Past Research Projects

2018 Consortium Projects

California Consortium on Thirdhand Smoke Exposure and Health Effects

Neal Benowitz, MD
University of California San Francisco

The term thirdhand tobacco smoke (THS) refers to tobacco smoke residues in indoor environments that remain, react and/or re-emit from materials and/or re-suspend from surfaces (dust). Over the last decade the concept that THS is a distinct harmful entity has emerged, as well as concern about its health risks for adults and especially children.

For the last 6 years, the Consortium has been supported by TRDRP to build a strong science base for understanding the health effects of exposure to THS. The Consortium now seeks funding for three more years, with an emphasis on carrying out laboratory and field studies that will help in translating THS research findings so that they are useful and accessible to health practitioners, public health organizations, advocacy groups, and the public. We also intend to increase dissemination of our THS research, supporting policy modeling and development.

We propose to extend our basic science research to identify biomarkers (tracers of harm inside the body) that will be useful in studies of human health effects, and examine factors associated with developmental and genetic vulnerability to THS-induced injury. We will investigate mechanisms of THS-induced adverse effects in cell models and in animal studies to further understanding the potential risks and pathways for THS damage.

We propose more detailed studies of human exposure and effects of THS, including applying biomarkers that were developed in preclinical studies, with a particular focus on exposure by contact through skin and uptake of THS in children. To answer the need for diagnostic tools and solutions to THS contamination, we propose to develop standard methods for assessing THS contamination in various environments, and test the effects of various remediation methods on THS constituents.

We propose to disseminate our research widely, and engage with public and private stakeholders to develop, implement and promote tobacco control policies that will reduce the harmful impacts of THS on public health. Cost-effectiveness of prevention, remediation and treatment strategies will also be considered.

These efforts will inform new basic as well as applied science and, with involvement of the public, may nurture widespread understanding of risks related to thirdhand smoke contamination and exposures.

Genetic Susceptibility to Thirdhand Smoke Effects and Biomarker Studies

Bo Hang, MD, PhD
Lawrence Berkeley National Laboratory

Thirdhand smoke (THS) has gained public attention in recent years but its health impact is largely unknown. During the last six years of research within the Consortium, we have shown that exposure to THS or its specific chemical constituent NNA causes damage to cellular genetic material (DNA) with covalent binding, changes in cellular metabolites and tumor-forming properties; exposure at early age in mice resulted in significant developmental, immunological abnormalities and lung cancer formation. The goal of this sub-project is to study genetic factors for individual susceptibility to THS-induced effects, especially tumor development, and to develop biomarkers in animal studies and human samples, with an emphasis on carrying out laboratory studies that will help in translating findings to human practice and the public. We hypothesize that (I) genetic susceptibility influences disease risk of THS exposure; (2) the NNA-induced deoxyguanosine (dG)-binding adduct is a biomarker (a tracer of harm inside the body) for THS exposure and the adduct level is associated with disease risk. We will employ an integrative systems biology approach for this project by using the generally diverse Collaborative Cross (CC) mouse resource that recapitulates the genetic diversity of the human population. In Aim 1 we will first investigate the tumor incidence across 20 THS-exposed CC strains and then perform genetic analysis to identify individual genetic susceptibility to these biological and health effects of THS exposure. Translational implications include (1) developing useful animal models for predicting health risk of THS exposure in human population; (2) using genome-wide association analysis together with RNA-sequencing, we will identify gene biomarkers for genetic susceptibility to THS effects, and find related human homologs of these mouse genes that can be used to predict and study human disease risk induced by THS; and (3) identify molecular mechanism for THS-induced disease for ultimately developing strategies for disease prevention and treatment.  In Aim 2, we will develop a quantitative LC-MS/MS method for the validation of the NNA adduct 1,N2-NNA-dG as a specific biomarker for THS exposure and indicator of cancer risk in both animals and humans. We will determine whether iso-NNAC is an NNA metabolite in vitro with human liver homogenates. For biomarkers we will perform proof-of-concept studies using available human specimen. Such translational studies will provide novel evidence of THS impact on cancer development in general population and how individual genetics affect vulnerability, and to use both basic science and translational findings to reduce the THS impact in exposed humans and to support policy modeling and development.

Thirdhand Smoke Chemistry: Exposure assessment, detection tools and remediation

Hugo Destaillats, PhD
Lawrence Berkeley National Laboratory

Our group has contributed to the development of the Thirdhand Smoke (THS) Consortium by describing indoor chemical processes that dominate exposures to THS. We have studied the critical role of indoor reactions of THS constituents with oxidants and nitrosating agents commonly found indoors. The identification and quantification of inhalable THS constituents enabled the assessment of their health impacts. Sampling and analytical methods were developed to expand the scope of THS constituents that can be measured. More recently, we initiated a study to evaluate the efficiency and potential risks associated with releasing high levels of ozone as a THS remediation approach. We have also collaborated with other Consortium partners in the evaluation of THS biological effects. On this new phase of the Consortium, we intend to extend our research on these areas by:

Aim 1: Quantify intake of carcinogenic tobacco-specific nitrosamines (TSNAs) via the main exposure routes (dermal, dust ingestion, inhalation), to better implement protective measures and THS remediation strategies. Our approach will involve a combination of experimental methods to assess dermal uptake, and modeling incorporating recently published information on TSNA levels in THS.

