Cytokines and other immunological biomarkers in children's environmental health studies

Abstract

Environmental exposures (e.g. pesticides, air pollution, and environmental tobacco smoke) during prenatal and early postnatal development have been linked to a growing number of childhood diseases including allergic disorders and leukemia. Because the immune response plays a critical role in each of these diseases, it is important to study the effects of toxicants on the developing immune system. Children's unique susceptibility to environmental toxicants has become an important focus of the field of immunotoxicology and the use of immune biomarkers in molecular epidemiology of children's environmental health is a rapidly expanding field of research. In this review, we discuss how markers of immune status and immunotoxicity are being applied to pediatric studies, with a specific focus on the various methods used to analyze T-helper-1/2 (Th1/Th2) cytokine profiles. Furthermore, we review recent data on the effects of children's environmental exposures to volatile organic compounds, metals, and pesticides on Th1/Th2 cytokine profiles and the associations of Th1/Th2 profiles with adverse health outcomes such as pediatric respiratory diseases, allergies, cancer and diabetes. Although cytokine profiles are increasingly used in children's studies, there is still a need to acquire distribution data for different ages and ethnic groups of healthy children. These data will contribute to the validation and standardization of cytokine biomarkers for future studies. Application of immunological markers in epidemiological studies will improve the understanding of mechanisms that underlie associations between environmental exposures and immune-mediated disorders.

Figure 1

Role of biomarkers in children's environmental studies

Figure 2. Flow cytometric detection of intracellular Th1/Th2 cytokines in childhood leukemia

Th1/Th2 cytokines were detected by flow cytometry (Duramad et al., 2004) in samples from patients with acute lymphoid leukemia (ALL) and acute myeloid leukemia (AML). Whole blood stimulated with PMA/ionomycin was stained with cytokine-specific antibodies (IFN -γ-FITC for T-helper-1 cells and IL-4-PE for T-helper-2 cells). In the scatter plot of all cells (A), a circular gate was placed around the live lymphocyte population, based on size and granularity. The granulocyte population is more pronounced in the AML sample. Of total lymphocytes, only the CD4+ T-helper cells were selected (right peak, B). These T-helper cells were examined for Th1/Th2 cytokine production (C), with cells that stained positive for IFN-γ only (lower right quadrant) classified as Th1 cells and those that stained positive for IL-4 only (upper left quadrant) classified as Th2 cells.

Figure 3. Th1/Th2 cytokines in patients with childhood acute lymphoid leukemia (ALL) and acute myeloid leukemia (AML)

Intracellular Th1/Th2 cytokines were detected by flow cytometry (Duramad et al, 2004a) in a subset of pediatric peripheral blood samples (n=32, age 0-14 years) collected by the Northern California Childhood Leukemia Study (Buffler et al., 2005). Acute lymphoid leukemia (ALL, n=26) subjects had consistently higher %Th1 but slightly lower %Th2 compared to subjects diagnosed with acute myeloid leukemia (AML, n=6), however these differences were not statistically significant (Duramad et al. 2004b). Females (n=20) and males (n=12) had similar %Th1 (1.6 vs. 2.0; p=0.8), and %Th2 (0.3 vs. 0.6; p=0.2).

Figure 4. Changes in Th1 and Th2 with age

Intracellular Th1/Th2 cytokines were detected by flow cytometry (Duramad et al, 2004) in a subset of peripheral blood samples (N=39 for 12 months old, N=239 for 24 months old, and N=153 for 60 months old) from Latino children enrolled in CHAMACOS study (Eskenazi et al. 2003). A statistically significant increase in Th1 and Th1/Th2 ratio but decrease in Th2 was observed between ages 12 and 60 months. (* p<0.05 and ** p<0.001).