LOW PH AND HYPEROSMOLALITY DETERMINE THE DIFFERENTIATION OF MDCK CELLS TYPICAL FOR THE OUTER MEDULLARY COLLECTING DUCT

The Madin-Darby canine kidney (MDCK) cell line resembles cortical collecting duct epithelium, forming cells with properties corresponding to principal (P) and intercalated (IC) cells. Cells express carbonic anhydrase (CA) activity, and show peanut lectin (PNA) and wheat germ agglutinin (WGA) binding to the apical membrane domain. In order to further characterize the nature of MDCK cells, and to answer the question whether or not their IC-like cells emerge from a precursor cell by differentiation they were cultured on solid (plastic dishes) or semipermeable supports (collagen-coated filters) and acutely exposed to hyperosmotic and acidic media (600 mosm/1 by NaCl, pH 6.5) over a period of 48 h. Upon exposure, domes disappear and both CA activity and PNA-binding capacity decrease. In addition, cell morphology changes markedly. As revealed by scanning electron microscopy, apical membrane microvilli largely disappear and are redistributed to the cell borders. MDCK epithelia cultured on collagen-coated filters exhibit a higher degree of differentiation and the occurrence of a large number of dark cells. In transmission electron microscopy, filter-grown MDCK epithelia closely resemble the morphological characteristics of a mammalian cortical collecting duct. Following exposure to culture media simulating the inner stripe of outer medulla (600 mosm/l and a pH of 6.5), the number of dark cells increases by a factor of 2. Surface pH measurements on IC-like cell apical membranes show an alkalinization by 0.1 unit for IC-like, dark and PNA-binding cells in isoosmotic cultures, whereas in hyperosmotic and low-pH cultures an acidification can be determined for the dark IC-like but PNA-negative cells. The results obtained indicate the presence of mainly B-IC equivalent cells in isoosmotically and A-IC like cells in cultures adapted to hyperosmotic and acidic conditions. Short-term exposure of MDCK cells to hypertonic environment leads to conversion of the monolayer, which is similar to a situation found in the mammalian outer medullary collecting duct, where mainly A-IC cells are present and the ratio of IC to P cells is found to be twice as high as in the renal cortex.