Aim 2: Develop a THS diagnostic tool for untrained non-professional users, based on measurement of airborne nicotine. Accurately quantifying very low levels of nicotine in indoor air is a useful tool to assess THS contamination, because it integrates emissions from multiple sources. Samplers will trap nicotine on a bisulfate-coated filter connected to a miniature battery-operated pump and flow sensor with continuous logging capabilities. The method will be validated in the lab, and deployed alongside other THS diagnostic tools in field measurements led by SDSU. We intend to increase our ability to detect trace levels of nicotine by 1-2 orders of magnitude with respect to the current practice using bisulfate-coated filters.

Aim 3: Contribute to Consortium-wide efforts to assess remediation strategies by evaluating the efficacy of carpet replacement in THS-contaminated homes that are being cleaned and conditioned for rent or sale. Air and surface samples will be measured before and after a new carpet is installed, to quantify the impact of source elimination. The THS-contaminated carpet and padding materials will be installed at LBNL’s room-sized chamber and used in ozonation experiments. An ozone generator commonly used in homes by companies providing cleaning and restoration services will be used to assess short- and long-term effects, including formation of airborne and surface-bound oxidation byproducts.

Human Exposure, Toxicology and Biomarkers Analytical Chemistry Laboratory

Peyton Jacob, PhD
University of California San Francisco

This subproject is part of a consortium of California researchers who are seeking to understand the risks of exposure to aged secondhand smoke, which has been termed “thirdhand smoke” (THS). Past studies have shown that secondhand smoke becomes more toxic as it ages. It is particularly concerning that levels of potent cancer-causing substances (carcinogens) called tobacco-specific nitrosamines (TSNAs) increase as smoke ages. Studies carried out by this consortium have identified numerous toxic substances in THS, and that many of them persist in homes and other places for long periods after smoking has ceased.

This third phase of the Consortium will focus on studies that are “translational” in nature, using the information from previous basic science studies, and applying this knowledge to studies that relate more directly to human health issues, and to remediation (detoxification) of THS contaminated homes, automobiles, and other places contaminated by THS.

The researchers in this subproject are chemists who have been studying human exposure to toxic substances in tobacco smoke for many years. For this subproject they will continue to develop methods for measuring human exposure to toxic substances in THS, needed to augment those developed in the past. To assist in achieving the goals of the Consortium, they will analyze samples from studies carried out by researchers in other subprojects that will assess the toxicity of THS, and analyze samples from other subprojects to measure human exposure in real-world settings.

These studies will expand our knowledge about the chemical nature of THS and its impact on public health. It is expected that the research carried out by this consortium will be useful in developing policies for protecting people from THS exposure, and will help in developing methods for detoxifying places contaminated by THS.

Translating Mouse Exposure Studies into Human Health Effects

Manuela Martins-Green PhD
University of California Riverside

Third Hand Smoke (THS) has been identified as a potential danger to human health and society, however, more work still needs to be done to make the public aware of this stealth toxicant. THS consists of a complex mixture of toxins that are found on the surfaces of homes, cars or public places where cigarettes have been smoked. These toxins land and remain on surfaces, can persist for months to years and cannot be removed with general household cleaning products. Furthermore, these toxins can be re-emitted into the air, and can react with other chemicals in the ambient environment to result in their conversion into carcinogens. Many of the toxins can then enter the body, because they are easily absorbed through the skin, and can be inhaled and ingested. This is particularly detrimental to children and infants living in homes of smokers because they crawl on the absorption through the skin and in breathing the volatile chemicals released from the surfaces.

Our data show that THS-exposed mice have alterations in multiple organ systems and excrete levels of a tobacco-specific carcinogens that are similar to those found in children exposed to SHS (and consequently to THS). Specifically, (1) In liver, causes increased lipid levels causing fatty liver disease, a precursor to cirrhosis and cancer and a contributor to heart disease. (2) In lung, it stimulates fibrosis and high levels of inflammatory molecules, with implications for diseases such as asthma. (3) In wounded skin, it causes problems found in the poor healing of surgical incisions in human smokers. (4) In behavior, THS-exposed mice become hyperactive. Also, (5) we determined the minimum exposure time and dose of THS needed to detect differences in specific biological markers of harm to health; (6) we have shown that THS-exposure affects insulin metabolism; (7) we also found that exposure to THS in utero and during lactation results in smaller litters and smaller mice.

