Pulmonary Effects of Engineered Nanoparticles

Hasan Bayram MD,PhD.

Department of Respiratory Medicine, School of Medicine, University of Gaziantep

E-mail: bayram@gantep.edu.tr

TTS 14th Annual Congresss, Antalya 13-17 April 2011.

2

Conflicts of Interest• Abdi Ibrahim Drug Company sponsored

my attendance at international congresses twice in 2008 and 2010.

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PubMed Publications: “Nanoparticles” AND “Lung” AND “Toxicity”

Publications

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Content• Nanoparticles entering the lung and the cell• Histopathological and inflammatory changes• Interaction with allergens and LPS• Effects of cell permeability and toxic parameters• Effects on cell viability, apoptosis, cell cycle (cell

lines, primary cells, COPD cells)• Signaling pathways, transcriptional effects• Effects on oxidative stress• Summary

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Nanopartcle Application at Experimental Settings

• Pharyngeal aspiration

• Intra-tracheal instillation

• Inhalation of aerosols

• Exposure to freshly generated nanoparticles

• Oral application

• Intraarticular injection

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6

MWCNT in the Conducting Airways

ciliaMWCNT

MWCNT

Courtesy of John Howard, NIOSH

7

MAKFJPARK

BRONCHİOL

Ryman-Rasmussen JP et al. AJRCMB, 2009 .

MWCNT Distribution in Lung Tissue of Mice 1d After Inhalation of Aerosol (100mg/m3) for 6hrs

8

8

Migration of MWCNTs from Lung: Pleural Penetration

Courtesy of John Howard, NIOSH

9

Polystyrene Nanoparticle Trafficking Across Alveolar epithelium

Yacobi NR et al, 2008

1010

Effects of Pharyngeal Aspiration of Single Wall Carbon Nanotube on Respiratory Tract in Mice

Shvedova AA et al, 2005

1 day post exposure 40 g/mouse SWCNT

28 days post exposure 40 g/mouse SWCNT

20 microns 20 microns

Acute inflammation and fibrosis Granulomas

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Percentage of Neutrophils in Lung Lavage of Rats and Mice 24 hr After Intratracheal Instillation

Oberdörster G et al, 2005

13

Effects of Titanium Dioxide Nanoparticles (TNP, 20/mg/kg) on Cytokine Levels in BAL After a

Single Intratracheal Instillation

Park EJ et al. Toxicology. 2009

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TiO2 Particles Coated with Silica Induce Level of BAL Cells of Mice Following 2h, 2h/4d, 2h/4wks

Exposure

Rossi FM et al, Toxicol Sci. 2010

A=2hrsB=2hrs/4dysC=2hrs/4wks

• CXCL1 and TNF-α mRNA levels increased

16

Effects of Multi-walled Carbon Nanotubes (100nm; 0.4mg/ml) on Cytokine Release From Human Keratinocytes

Witzmann FA et al, 2006

18

Effect of 300µg/ml cnTiO2 on Expression of TNF-α and CXCL 1 mRNA in Human Macrophages after 3-24hrs

Rossi FM et al, Toxicol Sci. 2010

19

Effects of Inhaled TiO2 Particles (42mg/m3,11days) on microRNA Expression in Lungs of Mice

Halappanavar S, et al. Environ Mol Mutagen. 2011

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Effects of Gold NPs ± LPS on Gene Expression of TNF-α, IL-8 and iNOS in a Triple-Culture Model of Macrophages, A549 and Dentritic

Cells

•No effect on inflammatory and anti-inflammatory cytokine release, either.

Brandenberger C, et al. Toxicol Appl Pharmacol, 2010.

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Effects of Titanium Dioxide Nanoparticles (TNP, 20/mg/kg) on Levels of IgE in BAL and Blood After

a Single Intratracheal Instillation

Park EJ et al. Toxicology. 2009

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IL-13 mRNA and Protein Levels at 1d After MWCNT Inhalation

Ryman-Rasmussen JP et al. AJRCMB, 2009 .

• Protein levels of PDGF and TGF-β1, • mRNA levels of IL-5, CCL2/MCP-1, CCL11/eotaxin and CXCL9 were increased by OVA + MWCNT

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Lung Histopathology in Rats Exposed to MWCNT for 21d (blue indicates collagen).

MWCNT MWCNT + LPS

Cesta MF et al. AJRCMB’09

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Effects of CB or MWCNT on Lung Injury

Cesta MF et al. AJRCMB’09

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Tight Junction Expression and Transepithelial Electrical Resistance of A549 Cells

Rothen-Rutishauser B et al. Environ Sci Technol. 2009

10’=12µg/cm2

20’=19µg/cm2

30’=24µg/cm2

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Effects of Zinc Oxide NPs on Electrical Resistance (RT) of Rat Alveolar Epithelial Cell Cultures

Kim YH, et al. AJRCCM 2010.

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Effects of Zinc Oxide NPs on Lactate Dehydrogenase Release from Rat Alv. Epithelial Cell Cultures

Kim YH, et al. AJRCCM 2010.

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Effects of Tungsten Carbide (WC) and Cobalt-doped Tungsten Carbide (WC-Co) NPs on Viability of Mammalian Cells.

Bastian S et al, 2009.

