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Arsenic and Human Health Arsenic is considered the number one environmental chemical of concern for human health both in the U.S. and worldwide. As many as 25 million people in the U.S. alone may be exposed to excess arsenic at levels that have been associated with increased risk of disease, and worldwide such exposures may include several hundred million people. People can be exposed to arsenic through drinking water, air, food and other sources, but the primary concern is consumption of arsenic in drinking water, primarily from contaminated private wells. For example, in New Hampshire, approximately half the population gets its drinking water from private wells, and as many as one fifth of these contains excess arsenic, representing a tenth of the population or about 130,000 people. Similar situations exist in neighboring Maine and other New England states, in Michigan and other northern Midwest states, in Arizona, New Mexico and other Southeast states, in the Rocky Mountains, and in California. Studies in areas with high levels of drinking water arsenic, such as Taiwan, Argentina and Bangladesh have shown that chronic exposure to arsenic, even at levels that are not associated with clinical signs of arsenic poisoning, is associated with a greatly increased risk of vascular and heart disease, diabetes, reproductive and developmental problems, and a wide variety of cancers including skin cancer, bladder cancer, lung cancer, liver cancer and kidney cancer. How arsenic exposure is able to affect the risk of so many different diseases is not well understood. Research led by Joshua Hamilton is examining a specific mechanism that Hamilton and colleagues believe is one of the primary ways that arsenic affects human health. Their laboratory discovered that arsenic can act as a potent endocrine disrupting chemical (EDC). Previously, most work on EDCs focused on pesticides and other organic chemicals in the environment that mimic hormones. However, Hamilton and colleagues demonstrated that arsenic can also affect hormone processes, but by a unique mechanism that does not involve hormone mimicry. The primary goal of the research is to understand this mechanism of action at a more detailed level, and to investigate the health consequences of such hormone disruption. Hormone Receptors as Targets for Arsenic Hamilton’s laboratory had previously demonstrated that extremely low levels of arsenic could alter hormone-mediated pathways in the cell. Arsenic was first shown to disrupt the ability of a particular receptor, the glucocorticoid hormone receptor, to regulate expression of genes that are normally responsive to glucocorticoid hormone. Glucocorticoids control many different functions in the body, including the blood sugar, glucose (after which they are named), cell growth and differentiation, inflammation (which is what topical cortisol is used for), embryonic and fetal development, and many other functions. So it would not be surprising that disruption of this key hormone pathway might affect a wide variety of diseases including cancer, diabetes, heart disease, and development. Hamilton’s group then went on to show that arsenic has similar effects on all five steroid receptor pathways. These pathways include not only the receptors for glucocorticoids but also those for the sex steroids, estrogen, progesterone, and testosterone, as well as the receptor for mineralocorticoids (aldosterone, which controls salt concentrations and other kidney functions). Through findings in more recent studies, Hamilton’s laboratory has extended this list of affected pathways to include the receptors for retinoic acid and thyroid hormone, which are part of the same large family of nuclear hormone receptors. Collectively, these hormone receptor regulated pathways control a myriad of body functions, and so disruption of all of these by arsenic would be expected to have profound consequences on disease risk. One of the principal aims of the current work is focused on how arsenic is able to disrupt all of these pathways, examining these processes at the molecular level inside cells. Alterations in Patterns of Gene expression A second focus of the current research is to determine the “downstream” consequences of these endocrine disrupting effects of arsenic. In other words, what happens to cells or animals when such altered hormone signaling occurs, and is this directly related to how arsenic affects disease risk? Hamilton and his colleagues are studying these downstream effects using a new tool in biology called genomics. Hamilton’s laboratory, like many other labs doing similar work, had previously investigated the effects of arsenic, chromium and other toxic metals on expression of individual genes that they hypothesized might be affected. They showed in this manner that some genes were very specifically affected by arsenic while many others were not, and in this manner they were led to investigate the regulation of some of these genes, which in turn revealed the endocrine disrupting potential of arsenic. While it has often been fruitful to investigate expression of individual genes in this manner, such research required the investigator to know, or try to guess, that a particular gene might be altered by a given treatment. However, given that there are some 30,000 genes in the human genome, this is at best a hit-or-miss strategy. Even with success at finding some genes this way, Hamilton describes the process as much like three blind men feeling an elephant: one feels the tail and pronounces that the animal is a rat, another feels the legs and says it is a rhinoceros, and a third feels the trunk and says that it is a snake. It is often only when the entire picture is revealed that scientists see the true nature of the beast they are studying. Thus, for gene expression one would like to know all the genes that are altered by arsenic, and also rule out all the genes that are unaffected. Using what is called a “functional genomics” approach, the