The goals of this research project are: Project 1 Determine dose-dependent effects of THS exposure in mice at different ages; Project 2, identify whether effects of THS-exposure damage mitochondrial DNA so that future studies can be performed to determine whether that damage is passed on to the next generation. Mitochondria are critical parts of cells because they produce the energy the cells use to function; Project 3 perform Proof-of-Concept studies in children using Biological Markers of Harm induced by THS exposure that we identified in mouse tissues, serum and urine. We will also measure levels of cotinine in the saliva of the parents and their children measured in the Dr.’s office during the visit to provide in real-time feedback to the parent/caregiver so that smoke cessation measures could be put in place.

This is the first-time research will be done to determine whether the effects of THS dose are age dependent, to determine if younger ages more susceptible to the exposure. We will also show if the parts of the cell that are critical to proper function are affected by THS exposure. We will detect cotinine levels in the saliva for doctors to advise parents in real-time that their children are being exposed to THS and are being affected by their smoking.

The impact of these studies is three-fold. We will: (a) determine whether younger ages are more susceptible to different doses of THS-exposure; (b) determine whether THS exposure leads to malfunction of cells in the body that can lead to disease; (c) provide toxicological evidence to design clinical trials that can be performed in humans; (d) help in the design of policies that mitigate exposure of infants, children, the elderly and workers in places where smoking is allowed.

Controlled Human Exposure and Thirdhand Smoke Generation Core

Suzaynn Schick, PhD
University of California San Francisco

The goals of the Controlled Human Exposure and THS Generation Core are to identify the health effects of acute exposures to THS, to test strategies for identifying members of the public who are exposed to THS and to continue to provide other researchers with high quality THS samples to study. In the last funding cycle, we found that wearing cotton clothing that had been exposed to as much smoke as would be in a heavy smokers’ house caused large increases in the metabolites of nicotine (cotinine) and the tobacco-specific carcinogen NNK (NNAL). These increases were 2-10 times higher than we have seen in previous respiratory exposure experiments. The skin is an important exposure route for THS exposure because the majority of THS in the indoor environment is sorbed to surfaces.

Consortium research has shown that THS exposure causes numerous disorders and diseases in laboratory animals and in vitro systems, but we have not yet demonstrated a connection between THS exposure and health problems in humans. In our new controlled human exposure experiments, we will compare inhalational exposure to SHS, inhalational exposure to THS and dermal exposure to THS, testing flow-mediated dilation, reactive hyperemia, peripheral arterial tonometry, and heart rate variability in healthy nonsmokers. We will compare the effects of dermal and respiratory exposure to THS on uptake and metabolism of nicotine, NNK, volatile organic compounds and polycyclic aromatic hydrocarbons. We will measure epidermal retention of THS and effects of exposure on gene activation and the protein synthesis in epidermal, buccal and nasal cells. Our goal is to identify any changes in bodily function that are caused by acute THS exposure.
Though it is important to identify the effects of acute THS exposure, chronic exposures are more likely to cause harm. We will test two different methods to identify people who are exposed to THS. First, in our studies we always find people with higher baseline levels of THS biomarkers. We will ask participants to collect dust and wipe samples in their homes for testing and to join a registry for future research. To see how children are exposed, we will advertise for parents who think they may be living with THS. If wipe and dust samples show their homes are contaminated, we will give their children custom-made teddy bears to play with for one month. The bears will be made of plush cloth to absorb THS. We will test the cloth, learn more about the children’s exposure and invite the families to join a registry for future research and remediation.
Finally, we will continue to generate THS in the laboratory and share THS samples and biological samples from our study participants with other laboratories that study THS. By identifying the rapid health effects of THS, reaching out to people who live with THS and sharing research materials, we will improve understanding of the nature and biological effects of THS.

