Air Pollution in a Lifetime Health Perspective

Vision

Background

Study Plans

Work packages

Perspectives

References

The multidisciplinary research approach in AIRPOLIFE addressing health effects of air pollution throughout the life time.

Problem statement

Air pollution is probably the most important environmental factor affecting health in urban societies[1]. It is estimated that in Denmark particulate air pollution cost 3400 lives yearly corresponding to a reduction in mean life expectancy of ½-1 year[2;3]. The major outdoor source is vehicle emissions and additional sources may contribute to indoor air pollution. Man is exposed to air pollution throughout life. Intrauterine exposure may be particularly important for birth outcome and early life. Cumulated exposure may contribute to the development of e.g., cardiovascular and airway disease, and cancer. Peak exposures may trigger acute events such as asthma attacks and acute myocardial infarction or cardiac insufficiency, especially in high-risk groups.

The public health impact of air pollution is large because the majority of the population is exposed although the individual risk may be limited. Moreover, air pollutants interact with diet, life style, social factors and drug treatment, and genetic factors influence susceptibility. Whereas the potential for prevention and health promotion by reducing air pollution is large, the costs of unfocused interventions may be high. Risk communication is complicated because many sources and exposure determinants are integrated parts of our daily life and the individual risk is limited. An integrated research effort addressing the health aspects of air pollution throughout life is needed for optimum prevention based on proper risk assessment and management. 

Aims and vision

AIRPOLIFE will characterise health risks related to air pollution in a lifetime perspective with focus on generation of new knowledge for understanding mechanisms, risk assessment and prevention. Our working hypothesis involves a unifying mechanism of air pollution effects relating to inflammation and generation of reactive oxygen species (ROS). In a truly cross-disciplinary approach AIRPOLIFE will address exposure and systemic and target tissue effects of air pollutants, especially particles, in foetal life, childhood and adult populations. This will be accomplished by the following research tasks:

• Develop and apply new experimental models for characterisation of deposition, systemic translocation and health effects of particles in animals and humans.

• Characterise chemical and physical determinants of effects of particles in relation to sources.

• Develop and apply mathematical models, individual monitoring and biomarkers for assessment of exposure.

• Characterise relationships between exposure to air pollution and health outcomes using unique Danish population cohorts and disease registers.

• Study interactions between exposure to air pollution and genetic susceptibility, adaptive responses, diet, life style, social factors and drug treatment.

• Develop and apply biomarkers based on novel technologies, e.g. genomics and proteomics.

• Develop the study of cost-effectiveness of interventions.

It is the vision that the multiple involved groups, disciplines and approaches will create great synergy and provide research at the highest international level. AIRPOLIFE includes basic experimental research related to cancer, inflammation, nutrition and cardiovascular function, molecular genetics, epidemiology, biostatistics, clinical physiology, particle physics and chemistry, aerosol and atmospheric science, public health, environmental impact assessment etc. AIRPOLIFE will constitute a virtual centre of excellence with respect to particulate air pollution and will be an ideal forum for research training in this area. AIRPOLIFE will provide expert advice and communicate with stakeholders.

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Deliverables

AIRPOLIFE will deliver the following innovations required for targeted prevention of health effects of air pollution and evaluation of interventions:

• Provide understanding of the mechanisms and adaptive responses involved

• Characterisation of factors determining relevant exposure, deposition and effects

• Dose-response relationships of health effects of air pollutants

• Provide tools of exposure modelling, monitoring and socio-economic predictions

Background

Particle sources and characteristics. Ultrafine (<100nm) particulate matter (PM) from vehicle exhausts may be responsible for a large part of the health effects ascribed to air pollution in the urban environment[4;5]. Secondary particles, from SO₂ and NO_x contribute significantly to fine particles (PM_2.5)/PM₁₀ in urban air. PM_2.5 and in particular PM₁ and ultrafine PM penetrate easily to the indoor environment, although indoor sources and activities such as cooking, tobacco smoke, candles, stoves and materials contribute significantly to the indoor concentrations[6]. Additionally, microbial growth due to poor cleaning or humid/water-damaged buildings may play a major role in development of airway disease. The composition of particulate air-pollution is complex and naturally controlled by the predominant sources. The ultrafine soot particles at street-level mainly consist of elemental carbon associated with small nanoparticles of metal compounds (mainly Fe-, Ni-, and Zn-oxides) as well as different organic substances, e.g. PAHs[7;8]

