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Oct 29, 2022 29 likes •310 views

10 th US Korea Conference on XX Destination Nano: Creating the Next Industrial Revolution Integrated Approaches to Meet NanoEHS Challenges Along the Life Cycle of Nano- Enabled Products : Nanoparticles from Photocopiers as an Example D.

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SLIDE 1

Destination Nano: Creating the Next Industrial Revolution

Integrated Approaches to Meet NanoEHS Challenges Along the Life Cycle of Nano- Enabled Products :

Nanoparticles from Photocopiers as an Example

  • D. Bello,1,2,4 M. Khatri,3 A. Pal,1,3 J. Martin, S.

Pirella,2,4 P. Gaines 3 P. Demokritou2,4

1

100 nm

  • 1. University of Massachusetts Lowell; Department of Work Environment;
  • 2. Harvard School of Public Health; Molecular and Integrative Physiological

Science

  • 3. Biomedical Engineering and Biotechnology Program
  • 4. Harvard Center for Nanotechnology & Nanotoxicology

Center for Nanotechnology and Nanotoxicology at Harvard School of Public Health

10th US‐Korea Conference on XX

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SLIDE 2

Destination Nano: Creating the Next Industrial Revolution

Disease – A Complex Process

2

Disease Morbidity & Mortality Exposure Tissue Dose Biological interaction Perturbation

System Inputs Biological Function Impaired function

Adaptation / Repair

Andersen et al, Trends in Biotech, 2005,23,3, 122‐127

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SLIDE 3

Destination Nano: Creating the Next Industrial Revolution What Do We Want to Know? What Test Systems Should Be Used?

DHIMITER_BELLO@UML.EDU 3

Proportion of Population Affected Death Permanent impairment Progressive disease Episodic incapacitating illness Episodic illness limiting activity Episodic illness not limiting activity Subclinical pathophysiological changes Physiological responses potentially contributing to pathology Homeostatic physiological responses Dose of pollutant or reactive product to target organs Exposure

Region of Uncertainty

Degree of Adversity

Mauderly et al., Chapt. 4 in: Technical Aspects of Multipollutant air Quality Management, Springer. 2011

ACELLULAR

HUMAN

ANIMAL

CELLULAR

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SLIDE 4

Destination Nano: Creating the Next Industrial Revolution

  • ~60 studies in all, 25 studies in workplaces
  • Area Samplers
  • Number concentration & size distribution + STEM/EDS
  • Need for standardization & harmonization of

measurements & approaches

  • Collection of systematic contextual information
  • Tiered Approach to understanding nature of exposures

dhimiter_bello@uml.edu 4

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SLIDE 5

Destination Nano: Creating the Next Industrial Revolution

Nanomanufacturing: Diverse, Dynamic & Challenging to Study

dhimiter_bello@uml.edu 5

Bekker et al, AnnHyg 2012

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Destination Nano: Creating the Next Industrial Revolution

An Integrated Approach to Fast Track NanoEHS

VALIDATE OBSERVATIONS in ANIMALS / HUMANS VALIDATE OBSERVATIONS in ANIMALS / HUMANS

IN‐VITRO Toxicity Assessment IN‐VITRO Toxicity Assessment

Equivalent Dose Estimates Mechanistically Endpoints

CALCULATE EQUIVALENT DOSE to HUMANS CALCULATE EQUIVALENT DOSE to HUMANS

Regional Deposition Model Sensitivity Analysis

CHARACTERIZE REALISTIC EXPOSURES CHARACTERIZE REALISTIC EXPOSURES

Levels; PCM Variability

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SLIDE 7

Destination Nano: Creating the Next Industrial Revolution

We started here….

FMPS APS CPC

OZONE METER VOC METER

X 2012 DHIMITER_BELLO@UML.EDU 7

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SLIDE 8

Destination Nano: Creating the Next Industrial Revolution

8

Nanoparticle Emissions from Commercial Photocopiers

Bello et al Nanotoxicology 2012

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Destination Nano: Creating the Next Industrial Revolution

Morphology & Chemistry

9

Bello et al Nanotoxicology 2012

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Destination Nano: Creating the Next Industrial Revolution Deposition Models & Dose: Linking in Vivo with In Vitro Dosimetry