Thirdhand Smoke Dissemination, Outreach, and Resource Center

Georg Matt, PhD
San Diego State University

Thirdhand smoke (THS) refers to the residue that is left behind in dust and on surfaces (such as furniture, carpet, and walls) long after someone smoked tobacco in an enclosed space. THS is made up of a mixture of hundreds of toxic chemicals that have been found to be harmful to humans. Over the past 15 years, researchers from the California THS Research Consortium and elsewhere have found THS in many different indoor spaces where tobacco had been used. This includes apartments, homes, cars, hotels, casinos, and hospitals. THS has been detected years after cigarettes had been smoked regularly in a home. Thus, it is important to raise awareness about the long-term consequences of smoking indoors and how it may affect nonsmokers. This project will inform Californians about where THS can found, how one can get exposed to it, who is at risk of harmful effects, and how to prevent THS. This includes making people aware that renting or buying a home where people smoked could lead to chronic THS exposure, helping people test their homes for THS, and encouraging everybody to put into action strong smoking bans. The goal of this project is to increase public awareness among California residents, communities, tobacco control groups (such as American Lung Association), health professional (such as health departments, pediatricians), and relevant business owners (such as apartment managers, real estate agents, rental car agencies, hotel managers). To achieve this goal, we will establish a THS Resource Center to connect the THS researchers and the community. This will include a website to share THS information and outcomes from Consortium supported research with California residents, communities, tobacco groups, and business owners (Aim 1). To increase awareness among California residents, we will post educational messages on Facebook and study how well they help people become more aware of THS (Aim 2). We will share what we know about THS with community groups, health professional, and relevant business owners through a series of online workshops, and evaluate how well the workshops help to inform groups about THS risk (Aim 3). Finally, we will work with community groups, health professional, and relevant business owners on creating THS policies and collecting samples from homes and business to measure THS. This will be done to help people and organizations understand how widespread THS pollution is, what can be done to prevent THS from building up, and what options are available for cleaning up THS after it has become firmly embedded (Aim 4). By increasing awareness of THS pollution and exposure risks, this project will help reduce tobacco use in indoor spaces, create a better understanding about why enforcing tobacco-free policies is important, and expand tobacco-free policies to places that currently do not have them. This project has the potential to lead to indoor spaces that are 100% free of toxic tobacco smoke and to encourage people not to smoke.

Reducing Exposure to Thirdhand Smoke in Multiunit Housing

Penelope J.E. Quintana, PhD
San Diego State University

From our previous studies, we know that homes can be contaminated by previous smoking in the home as well as by drifting tobacco smoke, even if current residents do not and have never smoked. One of the most common questions from the general public, realtors, and others is how to clean homes that have been smoked in so that nonsmokers are not exposed to toxic tobacco smoke residue (also known as thirdhand smoke). This residue remains on walls, floors, carpet, and other surfaces, and in dust and air.

This study will recruit nonsmoking residents who live in multiunit housing homes. We will collect samples to measure thirdhand smoke in their homes. First, we will screen homes for thirdhand smoke residue. Then in highly polluted homes identified by screening, as well as low-polluted homes as a comparison group, we will collect more samples to carefully measure levels of thirdhand smoke chemicals in dust, air and surfaces. We will test simple devices being developed by the Thirdhand Smoke Consortium members for measuring thirdhand smoke contamination on surfaces and in air, and measure thirdhand smoke in a silicone wristband worn by a participant. We will also measure personal exposure of residents to thirdhand smoke by examining their urine and saliva samples. In the most contaminated homes, we will also clean the homes to see if our method removes the thirdhand smoke residue. Commercial companies will apply a cleaning and remediation protocol developed by the THS Consortium to the homes. We will then evaluate how effective this cleaning was for removing thirdhand smoke.

The purpose of this project is to provide answers to current knowledge gaps about thirdhand smoke, including identifying chemicals of concern in thirdhand smoke, how to test for them, and how to remove them. Our results will be immediately useful to concerned residents, landlords, realtors, tobacco control advocates, and others. We will be able to give practical advice about how to test for thirdhand smoke, which surfaces and areas in a home are most polluted, and how many samples should be collected to obtain an accurate measure. We will test simple methods for measuring thirdhand smoke residue that could be used by people to check their own homes. We will disseminate our findings in collaboration with the THS Consortium subproject 7, the Dissemination, Outreach, and Resource Center Core.

2011 Consortium Projects

Thirdhand Smoke in Homes and Tobacco-specific Nitrosamine Exposure in Children

Neal Benowitz, MD
University of California San Francisco

Secondhand smoke (SHS) is a known major risk factor for illness including asthma, respiratory tract infections and ear infections in infants and children. Third hand smoke (THS, the byproduct of SHS on surfaces and dust in rooms where cigarettes are not currently being smoked) contains many of the same toxic substances as SHS (and probably some additional ones formed when SHS “ages” over time) and thus could also be a significant cause of disease, especially in infants and children and in adults with chronic illnesses.

In California today, after new regulations limiting smoking in public places, the home is the major place where non-smokers, including children, are exposed to the toxic substances derived from tobacco smoke. Even in homes where smokers do not smoke in the presence of children, the children may live and play in rooms in which smoking has previously occurred, or (in multiunit buildings) smokers may live in other units or the unit may have been formerly occupied by smokers.

Our subproject consists of two exploratory studies which specifically address the TRDRP goals of directly measuring exposure to THS in residential housing (with specific emphasis on housing affecting children) and assessing the extent of exposure of residents by measuring urine levels of substances resulting from that exposure.

