A biomarker is a measurable substance, typically a chemical or biomolecule (protein, lipid, nucleic acid), found in biological samples that is indicative of a normal or abnormal condition of a living organism. But how can biomarkers be used in deciphering human health impacts of environmental contaminants?
Biomarkers have long been used in medicine to diagnose and monitor disease progression and response to therapy. Biomarkers are also a critical tool for evaluating human exposure and response to chemicals in the environment. Diverse biological samples are used by environmental health scientists to measure biomarkers in humans, including blood, urine and other bodily fluids; tissues, such as adipose, bone, teeth and hair; and images of living tissue, such as X-rays.
Biomarkers used in environmental health studies are broadly classified as biomarkers of exposure, effect, or susceptibility. Biomarkers of exposure are used to evaluate whether an individual has been exposed to a chemical, and, if so, how much. Biomarkers of exposure are used in occupational settings to ensure that workers are not exposed to unsafe levels of chemicals in the workplace. They are also used in epidemiological studies to evaluate associations between exposure to environmental chemicals and adverse human health effects. The use of biomarkers of exposure enables epidemiologists to rank subjects according to their exposure level and determine whether the health outcome is significantly correlated with the magnitude of exposure, e.g., is the health effect more prevalent or more severe in individuals with higher exposures relative to those with lower or no exposures?
Biomarkers of effect are used to measure biological responses to chemical exposures. Like biomarkers of exposure, biomarkers of effect are used in the workplace to monitor the health of workers with potential occupational exposures to toxic chemicals and in epidemiological studies to assess health conditions in exposed versus control (unexposed) populations. An example of a biomarker of effect would be altered blood levels of a liver enzyme as a biomarker of liver toxicity. Biomarkers of effect that are based on the molecular mechanisms by which the chemical causes toxicity can also provide insights on how chemical exposure influences adverse health outcomes. For example, biomarkers of mitochondrial dysfunction have been used to identify chemicals that increase the risk of Parkinson’s disease by interfering with the function of mitochondria.
In some cases, a biomarker can be a measure of both exposure and effect. An example is the activity of acetylcholinesterase (AChE) in blood. AChE is an enzyme that inactivates the neurotransmitter acetylcholine to terminate signalling between upstream and downstream cells in a neural circuit. Organophosphate (OP) insecticides cause neurotoxicity by inhibiting AChE in the nervous system. AChE is also expressed by red blood cells, and red blood cell AChE is similarly inhibited by OPs. Thus, AChE activity in blood samples is widely used to monitor exposure to OP pesticides in agricultural workers and to confirm OP exposure in individuals exposed to these compounds either accidentally or intentionally (e.g., terrorism, warfare, murder or suicide). Because this biomarker measures a biological response to OPs (inhibition of AChE), it can also be considered a biomarker of effect, and the magnitude of AChE inhibition in the blood is used to predict the likelihood that an exposed individual will develop severe symptoms of OP poisoning.
Biomarkers of susceptibility are used to assess an individual’s vulnerability to the toxic effects of chemical exposure. These can include genetic factors that increase or decrease the toxicity of environmental exposure or other biological factors, such as metabolism, nutrition/diet, age, health status, etc., that influence individual response to chemical exposure. One example is elevated cholesterol levels, which are associated with an increased risk of developing numerous diseases following toxic exposures. Biomarkers of susceptibility are also useful for understanding gene-environment interactions involved in disease pathogenesis and provide potential answers for explaining the variable responses often observed among individuals in an exposed population.
Below, we discuss examples of well-established and emerging biomarkers used in environmental health to assess exposure to and effects of chemicals that target the gut, respiratory system and brain. The gut and respiratory systems are the first organ systems impacted following exposure to environmental chemicals via ingestion or inhalation, which are primary routes of human exposure to chemicals. A number of non-invasive or minimally invasive approaches are available to assess environmental impacts on these organ systems. In contrast, there have historically been few to no biomarkers for assessing environmental impacts on the living brain; however, recent advances in technology for imaging the living brain suggest exciting new approaches for overcoming that challenge.