Dartmouth group can now do that. With the advent of modern molecular biology techniques, and the completion of the human genome project, it is now possible to simultaneously examine the expression of virtually all the genes in a cell at the same time. This global approach to gene expression is a new field that is referred to as genomics. A similar approach that aims to look at more global patterns of protein expression is called proteomics. Hamilton and colleagues are currently using these approaches to examine the overall pattern of altered gene expression with arsenic, with the aim of being able to understand the consequences of arsenic exposure from these altered patterns. In a recent study, they reported that arsenic, chromium, cadmium, nickel and an organic carcinogen, mitomycin C, each produced a unique pattern of altered gene expression. What’s more, the patterns of altered gene expression showed very little overlap, with almost no gene represented on more than one list. The results show that the list of genes affected by each chemical is almost completely unique, an indication that each chemical behaves very differently inside the cell. Moreover, the number of genes that were affected by each treatment was very small compared to the total genes analyzed, suggesting that these gene expression patterns might represent a unique “fingerprint” that could be used to identify exposed individuals in the population. The individual genes that were affected also led to new hypotheses about each toxic metal’s mechanism of action. Those hypotheses are now under further investigation by Hamilton and his group. More detailed investigations of arsenic’s effects further supported the notion that these patterns were both unique and diagnostic, and also could be used to generate new mechanistic hypotheses. In one experiment, a low dose of arsenic (where there were no obvious signs of toxicity) was compared to a higher dose (where there was obvious toxicity and cell stress). The two patterns produced were almost completely unique, almost as if they represented two different chemicals. This was somewhat surprising and has interesting implications. One of the founding principles of toxicology is that “the dose makes the poison.” This appears to be the case with arsenic and gene expression, suggesting that it is not simply that higher doses do the same thing as lower doses, only more so, but that there is a different biological response to low doses than to higher doses. This further suggests that one might see different patterns of disease in different populations if their levels of exposure are different, for example, comparing the very high doses or arsenic exposure in Bangladesh — where there are obvious clinical signs of arsenic poisoning — with the much lower but still elevated doses more typically found in the U.S. where there is rarely any clinical sign of arsenic toxicity. Another interesting difference in patterns was seen in a series of experiments in mice given arsenic in drinking water at doses similar to the elevated levels typically encountered in the U.S. Hamilton and colleagues found that there were differences in the patterns of response at different doses, at different times after initial exposure, and in different tissues of the same animal. This may be why arsenic can cause so many different diseases in people, by altering different genes in different exposure settings. This will require much further study in order to understand these patterns and their implications, and is the other principal focus of the current work. Consequences of Exposure: Frogs, Fish, Water Fleas and People The Hamilton laboratory is performing other experiments and is also collaborating with other groups within the Dartmouth Toxic Metals Program in order to more fully understand the health consequences of arsenic exposure. One project in the Hamilton lab is examining the effects of low-level arsenic exposure on frog tadpole development. The rapid metamorphosis of frogs from a water-swimming, gill-breathing, fish-like tadpole to a land-based, lung-breathing frog is a remarkable process that is tightly regulated by hormones, in particular the thyroid hormone, T3. Since this hormone receptor pathway has been shown by the Hamilton lab to be perturbed by arsenic, they are currently investigating the effects of arsenic on frog morphogenesis in order to determine the developmental consequences of such exposure. Preliminary results indicate that tadpole development is profoundly affected by low levels of arsenic. If this turns out to be the case, this has important implications for human fetuses who may be exposed to arsenic through their mothers during development. A second project, in collaboration with Bruce Stanton’s laboratory at Dartmouth, is examining the effects of arsenic on killifish physiology. Killifish (or mumichogs, Fundulus heteroclitus) have a remarkable ability to go from freshwater to saltwater within hours. Most other fish are adapted to one environment or the other but cannot rapidly transition. But killifish can increase expression of a protein in their gills that helps them adjust and maintain their internal salt concentrations, called CFTR. The CFTR protein is the same in killifish as in humans. Mutations in the CFTR gene, leading to mutated forms of the protein, are responsible for the human genetic disease, cystic fibrosis, making this is an excellent model to study this disease. The ability of killifish to change expression of CFTR in their gills is regulated by the glucocoorticoid receptor, the same steroid receptor that Hamilton’s group showed is highly sensitive to arsenic. Interestingly, killifish have the same response to arsenic when exposed in either freshwater or in seawater. But if killifish that are exposed to arsenic in freshwater are then placed in seawater, they are unable to adapt to the new environment, and they die. Hamilton and Stanton have hypothesized that this is primarily because of the effects of arsenic on the glucocorticoid receptor. |
