Hyaluronan is apically secreted and expressed by proliferating or regenerating renal tubular cells.
BACKGROUND
Hyaluronan has diverse biologic functions in the body, varying from structural tasks to cell stress-induced CD44-mediated activation of intracellular signaling pathways. Hyaluronan biology is relatively unexplored in the kidney. Previously, we identified hyaluronan as binding molecule for crystals in the renal tubules. Crystal retention is a crucial early event in the etiology of kidney stones. The present study was performed to determine the polarized distribution of hyaluronan and CD44 by renal tubular cells.


METHODS
Madin-Darby canine kidney (MDCK) strain I and primary cultures of human renal tubular cells were grown on permeable supports in a two-compartment culture system. Studies were performed during growth and after scrape-injury. Metabolic labeling studies and an enzyme-linked hyaluronan -binding assay were used to measure the molecular mass and the amount of secreted hyaluronan in apical and basal medium. Confocal microscopy was applied to detect membrane hyaluronan and CD44. Hyaluronan synthase (HAS) mRNA expression was studied with reverse transcriptase-polymerase chain reaction (RT-PCR). The in vitro expression profile of hyaluronan was compared with that in biopsies of transplanted human kidneys with acute tubular necrosis.


RESULTS
Proliferating cells produced more hyaluronan (M(r) > 10(6) Da) than growth-inhibited cells in intact monolayers and up to 85% was targeted to the apical compartment, which was accompanied by increased HAS2 mRNA expression and slightly decreased HAS3 mRNA, while HAS1 mRNA remained undetectable. Hyaluronan and CD44 were exclusively expressed at the apical surface of proliferating/regenerating cells. After (re)establishment of tight junctions, hyaluronan was no longer detectable while CD44 was targeted to basolateral membrane domains. In vivo in inflamed human kidneys hyaluronan was abundantly expressed in the cortical tubulointerstitial space as well as at the luminal surface of regenerating renal tubular cells.


CONCLUSIONS
These results demonstrate that the production of hyaluronan by renal tubular cells is activated during proliferation and in response to mechanical injury and that hyaluronan and CD44 expression is highly polarized. The targeted delivery of hyaluronan to the apical compartment suggests that hyaluronan produced by renal tubular cells supports proliferation/regeneration in the renal tubules, but that it does not contribute to hyaluronan accumulation in the renal interstitium. These data further support the concept that mitogen/stress-induced hyaluronan deposition in the renal tubules increases the risk for crystal retention and stone formation.
Asselman
Marino
M
Department of Urology, Erasmus Medical Center, Rotterdam, The Netherlands.
Verhulst
Anja
A
Van Ballegooijen
Eddy S
ES
Bangma
Chris H
CH
Verkoelen
Carl F
CF
De Broe
Marc E
ME
D000818:Animals; D000963:Antimetabolites; D001483:Base Sequence; D001973:Bromodeoxyuridine; D002455:Cell Division / Q000502:physiology; D002460:Cell Line; D016764:Cell Polarity / Q000502:physiology; D004285:Dogs; D015971:Gene Expression Regulation, Enzymologic; D014453:Glucuronosyltransferase / Q000235:genetics; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism; D000076002:Hyaluronan Synthases; D006820:Hyaluronic Acid / Q000096:biosynthesis / Q000378:metabolism*; D007683:Kidney Tubular Necrosis, Acute / Q000378:metabolism* / Q000503:physiopathology; D007684:Kidney Tubules / Q000166:cytology / Q000378:metabolism*; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D012038:Regeneration / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014316:Tritium
D000963:Antimetabolites; D018960:Hyaluronan Receptors; D014316:Tritium; D006820:Hyaluronic Acid; D014453:Glucuronosyltransferase; D000076002:Hyaluronan Synthases; D001973:Bromodeoxyuridine