In the first study (Project A: THS Household Characterization Study), we will determine in dust and surface wipe samples concentrations of various components of THS in different types of homes. We will study urine samples of nonsmoker adult residents of these homes and compare them to the concentrations of THS we found in the home. We will also look at ways to distinguish THS from SHS exposure using urine specimens.

In the second study (Project B: THS and TSNA Exposure in Children – Pediatric Exposure Study) we will take urine samples from infants and young children who might be exposed to THS, and compare levels of THS components in the urine to levels we find in dust and surface wipes in their homes.

We expect our studies to produce preliminary data on the levels of THS substances in different types of homes and how these levels are related to the actual measures of THS exposure in adults and children in these environments. Our projects could lead to additional studies which would define more clearly the relationship between human exposure to THS and subsequent health effects, which in turn is likely to result in public demand for legislative policies to protect nonsmokers from THS.

Genotoxocity of Thirdhand Smoke in Mammalian Cells and Tissues

Bo Hang, M.D., PhD
Lawrence Berkeley National Laboratory

Exposure to thirdhand smoke (THS) – the smoke residue that lingers on surfaces, remains, reacts and/or re-emits, has recently become a public health concern, because it may become a long-term source of exposure to tobacco toxicants, especially for toddlers and occupational workers who may have close contact with contaminated surfaces. Previous studies from the cigarette industry showed that the aging of SHS leads to its increased toxicity and research under laboratory conditions demonstrated that new toxicants are produced from reactions of nicotine with common atmospheric pollutants ozone and nitrous acid, which remain on the indoor surfaces or enter the air. Therefore, THS is expected to contain newly formed toxicants under realistic conditions that may impose adverse effects on health. However, little experimental evidence is available about the chemical composition and actual biological impacts of components produced de novo in THS, especially what harm may be caused to structure and function of the genetic material DNA, which may ultimately lead to diseases such as cancer. The aim of this subproject is to detect bulky DNA adducts (caused by covalent binding to DNA of tobacco toxicants with large size) in order to assess such genotoxic nature of the THS samples. It is well accepted that DNA adducts play a central role in smoke-induced carcinogenesis. We believe that the timely and unequivocal identification of such genotoxicity could be critical for framing and enforcing new policies against indoor smoking such as in homes and passenger cars in California and elsewhere in order to protect vulnerable populations.

Our main hypothesis is that THS contains genotoxic chemicals capable of forming specific DNA adducts in mammalian cells as well as tissues of experimental animals exposed to THS. To test this hypothesis, multiple collaborations will be carried out within this consortium and the following specific aims will be pursued: (1) Using calf thymus DNA, we will investigate the unknown adduct-forming potential of THS components, including 4-(N-methyl-N-nitrosamino)-4-(3-pyridyl)-1-butanal (NNA, a major tobacco-specific N-nitrosamine found in nicotine reaction with indoor nitrous acid) and selected new THS constituents to be identified in other subprojects. THS samples/compounds will be collected and characterized by investigators at Lawrence Berkeley National Laboratory/University of California at San Francisco. In this study, highly sensitive methods involving 32P-postlabeling and Liquid chromatography-mass spectrometry (LC-MS) will be used to detect putative bulky adducts on the basis of their characteristic chemical/structural features. It should be noted that the current funded project (to B.H), which is focused on using calf thymus DNA to analyze genotoxicity of whole THS extracts in the first two years (2010-2012), has no overlap in specific aims and time with this proposed research that will begin in 2013, if funded; and (2) We will examine the cellular genotoxicity of THS by measuring the formation of bulky DNA adducts resulting from exposure of cultured mammalian cells and tissues of rats to THS, including whole extracts with and without exposure to nitrous acid, and isolated chemicals identified in THS. Cells studied will include HepG2 cells (containing P450s), as well as other types of cells from the labs of the participating subprojects, including human/rat lung epithelial cells and pulmonary fibroblasts, and human embryonic stem cells. These experiments will help establish the genotoxic nature of THS under various conditions using biologically effective doses of THS and its components.

Identification and Characterization of Toxicants in Thirdhand Tobacco Smoke

Hugo Destaillats, PhD
Lawrence Berkeley National Laboratory

In this application the term thirdhand tobacco smoke (THS) refers to tobacco smoke residues that remain on indoor surfaces, react and/or re-emit to indoor air long after active smoking has ended.

Our group has recently found that tobacco residues can become more hazardous when they react with common air pollutants like nitrous acid (HONO) or ozone. HONO is formed indoors when combustion sources are operating, and about half of the ozone outdoors typically makes its way inside homes. Ozone may also be generated indoors with “air purification” purposes. Our recent laboratory work revealed the formation of carcinogenic (cancer-causing) tobacco-specific nitrosamines (TSNAs) from reactions of nicotine (adsorbed to paper, used as a model indoor material) with HONO. Our results predicted that the amounts of TSNAs on typical surfaces were high enough to justify finding out how dangerous THS is by discovering what its health effects could be. Our other recent study showed that very small (nano-sized) particles form when nicotine by itself or secondhand tobacco smoke reacts with ozone. The major compounds in the new particles turned out to include many chemicals that could cause or exacerbate asthma. Both types of toxic products could greatly increase the harmful effects of exposure to second- and thirdhand smoke.

