Herpes Simplex Virus, Nerve Damage; Melanoma, Tumor Pathogenesis

Anatomy of the Skin

The skin is composed of three layers: epidermis, dermis, and hypodermis. The epidermis is the outermost layer of the skin and is primarily composed of keratinocytes, the cells from which basal cell carcinoma develop. As keratinocytes mature, they become post-mitotic and lose their nucleus, becoming corneocytes, which are eventually shed from the epidermis. Also present in the epidermis are melanocytes, pigmented cells that contribute to skin color and are the cell-of-origin of melanoma. The dermis is the middle layer of the skin and primarily contains connective tissue composed of fibroblasts, macrophages, and adipocytes. Finally, the hypodermis (innermost layer) contains blood vessels and nerves and is responsible for securing the skin to the bone and muscles underneath. Together, the skin makes up our largest organ and is our first line of defense against the external environment.

The Lifeline® catalog includes various types of cells isolated from the skin, including:

• Dermal microvascular endothelial cells (adult and neonatal), optimized for VascuLife® medium
• Epidermal melanocytes (adult and neonatal), optimized for DermaLife® medium
• Dermal fibroblasts (adult and neonatal), optimized for FibroLife® medium
• Epidermal keratinocytes (adult and neonatal), optimized for DermaLife® medium

Recent Studies Using Lifeline® Skin Cells

Herpes simplex virus (HSV) is an infectious virus capable of latency, which occurs when the virus remains quiescent within peripheral sensory nerves. Despite recurrent infections that develop when the virus is reactivated from its latent state, nerve damage due to HSV infection is rare. Peng et al. set out to investigate further whether nerve protective mechanisms are activated by HSV infection, which would explain the lack of nerve damage observed in patients. The authors first examined how HSV infection affected peripheral nerve morphology. In skin biopsies from infected tissue, they found that nerve fibers in tissues taken from sites of HSV reactivation exhibited increased fiber density and length compared to those in control tissues, taken from areas without HSV reactivation.

The researchers then hypothesized that keratinocytes, the primary site of HSV infection, may play a role in preventing HSV-induced nerve damage. To determine how keratinocytes might affect nerve fibers upon HSV infection and recurrent reactivation, the researchers used laser capture microdissection to isolate keratinocytes from patient biopsy samples. Following gene expression analysis, they identified three genes that were significantly upregulated in keratinocytes from HSV-2 active infections and post-healed tissue, including IL-17c, CCL5, and TNFSF10.

To investigate further, the authors infected Lifeline® human keratinocytes in vitro and found that HSV-2 infection induced keratinocyte IL-17c expression. After discovering that IL-17c did not affect the ability of HSV-2 to infect keratinocytes, the group identified that the IL-17c receptor (IL-17RE) was expressed on nerve fibers in patient biopsies at sites of infection, suggesting that keratinocyte-derived IL-17c acts on nerve cells that express its receptor. Importantly, using an in vitro microfluidic culture system, they found that IL-17c treatment of differentiated SY5Y neuron-like neuroblastoma cells stimulated neurite growth. The same effect was illustrated for human sensory neurons isolated from fetal dorsal root ganglia. Finally, the authors tested whether IL-17c promoted neuronal survival by pretreating mouse primary cortical neurons with exogenous murine IL-17c before HSV-2 infection and measuring cleaved caspase 3 as a readout of apoptosis. They found that the proportion of neurons undergoing apoptosis was decreased following IL-17c pretreatment.

Together, the results of this study demonstrate that keratinocyte-derived IL-17c is a neurotrophic factor and has some anti-apoptotic activity.

Melanoma is a form of skin cancer that originates in melanocytes. Although surgical resection of a melanoma is relatively successful at early stages, melanoma is extremely difficult to treat once it metastasizes to other organs. Micro RNAs (miRNAs) are of interest in many cancer types as their ability to regulate gene expression can have significant effects on tumor pathogenesis. In particular, miR-373 is commonly upregulated in melanoma, but its role is currently unknown. To investigate whether miR-373 is involved in the pathogenesis of melanoma, Bai and colleagues first evaluated the expression of miR-373 transcripts in a panel of melanoma tissues and cell lines. They found that miR-373 was upregulated in melanoma tissues, as well as in WM115, WM75, and A375 melanoma cells lines, compared with Lifeline® normal human melanocytes. Next, the authors overexpressed miR-373 in A375 and WM115 cells and found that compared with cells transfected with a control miRNA, cells overexpressing miR375 had increased migratory potential.

The researchers then set out to identify targets of miR-373 that may be responsible for the increased cell migration phenotype observed in cells overexpressing miR-373. Using the miR-373 seed sequence and a fluorescence reporter assay, the authors discovered that salt-inducible kinase 1 (SIK1) was a suitable candidate. Supporting this hypothesis, they observed that SIK1 protein expression was decreased following miR-373 overexpression. Finally, the authors demonstrated that SIK1 knockdown resulted in increased migration of A375 and WM115 cells.
Together, the results of this study suggest that upregulation of miR373 in melanoma cells increases cell migration, potentially through decreased expression of SIK1.

How are you using Lifeline® cells in your research? Let us know and your study could be featured here on our blog! Check back every other week to see the research we are highlighting next!