Tissue tension supports the in vitro co‐culture of ras‐keratinocytes and myofibroblasts

Abstract

Tissue tension is necessary to support the myofibroblast, a cell derived from a fibroblast and present in wound healing and tumor stromas. Currently‐used skin equivalent models lack the ability to generate tension. Our goal was to determine whether a tension‐maintaining skin equivalent model supports myofibroblast growth and the growth of ras‐keratinocytes, precancerous cells previously shown to modulate their phenotype in response to their environment. We developed a dermal equivalent with a plastic ring within a liquid collagen/fibroblast mixture and converted it to a skin equivalent by overlaying with keratinocytes. Skin equivalents were cultured at the air/liquid interface. Replicate tissues were harvested weekly and prepared for immunofluorescence staining. Myofibroblasts in the dermis were identified by alpha‐sm actin staining in stress fibers; ras‐keratinocytes differentiated and stratified atop the dermal compartment. Proliferation of both cell types was identified by EdU incorporation and staining. Interestingly, the keratinocyte strata consisted of disorganized clumps of cells rather than organized layers even though invasion into the dermal compartment was inhibited. The tension‐maintaining skin equivalent supports ras‐keratinocytes and can be used to study the role of myofibroblasts during cancer progression in vitro.Grant Funding Source : The University of Central Oklahoma and Grant Number P20RR016478 from the National Center for Research Resources (NCRR), a component of the National Institutes of Health (NIH).