Deposition and translocation of particles. Deposition of inhaled PM in the airways is determined by size, anatomy and respiration [9]. Coarse particles are mainly deposited in the upper airways. Particles smaller than 2.5-10 µm can deposit in the bronchia, from where they are removed by cilia movements with resulting gastrointestinal exposure. Fine and in particular ultrafine particles can reach the alveoli [10], where no cilia are present, and must be removed by dissolution, phagocytosis and interstitialisation. Systemic translocation of ultrafine particles has been demonstrated in experimental systems and suggested by deposition in humans[11-13]. The deposited fraction of particles is increased with decreasing size and deeper respiration, e.g. exercise[14] and in airway disease[15]. The smallest ultrafine particles (often defined as nano-particles) may also deposit higher in the respiratory system due to their high mobility. However, also other properties are important for the deposition of the particles and their adverse health effects, e.g. solubility in water, state (liquid/solid), shape and chemical composition.

Health effects of air pollution. Mortality related to cardiovascular and airway disease has been linked to PM_2.5/PM₁₀ in two American cohort studies and a number of time-series studies[5;16]. The dose-response estimates of mortality appear much higher in cohort studies, presumably also including accumulated effects, as compared to time–series studies, which rely on associations between peak levels of air toxics and daily counts of deaths and hospital admissions. Time-series-based associations have been shown for daily PM_2.5/PM₁₀ or NO₂ levels and exacerbation of asthma and COPD in panel studies[17], whereas the role of particles in development of asthma is unclear[18]. Nevertheless, airway symptoms in the first year of life have been associated with estimated PM_2.5 levels at the residence[19]. Exposure with conditioning effects may also occur during pregnancy as shown for smoking and suggested by relationships between air pollution exposure and birth weight[20]. It should be recognized that most studies have estimated exposure as PM_2.5/PM₁₀, which have many sources with potentially very different health effects. The few available time-series studies of ultrafine particles have suggested that the association with cardiopulmonary mortality and asthma symptoms in adults is at least as strong as with PM_2.5/PM₁₀ whereas ultrafine appeared to have limited effect in childhood asthma[21]. Even short term exposure to air pollution can affect alveolar permeability and possibly mucociliar transport[22].

In almost all studies exposure has been estimated at residences by assuming common background levels in large urban areas or by simple proxies such as proximity to streets. It is likely that assessment of individual exposure, including a historical perspective and focusing on different sources, toxic components and particle size fractions will improve the possibility of demonstrating associations, identify risk groups and provide much better estimates of dose-response relationships. Moreover, optimum control of confounders by the use of well-described cohorts is required. Interpretation relies on the understanding of deposition, translocation molecular mechanisms of action, which, and can be studied by means of experimental models and biomarkers.  

Mechanisms of action. The suspected common mechanisms of action of PM involve inflammation and oxidative stress, relevant for at least cardiovascular disease, cancer and airway disease. In addition PM contains PAH, which can form adducts to DNA after metabolic activation [23]. This is together with oxidative DNA damage responsible for mutagenic effects, which may lead to cancer with promotion through inflammation per se.

The capacity of PM for induction of inflammation and oxidative stress depends on the size, surface area and chemical composition, including soluble materials. Ultrafine PM induces more pulmonary inflammation than the same mass of fine particles in animal models[24]. PM collected in Copenhagen streets induces far more DNA damage in isolated DNA than diesel exhaust (DE)PM do per unit mass (unpublished). Yet, DEPM induces DNA damage, oxidative stress response, DNA repair genes and IL6 release in vitro and/or in vivo after inhalation[25]. A number of genes and mediators are central in oxidative stress accompanying inflammation and adaptive responses. These include the potent pro-inflammatory cytokine TNFa, the nuclear transcription factor NFkB and the nuclear response factor Nrf2 [26;27]. NO from inducible nitric oxide synthase (iNOS) and superoxide are responsible for nitrosative damage, including nitrotyrosine. The adaptation and defence systems involve upregulation of antioxidant enzymes, heat shock proteins, including HSP32-heme oxygenase-1 (HO-1), and DNA repair enzymes, as recently shown e.g. in the lungs of mice subjected to ionising radiation or inhalation of DEPM[25;28].

Extrapulmonary cancer related to air pollution is suggested by associations between occupational exposure to diesel exhaust and colon or liver cancer[29;30;30;31;31]. Inhaled particles are transported to the gastrointestinal tract and foodstuff may be polluted with vehicle emissions [32]. Moreover, rats fed with relatively low amounts of diesel particles had increased DNA damage from oxidative stress and PAH’s as well as upregulation of DNA repair enzymes in colon and liver cells[33;34]. More DNA damage was found in the lungs from dietary exposure to DEPM in rats than from inhalation or instillation in mice or Guinea pigs, suggesting important systemic effect[25;35]. Iron and inflammation from particles may play a role in colorectal cancer. E.g. variant hemochromatosis gene alleles appears to be associated with an increased risk, whereas Cox-2 inhibition lowers the risk[36;37]. Furthermore, interactions with dietary constituents such as fruit and vegetable constituents, fat and sugar, which per se is mutagenic in our models [38], are likely to occur.