10

Mass Flux 0.072 µg/(m2min) & Exposure time of 480 min= 8hr

Estimated lung surface dose of 34.6 µg/m2 Nasal Cavity: 150 cm2 Deep Lungs: 140 m2 SA Lungs/Nasal Ratio = ~1.2 x 104 Deposited Fraction ~5x Nose/Alveolar Dose (cm‐2) ~ 2,500x

Khatri et al Part Fiber Tox 2013

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SLIDE 11

Destination Nano: Creating the Next Industrial Revolution

B) Nasal lavage fluid is collected by aspiration from both nostrils for 1 min each through a silicon tube 2.7 mm in diameter, and trapped in mucous trap

A) The nasal cavity is misted over with saline using a disposable plastic sprayer

C) Traps are connected with the small portable evacuator

A) B) C)

Sample collection

DHIMITER_BELLO@UML.EDU 11

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Destination Nano: Creating the Next Industrial Revolution

Biomarkers in Nasal lavage & Urine

Khatri, Bello et al 2012 Nanotoxicology

12

10 cytokines

  • verexpressed (10‐2X):

G‐CSF, IL‐8, VEGF, IL‐6, IL‐10, MCP1, Fractalkine, TNF‐α, EGF, IL‐1β IFN, MCP‐1, IL‐1α Total Protein Inflammatory cells 8‐OH‐dG in urine UP

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Destination Nano: Creating the Next Industrial Revolution

Particle collection from copy centers Human primary respiratory epithelial cells and THP‐1 cell line exposed PM0.1, PM0.1‐2.5

In vitro cytotoxicity and cytokine release

10ug/ml polymyxine B treatment for 60 min

In vitro exposure

6h 24 h Inflammatory cytokines DNA damage/ROS Apoptosis

30 ug/ml 100ug/ml 300ug/ml

Cell Viability

13

Gene Expression

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Destination Nano: Creating the Next Industrial Revolution

Delivered Dose is Critical

y = -2.8027x + 97.008 y = -8.2431x + 97.008 20 40 60 80 100 120 1 2 3 4 5 6 7 8 9 10 % Cell viability Mass (ug)

Printex 100

Administered Depsoited y = -2.4552x + 79.105 y = -13.64x + 79.105 20 40 60 80 100 1 2 3 4 5 6 7 8 9 10 % Cell viability Mass (ug)

SWCHN-ox

Administered Depsoited y = -1.7322x + 89.866 y = -1.7604x + 89.866 20 40 60 80 100 120 1 2 3 4 5 6 7 8 9 10 % Cell viability Mass (ug)

TiO2 P25

Administered Depsoited

Pal et al (manuscript in preparation)

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SLIDE 15

Destination Nano: Creating the Next Industrial Revolution

GCSF

6 h 2 4 h 5000 10000 15000 20000

Concentration in pg/ml

IL-I

6 h 2 4 h 2000 4000 6000

Concentration in pg/ml

GM-CSF

6 h 2 4 h 500 1000 1500

Concentration in pg/ml

TNF-

6 h 2 4 h 2000 4000 6000 8000

Concentration in pg/ml

IL-6

6 h 2 4 h 1000 2000 3000 4000 5000

Concentration in pg/ml

IL-1

6 h 2 4 h 20 40 60

Concentration in pg/ml

MCP-1

6 h 2 4 h 2000 4000 6000 8000

Concentration in pg/ml

VEGF

6 h 2 4 h 1000 2000 3000

Concentration in pg/ml

IL-8

6 h 2 4 h 5000 10000 15000 20000

Concentration in pg/ml

THP‐1 Cell Line‐ Inflammatory Cytokines

* * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * *

15

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Destination Nano: Creating the Next Industrial Revolution Delivered dose makes a difference

y = 93.21x + 1010.5 R² = 0.77

y = 12.929x + 371.43 R² = 0.97

5000 10000 15000 50 100 150 200

TNF‐a (pg/mL) Delivered Dose (ug/mL)

6 hr time point Ratio of Slopes Copier NP/CuO ~ 7x

Copier‐ NP CuO

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Destination Nano: Creating the Next Industrial Revolution

Apoptosis Results

C

  • l

l e c t e d N P C a r b

  • n

N P s C u

  • 0.0

0.5 1.0 1.5 2.0 2.5

30 100 300 Annexin V+ cells (Fold change over control)

* * * * * * *

17

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Destination Nano: Creating the Next Industrial Revolution

