A critical issue affecting the interpretation of upstream events is the relationship between biomarkers captured in clinical or animal studies and specific adverse outcomes. Studies involving upstream biomarkers are most useful when these biomarkers have been causally linked to downstream adverse outcomes. For example, interpreting studies of perchlorate and T4
are relatively straightforward because the only known toxic effect of perchlorate is interference with thyroid function (National Research Council 2005
); thus, any effects of perchlorate on the nervous system are necessarily interpreted to be subsequent to a reduction in serum THs.
Difficulties can arise when attempting to predict changes in upstream biomarkers based on adverse outcomes. For example, if the adverse outcome(s) of a specific toxicant or mixture is caused by more than one mechanism, then individual downstream outcomes (i.e., “effects”) are not diagnostic of upstream events, and causative links between a known exposure and outcome are difficult to discern. illustrates this by the alternative mechanisms activated by chemical X that may cause similar adverse outcomes. Indeed, some of these adverse outcomes may be caused by exposure to other chemicals (chemical Z). A key to using adverse outcomes in these cases is the use of patterns of outcomes that may be diagnostic.
PCBs offer a good example of the problems associated with inferring upstream changes in THs as the causative agent of downstream neurotoxic outcomes. PCBs produce changes in a number of behavioral domains in humans and animals (Rice 2000
; Schantz et al. 2003
). They also affect multiple neurochemical pathways (Kodavanti et al. 1993
; Kodavanti and Ward 1998
; Seegal 1996
; Seegal et al. 1991
) in addition to TH (Crofton and Zoeller 2005
). Although changes in THs during development predict specific behavioral changes, effects of PCBs on some specific tasks in animals or outcomes in epidemiologic studies may not necessarily be attributable to changes in THs.
Another example of the difficulty in linking serum TH to adverse outcomes is provided by the recent observation in humans of an abnormal TH profile in boys with a genetic mutation in the T3
-specific transporter mono-carboxylate anion transporter 8 (MCT8
). In all cases, serum T3
is elevated, but serum T4
, free T4
, and TSH may be low, normal, or elevated (Jansen et al. 2007
). Thus, the elevated serum T3
appears to be a biomarker of the MCT8 mutation among the patients evaluated, although it is not the only mechanism by which T3
can become elevated. In addition, all of the boys evaluated presented with severe psychomotor deficits, but it is unlikely that the elevated serum T3
itself was the root cause of their condition. Thus, environmental factors that influence T3
transport through MCT8 may represent a situation in which the profile of serum TH hormones is perturbed in ways that are not immediately recognizable as due to an endocrine disruptor, but may signal that adverse effects occur through a mechanism that interferes with TH signaling.
Recognition of the role of “critical windows of exposure” in characterizing causal relationships between toxicant effects on serum THs and downstream adverse effects is critical. Specifically, the role of TH in brain development changes as development proceeds (Zoeller and Rovet 2004
). Therefore, to establish a causal role of toxicant-induced low TH in the mechanism of neurotoxicity, it is important to show that T4
replacement can reverse the effects of toxicant. However, it is important to be cognizant of the relevant “windows” of vulnerability in the design of these experiments. For example, the impact of TH disruption on the development of auditory function in rats correlates well with circulating T4
levels during the second postnatal week (Crofton 2004
). This is entirely consistent with the known role of THs in auditory development (Uziel et al. 1981
), the critical postnatal ontogeny of auditory function (Rubel 1978
), and the pharmacokinetics of the chemicals tested (Crofton and Zoeller 2005
). In addition, this correlation establishes a prognostic power of early postnatal T4
for adverse consequence of developmental exposure to TDCs in rats (Crofton 2004
). An understanding of the role of THs in development, coupled with hormone level measurement during the critical window, allows the establishment of a developmental mode of action that assigns a key causative role to TH disruption in the adverse outcome ().
Studies designed to test for associations between toxicant exposures and circulating levels of TH in humans require careful consideration of confounding variables. For example, blood levels of TH vary among individuals (Andersen et al. 2002
), which will affect the number of samples required for such a study to be sufficiently powered to identify associations of interest. In the case of newborn TH levels, a number of maternal, infant, and delivery factors influence TH levels in cord blood and in infant serum (Herbstman et al. 2008
), and these must be carefully considered when attempting to identify associations between toxicant exposures and serum TH levels. A good recent example is that of Herbstman et al. (2008)
, who showed that PCB measures in cord blood were associated with circulating levels of TH only in those babies born via an unassisted vaginal delivery. Thus, these confounding variables may explain the studies in which PCB body burden has not been found to be associated with THs.