Iron nanoparticles increase 7-ketocholesterol-induced cell death, inflammation, and oxidation on murine cardiac HL1-NB cells.
OBJECTIVE
To evaluate the cytotoxicity of iron nanoparticles on cardiac cells and to determine whether they can modulate the biological activity of 7-ketocholesterol (7KC) involved in the development of cardiovascular diseases. Nanoparticles of iron labeled with Texas Red are introduced in cultures of nonbeating mouse cardiac cells (HL1-NB) with or without 7-ketocholesterol 7KC, and their ability to induce cell death, pro-inflammatory and oxidative effects are analyzed simultaneously.


METHODS
Flow cytometry (FCM), confocal laser scanning microscopy (CLSM), and subsequent factor analysis image processing (FAMIS) are used to characterize the action of iron nanoparticles and to define their cytotoxicity which is evaluated by enhanced permeability to SYTOX Green, and release of lactate deshydrogenase (LDH). Pro-inflammatory effects are estimated by ELISA in order to quantify IL-8 and MCP-1 secretions. Pro-oxidative effects are measured with hydroethydine (HE).


RESULTS
Iron Texas Red nanoparticles accumulate at the cytoplasmic membrane level. They induce a slight LDH release, and have no inflammatory or oxidative effects. However, they enhance the cytotoxic, pro-inflammatory and oxidative effects of 7KC. The accumulation dynamics of SYTOX Green in cells is measured by CLSM to characterize the toxicity of nanoparticles. The emission spectra of SYTOX Green and nanoparticles are differentiated, and corresponding factor images specify the possible capture and cellular localization of nanoparticles in cells.


CONCLUSIONS
The designed protocol makes it possible to show how Iron Texas Red nanoparticles are captured by cardiomyocytes. Interestingly, whereas these fluorescent iron nanoparticles have no cytotoxic, pro-inflammatory or oxidative activities, they enhance the side effects of 7KC.
Kahn
Edmond
E
INSERM U678/UMR - S UPMC, IFR 14, CHU Pitié-Salpêtrière, 75634 Paris Cedex 13, France. kahn@imed.jussieu.fr
Baarine
Mauhamad
M
Pelloux
Sophie
S
Riedinger
Jean-Marc
JM
Frouin
Frédérique
F
Tourneur
Yves
Y
Lizard
Gérard
G
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D007501:Iron / Q000009:adverse effects*; D007653:Ketocholesterols; D051379:Mice; D009205:Myocarditis / Q000139:chemically induced* / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000473:pathology*; D053758:Nanoparticles / Q000009:adverse effects*; D010084:Oxidation-Reduction / Q000187:drug effects
D007653:Ketocholesterols; D007501:Iron; C003001:7-ketocholesterol
Cytometry A. 2006 May;69(5):359-73
16604541
J Opt Soc Am A. 1987 Nov;4(11):2101-6
3681518
Adv Exp Med Biol. 2007;620:57-73
18217335
Science. 2001 Aug 17;293(5533):1289-92
11509722
Int J Immunopathol Pharmacol. 2007 Oct-Dec;20(4):685-95
18179741
FEBS Lett. 1998 Dec 4;440(3):434-9
9872417
Pathol Biol (Paris). 2001 Apr;49(3):194-8
11367552
J Biol Chem. 2009 Jun 5;284(23):15826-34
19351882
J Leukoc Biol. 1990 May;47(5):440-8
2159514
Science. 1998 Sep 25;281(5385):2013-6
9748157
Comput Med Imaging Graph. 1993 Jul-Oct;17(4-5):397-404
8306315
Science. 2005 Jan 28;307(5709):538-44
15681376
Hum Reprod. 2000 Apr;15(4):756-61
10739815
Mol Aspects Med. 2009 Jun;30(3):153-70
19248805
Redox Rep. 2006;11(6):255-62
17207307
Biochem Soc Trans. 2007 Jun;35(Pt 3):527-31
17511644
J Nutr Biochem. 2009 Jan;20(1):45-61
18495460
Nat Biotechnol. 2003 Oct;21(10):1166-70
14520401
Future Cardiol. 2009 Jul;5(4):343-54
19656059
Prog Biophys Mol Biol. 2006 Jan-Apr;90(1-3):270-98
16140363
Cytometry A. 2007 Aug;71(8):550-62
17458884
Circ Res. 2006 Sep 29;99(7):692-705
17008598
Nat Clin Pract Oncol. 2008 Jan;5(1):44-54
18097456
Cell Mol Life Sci. 2001 Nov;58(12-13):1950-60
11766890
Cell Biol Toxicol. 2005 Mar;21(2):97-114
16142584
J Investig Med. 2006 Nov;54(7):412-24
17169263
J Microsc. 1999 Mar;193(Pt 3):227-43
10199002
Science. 2000 Sep 8;289(5485):1757-60
10976070
Adv Exp Med Biol. 2007;620:168-80
18217343
Adv Drug Deliv Rev. 2006 Dec 1;58(14):1471-504
17116343
FASEB J. 1998 Dec;12(15):1651-63
9837855
Braz J Med Biol Res. 2008 Jul;41(7):545-56
18719735
Toxicol In Vitro. 2005 Oct;19(7):975-83
16125895
Methods Enzymol. 1994;233:539-48
8015489
Int J Nanomedicine. 2007;2(4):609-22
18203428
Proc Natl Acad Sci U S A. 1998 Mar 17;95(6):2979-84
9501201
Anal Quant Cytol Histol. 2006 Feb;28(1):14-26
16566276
Int J Nanomedicine. 2007;2(2):129-41
17722542
Free Radic Biol Med. 2004 Apr 15;36(8):976-84
15059638
Environ Health Perspect. 2005 Jul;113(7):823-39
16002369
Anal Cell Pathol. 1996 Oct;12(1):45-56
8933908
J Neurosci Res. 2000 Mar 15;59(6):788-96
10700016
Anal Quant Cytol Histol. 2004 Feb;26(1):1-6
15032074
Cytometry. 2000 May 1;40(1):42-9
10754516
Environ Health Perspect. 2004 Jul;112(10):1058-62
15238277
Atherosclerosis. 1999 Jan;142(1):1-28
9920502
Nat Nanotechnol. 2009 Oct;4(10):634-41
19809453
Toxicol Sci. 2006 Jan;89(1):42-50
16177233