Intra- and extracellular proteins in human normal and polycystic kidney epithelial cells

Intra- and extracellular proteins in human normal and polycystic kidney epithelial cells. A tissue culture method was established for the continuous growth of epithelial cells from the cortex of human normal kidney (HNC) and from the epithelial layer of kidney cysts from autosomal dominant polycystic kidney disease (ADPKD) patients. Primary cells were grown to 80 to 90% confluency from 1mm2 slices of tissue, and subcultured up to 10 times. The subcultured HNC and ADPKD cells retained characteristic epithelial polygonal and elongated shape and positive immunofluorescent staining for cytokeratin. The cell doubling time for both HNC and ADPKD epithelia was three to four days at a fetal calf serum (FCS) concentration of 5%. Using these culturing procedures 1 to 5 × 109 epithelial cells could be obtained from each kidney specimen. Profiles of 35S-methionine radiolabeled intracellular proteins of HNC and ADPKD cells qualitatively demonstrated a high degree of similarity, thus confirming a similarity of epithelial origin and protein biosynthesis. Both the underexpression of three proteins (a) protein p2, Mr ∼ 47 kDa, pI ∼ 6.0; b) protein p3, Mr ∼ 50 kDa, pI ∼ 5.9; and c) protein p4, Mr ∼ 44 kDa, pI ∼ 5.8) and the overexpression of several proteins (including: a) p5, Mr ∼ 56 kDa, pI ∼ 7.3; b) protein p6, Mr ∼ 32 kDa, pI ∼ 7.3; c) protein p7, Mr ∼ 33 kDa, pI ∼ 5.3; d) protein p8, Mr ∼ 45 kDa, pI ∼ 6.9; e) protein p9, Mr ∼ 35 kDa, pI ∼ 6.7; and f) protein p10, Mr ∼ 30 kDa, pI ∼ 6.6) were found in ADPKD cells. In addition, primary ADPKD cells, but not primary HNC cells, produced three extracellular proteins: Mr ∼ 220, 170 and 45kDa. Upon subculturing, identical extracellular proteins were biosynthesized by both HNC and ADPKD cells. This subculturing system provides an important methodological tool for the study of the molecular and biochemical abnormalities of ADPKD