , p19 ARF , Tyr , MelanA , Trp2 (16), IL-10 (18), IL-12p35 (19), IL-12p40 (19), IL-23p19 (20), IL-17 (20), IFNƔ (21) are described in detail elsewhere.…”