The mechanism relating air pollution, ultrafine PM and cardiovascular disease is uncertain. Acute cardiovascular disease may be triggered by pulmonary release of cytokines and increased blood viscosity[39;40]. Induction of atherosclerosis per se through oxidative stress is suspected from mechanistic and epidemiological studies[41-44].[45]Oxidative modification of lipoproteins is known to affect atherogenicity. Indeed, oxidative stress in plasma lipids and proteins correlated with exposure to air pollution in Copenhagen[46;47]. Inflammation in the vessel wall with generation of oxidative and nitrosative stress central in cardiovascular disease[48]. Inducible nitric oxide synthase (iNOS) is required for development of atherosclerosis and plasma lipid oxidation in ApoE-/- mice[49;50].  Nanoparticles can penetrate into the circulation [10] and could cause inflammation, iNOS induction, oxidative stress, plaque formation/alteration/rupture and endothelial dysfunction, although this has yet to be demonstrated experimentally. Endothelial (dys)function is central in the manifestations of cardiovascular disease[51].

Gene-environment interactions. Health effects of environmental exposures are extensively modified by defence systems controlled by genes, many of which show genetic polymorphisms. In experimental models with knockout of such defence genes and by characterising subjects in environmental epidemiology studies it is much more likely that associations can be demonstrated and susceptible individuals characterised. A number of genetic polymorphisms and haplotypes related to enzymes metabolising foreign compounds, defence against oxidative stress and repairing DNA have been shown to be risk factors in many different cancers in case-control studies[52;53].

A number of newly detected polymorphisms, in particular in genes related to oxidative stress and inflammation, may have relevance for effects of air pollution, e.g. GSTO1 and catalase [54;55]. Genetic predisposition is important in CVD disease, particularly regarding lipid metabolism[56-58]. Recently, chemokine receptor (CCR2) genotype was shown to predict the risk of MI [59]. Polymorphisms in the pulmonary defence genes α1-antitrypsin gene and in cystic fibrosis related genes are related to risk of airway disease[60;61]. Similarly, polymorphisms in GST’s have been related to development of asthma in childhood, in particular with exposure for environmental tobacco smoke, as well as in adults[62;63]. Also a number of genes related to inflammation, β2-receptors, nitric oxide etc. are candidates of modulators of risk in airway disease. New SNPs in candidate genes can easily found in open databases guided by experimental results.

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Study Plans

The AIRPOLIFE research program is structured in a series of 4 integrated Work packages (WP) divided according to a lifetime health perspective. A series of experimental model set up and cohorts are used across these 4 WP’s. In addition, one WP will develop characterisation of particle exposure and one will study the socio-economic aspects across the health effects. The partners are:

1. University of Copenhagen, Institute of Public Health (UCIPH)

2. University of Copenhagen, Department of Pharmacology (UCDP)

3. National Institute of Occupational Health (NIOH)

4. National Environmental Research Institute, Dept. of Atmospheric Environment (NERIAE)

5. National Environmental Research Institute, Dept. of Policy Analysis (NERIPA)

6. Institute of Cancer Epidemiology, Danish Cancer Society (ICE)

7. University of Aarhus, Institute of Environmental and Occupational Medicine (AU)

8. Institute of Food Safety and Nutrition (IFN)

9. The PET Centre, Rigshospitalet (RHPET)

10. Department of Clinical Biochemistry, Section for Molecular Genetics, Rigshospitalet (RHMG)

11. The COPSAC study, Gentofte Hospital (COPSAC)

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Work packages

Exposure assessment

WP2-5 Integrated health effect studies

Experimental models and biomarkers

Cohorts, registers and methods applied in the epidemiological studies.

WP2. Foetal development and conditioning

WP3. Airway disease in childhood

WP4. Cardiovascular and airway disease  

WP5. Cancer

WP6. Socio-economic aspects of air pollution.