One objective of this sub-project is providing well-characterized samples of THS to other investigators who will study its health effects. The research team will provide experimental protocols and standardized THS samples to other sub-projects. The number of cigarettes, ventilation rates and the length of sampling will be controlled. Two types of samples will be generated on sterile cellulose paper for studies of health effects: a) THS samples produced by smoldering cigarettes in sterile chambers, and b) THS samples exposed to HONO, in which a fraction of samples collected in (a) will be exposed to realistic HONO levels (50-100 ppbv) to produce TSNAs. The THS samples prepared will be characterized by chromatography at LBNL.

The other main objectives of this sub-project involve discovering more about the composition of THS, and how it forms, accumulates and reacts with indoor pollutants. To find out how THS is formed on surfaces, key constituents of secondhand smoke (included nicotine and other amines) will be mixed with HONO or ozone in chambers that hold model indoor materials, such as cotton, carpet fibers and indoor dust. We will also investigate THS formation on surfaces coated with human skin oils, to explore their role in inhibiting or promoting those reactions. After identifying and quantifying THS constituents, we will evaluate their distribution among indoor air, dust and surfaces, and we intend to relate those measurements to the exposure people could receive by inhalation or skin contact with contaminated clothing, furniture or carpets. Our team will also examine the role that household dust plays in exposure to THS because small children could be particularly exposed to THS-containing dust. We also plan to collaborate with other subprojects to identify new compounds that can originate in THS and be present in urine (biomarkers), allowing for tracking THS exposures.

The results from this sub-project will be used to predict the levels of THS constituents associated with the main pathways for THS exposure: inhalation, skin contact and ingestion of dust. Such knowledge is crucial to other consortium sub-projects that will probe associations between exposure to these compounds and adverse health effects.

Thirdhand Smoke: Exposure Risk, Policy and Economic Implications

Jonathan Samet, MD, MS
University of Southern California

Policies banning smoking in workplaces and public locations are now in-place in California and the majority of other states. These laws protect many non-smokers from toxic chemicals that cause death and disease.  However, we now have new information showing that toxic chemicals from smoke stay on carpets and other surfaces, even when smoke is no longer in the air. This new information raises concern that some people, including babies and small children, may be exposed to harmful toxins from cigarettes, just by being in contact with the contaminated surfaces. In addition to homes, places that may be contaminated with smoke toxins include rental housing, hotel rooms, and cars. The goal of this sub-project is to determine how common this contamination is in California and to understand if these toxins are entering the bodies of children and adults. Based on this information we will determine how dangerous thirdhand smoke is to the health of children and adults and what should be done to protect people. Based on this assessment, we will evaluate approaches to protect people from being exposed to thirdhand smoke, such as testing for its presence at the time of real estate transactions and requiring appropriate clean-up if needed. We will also evaluate the economic costs of thirdhand smoke.

UCSF Human Exposure, Toxicology, and Biomarkers Core Analytical Chemistry Laboratory

Peyton Jacob, PhD
University of California San Francisco

This subproject is part of a consortium of California researchers who are seeking to understand the risks of exposure to aged secondhand smoke, which has been termed “thirdhand smoke” (THS). Past studies have shown that secondhand smoke becomes more toxic as it ages. It is of particular concern that levels of potent cancer-causing substances (carcinogens) called tobacco-specific nitrosamines (TSNA) increase as smoke ages.

The researchers in this subproject are chemists who have been studying human exposure to tobacco smoke toxins for many years. For this subproject they will develop methods for measuring human exposure to toxins in THS, including carcinogens. They will also analyze samples from studies carried out by researchers in other subprojects that will assess the toxicity of THS, and samples from other subprojects to measure human exposure in real-world settings.

These studies will expand our knowledge about the chemical nature of THS and its impact on public health. It is expected that the research carried out by this consortium will help in developing policies for protecting people from THS exposure, and may help in developing methods for remediation (detoxifying) hotel rooms where smoking has occurred, and homes and cars of smokers that are for sale.

