title: Molecular size and origin do not influence the harmful side effects of hydroxyethyl starch on human proximal tubule cells (HK-2) in vitro.
abstract: BACKGROUND
  Recently, clinical trials revealed renal impairment induced by hydroxyethyl starch (HES) in septic patients. In prior studies, we managed to demonstrate that HES accumulated in renal proximal tubule cells (PTCs). The related pathomechanism has not yet been discovered. To validate our hypothesis that the HES molecule itself is harmful, regardless of its molecule size or origin, we conducted a comprehensive study to elucidate the influences of different HES preparations on PTC viability in vitro.
  
  
  METHODS
  Cell viability of human PTC was measured with a cytotoxicity assay, quantifying the reduction of tetrazolium salt to colored formazan. Experiments were performed by assessing the influence of different carrier solutions of HES (balanced, nonbalanced, culture medium), different average molecular weights (70, 130, 200 kDa), different origins (potato or corn derived), and various durations of incubation (2-21 hours). Furthermore, HES 130/0.4 was fractionated by ultrafiltration, and the impact on cell viability of average single-size fractions with <3, 3 to 10, 10 to 30, 30 to 50, 50 to 100, and >100 kDa was investigated. We also tested the possible synergistic effects of inflammation induced by tumor necrosis factor-α.
  
  
  RESULTS
  All tested HES solutions, regardless of origin or carrier matrix, decreased cell viability in an equivalent, dose-dependent manner. Coincubation with tumor necrosis factor-α did not reduce HES-induced reduction of cell viability. Minor differences were detected comparing 70, 130, and 200 kDa preparations. Analysis of fractionated HES revealed that each fraction decreased cell viability. Even small HES molecules (10-30 kDa) were significantly deleterious.
  
  
  CONCLUSIONS
  For the first time, we were able to show that only the total mass of HES molecules applied is responsible for the harmful impact on renal PTC in vitro. Neither molecular size nor their origin showed any relevance.
authors.affiliation: From the Institute of Anaesthesia and Critical Care, University Hospital Würzburg, Würzburg, Germany; and †Department of Medicinal Chemistry, University of Vienna, Vienna, Austria.
authors.forename: Raphael R
authors.initials: RR
authors.lastname: Bruno
authors.forename: Winfried
authors.initials: W
authors.lastname: Neuhaus
authors.forename: Norbert
authors.initials: N
authors.lastname: Roewer
authors.forename: Christian
authors.initials: C
authors.lastname: Wunder
authors.forename: Martin A
authors.initials: MA
authors.lastname: Schick
mesh_terms: D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D003102:Colloids; D000077324:Crystalloid Solutions; D004305:Dose-Response Relationship, Drug; D004337:Drug Carriers; D005562:Formazans / Q000737:chemistry; D006801:Humans; D006895:Hydroxyethyl Starch Derivatives / Q000009:adverse effects*; D007202:Indicators and Reagents; D018836:Inflammation Mediators / Q000378:metabolism; D007552:Isotonic Solutions; D007687:Kidney Tubules, Proximal / Q000187:drug effects / Q000473:pathology*; D008970:Molecular Weight; D019999:Pharmaceutical Solutions; D010952:Plasma Substitutes / Q000009:adverse effects*; D016133:Polymerase Chain Reaction; D012313:RNA / Q000096:biosynthesis / Q000235:genetics; D011198:Solanum tuberosum / Q000737:chemistry; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D003313:Zea mays / Q000737:chemistry
chemical_list: D003102:Colloids; D000077324:Crystalloid Solutions; D004337:Drug Carriers; D005562:Formazans; D006895:Hydroxyethyl Starch Derivatives; D007202:Indicators and Reagents; D018836:Inflammation Mediators; D007552:Isotonic Solutions; D019999:Pharmaceutical Solutions; D010952:Plasma Substitutes; D014409:Tumor Necrosis Factor-alpha; D012313:RNA