title: Effect of metals on β-actin and total protein synthesis in cultured human intestinal epithelial cells.
abstract: BACKGROUND
  As an important structural protein, β-actin is associated with anchoring of tight junctions (TJs) to the cell scaffold. Caco-2 cells, an immortal intestinal epithelial cell line, rely on β-actin to form intact monolayers with high transepithelial electrical resistance in cell culture inserts.
  
  
  METHODS
  We examined the effect of six metals on expression of β-actin mRNA and β-actin synthesis, on total and net production of newly synthesized proteins, on paracellular transport of TJ markers, and on cell viability in confluent monolayers. [(3)H]-glycine and [(3)H]-tyrosine were used as indicators of newly synthesized proteins in the absence or presence of increasing concentrations of arsenic, cadmium, copper, manganese, mercury and nickel. The monolayers were exposed to 24-h single exposures as well as continuous daily repeated doses of metals for 48-h and 96-h.
  
  
  RESULTS
  Results suggest that decreases in newly synthesized proteins, in which β-actin represents about 10%, correlated with 2- to 5-fold higher expression of β-actin mRNA for the higher concentrations of metals. Interestingly, IC(50)s calculated for each chemical for 24-h acute and 48- and 96-h repeated dosing experiments, using the MTT viability assay and paracellular permeability markers, decreased newly synthesized and total proteins to 10% and 40% of control, respectively.
  
  
  CONCLUSIONS
  Overall, the results indicate that, at equivalent concentrations, the metals affect β-actin mRNA and newly synthesized proteins before cell viability and paracellular permeability are compromised. Consequently the results help in elucidating mechanisms of metal cytotoxicity that lead to understanding the relationship between tight junction integrity, paracellular transport, and cell viability.
authors.affiliation: St. John's University, College of Pharmacy and Allied Health Professions, Department of Pharmaceutical Sciences, Toxicology Division, Queens, New York 11439, USA.
authors.forename: Anthony R
authors.initials: AR
authors.lastname: Calabro
authors.forename: Dmitry I
authors.initials: DI
authors.lastname: Gazarian
authors.forename: Frank A
authors.initials: FA
authors.lastname: Barile
mesh_terms: D000199:Actins / Q000032:analysis / Q000096:biosynthesis*; D001692:Biological Transport; D018938:Caco-2 Cells; D018929:Cell Culture Techniques; D002463:Cell Membrane Permeability / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D017097:Electric Impedance; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism; D005998:Glycine / Q000378:metabolism; D006801:Humans; D007413:Intestinal Mucosa / Q000187:drug effects*; D008670:Metals / Q000494:pharmacology*; D014176:Protein Biosynthesis / Q000187:drug effects*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D019108:Tight Junctions / Q000187:drug effects / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
chemical_list: D000199:Actins; D008670:Metals; D012333:RNA, Messenger; D014443:Tyrosine; D005998:Glycine
references.citation: Proc Natl Acad Sci U S A. 2002 Apr 16;99(8):5406-11
references.pmid: 11959995
references.citation: Toxicol In Vitro. 2009 Oct;23(7):1298-304
references.pmid: 19619637
references.citation: Toxicol Appl Pharmacol. 1999 Mar 1;155(2):117-26
references.pmid: 10053166
references.citation: Biochim Biophys Acta. 2005 Jun 30;1741(1-2):215-23
references.pmid: 15905073
references.citation: Pharmacol Ther. 2007 Apr;114(1):74-93
references.pmid: 17316817
references.citation: J Pharmacol Toxicol Methods. 2007 Mar-Apr;55(2):176-83
references.pmid: 16891127
references.citation: Toxicol Ind Health. 2004 Jun;20(1-5):57-68
references.pmid: 15807409
references.citation: Toxicol In Vitro. 2009 Dec;23(8):1510-5
references.pmid: 19540330
references.citation: Toxicol In Vitro. 2005 Aug;19(5):675-84
references.pmid: 15896555
references.citation: Proc Natl Acad Sci U S A. 1996 Sep 17;93(19):10366-70
references.pmid: 8816806
references.citation: Environ Health Perspect. 1987 Nov;75:59-64
references.pmid: 3319566
references.citation: Toxicol Sci. 2001 Sep;63(1):132-42
references.pmid: 11509753
references.citation: Toxicol In Vitro. 2006 Sep;20(6):942-53
references.pmid: 16387471
references.citation: Mol Cell Biol. 1995 May;15(5):2547-57
references.pmid: 7537850
references.citation: J Pharm Sci. 1999 Nov;88(11):1161-8
references.pmid: 10564065
references.citation: Toxicol In Vitro. 1996 Feb;10(1):27-36
references.pmid: 20650179
references.citation: J Occup Health. 2003 Nov;45(6):331-4
references.pmid: 14676411
references.citation: J Biol Chem. 1998 Nov 6;273(45):29745-53
references.pmid: 9792688
references.citation: Br Med Bull. 2003;68:167-82
references.pmid: 14757716
references.citation: Biochimie. 2006 Nov;88(11):1549-59
references.pmid: 17070979
references.citation: J Drug Target. 1998;6(1):37-43
references.pmid: 9769019
references.citation: Rev Environ Health. 2007 Apr-Jun;22(2):91-113
references.pmid: 17894202
references.citation: J Inorg Biochem. 2000 Apr;79(1-4):241-4
references.pmid: 10830873
references.citation: Biochem J. 1993 May 1;291 ( Pt 3):657-71
references.pmid: 8489492
references.citation: Arch Toxicol. 2004 Apr;78(4):201-6
references.pmid: 14661071
references.citation: Proc Natl Acad Sci U S A. 1985 Sep;82(18):6133-7
references.pmid: 2994062
references.citation: Crit Rev Oncol Hematol. 2002 Apr;42(1):25-34
references.pmid: 11923066
references.citation: Toxicol In Vitro. 2005 Dec;19(8):1071-8
references.pmid: 16023827
references.citation: Toxicol In Vitro. 2000 Jun;14(3):219-26
references.pmid: 10806372
references.citation: In Vitro Cell Dev Biol Anim. 1998 Sep;34(8):631-5
references.pmid: 9769147
references.citation: J Membr Biol. 2007 Aug;218(1-3):93-100
references.pmid: 17960321
references.citation: Mutat Res. 2003 Dec 10;533(1-2):67-97
references.pmid: 14643413
references.citation: Toxicol In Vitro. 2008 Aug;22(5):1273-84
references.pmid: 18468840
references.citation: Toxicol In Vitro. 1990;4(1):1-7
references.pmid: 20702277
references.citation: Mol Cell Biochem. 2004 Jan;255(1-2):47-55
references.pmid: 14971645
references.citation: Curr Top Med Chem. 2001 Dec;1(6):529-39
references.pmid: 11895129
references.citation: Chem Res Toxicol. 2008 Jan;21(1):28-44
references.pmid: 17970581
references.citation: Med Sci Monit. 2009 Jan;15(1):RA1-5
references.pmid: 19114980
references.citation: Toxicol In Vitro. 2002 Aug;16(4):399-404
references.pmid: 12110278
references.citation: Biochem Pharmacol. 2000 Sep 15;60(6):771-80
references.pmid: 10930531
references.citation: Prog Biophys Mol Biol. 2003 Jan;81(1):1-44
references.pmid: 12475568
references.citation: Chem Res Toxicol. 2005 Dec;18(12):1934-42
references.pmid: 16359184
references.citation: J Biol Chem. 2009 Jan 9;284(2):717-21
references.pmid: 18757361