Post on 01-Dec-2021
a tour of new features
Navigating Environmental Public
Health Behind Health Impacts of Metal
Nanoparticles
Jong Sung Kim, MSc, PhD, Professor & Director of HERC Laboratory
Department of Community Health & Epidemiology
Department of Microbiology & Immunology
Faculty of Medicine, Dalhousie University
HERC Laboratory was established as one of three national centres (UT,
Dalhousie, UBC) of the Canadian Aerosol Research Network (CARN) to
enable a collective study of climate, air quality, population exposures, and
human health. It was created using research awards and matching resources
($4.2 million) from the Canada Foundation for Innovation (CFI), the Nova
Scotia Research and Innovation Trust (NSRIT), Dalhousie Faculty of
Medicine, and industrial partners.
Analytical instrumentation: LC-MS, ICP-MS, GC-MS, HPLC, IC
DALHOUSIE HERC LABORATORY
RESEARCH PROGRAM
To better understand how emerging hazards and exposures lead to
adverse health outcomes at various levels of biological organization
(from cellular and molecular levels to populations) and how human
body modify these responses to maintain homeostasis (host-defense).
Healthy environments for healthy people
WHAT WE DO
• Advance knowledge of the relationship between health and the environment
(evaluating, tracking, and preventing environmental health hazards).
Hazard
Exposure
Health outcome
Dissemination Prevention
Data Stakeholders*
Improved health
*Stakeholders include: Governmental agencies
academia, health care system, policy makers, media
public, business & industry, non-governmental organizations
We have features for every step of the way
WHERE WE LIVE? EMERGING RISKS…
Nanotechnology is seen as the way
of the future will bring a lot of
benefits nothing is ever perfect!
Cartoon source: www.cartoonstock.com
WHAT IS NANO?
Source: National Nanotechnology Initiative (NNI)
• “Nano” is a prefix that comes from the Greek word for dwarf.
• It simply means one billionth (1 nm = 10-9 m).
Materials designed and produced to have structural features with
Nanoscale
Think really, really small
1-100 nm
WHAT’S SO SPECIAL ABOUT THE NANOSCALE?
Nanoscale-associated behavior
Scale at which Quantum Effects dominate properties of materials
- The materials’ properties change at the nanoscale (quantum effects).
- Size-dependent properties are the major reason that nanoscale objects have such amazing potential.
Source: National Nanotechnology Initiative (NNI)
WHAT’S SO SPECIAL ABOUT THE NANOSCALE?
Nanoscale-associated behavior
Scale at which surfaces & interfaces play a large role
- Nanomaterials have far larger surface areas than larger-scale materials.
- As surface area of a material increases, a greater amount of the material can come into contact with surrounding materials, thus affecting reactivity.
High Surface area to volume ratio
Surface area increasing
S.A.=h*w*#
• 6 cm2 = (1 cm)2 * 6
• 60 cm2 = (1/10 cm)2 * 6 * 1000
• 6*107 cm2 = (1/10 cm)2 * 6 * 109
Source: National Nanotechnology Initiative (NNI)
WHAT’S SO SPECIAL ABOUT THE NANOSCALE?
Higher Surface area to volume ratio
NMs can be made to be stronger, lighter, more durable, water-repellent, antimicrobial, self-cleaning, better electrical conductors among other traits.
Benefits of Small Systems
Doing more with less
Multi-functionality
New applications
Quicker performance
Better performance
NANOTECHNOLOGY APPLICATIONS
Source: National Nanotechnology Initiative (NNI)
NT Applications
US $76 billion by 2020
POTENTIAL HEALTH IMPACTS
The Bio-Nano Interface: Nanoparticle-Bio Interactions
Scale at Which Much of Biology Occurs
- Hemoglobin, the protein that carries oxygen through the body,
is 5.5 nm in diameter.
- A strand of DNA, one of the building blocks of human life, is
only about 2.5 nm in diameter.
Interfacing engineered NPs with biological systems:
Anticipating adverse nano-bio interactions
Source: National Nanotechnology Initiative (NNI)
POTENTIAL HEALTH IMPACTS
Risk = Function of Hazard and Exposure.