Biomarkers Associated with dietary exposures and toxic effects on the gut
As indicated above, humans are commonly exposed to toxic contaminants in the environment via ingestion of contaminated food and water. The gut is a primary line of defense against these contaminants. The mucus-coated epithelial cell lining of the gut provides a physical barrier that restricts uptake of many substances, while diverse enzymes in the gut can metabolize (alter) toxic substances to make them less toxic and/or easier to eliminate from the body. Another important defense against toxic substances in the diet is the community of microorganisms that live in the GI tract, also known as the gut microbiome. The gut microbiome plays a crucial role in human health by aiding in digestion, releasing nutrients, and producing metabolites that are taken up into the circulatory system and distributed throughout the body to influence the health and function of diverse organ systems. The gut microbiome also facilitates the breakdown and elimination of toxic substances in the diet.
The waste material produced by the gut and the gut microbiome is eliminated as feces. Thus, stool samples are an important source of biomarkers of exposure to environmental contaminants. Stool samples can be analyzed for specific environmental contaminants or their metabolites, for example, pesticides, metals (arsenic, lead, cadmium, mercury), persistent organic pollutants (PFAS, PCBs, PBDEs) and, more recently, microplastics and nanoplastics. However, this approach will yield exposure data only for the specific substances that are being measured, and it is well known that we are exposed to tens of thousands of different environmental chemicals in our diet.
An emerging approach for generating a more holistic assessment of dietary exposure to toxic substances is to measure the diversity of microbes in the feces. Gut microbes are continuously shed in the feces, and both human and animal studies indicate that the profile of microbes detected in the feces reflects the profile of microbes in the gut itself. Experimental animal models have also demonstrated that dietary exposure to environmental chemicals can change the microbial composition and metabolites in the gut microbiome and feces. The limitation of this approach is that it does not identify the specific dietary substance(s) causing the change. It is hypothesized that different classes of toxic chemicals generate a unique microbial “fingerprint”, e.g., there are chemical-specific shifts in microbial diversity or metabolite profile, but this has yet to be proven.
While the gut typically functions to defend us from harmful environmental contaminants in the diet, it can be adversely impacted by dietary exposures. For example, dietary exposure to arsenic is linked to intestinal inflammation and cancer, while PFAS in the diet is associated with significant gut dysbiosis, a disruption of the gut microbiome that changes its functional composition and/or metabolic activities. Imbalance of the gut microbiome is linked to a number of adverse health effects, including not only digestive disturbances but also adverse systemic effects ranging from fatigue to skin problems to inflammation and aching joints.
The composition of the stool, including the microbial composition and metabolite profile, can also serve as biomarkers of effect, providing clues about gut health. For example, the U.S. Food and Drug Administration has approved several tests for the detection of colon cancer based on detection of inflammatory molecules in the stool. Some microbial metabolites are also being used as biomarkers of disease. For example, the microbiome produces unique bile acids, like lithocholic acid (LCA) and deoxycholic acid (DCA). A high fecal LCA:DCA ratio is a biomarker of susceptibility for colorectal cancer, breast cancer, and gallstones. Other microbial metabolites are biomarkers of other types of gut dysbiosis, suggesting the exciting possibility that a fecal microbial metabolite panel could provide a comprehensive assessment of environmental exposures and individual risks of specific disease states associated with gut dysbiosis.
Biomarkers associated with inhalation exposures and respiratory toxicology
Inhalation is another common route of human exposure to environmental contaminants and as of 2021, air pollution caused over 12 million deaths per year worldwide. The respiratory system, which includes the nasal cavity, trachea and lungs, is a primary target of inhaled environmental contaminants, but it is important to note that inhalation is not the only way in which the respiratory system is exposed to toxic chemicals in the environment. For example, ingestion of the herbicide paraquat is linked to pulmonary fibrosis that can lead to respiratory failure and 6 even death and injected chemotherapy drugs can have serious respiratory side effects. It is equally important to note that inhalational exposures do not solely impact the respiratory system. Secondary effects from inhaling air pollutants include increased risk of cardiovascular diseases, metabolic disorders, immune dysfunction and even neurological disorders. Interestingly, inhaled substances can reach the brain directly via the olfactory nerves that connect the nose to the brain.