Dissemination of results

Program feasibility

Ethics

WP 1. Exposure assessment

AIRPOLIFE will develop large-scale assessment of individual exposure to specific air pollutants, including a historical perspective, required for innovation of air pollution epidemiology. The models will be applied in a series of health effect studies (WP2-5).  Although exposure assessment based on ambient concentrations at the residence is usually the only possible approach in large-scale epidemiological studies, indoor sources may be important contributors and risk groups may have different exposures, deposition and mucociliary function. Personal monitoring and biomarkers will be used to characterise individual exposure and identify hot spots and significant sources to air pollution with relevant biological effects. Moreover, such data are highly valuable for validation of exposure models used in epidemiological studies.

WP1A. Physical-chemical characterization of particulate air-pollution and exposure assessment.

The physical-chemical characterizationincludes measurements of particle size distributions, mass concentrations (PM₁, PM_2.5 and PM₁₀) as well as quantification of selected toxics and key-chemical compounds in the regional and urban background, and in streets and houses. Analyses include emission characteristics, chemical composition and toxicity of particles from, e.g., candlelight, gas stoves and wood burning in heat stoves. Particles in relevant size fractions will be sampled for detailed physical-chemical analysis and bioassays. The focus of the size-fraction analyses is exposure assessment of soot and specific elements e.g. V, Fe and Zn playing a major role on the induction of proinflammatory cytokines. Metals in ambient particles mainly occur in the 0.5 to 1.0 m-size fractions, but as much as 10-14% are seen in the ultrafine fraction[7;31]. We will also focus on content of organics, e.g. nitro-PAH, nitrobenzanthron, and oxy-PAH. Organic compounds in particles will be estimated from automated measurements of the their sum (time resolution 1 hour). Parts of the air pollution data are obtained from the Air Quality Monitoring Programme.

The exposure assessment is established using well-tested models combined with measurements obtained from the urban and regional air pollution monitoring network. Regional and urban air pollution data will be obtained from the urban background model UBM[64], and other models under the prognostic system THOR[65] as well as from monitoring data. The AirGIS system [66] will generate input data for air pollution calculations of nitrogen oxides, ozone, fine and ultrafine particles with the street pollution model OSPM[67]. These assessments are compared to a simple assessment of local exposure based on distance to busy streets inferred from GIS operation on geo-coded addresses and digital maps. The AirGIS system will be further developed to account for the changes in urban building structure over time using information from the building and housing registry. Historical records for regional and urban background levels of fine and ultrafine particle concentrations are derived together with historical records of emissions factors. As the composition of the car fleet has changed considerably over time, data are collected in order to improve emission estimates back to the 1960’ties. For the regional background, emission inventories are available back to the 1980’ties. Best available estimates are applied for going back to the 1960’ties. PI Ole Hertel; key researchers: Keld Alstrup Jensen^NIOH, PhD students and Post.doc.s, Finn Palmgren^NERIAE.

WP1B. Personal monitoring.AIRPOLIFE will study individual exposure by monitoring and biomarkers in risk groups including elderly subjects, young subjects exercising in polluted areas, and pregnant women. Personal samplers of ultrafine particles and PM_2.5 inlet are fitted with GPS devices into backpacks[46;68] to be carried or placed close to the study subject. Biomarkers include DNA damage and repair enzymes as well as oxidative stress response genes, such as HO-1 and inflammation by means of cytokine arrays. The effect of exposure during exercise on alveolar and mucociliar function will be studied by 99mTc-labeled diethylenetriaminepentaacetic acid and albumin and by spirometry[69]. The influence of size on deposition of real life street ultrafine particles will be determined in subjects with and without preceding exercise. The size distribution of particles in exhaled air and air inhaled at a busy street intersection will be measured. PI Steffen Loft^UCIPH; key researchers: PhD student^UCIPH, Peter Vinzentz^UCIPH, Herman Autrup^AU, Finn Palmgren^NERIAE, Jan Mortensen^RHPET.

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WP2-5  Integrated health effect studies

Experimental models and biomarkers

Experimental models in AIRPOLIFE include susceptibility by knockout and reporter gene transgenics with exposure by newly developed nose-only inhalation in mice, intratracheal instillation, intravenously, ingestion or gavage. MutaMice and BigBlue mice and rats harbour a cII mutation reporter gene. Activation of NFkB in the any target tissue is detected by using luminescence in mice transgenic for NFkB-enhancers in front of luciferase[15]. Susceptibility models, including knockout of OGG1, CSB, TNF-a, ApoE-, iNOS-, nrf2, - are already bred in our laboratories or available commercially or from collaborators. Double knockouts or knockouts with reporter genes are available for our studies from collaborators, e.g.,  (OGG1-/--CSB-/-; OGG1-/- Bigblue). In vitro models include A549 human lung epithelial cells MutaMouse lung epithelial cell expressing the cII reporter gene and lymphocytes.