Human exposure to NPs: skin (dermal), lungs (inhalation), gastrointestinal
tract (ingestion), or by injecting as a formulated medicine.
The most critical concern over health & environmental effects: when NPs
are aerosolized (highly mobile & enter the human body via inhalation).
Source: Card et al (2008) Am J Physiol Lung Cell Mol Physiol 295: L400-411.
THE CHALLENGE OF TESTING NP TOXICITY
Toxicity test methods, which are routinely applied to testing of nanomaterials, were originally developed for soluble chemicals.
Nanomaterials are different than chemicals.
We should not treat them as chemicals.
Nanomaterials are fundamentally different from many 'conventional' chemicals as they often have limited or no solubility at all and are potentially released to the environment.
Only limited nano-specific guidance on ecotoxicity testing is currently available (OECD)
TOXICITY TESTING MODELS
In Vitro (Cells) Study
In Vivo (Animal) Study
Nanomaterials
Nanoscale-
associated behavior
Human
DESIGN CRITERIA
Design Criteria:
Toxicity Testing of Environmental Agents
Toxicity Testing in the 21st Century: a vision and a strategy. US National Research Council of the National Academies 2007
OPTIONS FOR FUTURE TOXICITY TESTING STRATEGIES
Toxicity Testing in the 21st Century: a vision and a strategy. US National Research Council of the National Academies 2007
PULMONARY TOXICITY ASSESSMENT
In Vivo Model (using animals)
Allow realistic route with reproducible NP dosing & lung
distribution
Inhalation & instillation exposures
Bronchoalveolar lavage fluid evaluation
• Cell differential analysis
• Lactate dehydrogenase
• Cytokine/chemokines
General Components of the Pulmonary Bioassay
Lung tissue analysis
• Lung histopathology
• Dosimetry
• Enzyme activity
BRONCHO ALVEOLAR LAVAGE (BAL)
Cytokines/chemokines, total protein &
lactate dehydrogenase (LDH)
Saline solution
Fluids Cells
Total cell & Differential cell counts
(macrophages, neutrophils, lymphocytes etc)
A saline wash of the
airways (broncho) and air
sacs (alveolar) for
recovery of inflammatory
cells.
Indicate inflammatory
responses and
cytotoxicity induced by
toxicant
IN VIVO NANOTOXICOLOGY
• Human exposure to NPs & environmental bacteria can occur
simultaneously.
• The purpose of this study was to determine if host defense
against bacterial infection is enhanced or impaired by Cu NPs in
a murine pulmonary infection model.
BRONCHO ALVEOLAR LAVAGE (BAL)
Schematic of the pulmonary bacterial
clearance model. We established a murine
pulmonary infection model of Klebsiella
pneumoniae (K.p.) to determine if pulmonary
bacterial clearance is impaired by NP
exposure. Following both sub-acute inhalation
and intratracheal instillation, mice were
intratracheally challenged with K.p. bacteria at
a dose of 1.4 ± 0.1 × 105 CFUs/mouse.
IN VITRO MODEL IN NANOTOXICOLOGY
In vitro assays can serve as a screening method for assessing NP
toxicity.
opportunity for extensive investigation of
(can’t be conducted in vivo).
Alternative and the 3 Rs (Replace, Reduce, Refine)
Submerged System
Particle suspension in medium & Exposure of
immersed cells
In Vitro Exposure of Lung Cells to Nanoparticles
Air-Liquid Interface (ALI)
Air delivery of NPs to lung cells at the ALI
NP INTERACTION WITH ALVELOAR EPITHELIUM
Source: Card et al (2008) Am J Physiol Lung Cell Mol Physiol 295: L400-411.
into the systemic circulation is
most likely to across with its very large
surface area (>100 m2 in humans) and thin barrier thickness.
However, interactions between
LIMITATIONS: IN VITRO STUDY
Medium
Proteins
Cells
Agglomeration
Air-Liquid Interface Exposure
NPs
Basolateral side: Medium
Airway Surface Liquid (ASL)
Apical side: Lung cells
Semi-permeableMembrane
Exposure chamber
Transwell
Drawbacks • Not mimicking alveolar epithelial conditions in vivo.
