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Tr ai l of b re ad cr um bs Discovering the molecular mechanisms of nanotoxicity in fish. Christopher Anthony Dieni Department of Chemistry and Biochemistry Mount Allison University. UNB Biology Seminar Series Friday, March 28 th , 2014. Michael Owens. Nanotechnology - PowerPoint PPT Presentation

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Michael OwensTrail of breadcrumbsDiscovering the molecular mechanisms of nanotoxicity in fishChristopher Anthony DieniDepartment of Chemistry and BiochemistryMount Allison UniversityUNB Biology Seminar SeriesFriday, March 28th, 2014

Wikimedia CommonsNanotechnology

Origins traced back to the mid-20th centuryPhysicist Richard Feynman delivers his talk Theres Plenty of Room at the Bottom American Physical Society meeting at Caltech, December 29, 1959

In more than a half-century since then, we have become dependent on nanotechnology for:BiosensorsAntimicrobial agentsDrug deliveryMolecular scale electronicsNanorobotics and much more!Todays talk

Design of nanomaterials and constituent materials

Synthesis/engineering of nanomaterials

Functionalization/conjugation of nanomaterials for specific purposes (e.g. drug delivery)Todays talk

Design of nanomaterials and constituent materials

Synthesis/engineering of nanomaterials

Functionalization/conjugation of nanomaterials for specific purposes (e.g. drug delivery)

Release of nanomaterials in the environment and interaction with indigenous organismsNanoparticle toxicity

Nel, A. et al. (2006) Toxic potential of materials at the nanolevel. Science 311: 622-627Nanoparticle toxicity

Nel, A. et al. (2006) Toxic potential of materials at the nanolevel. Science 311: 622-627

Nanoparticle-protein interactions

University of Massachusetts

Nanoparticle-protein interactions

University of Massachusetts

Nanoparticle-protein interactions

University of Massachusetts

Nanoparticle toxicity

Nel, A. et al. (2006) Toxic potential of materials at the nanolevel. Science 311: 622-627Model nanoparticle: nanoscale zinc oxide (nZnO)

WebElements.comWikimedia commons25 nm nZnO

Dieni et al. Comp Biochem Physiol Toxicol Pharmacol in pressScale bar = 1 m25 nm nZnO

Dieni et al. Comp Biochem Physiol Toxicol Pharmacol in pressScale bar = 1 m

Wikimedia commons

Uses of nZnOUK Daily Mail

In vivo exposureIn vitro exposureLive physiological/systemic (e.g. cardiorespiratory physiology)Postmortem biochemical assays(e.g. antioxidant enzymes, damage markers)Complex media(e.g. pooled rat blood plasma)Simplified conditions(e.g. BSA solution)Nanotoxin?In vivo exposureIn vitro exposureLive physiological/systemic (e.g. cardiorespiratory physiology)Postmortem biochemical assays(e.g. antioxidant enzymes, damage markers)Complex media(e.g. pooled rat blood plasma)Simplified conditions(e.g. BSA solution)Nanotoxin?

In vivo exposureIn vitro exposureLive physiological/systemic (e.g. cardiorespiratory physiology)Postmortem biochemical assays(e.g. antioxidant enzymes, damage markers)Complex media(e.g. pooled rat blood plasma)Simplified conditions(e.g. BSA solution)Nanotoxin?

The white sucker, Catostomus commersonii

Benthic (bottom-feeding)

Likely to come into contact with well-dispersed or sedimentary nanoparticles

Easily accessible (Silver Lake)

1 mg/L nZnO30 hours

Dr. Tyson J. MacCormackKathryn M. A. Butler, B.Sc. Biochem (Hons) 2013

Live physiological/systemic level

ElectrocardiographyRespirometry (resting MO2)

Live physiological/systemic level

ElectrocardiographyRespirometry (resting MO2)

Heart rate decreases by 25% (temporarily)No change in resting MO2Live physiological/systemic level

Two schools of thought:Physiological changes overt enough to affect a whole, live organism are most meaningful

Is a toxic or pathologic response grave enough?

Is a therapeutic good enough?Live physiological/systemic level

Two schools of thought:Physiological changes overt enough to affect a whole, live organism are most meaningful

Is a toxic or pathologic response grave enough?

Is a therapeutic good enough?Changes at the biochemical level may not reveal themselves at the systemic level yet

Incubation period of an infectious disease before virulence and immune response

Initial mutations leading to cancer

EtcIn vivo exposureIn vitro exposureLive physiological/systemic (e.g. cardiorespiratory physiology)Postmortem biochemical assays(e.g. antioxidant enzymes, damage markers)Complex media(e.g. pooled rat blood plasma)Simplified conditions(e.g. BSA solution)Nanotoxin?

In vivo exposureIn vitro exposureLive physiological/systemic (e.g. cardiorespiratory physiology)Postmortem biochemical assays(e.g. antioxidant enzymes, damage markers)Complex media(e.g. pooled rat blood plasma)Simplified conditions(e.g. BSA solution)Nanotoxin?

Nanoparticle toxicity

Nel, A. et al. (2006) Toxic potential of materials at the nanolevel. Science 311: 622-627Reactive oxygen species (ROS)

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)

Neal I. Callaghan, Honours Biochemistry student

Wikimedia commons

Armstrong JS et al (2004) Bioessays 26: 894-900

Wikimedia commons

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)Dieni et al. Comp Biochem Physiol Toxicol Pharmacol in press

aControlnZnOG6PDH activity decreased with nZnO exposure (~29%)

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)GR remained unchangedDieni et al. Comp Biochem Physiol Toxicol Pharmacol in pressControlnZnO

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)Dieni et al. Comp Biochem Physiol Toxicol Pharmacol in pressControlnZnO

aTotal glutathione levels increased with nZnO exposure (~56%)

Neal I. Callaghan, Honours Biochemistry student

Wikimedia commons

Armstrong JS et al (2004) Bioessays 26: 894-900

Wikimedia commons

Neal I. Callaghan, Honours Biochemistry student

Wikimedia commons

Armstrong JS et al (2004) Bioessays 26: 894-900

Wikimedia commonsDieni et al. Comp Biochem Physiol Toxicol Pharmacol in pressControlnZnO

bcAconitase activity decreased with nZnO exposure (~65%)

Reactivated by supplementation with Fe(NH4)2SO4 (source of Fe2+)

Neal I. Callaghan, Honours Biochemistry student

Wikimedia commons

Armstrong JS et al (2004) Bioessays 26: 894-900

Wikimedia commons

Neal I. Callaghan, Honours Biochemistry student

Wikimedia commons

Armstrong JS et al (2004) Bioessays 26: 894-900

Wikimedia commonsDieni et al. Comp Biochem Physiol Toxicol Pharmacol in pressControlnZnO

Malondialdehyde (MDA) levels remained unchangedRND systemsG6PDH~29% decreaseGR-Glutathione~56% increaseAconitase~65% decrease (reactivated by Fe2+)MDA-Explanation please?Hepatic responses to 1 mg/L nZnO exposure

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (G6PDH)XX

Superoxide radical anionHydroxyl radical

Neal I. Callaghan, Honours Biochemistry student

OROR others

Reduced glutathione (GSH)Oxidized glutathione (GSSG)NADPHNADP+Glutathione reductase (GR)G6P6PGLGlucose-6-phosphate dehydrogenase (