Assays for oxidative damage to DNA, lipids and proteins as well as PAH adducts are functional[34]. Inflammatory and adaptive responses are studied at the mRNA and protein levels. Assays based on real-time PCR and immunobased assays of key genes of DNA repair enzymes (e.g. OGG1 and ERCC1), cytokines (e.g. a series of IL’s, NFkB) and oxidative stress response (e.g. HO-1) have been developed[25;28] and will be supplemented guided by whole genome cDNA and selected protein (e.g. cytokine-growth factors) arrays experiments. Immunohistochemical and in situ hybridisation assays will be used to verify cellular localization of damage and gene responses (initially nitrotyrosine, 8-oxodG, PAH-adducts, iNOS, HO-1). We will further implement assays for activity of the related response enzymes, e.g. OGG1, HO-1 and GPx activity. Endothelial function is studied in mouse aorta in myographs by the response to acethylcholine after precontraction with prostaglandin.

Cohorts, registers and methods applied in the epidemiological studies.

High quality Danish cohort data and state of the art individual exposure data from DMU enable the use of the variety of the latest statistical methods in studying the relationship between air pollution exposure and various health outcomes.

Cohorts. Between 1993 and 1997, the Diet, Cancer and Health study (DCH) enrolled 57,053 persons between age 50 and 64 and with no previous cancer diagnosis. At baseline, data were collected for each individual on diet, smoking and passive smoking history, medication, body composition, reproductive and many other environmental, occupational and lifestyle factors. Several other cohorts studies conducted in the Greater Copenhagen area between 1964 and 1992 have been united in the Copenhagen Centre for Prospective Population Studies to obtain standardized data for the altogether 31,194 participants on smoking habits, alcohol consumption, BMI, physical activity and other life style factors. Recently, the enrolment of 100,000 pregnant women into The Danish National Birth Cohort (DNBC) was completed with data on medical conditions, life style, dietary patterns, smoking and environmental exposures recorded several times during the pregnancy. The COpenhagen Prospective Study on Atopy in Childhood (COPSAC) has established a cohort of 400 children of asthmatic mothers and a large number of exposures in utero and during childhood have been registered. These include repeated measurements of (PM_2.5), NO₂, NO_x, formaldehyd, acetaldehyd and acrolein in the bedroom of each child. Health outcomes are registered by use of clinical examinations and self-administrated diary notes.

Registers. Since the Central Population Registry (CPR) began on April 1, 1968, each resident in Denmark has been assigned a unique 10-digit personal identification number (PIN). The CPR can provide information on dates of death, emigration or disappearance as well as residential addresses presently and back to 1972. Moreover, the PIN can be used to link up a study population with a wide range of population-based health registers, e.g. Cancer Register, Register of Causes of Death, and the National Hospital Register.

Design and methods. Mostly, the epidemiological studies in AIRPOLIFE will use traditional cohort designs or the case-cohort design, analysing associations between air pollution and health outcomes by Cox-proportional hazard models, but also time-series analyses relating day-to-day variation in air pollution to similar variation in health outcomes will be used. Moreover, the relatively new case-crossover design for acute effects will be compared with the traditional nested case-control design. The risk estimates will be adjusted for the effect of potential confounders like smoking, passive smoking, occupational exposures and diet. 

WP2. Foetal development and conditioning

Maternal exposure to air pollution may adversely affect foetal development as indicated by the observed negative relationships between heavy exposure of pregnant women to air pollution in the Czech Republic and neonatal birthweight[70]. Other studies have confirmed the transport of PAH from maternal blood to foetal blood[71]. Eexperimental exposure to DEPM during gestation affects fetal development[72] and the developing nervous system may be particularly sensitive. The hypotheses that ultrafine particles can translocate from exposure trough airways or ingestion to the foetus and that air pollution affect development are addressed by 5 different approaches

WP2A. Transport of particles cross the human blood-placenta barrier will be studied by use of the isolated perfused human placenta system developed in Finland[73] and established at UCIPH as part of the European Network on children's susceptibility and exposure to environmental genotoxicants. Passage of PM (initially of colloidal gold, later genuine) through the placenta barrier will be detected by electron microscopy. DNA damage and gene activation induced ex vivo will be studied in maternal and cord blood by postlabelling and immunochemistry and in situ hybridization. PI: Lisbeth Knudsen^UCIPH; key researchers: PhD student^UCIPH, Steffen Loft^UCIPH.