• Interaction between NPs & media.
• Uncertainty of dosimetry.
• NP agglomeration/dispersion problems.
Conventional in vitro study: Need new approach
IN VITRO NANOTOXICOLOGY
• The objective of this study was to overcome the limitations of
conventional in vitro exposure of submerged lung cells to NPs for
NP toxicity assessment.
• We developed a dynamic in vitro exposure system (DIVES) capable
of generating and depositing airborne NPs directly onto lung cells at
an ALI (simulation of human pulmonary exposure to NPs).
Aerosol Inlet
Cell culture insert
Cells on membrane
Culture medium
Air o
ut
Air o
ut
Air
Cellular Responses
NANOPARTICLE IN VITRO EXPOSURE SYSTEM
Figure: A Schematic of a NP In Vitro Exposure System. This in vitro approach simulates particle deposition in the human lung more realistically than does submerged cell exposure(without an apical air interface), and it preserves the inherent properties of the particles.
(Vitrocell)
CELLULAR DOSIMETRY (ICP-MS)
Table: The mass concentration of Cu after exposure of A549 cells to Cu NPs at the ALI.
Adjusted Cu massin/on the cells
Adjusted Cu massin basal medium
Total air-delivered Cu mass concentration
(4.7 cm2) (18 mL) (Transwell) (µg NP/cm2)
---------- µg ± SE ----------
1.7 ± 0.1 2.9 ± 0.1 4.6 ± 0.1 1.0 ± 0.02
Cellular Dosimetry: large amount of Cu was dissolved and released to the
medium (62% of total mass) during continuous air-delivery of Cu NPs.
Digest A549 cells + Cu NPs
HCl + HNO3 at 95 °C (~ 4 h)• Measure deposited Cu by ICP-MS
• [Cells + Cu NPs] – [Cells]
• A549 cells (a human alveolar type-II-like cell line)• Type II alveolar epithelial cells: first lung cells to be exposed to inhaled NPs.
S.P DISEASE BURDEN
Most people have colonization of S.p in nasopharynx, sinuses, nasal cavities, and have no
symptoms.
Infants, elderly, and chronically ill at highest risk for pneumonia, and other diseases.
Pneumonia (and related diseases) result in more than 4 million deaths per year (mostly by
S.p).
Big picture question: What are the underlying mechanisms that make S.p pathogenic
and cause disease? What makes certain people/groups susceptible?
Member of the streptococcus family
Gram positive diplococci
NPs in Welding Fume
†Graczyk, H., et al., Characterization of Tungsten Inert Gas (TIG) Welding Fume Generated by Apprentice Welders. Annals of
Occupational Hygiene, 2016. 60(2): p. 205-219.
A recent welding fume characterization study at the
breathing zone across welders (n=20)† indicated 92%
of the particles were <100 nm, with 50% of the
particles <41 nm. Inhalation of these metal fume
NPs can result in particle deposition onto the alveolar
epithelial surface of the lungs, compromising the
respiratory and circulatory systems
Exposure to welding fumes increase the risk of S.p infections in welders.
Pneumonia was associated with a reported occupational exposure to metal fumes in
the previous year (OR = 1.96).
To date, the mechanism by which welding fumes increase susceptibility to S.p
infection is not well known.
Are the metal NPs in the welding fumes to blame?
S.P DISEASE BURDEN
Goal: to determine the relationship between metal NP exposure (e.g.: copper) and the
increased susceptibility of welders to infection by Streptococcus pneumoniae.
Do copper NPs promote cytotoxicity in human alveolar epithelial cells (A549 cells) to
enhance adhesion of S.p?
S.p adhesion to lung cells is increased after Cu NP exposure
The Power of Innovation!
DALHOUSIE HERC LABORATORY
Contact:Dr. Jong Sung Kim, Laboratory DirectorEmail: jskim@dal.caTel: 902-494-4225