Effect of Thirdhand Smoke (THS) on Oviduct Function and Embryonic Development

Prudence Talbot PhD
University of California Riverside

Tobacco smoke deposited on surfaces or “thirdhand smoke” (THS) is present in many locations including homes and cars of smokers, offices, and hotel rooms. Our prior work showed that sidestream smoke, which burns off the end of cigarettes and is the main contributor to THS, is highly toxic. A consortium partner recently showed that nicotine in THS in the presence of nitrous acid is converted into several carcinogens. While these studies suggest that THS is dangerous, its biological effects on cellular processes have not yet been investigated. We propose to test the hypothesis that THS contains toxicants capable of adversely affecting a spectrum of dynamic cellular processes. Experiments will be done using assays, developed in our lab, that provide multiple physiological, cellular, and molecular endpoints. We will specifically investigate the effects of THS on functioning of the female reproductive tract (specifically the oviduct, an organ vital for normal fertility). We will also examine THS’s effect on cultured human lung fibroblasts which are likely to receive significant levels of THS exposure through inhalation and on human embryonic stem cells (hESC) which model the earliest stages of human prenatal development and are likely to be highly sensitive to THS. In Specific Aim #1, we will perform dose response experiments to characterize the effects of THS on the female reproductive tract using an oviductal bioassay that provides multiple functional endpoints. The endpoints that are measured include ciliary beat frequency, the rate at which the oocyte is picked-up by the oviduct, the strength of adhesion between the oocyte and oviduct, and the rate of oviductal smooth muscle contraction. This Aim will provide novel information on THS’s effect on the female reproductive tract and multiple cellular processes important in many organ systems. In Specific Aim #2, we will characterize dose dependently the effects of THS on cellular processes using in vitro assays with human cells. Morphological and molecular endpoints for blebbing, attachment, survival, proliferation, cell death, and pluripotency/differentiation will be studied using novel video technology combined with software tools for mining data from video images. Quantitative evaluation of gene expression will yield complementary molecular endpoints. Our Stem Cell Core has excellent instrumentation for collecting (BioStation CT) and mining (software) time lapse video data on dynamic cellular processes. Our campus is well known for development of video bioinformatics software, and our project will be supported by resources provided through our Stem Cell Core Facility, NSF IGERT training grant in Video Bioinformatics, as well as through our collaborative work with DR Vision (a video bioinformatics software development company in Seattle). Experiments in Aim #2 will be done using hESC which model early human development and are likely to be very sensitive to THS and with human lung fibroblasts that will serve as an adult comparison from an organ likely to receive significant doses of THS. It is important to use an embryonic model as this stage of the life cycle is likely to be the most sensitive to environmental chemicals. In both Aims, THS will be produced at the Lawrence Berkeley National Laboratory by Dr. Destaillats group. We will test THS, THS smoke treated with nitrous acid, and individual chemicals in THS as they are identified by other consortium labs including Dr. Jacob’s group at UCSF. For each endpoint, we will determine the lowest concentration that produces an effect, the concentration that reduces an effect by 50%, and the maximum percentage reduction of an effect. In the experiments involving stem cells and lung fibroblasts, we will measure gene expression in cells at the end of the time lapse experiments to correlate molecular data with the morphological endpoints obtained using BioStation technology. Finally, we will do reversal experiments by exposing oviducts/cells to THS or individual chemicals, washing out the treatment compound, and examining recovery. Our data will establish the toxicological effects of THS and its components on a spectrum of cellular processes, the doses that are effective, the efficacy of each treatment, and the feasibility of reversal of effects after exposure. Effective doses for each endpoint will be correlated with concentrations of THS biomarkers found in humans as this information becomes available through other consortium labs. In summary, our study will provide new quantitative data on the general toxicity of THS and its constituents and give new insight into how THS could impair human health by adversely affecting various cell types, cell processes, organs, and adult vs. embryonic cells.

Controlled Exposure of Human Subjects to Thirdhand Smoke

Suzaynn Schick, PhD
University of California San Francisco

There is growing evidence that compounds from thirdhand smoke can be detected in human subjects who are exposed in their homes. We don’t yet know whether the route of exposure is inhalation, dermal absorbtion, oral uptake, or a combination of the three. Nor do we know whether thirdhand smoke exposure has detectable health effects. The purpose of this subproposal is to test whether we can detect any chemicals from thirdhand smoke in people who breathe a known concentration of thirdhand smoke under controlled conditions. To generate controlled exposures to thirdhand smoke we will use a system designed for exposing human subjects to secondhand smoke. The system uses a machine to smoke cigarettes and then dilutes the sidestream smoke and ages it in a 6 cubic meter stainless steel aging chamber so that it is like the mixture of fresh and aged smoke that encountered under realistic ventilation conditions in smoking permitted indoor spaces. On average, 40% of the secondhand smoke particles deposit in the aging chamber during the 60-minute aging for a secondhand smoke exposure. To generate thirdhand smoke, we will just fill the aging chamber with smoke, close it up for 12-48 hours and then run clean air through it to pick up desorbing smoke compounds for the exposure. The subjects will wear a modified pressurized air purifying respirator hood over their heads and breathe the thirdhand smoke while at rest.

