INDUCTION OF CALBINDIN D-28K IN MADIN-DARBY BOVINE KIDNEY-CELLS BY 1,25(OH)2D3

Renal calbindin D-28K is a calcium binding protein, localized to the d istal nephron, whose expression is reduced in vitamin D deficiency and increased upon administration of 1,25(OH)2D3, the active form of vita min D. Investigation into the molecular regulation of renal calbindin D-28K expression has been limited by the lack of an established cell l ine which expresses calbindin D-28K in a vitamin D responsive manner. In the studies described here, we compared the expression of calbindin D-28K and the vitamin D receptor (VDR) in four renal cell lines: Madi n-Darby bovine kidney (MDBK) cells, Madin-Darby canine kidney (MDCK) c ells, LLC-PK, pig kidney cells and opossum kidney (OK) cells. We repor t that MDBK cells express the highest relative levels of calbindin D-2 8K and the VDR, and that 1,25(OH)2D3 increases calbindin D-28K express ion in these cells. No expression of calbindin D-28K was detected in M DCK, LLC-PK, or OK cells. Kinetic studies indicated that calbindin D-2 8K protein increased with time for up to 24 hours after a single dose of 1,25(OH)2D3 (10(-7) M) in MDBK cells. Immunofluorescence studies in dicated that in control MDBK cells, the majority of calbindin D-28K wa s localized in cytosol, with a definite concentration in the peri-nucl ear region. In 1,25(OH)2D3 treated cells, calbindin D-28K was enhanced in cytosol and was detected within the nucleus. In contrast to hetero geneous primary culture systems, in which a minority of cells express calbindin D-28K, virtually all MDBK cells expressed calbindin D-28K, e ven in the absence of 1,25(OH)2D3. In summary, MDBK cells represent th e first established renal cell line identified which expresses calbind in D-28K in a vitamin D responsive manner, providing a model in which to examine the molecular regulation of calbindin D-28K by 1,25(OH)2D3.