DNA damage‐ Comet assay

18

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Destination Nano: Creating the Next Industrial Revolution

Gene Expression (RT‐qPCR), 5ug/mL

April Gu’s group

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Destination Nano: Creating the Next Industrial Revolution

PM0.1‐2.5, Cytokines in THP‐1

Khatri et al Inh Tox 2013

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Destination Nano: Creating the Next Industrial Revolution

PM0.1‐2.5, Apoptosis in THP‐1

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Destination Nano: Creating the Next Industrial Revolution

Lavaged neutrophils 24 hours post-particle exposure

BALF Neutrophils (millions)

R P M I / 1 % F B S P M < . 1 ( . 2 m g / k g ) P M < . 1 ( . 6 m g / k g ) P M < . 1 ( 2 m g / k g ) P M . 1

  • 2

. 5 ( . 2 m g / k g ) P M . 1

  • 2

. 5 ( . 6 m g / k g ) P M . 1

  • 2

. 5 ( 2 m g / k g ) F e 2 O 3 ( . 2 m g / k g ) F e 2 O 3 ( . 6 m g / k g ) F e 2 O 3 ( 2 m g / k g ) W F ( . 2 m g / k g ) W F ( . 6 m g / k g ) W F ( 2 m g / k g ) 0.0 0.5 1.0 1.5 2.0 2.5

*

#

PM0.1 (mg/kg bw) PM0.1-2.5 (mg/kg bw) Welding fumes (mg/kg bw) Fe2O3 (mg/kg bw) RPMI/10% FBS

In vivo instillation study in mice done at Harvard, Prof. Demokritou’s group Pirella et al Inh Tox 2013

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Destination Nano: Creating the Next Industrial Revolution

Test System Cytokine

In Vitro (Fine particle dosing) In Vitro (Nanoparticle dosing)

Khatri et al 2013

In Vivo

THP-1 Nasal Epithelial Cells Small Airway Epithelial Cells THP-1 Nasal Epithelial Cells Small Airway Epithelial Cells Nasal lavage of Healthy Volunteers (Khatri et al 2012)

     

IL-8

      

IL-6



  • 
  • 

IL-1

  • 



GM-CSF

  • 
  • TNF-

  • 

  • 

MCP-1

  • 
  • 

VEGF

  

IL-1

  • 

  • G-CSF
  • 

Fractalkine

  • 
  • 

EGF

  • 

 

IFN-

  • Summary of cytokine production

Khatri et al 2013, Inh Tox

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Destination Nano: Creating the Next Industrial Revolution

Control Exposed week 1 Exposed week 2 Exposed week 3

  • 1

1 2 3 4 5 lnIl_6 week_type

W1 W2 W3 C

Chronic Exposures, NL

5 exposed in 3 centers; 2‐3 weeks & 8 controls

Same picture for:

Total Protein

IL‐8 TNF‐α MCP‐1 8‐OH‐dG

 G‐CSF Under‐expressed Acute Exposure, NL 6

24

Ln(IL‐6) No change: VEGF, IL‐10, Fractalkine, EGF, IL‐1β

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Destination Nano: Creating the Next Industrial Revolution

Conclusions

  • Approach is generalizable
  • Suitable for a large number of life cycle

scenarios of nano‐enabled products

  • It provides important context for risk and

hazard assessment to Nanotech companies

  • Important methodological insights for in vitro

testing and comparative assessment

  • Currently used to validate FRAS BOD as a

screening assay

Destination Nano 2013: Creating the Next Industrial Revolution

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Destination Nano: Creating the Next Industrial Revolution

Questions?

X 2012 DHIMITER_BELLO@UML.EDU 26

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Destination Nano: Creating the Next Industrial Revolution

Laser Printer Emissions & Human Health

“International foundation nano Control” database in Germany:

  • 2,500 persons who claim to be sick from laser printer

emissions

  • 90% report respiratory tract complaints, glossalgia,

chronic cough, rhinitis, and inflammation of the throat, tongue and paranasal sinuses

  • ~ 30% developed asthma
  • Inflammation of eyes, skin, diffuse pain and loss of

hair

DHIMITER_BELLO@UML.EDU 27

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Destination Nano: Creating the Next Industrial Revolution

Nel et al. Science 311, 622 (2006)

The hierarchical oxidative stress model