Specific Aim 1: Chemically characterize the sorbed and desorbing THS compounds in the SHS exposure system at the Human Exposure Laboratory at UCSF by testing for volatile organic compounds (VOCs), aldehydes and ketones, polycyclic aromatic hydrocarbons, nitrosamines, nicotine and other nicotine reaction products under two conditions: no added materials in the aging chamber and wallboard, paper, wool cloth and cotton cloth added.

Specific Aim 2: Expose 15 human subjects with nasal allergies to THS for 4 hours, test for exhaled nitric oxide, collect blood and urine samples and test for VOCs, VOC metabolites, nicotine and nicotine metabolites, nitrosamines and nitrosamine metabolites and markers of oxidative stress. Collect buccal and nasal epithelial cells and bank to analyze for protein expression. Repeat exposure using filtered conditioned air and compare results.

Specific Aim 3: Provide materials loaded with THS, extracts of THS contaminated materials and THS-conditioned media to other members of the consortium for chemical analysis and toxicity testing.

The outcome of the proposed chemical analyses will help bridge the gap between the small-scale chamber studies of THS chemistry and the collection of THS from homes and other real-world environments proposed by other researchers in this consortium. This will be the first human exposure experiment with known concentrations of thirdhand smoke and will yield information on the effects of THS exposure without the confounding factor of exposure to SHS seen in most realistic exposures to THS. We recruit subjects with nasal allergies because they are usually more sensitive to air pollutants. Exhaled nitric oxide concentrations often increase when the lungs are under physiological stresses like infection or air pollution. We will measure
the concentrations of markers of oxidative stress including malondialdehyde (MDA), oxidized glutathione (GSSG), C-reactive protein (CRP), GM-CSF, IL-1beta, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IFN-gamma and TNFalpha because concentrations of these compounds are also associated with damage to the respiratory tract. Creating a supply of well-characterized THS samples generated under controlled conditions will expand the materials available for other members of the consortium to test and ensure that the resultant toxicity results are as generalizable and meaningful as possible. We expect that the results of this project will help determine how much of an effect breathing thirdhand smoke has on human health.

Exposing Children to Secondhand Smoke in Contaminated Homes

Georg Matt, PhD
San Diego State University

When children are exposed to secondhand smoke (SHS) they are more vulnerable than adults to suffer negative health effects. The vast majority of SHS exposure among children takes place at home, where protecting children is particularly challenging. This is the case because public health policies for the protection of nonsmokers do not apply to the private homes of smokers and nonsmokers. Our recently completed TRDRP IDEA study found that air, dust, and surfaces in the homes of smokers were contaminated with ETS even when smokers tried to protect their children by not smoking at home. Because sources of SHS contamination do not vanish from a home when smokers move out, homes are likely to remain contaminated. Consequently, nonsmokers moving into smoker homes may be exposed to SHS produced by the former tenants.

The proposed study will be the first to examine whether private homes of smokers remain contaminated when they move out and nonsmokers move in. Moreover, this will be the first study to examine whether nonsmokers moving into former smoker homes are exposed to SHS through contaminated dust, air, and surfaces in their homes. The proposed study has four major aims: First, we will determine whether the dust, air, and surfaces in the homes of smokers with young children are contaminated with SHS before they move out. Second, we will study how much contaminated dust, surfaces, and air contribute to the SHS exposure in children of smokers. Third, we will investigate whether homes remain contaminated after smokers move out and nonsmokers move in. Fourth, we will examine whether nonsmokers who move into smoker homes are exposed to SHS through the contaminated home environments.

To examine these questions, we will study 150 homes with children ≤5 years during a change of occupancy. In the first part of this study, homes of 100 smokers and 50 nonsmokers will be visited before the current tenants move out. Household dust, air, and surfaces will be examined for evidence of SHS. Urine samples of children will be investigated for signs of SHS exposure. Parents will be interviewed to learn about their smoking histories, SHS exposure, and cleaning habits. In the second part of the study, the new tenants who move into the smoker homes will be contacted and recruited into the study if they are nonsmokers. Homes will be visited again, and household dust, air, and surfaces will be examined again for evidence of SHS. Moreover, urine samples of the youngest resident in each home will be investigated for signs of SHS exposure. The new tenants will be interviewed to collect data on their smoking histories, SHS exposure, and cleaning habits. Data will be analyzed to examine the contamination of smoker homes and the SHS exposure of children living in these homes. Data will be analyzed to determine if homes remain contaminated after smokers move out and nonsmokers move in. Finally, data will be analyzed to determine if nonsmokers moving into smoker homes are exposed to SHS smoke through the contaminated homes. This study will provide much-needed data on the importance of dust, surfaces, and air as sources of SHS exposure among young children. Findings from this study will also help better understand the level of contamination of apartments vacated by smokers. This study will help inform public policies for protecting nonsmokers from SHS exposure in their homes.

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