A novel strategy to facilitate uniform epithelial cell maturation using liquid-liquid interfaces.

Epithelial tissue forms and maintains a critical barrier function in the body. A novel culture design aimed at promoting uniform maturation of epithelial cells using liquid materials is described. Culturing Madin-Darby canine kidney (MDCK) cells at the liquid-liquid interface yielded reduced migration and stimulated active cell growth. Similar to solid-liquid interfaces, cells cultured on a fibronectin-coated liquid-liquid interface exhibited active migration and growth, ultimately reaching a confluent state. These cells exhibited reduced stress fiber formation and adopted a cobblestone-like shape, which led to their even distribution in the culture vessel. To inhibit stress fiber formation and apoptosis, the exposure of cells on liquid-liquid interfaces to Y27632, a specific inhibitor of the Rho-associated protein kinase (ROCK), facilitated tight junction formation (frequency of ZO-2-positive cells, F Z  = 0.73). In Y27632-exposed cells on the liquid-liquid interface, the value obtained by subtracting the standard deviation of the ratio of nucleus densities in each region that compartmentalized a culture vessel from 1, denoted as H LN , was 0.93 ± 0.01, indicated even cell distribution in the culture vessel at t = 72 h. The behavior of epithelial cells on liquid-liquid interfaces contributes to the promotion of their uniform maturation.