WP2B. Intrauterine exposure models. Nervous system function will be assessed in offspring of mice exposed to DEPM during gestation, by use of the Morris water maze, a sensitive method for detecting changes in learning and memory functions. Other endpoints are length of gestation, litter size, birth weight, organ weights, sexual maturation etc. NFkB activation in foetal tissue will be studied in reporter mice. PI: Håkan Wallin^NIOH; key researchers: Karin Sørig^NIOH.

WP2C Target organ exposure in humans will be assessed in a biomarker study with personal monitoring. 50 healthy women scheduled for delivery by elective ceaserean section will be monitored in and around their home with respect to level of fine and ultrafine particles during two days before delivery. Placenta and cord and maternal blood will be collected for measurement of biomarkers of exposure and effect, including DNA damage (bulky adducts, strand breaks and base oxidations), expression of DNA repair inflammation and oxidative stress response genes, NFkB nuclear translocation and possible others). Subjects will be genotyped for the most common polymorphic genes involved in foreign compound defence. PI Steffen Loft^UCIPH; key researchers: PhD student^UCIPH, Peter Vinzentz^UCIPH, Lars Dragsted^IFN, Herman Autrup^AU, Ole Hertel^NERIAE.

WP2D Early exposure and early childhood airway disease.A subcohort of approximately 15000 births from the DNBC is under study for relationships between exposure to air pollution during pregnancy and low birth weight, perinatal mortality and complications. The study will be extended such that ambient air pollution at the residence at the time of birth will be related to data on airway disease and a doctoral diagnosis of asthma obtained by interviews of the mothers when the children were 6 and 18 months old. At 18 months age, 16% of the children had a doctoral diagnosis of asthma. PI: Ole Raaschou-Nielsen^ICE; key researcher: Anne-Marie Nyboe Andersen^UCIPH, Sjurdur F. Olsen^DNBC.

WP3. Airway disease in childhood

Asthma is a common increasing disease among children with a lifetime prevalence of 15-20 % in young Danish adults[74]. Exposure to air pollution is a suspected cause of childhood asthma and is known to aggravate established asthma. Asthma is an inflammatory disease and the capacity of air pollution to induce inflammation is suspected to play an important role. It is likely that polymorphic gene products involved in inflammation and defence against oxidative stress and foreign compounds will be effect modifiers in relation to air pollution and asthma.

WP3A. Indoor and outdoor air pollution and childhood asthma. Associations between repeatedly measured indoor airway irritants, PM_2.5, NO₂, NO_x, formaldehyd, acetaldehyd and acrolein, model assessed outdoor concentrations and asthma symptoms registered in the family-administered diaries of the children of the COPSAC study will be analysed and reported. PI: Ole Raaschou-Nielsen; key researhcers: Hans Bisgaard^COPSAC, Ole Hertel^NERIAE

WP3B. Endotoxins and inflammatory potential of particles, and childhood asthma.  In the COPSAC cohort, PM_2.5 were collected and stored on filters 3 times from the bedroom of each child during the first years of life. We will study the risk of asthma in relation to the filter material and 1) the concentration of endotoxins and transition metals, 2) the inflammatory potential measured in vitro by cytokine release (IL8) in A549 cells and/or 3) the potential to induce ICAM-1 (a surface molecule upregulated in asthmatics) measured in vitro in A549 cells. PI: Ole Raaschou-Nielsen^ICE ; key researchers: Post.doc., Hans Bisgaard^COPSAC, Keld Alstrup Jensen^NIOH.

WP3C. Gene-environment interactions in childhood asthma. In the COPSAC study mentioned above, a limited set polymorphic genes involved in foreign compound metabolism (GSTs) are currently being assayed. This set will be expanded by genes involved in the inflammatory response, and defence against oxidative stress, (e.g. iNOS, eNOS, MnSOD, CF-genes). Genes showing promising effect modifying capabilities will then be tested in a larger sample in the DNBC cohort
PI: Steffen Loft; key researchers: PhD student, post.doc., Anne Tybjærg-Hansen^RHMG,Hans Bisgaard^COPSAC.


WP4. Cardiovascular and airway disease

AIRPOLIFE will address relationships between particulate air pollution and cardiovascular disease in experimental models as well as in epidemiological studies with focus on mechanisms involving inflammation and oxidative stress.

