title: Crystals cause acute necrotic cell death in renal proximal tubule cells, but not in collecting tubule cells.
abstract: BACKGROUND
  The interaction between renal tubular cells and crystals generated in the tubular fluid could play an initiating role in the pathophysiology of calcium oxalate nephrolithiasis. Crystals are expected to form in the renal collecting ducts, but not in the proximal tubule. In the present investigation, we studied the damaging effect of calcium oxalate crystals on renal proximal and collecting tubule cells in culture.
  
  
  METHODS
  Studies were performed with the renal proximal tubular cell lines, porcine proximal tubular cells (LLC-PK(1)) and Madin-Darby canine kidney II (MDCK-II) and the renal collecting duct cell lines, RCCD(1) and MDCK-I. Confluent monolayers cultured on permeable growth substrates in a two-compartment culture system were apically exposed to calcium oxalate monohydrate crystals, after which several cellular responses were studied, including monolayer morphology (confocal microscopy), transepithelial electrical resistances (TER), prostaglandin E(2) (PGE(2)) secretion, DNA synthesis ([(3)H]-thymidine), total cell numbers, reactive oxygen species [hydrogen peroxide (H(2)O(2))] generation, apoptotic (annexin V and DNA fragmentation), and necrotic (propidium iodide influx) cell death.
  
  
  RESULTS
  Crystals were rapidly taken up by proximal tubular cells and induced a biphasic response. Within 24 hours approximately half of the cell-associated crystals were released back into the apical fluid (early response). Over the next 2 weeks half of the remaining internalized crystals were eliminated (late response). The early response was characterized by morphologic disorder, increased synthesis of PGE(2), H(2)O(2), and DNA and the release of crystal-containing cells from the monolayers. These released cells appeared to be necrotic, but not apoptotic cells. Scrape-injured monolayers generated even higher levels of H(2)O(2) than those generated in response to crystals. During the late response, crystals were gradually removed from the monolayers without inflammation-mediated cell death. Crystals did not bind to, were not taken up by, and did not cause marked responses in collecting tubule cells.
  
  
  CONCLUSIONS
  This study shows that calcium oxalate crystals cause acute inflammation-mediated necrotic cell death in renal proximal tubular cells, but not in collecting tubule cells. The crystal-induced generation of reactive oxygen species by renal tubular cells is a general response to tissue damage and the increased levels of DNA synthesis seem to reflect regeneration rather than growth stimulation. As long as the renal collecting ducts are not obstructed with crystals, these results do not support an important role for crystal-induced tissue injury in the pathophysiology of calcium oxalate nephrolithiasis.
authors.affiliation: Department of Urology, Erasmus Medical Center, Rotterdam, The Netherlands.
authors.forename: Marieke S J
authors.initials: MS
authors.lastname: Schepers
authors.forename: Eddy S
authors.initials: ES
authors.lastname: van Ballegooijen
authors.forename: Chris H
authors.initials: CH
authors.lastname: Bangma
authors.forename: Carl F
authors.initials: CF
authors.lastname: Verkoelen
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002129:Calcium Oxalate / Q000737:chemistry / Q000633:toxicity*; D003460:Crystallization; D003470:Culture Media; D015232:Dinoprostone / Q000378:metabolism; D004285:Dogs; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D007669:Kidney Calculi / Q000737:chemistry / Q000378:metabolism / Q000473:pathology*; D007685:Kidney Tubules, Collecting / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D007687:Kidney Tubules, Proximal / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D018374:LLC-PK1 Cells; D009336:Necrosis; D013552:Swine; D013936:Thymidine / Q000493:pharmacokinetics; D014316:Tritium
chemical_list: D003470:Culture Media; D014316:Tritium; D002129:Calcium Oxalate; D006861:Hydrogen Peroxide; D015232:Dinoprostone; D013936:Thymidine