Human Keratinocyte Cells A549 Alveolar-epithelial Cells

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Effects of TiO2 and MWCNT on BEAS-2B Cell Viability

0 3 10 30 100

300

0.0

0.5

1.0

*

*

*p<0,0001

TiO

224. hour

Op

tica

l D

ensi

ty

0 3 10 30 100

300

0.0

0.5

1.0

1.5

*

*p<0,0001

*

48. Hour

0 3 10 30 100

300

0.0

0.2

0.4

0.6

0.8

1.0

MW

CN

T

*p<0.0001

* *

(g/ml)

Op

tica

l D

ensi

ty

0 3 10 30 100

300

0.0

0.2

0.4

0.6

0.8

1.0*p<0,0001

**

(g/ml)

Op

tica

l D

ensi

ty

Gögebakan B, et al. TTS Congress, 2011.

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Effects of TiO2 on Viability of Bronchial Epithelial Cells from Smokers and COPD Patients

0 3 10 30 100 3000

1

2

3

*

#*p<0,0001#p<0,0001

*

SM

OK

ER

S24. Hour

Op

tica

l D

ensi

ty

0 3 10 30 100 3000.0

0.5

1.0

1.5

2.0

2.5

48. Hour

0 3 10 30 100 3000.0

0.5

1.0

1.5

2.0

2.5*p<0,001**p=0,0016

CO

PD

TiO2 (g/ml)

** ** ** *

Op

tica

l D

ensi

ty

0 3 10 30 100 3000.0

0.5

1.0

1.5

2.0

*

*p<0,001

* *

TiO2 (g/ml)

Gögebakan B, et al. TTS Congress, 2011.

33

Effects of MWCNT on Viability of Bronchial Epithelial Cells from Smokers and COPD Patients

0 3 10 30 100 3000.0

0.5

1.0

1.5

**

*p<0,0001

SM

OK

ER

S24. Hour

Op

tica

l D

ensi

ty

0 3 10 30 100 3000.0

0.5

1.0

1.5

**

*p<0,0001

48. Hour

0 3 10 30 100 3000.0

0.5

1.0

1.5

*

*p=0,0418

CO

PD

MWCNT (g/ml)

Op

tica

l D

ensi

ty

0 3 10 30 100 3000.0

0.2

0.4

0.6

0.8

1.0

* * *

*p=0,0068

MWCNT (g/ml)

Gögebakan B, et al. TTS Congress, 2011.

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Effects of 48h Incubation With TiO2 and MWCNT on BEAS-2B Cell Apoptosis

LL LL LR LR UR UR UL UL0

2

4

6

885

90

95

100

*

*p= 0,0009**p=0,0169

*

**

SF (0g/ml TiO2)

300 g/ml TiO2

Viable Early Apo Late Apo Necrotic

Cel

ls %

LL LL LR LR UR UR UL UL

0

5

10

1575

80

85

90

95

100

*

*p<0,0001**p=0,0003***p<0,0001

*****

SF (0g/ml MWCNT)

300 g/ml MWCNT

Viable Early Apo Late Apo Necrotic

Cel

ls %

TiO2MWCNT

Gögebakan B, et al. TTS Congress, 2011.

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Effects of Single-walled Carbon Nanotubes and C(60) Fullerenes on Cell Cycle of Mouse Lung Epithelial Cells

Jacobsen NR et al, 2008

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Effects of 100µg/ml Polyamidoamine Dendrimer(PAMAM) Nanoparticles and 3-Methyladenine (Autophay Inhb.)

on Autophagy in A549 Cells After 24hrs

Li FT et al. J Mol Cell Biol. 2009 autophagosomes

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Effects of Different Sized CeO2 Nanoparticles on pERK, p38 and pJNK in BEAS-2B Cells after 24 h

Eom HJ et al. Toxicol Lett. 2009

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Expressions of NF-B, I-B and Nrf-2 in Cytosolic and Nuclear Fractions of BEAS-2B Cells Exposed to Different

Sized CeO2 Nanoparticles for 24h.

Eom HJ et al. Toxicol Lett. 2009

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Effect of ZnO on Expression of IL-8 mRNA in Human BECs

Wu W, et al. EHP, 2010

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Effect of ZnO Nanoparticles ± NAC, on IntracellularCalcium Concentrations

Huang CC et al. Toxicol In Vitro. 2009

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Effects of 20µg/cm2 Carbon Black and TiO2 NPs on Caspase 8 and Caspase 3/7 Activity in 16HBE14o- cells

Hussain S, et al. Part Fibre Toxicol. 2010

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Non-cellular Nanoparticle ROS Activity

Elder A, et al 2008 (http://www.epa.gov)

Surface AreaParticle Mass

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Effects of Cerium Oxide NPs (30nm) on ROS Induction in BEAS-2B Cells.

Park EJ et al, 2008

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Summary• Nanoparticles enter into airways, parenchyme, cells• Causes collagen deposition, fibrosis, granuloma,

inflammation• Modulate allergic response, synergism with LPS• Increases in epithelial permeability• Decreases in cell viability and cell cycle progression,

increases in apoptosis and autophagy• Diseased cells (COPD) may be more susceptible• Activation of signaling mechanism and transcription

factors that are sensitive to oxidative stress• ROS formation and induce oxidative stress

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Acknowledgements

Prof. Günter Oberdörster

Director, University of Rochester Ultrafine Particle

Center

Thank You for Your Attentionbayram@gantep.edu.tr