Exposure to airborne toxic substances is strongly associated with respiratory distress and disease. Air pollutants can result from anthropogenic (human-influenced) sources, such as tobacco smoke and traffic, or natural sources, such as wildfires. Environmental air pollution includes particulate matter (PM) and various chemicals, including nitric oxides (NOx), volatile organic compounds (VOC), and ozone (O3). It also includes indoor and outdoor allergens such as house dust mites, pollen, pet dander, mold spores and mycotoxins that can trigger respiratory symptoms and exacerbate conditions like asthma. Additionally, certain occupational exposures have been identified as having detrimental effects on the respiratory system. For example, inhalation of asbestos fibers or silica dust can cause lung fibrosis, scarring and cancer. Pesticides such as OPs and carbamates can also cause respiratory symptoms and lead to asthma and chronic obstructive pulmonary disease (COPD).
Toxic chemicals can differentially impact the respiratory system depending on the duration and intensity of exposure. Therefore, biomarkers of exposure, effect, and susceptibility are essential for monitoring and mitigating exposures to prevent diseases and improve overall lung health. The need for biomarkers of respiratory exposures is particularly important because it is difficult to sample the lung directly. Several different biological samples can be used to determine what is happening in the lungs following exposure. These include exhaled breath condensate (EBC), induced sputum, blood, and urine, all of which are minimally invasive approaches often used in combination with measures of lung function to understand onset or worsening of lung disease. More invasive approaches, such as bronchoscopy (in which a small sample of lung tissue is obtained) or bronchoalveolar lavage (collection of fluid from the lungs) have more restricted use and are only obtained in certain clinical settings. The collection of EBC is increasingly used because the instrumentation for collecting and measuring EBC has become more widely available. During EBC collection, exhaled breath passes through a cooling device resulting in the accumulation of substances in exhaled air. Concentrations of these substances, which can include environmental chemicals and biomolecules found in cells of the respiratory system, are influenced by lung disease and, thus, can be used as biomarkers of not only chemical exposure but also the pulmonary response to inhaled respiratory toxicants.
Similarly, induced sputum allows for the analysis of different types of cells in the airways and is widely used for characterizing airway inflammation. Sputum induction entails inhalation of sterile saline (a salt solution that is clean and free of germs), followed by coughing up and spitting out airway secretions. The elevated presence of white blood cells, immune cells important in the body’s defense against bacterial and parasitic infections as well as allergens, is an indication of inflammatory responses to environmental exposures. These cellular biomarkers can also be evaluated using the more invasive techniques of bronchoscopy and bronchoalveolar lavage (BAL) fluid collection. During these processes, a tube with a tiny camera on the end is inserted through the nose or mouth and into the lungs. In a bronchoscopy, a small piece of lung tissue is collected. In BAL, sterile saline is introduced into the lung and immediately suctioned to collect the saline solution for analysis. Compared to sputum analysis, which provides information about the nose and mouth, bronchoscopy and BAL allow for the sampling of deeper sections of the respiratory tract with less microbial contamination and can be useful for understanding local inflammation within the lungs.
When significant respiratory toxicity has occurred, or there is widespread inflammation throughout the body following an inhalational exposure, it is common to analyze specific lung-related proteins in the blood or urine as biomarkers of these effects. An early response to environmentally induced lung injury is increased permeability of the lung epithelium, the layers of cells that line the lung that functions in barrier protection (much like the epithelial cells that line the GI tract). Increased permeability allows epithelial proteins to leak into the blood and then subsequently pass into the urine. CC16, a lung cell protein, is an example of a blood and urinary biomarker of increased lung permeability. In children, urinary levels of CC16 are a validated biomarker of the development of wheezing and were found to be elevated following acute exposure to PM in wildfire smoke.
Biomarkers of susceptibility have provided important clues for understanding why certain individuals may be more susceptible to respiratory problems in response to toxic exposures and how environmental exposures may contribute to the development or exacerbation of respiratory disease. For example, specific genetic variations in the detoxification gene GSTM1 and the inflammatory cytokine gene TNF-α are associated with COPD, while genetic variations in the inflammatory cytokine gene interleukin-33 (IL-33) and the IL-33 receptor gene, ST2/IL1RL1 are linked to asthma.
