Department of Pathology, College of Graduate Studies, Midwestern University, Downers Grove, IL 60515, USA.
Department of Biomedical Sciences, College of Graduate Studies, Midwestern University, Downers Grove, IL 60515, USA.
Int J Mol Sci. 2021 Oct 29;22(21):11719. doi: 10.3390/ijms222111719.
Melanoma is a lethal form of skin cancer triggered by genetic and environmental factors. Excision of early-stage, poorly aggressive melanoma often leads to a successful outcome; however, left undiagnosed these lesions can progress to metastatic disease. This research investigates whether the exposure of poorly aggressive melanoma to certain normal skin cells can explain how non-metastatic melanoma becomes more aggressive while still confined to the skin. To this end, we used a serial co-culture approach to sequentially expose cells from two different, poorly aggressive human melanoma cell lines against normal cells of the skin beginning with normal melanocytes, then epidermal keratinocytes, and finally dermal fibroblasts. Protein extraction of melanoma cells occurred at each step of the co-culture sequence for western blot (WB) analysis. In addition, morphological and functional changes were assessed to detect differences between the serially co-cultured melanoma cells and non-co-cultured cells. Results show that the co-cultured melanoma cells assumed a more mesenchymal morphology and displayed a significant increase in proliferation and invasiveness compared to control or reference cells. WB analysis of protein from the co-cultured melanoma cells showed increased expression of Snail and decreased levels of E-cadherin suggesting that epithelial-to-mesenchymal transition (EMT) is occurring in these co-cultured cells. Additional WB analysis showed increased levels of Nodal protein and signaling and signs of increased Wnt activity in the co-cultured melanoma cells compared to reference cells. These data suggest that interaction between poorly aggressive melanoma cells with normal cells of the skin may regulate the transition from localized, poorly aggressive melanoma to invasive, metastatic disease via Nodal and/or Wnt induced EMT.
黑色素瘤是一种由遗传和环境因素引发的致命皮肤癌。早期、侵袭性弱的黑色素瘤通过切除通常可以获得良好的治疗效果；然而，如果未被诊断，这些病变可能会进展为转移性疾病。本研究旨在探讨侵袭性弱的黑色素瘤与正常皮肤细胞的相互作用是否可以解释非转移性黑色素瘤在皮肤内仍局限时如何变得更具侵袭性。为此，我们使用连续共培养方法，将来自两种不同、侵袭性弱的人黑色素瘤细胞系的细胞依次暴露于正常皮肤细胞，从正常黑素细胞开始，然后是表皮角质形成细胞，最后是真皮成纤维细胞。在共培养序列的每一步都对黑色素瘤细胞进行蛋白质提取，用于 Western blot (WB) 分析。此外，还评估了形态和功能变化，以检测连续共培养的黑色素瘤细胞与未共培养细胞之间的差异。结果表明，与对照或参考细胞相比，共培养的黑色素瘤细胞呈现出更具间质形态的特征，并且增殖和侵袭性显著增加。对共培养的黑色素瘤细胞的蛋白质进行 WB 分析显示，Snail 的表达增加，而 E-cadherin 的水平降低，表明上皮-间质转化 (EMT) 正在这些共培养细胞中发生。另外的 WB 分析显示，与参考细胞相比，共培养的黑色素瘤细胞中 Nodal 蛋白及其信号的水平增加，并且 Wnt 活性增加的迹象。这些数据表明，侵袭性弱的黑色素瘤细胞与皮肤正常细胞之间的相互作用可能通过 Nodal 和/或 Wnt 诱导的 EMT 调节从局部、侵袭性弱的黑色素瘤向侵袭性、转移性疾病的转变。
