Isolation, Culture and Characterization of Human Renal Tubular Cells

Abstract

Tubular cells have been isolated, characterized and cultured from more than 70 adult cadaver kidneys (postmortem time less than or equal to 12 hr.). Confluent monolayers were observed at 7 days after seeding (10(6) cells/ml.) and cells demonstrating normal human karyotypes have been passaged up to 6 times. Primary isolates and monolayer cultures were negative for Factor VIII activity, and strongly positive for gamma-glutamyltransferase activity and keratin. Ultrastructurally primary isolates consisted of cells with numerous mitochondria, microvilli, cytoplasmic filaments and well-developed endocytotic apparati. Monolayer cultures examined at 7, 14, 21 and 72 days demonstrated less prominent microvilli and the additional structures of desmosomes and cell junctions. Membrane-associated and cytosolic enzyme activities were measured up to 28 days in culture. The membrane-associated enzymes gamma-glutamyltransferase and alkaline phosphatase both exhibited approximately 10-fold decreases in activity during the 1st 7 days in culture. There was an approximately 5-fold increase in pyruvate kinase activity during the same time period, while fructose-1,6-bisphosphatase activity exhibited a 5-fold decrease. Glucose-6-phosphatase activity did not change during the 28 day culture period examined. From 7 to 28 days no further changes were noted in any of the enzyme activities measured. Decreased membrane-associated enzyme activity corresponded to the ultrastructural observation of less prominent microvilli. Increases in glycolytic enzyme activity and decreases in gluconeogenic enzyme activity may reflect the presence of glucose in the culture medium. The morphologic and biochemical evidence suggests that primary isolates and cultures are proximal tubule cells which should provide a well-defined in vitro human system for future studies.

Expression and Cytoplasmic Localization of SAM68 Is a Significant and Independent Prognostic Marker for Renal Cell Carcinoma

“…Normal renal tubular epithelial cells were established from tissue obtained during a nephrectomy for a benign lesion and cultured as described previously (19). The renal cell carcinoma cell lines 769-P, SN12C, SKRC-39, and ACHN were maintained in DMEM (Invitrogen) supplemented with 10% fetal bovine serum (HyClone).…”

Inflammasome-Independent NLRP3 Augments TGF-β Signaling in Kidney Epithelium

“…To study the biology of TECs, we established both human proximal tubular epithelial cell (HPTC) and mouse primary proximal TEC cultures, as previously described (15,27). In short, human kidneys were collected from patients undergoing nephrectomy for the usual surgical indications.…”

Identifying Modulators of hERG Channel Activity Using the PatchXpress® Planar Patch Clamp

“…The cells used were primary human renal proximal tubule epithelial cells (HRPTECs). 25 Cells were exposed to compounds for 24 h before end point measurements. Cytotoxicity is defined as the effective concentration that causes a 50% reduction in cellular ATP levels relative to solvent controls (EC 50 , µM).…”