WP4A. CVD models. Mice will be exposed to particles by injection, inhalation/instillation and/or orally. Activation of NFkB in target organs, including arteries, is detected luciferase reporter. In ApoE-/- mice plaque progression, cell infiltration, iNOS expression, nitration of tyrosine, cytokines, growth factors, adhesion molecules, oxidative stress and response genes (e.g. HO1), DNA damage and NFkB nuclear translocation are measured in the vessel wall and/or plasma. Full gene expression patterns are assessed by microarrays in selected exposures. Further mechanistic studied may be conducted in crossbred mice. Endothelial function is assessed in aorta rings mounted in myograph and L-NMMA is used to manipulate NO production. PI Peter Møller^UCIPH; key researchers: PhD student, Steffen Loft^UCIPH, Thomas Jonassen^UCDP, Ole Amtorp^UCDP, Håkan Wallin^NIOH

WP4B. Cause specific mortality and hospital admission in relation to air pollution.  Relationship between cumulated exposure to air pollution and cause specific mortality will be assessed in the 57.053 members of the DCH cohort.  Historical addresses for all cohort members will be traced back to 1972 and exposure at the more than 100.000 historical addresses will be assessed by AIRGIS and distance to densely trafficked roads. Outcomes will be identified in the Register of Causes of Death, Cancer Registry and Hospital Register. Interactions with diet and other life-style factors will be investigated. PI: Ole Raaschou-Nielsen^ICE, keypersons: Ole Hertel^NERIAE, Post.docs.

WP4C. Time-series analysis of CVD and respiratory disease. Daily hospital admissions and cause specific mortality is related to air pollution levels in periods up to 40 days preceding the event in all major Danish cities with air quality monitoring programs. Changes over time can be related to interventions affecting e.g. traffic generated air pollution, such as driving restrictions, fuel changes, filters for diesel vehicles. PI: Steffen Loft^UCIPH, key researchers: Finn Palmgren^NERIAE, , PhD student, Thomas Scheike^UCIPH.

WP4D. Gene-environment interactions, oxidative stress and DNA damage in acute CVD and airway disease. In an ongoing time-series study of admissions and deaths of CVD and airway disease (totally 16000) in subjects from Copenhagen population studies with a population of 70000 at risk during 1999-2000 are identified. In case-cross-over and nested case-control design the impact of air pollution assessed by AIRGIS in periods prior to admission is studied. AIRPOLIFE will assess effect modification with respect to polymorphic genes involved in lipid metabolism, oxidative stress and inflammation (e.g. NQO1, iNOS, MPO, catalase, CCR2, α1-antitrypsin) and markers of exposure in terms of protein and lipid oxidation. PI: Steffen Loft^UCIPH; key researchers: Anne Tybjærg-Hansen^RHMG, PhD student, post.doc., Lars Dragsted^IFN, Ole Raaschou-Nielsen^ICE.

WP5. Cancer

Particulate air pollution is suspected of increasing the risk for cancer in lungs, colon and possibly other organs. Moreover, enzymes involved in inflammation, defence against oxidative stress, PAH’s and DNA damage may modify this risk. AIROLIFE will address these hypothesis and study interactions with dietary constituents such as fruit and vegetable constituents, fat and sugar, experimentally and in cohort studies.

WP5A. Experimental study of cancer mechanisms. Mice and rats will be exposed to DEPM and ultrafine model compounds by nose-only inhalation, through the diet and single gavage. Activation of NFkB is detected in lung, liver and colon. TNA-a-/- mice are used to study the role of inflammation in DNA damage, showing surprising enhanced susceptibility initially. Importance of DNA repair mechanisms are studied in OGG1-/- and CSB-/- (required for transcription coupled DNA repair) double knockouts and  OGG1-/-BigBlue MutaMouse. The Nrf2 based adaptive response is addressed in Nrf2-/- mice. Lungs, liver, colon mucosa, blood and bronchioalveolar lavage fluid are isolated and analysed for oxidative stress markers, DNA damage, nitrotyrosine, oxidative stress response (e.g. HO-1), DNA repair, cytokine and inflammation (iNOS, Cox2) gene expression, DNA repair protein and activity. Further gene regulation aspects will be directed by cDNA and protein arrays in strategic experiments. The mutagenicity of ultrafine and other size fractions of PM from Copenhagen and components of urban air pollution will be tested in Mutamouse lung epithelial cell line by the Ames assay and in vivo in mice expressing the cII reporter gene for selected materials. Interactions with diet in terms of fruit and vegetable products, sugars and fat will be performed according to the initial results. PI: Håkan Wallin^NIOH, PhD student, Herman Autrup^AU, Steffen Loft^UCIPH, Peter Møller^UCIPH  

WP5B. Case-cohort study of colorectal cancer. 500 colorectal cancer cases developed among the members of the DCH cohort will be identified by linkage to the Cancer Registry, and a sub-cohort of a similar size will be selected as comparison group. Exposure at the historical addresses will be assessed by AIRGIS. The association between exposure and colorectal cancer and possible effect modification by diet, polymorphic DNA repair (e.g. XPD, XPC, RAI, ASE, ERCC1, OGG1) and biotransformation (e.g. NAT2) genes will be estimated. PI: Ole Raaschou-Nielsen^ICE; key researchers: Ulla Vogel^NIOH, PhD student, Anne Tjønneland^ICE, Bjørn Nexø^{NIOHassociate}, Herman Autrup^AU.

WP5C. Case-cohort study of lung cancer.  A similar ongoing study include 500 lung cancer cases from the DCH and 500 individuals as the comparison group addressing the relationship between cancer risk and genotype and gene expression of DNA repair enzymes. The study will be extended to include assessment of exposure to air pollution at historical addresses, polymorphic genes involved in metabolism of xenobiotics, PAH adducts analysed in lymphocyte DNA and oxidative DNA damage products in urine. PI: Ulla Vogel^NIOH; key researchers:  Post.doc., Ole Raaschou-Nielsen^ICE, Bjørn Nexø^{NIOHassociate}, Herman Autrup^AU, Steffen Loft^UCIPH.

WP6. Socio-economic aspects of air pollution.

The impact of air pollution on public health is profound, and its socio-economic implications are perceived to be significant. Costings of mortality and morbidity impacts have been undertaken in various EU RTD projects, but the methodological rigour needs considerable improvement. The linkage to medical research results remains sketchy and must be developed.

WP 6.0 By accounting for the imposed health costs, marginal benefits of air pollution control can be assessed. This accounting needs to rely on the multiple-pathway approach and must link exposure with dose-response and with monetary estimates of morbidity and mortality costs. Previous dose-response functions relate to a set of classical air pollution related diseases; more health impacts have been identified in recent years, and the palette of dose-response functions need to be broadened, this should be done in close cooperation with medical expertise, hence synthesizing the knowledge base. The implications of averaging exposure values, as opposed to considering peak value exposure have been ignored in previous economic analysis and needs more careful analysis. Non-market costs relating to loss of spare time and well-being need to be quantified and valued. Monetization of both acute and chronic morbidity effects require sophistication and application of the WHO lifetime table method to Danish age cohorts. An original contingent valuation study will be carried out in order to reveal the preferences for reductions in mortality related specifically to air pollution exposure; a collective and involuntary risk type differing in fundamental ways from the individual risks accepted in the traffic sector. PI: Mikael Skou Andersen^NERIPA, PhD student.

Dissemination of results

AIRPOLIFE will form a virtual centre of excellence in order to facilitate information exchange and coordinate collection of knowledge on air pollution. Results will be disseminated through international peer reviewed publications, international conferences and international seminars organized by AIRPOLIFE twice per year. The advisory board will participate at least in one of these and evaluate progress. AIRPOLIFE will provide stakeholders, including regulatory and health authorities, NGO’s and others with knowledge and advice through publication of a newsletter and in relevant journals, seminars and a formal conference every year. Communication of risk will be highlighted. AIRPOLIFE will participate in information exchange and results dissemination in AIRNET (airnet.iras.uu.nl) and APHEIS (www.apheis.net), the two relevant European forums. A website will be used for internal and external communication.

Program feasibility

AIRPOLIFE is well suited within the strategies and framework of the research partner organisations as documented. No heavy equipment is required for the research program. Many of the partners have previously collaborated successfully in research large programs, e.g. the Strategic Research Programs and Health Promotion and Prevention from the Research Councils.  

Ethics

Important ethical issues relating to information of study persons, consent, banking of biological material and data protection will be taken care of in accordance with the current ethical guidelines by submitting the protocols and the information materials and questionnaires to the local ethical committee for approval. The program implies genetic testing of children and families, which will be handled according to current guidelines and from recommendation developed within the European network on children's susceptibility and exposure to environmental genotoxicants. All animal studies will be justified and follow the guidelines developed by the authorities as will be described in the application for approval. 

 Perspectives

AIRPOLIFE focus on providing knowledge for prevention in relation to the quantitatively most important environmental factor affecting human health at present. AIRPOLIFE will provide understanding of the relevant exposures and involved mechanisms as well as interactions with life style factors. This is mandatory for focused interventions, forcing new technology and their evaluation. The effects of interventions can be estimated and evaluated by the tools of modelling, monitoring, biomarkers, experimental models and economic predictions developed in the program. The tools can be of immense value in future international preventive effort. The partners have established collaborations or intense contact with relevant groups in third world urban centres with severe air pollution problems. The developed mechanistic tools are also highly relevant for study of potential adverse effects of nanotechnology, which may pose serious toxic problems[75]. 

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