Faculty Bibliography

Early pregnancy is a powerful negative risk factor for breast cancer (BCa) in women. Pregnancy also protects rats against induction of BCa by carcinogens such as N-methyl-N-nitrosourea (MNU), making the parous rat a useful model for studying this phenomenon. Smoking during early pregnancy may lead to an increased risk of BCa in later life, possibly attributable to carcinogens in cigarette smoke (CS), or to reversal of the parity-related protection against BCa. To investigate these possibilities, 50-day-old timed first-pregnancy rats were exposed to standardized mainstream CS (particle concentration = 50 mg/m3) or to filtered air (FA) 4 h/day, Day 2-20 of gestation. Age-matched virgin rats were similarly exposed to CS or FA. At age 100 days, the CS or FA-exposed, parous and virgin rats were injected s.c. with MNU (50 mg/kg body wt), or with MNU vehicle. Mammary tumors (MTs) first appeared in virgin rats 9 weeks post-MNU injection. While no MTs were detected in FA-exposed parous rats until 18 weeks post-MNU, MTs appeared in the CS-exposed parous rats as early as 10 wks (P < 0.02). As no MTs developed in CS-exposed rats not injected with MNU, CS did not act as a direct mammary carcinogen. Serum prolactin concentration on Day 19 of pregnancy in CS-exposed dams was reduced by 50% compared with FA-exposed dams (P < 0.005). CS exposure during a pregnancy may thus 'deprotect' rats, enhancing their vulnerability to MNU-induced BCa. Prenatal CS exposure had no detectable effect on the immune responses of the pups examined at 3, 8 or 19 weeks of age. However, prolactin concentration in stomach contents (milk) of 3-day-old pups suckled by CS-exposed dams was decreased when compared with that of FA-exposed dams (P < 0.032). As milk-borne prolactin modulates development of the central nervous and immune systems of neonatal rats, CS exposure of the dams could adversely affect later maturation of these systems by reducing milk prolactin

Abundant literature exists demonstrating the immunomodulating effects of polychlorinated biphenyls (PCBs). To date, most of the research has focused on dioxin-like coplanar PCB congeners because of their high affinity for the aryl hydrocarbon receptor (AhR) and cytochrome P450-inducing capability. For this study, the impact of two structurally different PCB congeners on the immune responsiveness of bluegill sunfish (Lepomis macrochirus) was examined to evaluate the immunotoxic potential of each congener (as separate entities) and to relate effects on immune function with hepatic CYP1A induction. Fish received a single intraperitoneal injection of the: coplanar congener, PCB 126 (0.01 or 1.0 mug/g BW); noncoplanar PCB 153 (5.0 or 50.0 mug/g BW); or, the corn oil vehicle. PCB-induced effects on innate and cell-mediated immune parameters, and on hepatic CYP1A protein induction were evaluated in fish sacrificed 1, 3, 7, 14 or 21 days post-injection. In the absence of CYP1A induction, PCB 153 increased kidney phagocyte-mediated superoxide production 3 d post-injection, and at the highest dose suppressed B- and T-lymphocyte proliferation after 3 and 7 days, respectively. Treatment of fish with PCB 126 had no effect on oxyradical production, but altered B-lymphocyte proliferation after 1 day, also in the absence of CYP1A induction. Hepatic CYP1A was only induced in fish exposed to the highest PCB 126 dose; protein induction appeared at 3 d post-injection and persisted for up to 21 days. Taken together, these results demonstrate that exposure to different PCB congeners can alter immune function in the absence of CYP1A induction, suggesting that mechanisms other than the AhR pathway may play a role in PCB-induced immunotoxicity, particularly for the noncoplanar congeners

Epidemiologic evidence suggests that prenatal exposure to intact (unfractionated) cigarette smoke (CS) increases the incidence of cancer in the offspring. A toxicology study was carried out to examine the effects and underlying mechanisms of prenatal exposure to mainstream cigarette smoke (MCS) on offspring resistance to tumor challenge and surveillance mechanisms critical for the recognition and destruction of tumors. Pregnant B6C3F1 mice were exposed by inhalation to MCS for 5 days/week (4 h/day from gestational day 4 to parturition). Smoke-induced effects on offspring-host resistance to transplanted tumor cells; natural killer (NK) cell and cytotoxic T-lymphocyte (CTL) activity; cytokine levels; lymphoid organ immune cell subpopulations; and histology-were examined in 5-, 10- and 20-week-old male and female offspring. At a concentration of smoke roughly equivalent to smoking <1 pack of cigarettes/day, prenatally exposed male offspring challenged at 5 week of age with EL4 lymphoma cells demonstrated a greater than two-fold increase in tumor incidence (relative to age-/gender-matched air-exposed offspring); tumors in prenatally smoke-exposed pups also grew significantly faster. Cytotoxic T-lymphocyte activity in the smoke-exposed 5- and 10-week-old male pups was significantly less than that of the age- and gender-matched controls. No effects of prenatal CS exposure were observed on offspring NK activity, cytokine levels, lymphoid organ histology, or immune cell subpopulations. Results demonstrated that exposure of pregnant mice to a relevant dose of MCS decreased offspring resistance against transplanted tumor cells and persistently reduced CTL activity in prenatally exposed pups. This study provides biological plausibility for the epidemiologic data indicating that children of mothers who smoke during pregnancy have a greater risk of developing cancer in later life

Exposure to the environmental contaminant benzo[a]pyrene (BaP) results in suppression of immune function in both mammalian and fish species. This laboratory has previously demonstrated that a single intraperitoneal (IP) injection of BaP reduced lymphocyte proliferation, phagocyte-mediated superoxide generation, and antibody-forming cell (AFC) numbers in Japanese medaka (Oryzias latipes). The objective of the current study was to determine the role of BaP metabolism in the observed immunosuppression. Results from rodent studies have suggested that BaP elicits its immunotoxic effects via upregulation of cytochrome P4501A1 (CYP1A1) and the subsequent production of immunosuppressive BaP metabolites. In this study, exposure of medaka to 200 microg BaP/g BW significantly induced CYP1A expression or activity within lymphoid tissue 48 h post-IP injection; induction was observed specifically within distinct subpopulations of kidney mononuclear cells. Concurrent injection of fish with BaP and the CYP1A1 inhibitors alpha-naphthoflavone (ANF) or dehydroepiandrosterone (DHEA) resulted in inhibition of renal EROD activity and amelioration of BaP-induced suppression of medaka AFC numbers. Results of this study suggest that (1) BaP-induced suppression of medaka humoral immunity relies upon the CYP1A-catalyzed production of immunotoxic BaP metabolites and (2) BaP metabolites may be created in situ, directly by specific cells within kidney lymphoid tissue. Thus, apparently, mechanisms involved in BaP-induced immunosuppression have been phylogenetically conserved from fish to mammals

Knowledge gained through the use of alternative animal models has significantly enhanced our understanding of life at all levels of biological organization. The discipline of toxicology is under considerable pressure to develop such models due to increasing public concern regarding the experimental use of mammals. Studies in this laboratory have focused on the utility of a small laboratory fish model, the Japanese medaka (Oryzias latipes), to investigate immunotoxicological effects of benzo[a]pyrene (BaP). BaP is a ubiquitous environmental contaminant and known mammalian immunotoxicant. This laboratory has demonstrated that in vivo exposure of medaka to BaP (2-200 microg/g BW) significantly depresses both innate and humoral immunity. Further studies have indicated that BaP activates its own biotransformation pathway within medaka immune cells following both in vivo and in vitro exposure. In addition, reduction of BaP metabolism with alpha-naphthoflavone results in the reversal of BaP-induced suppression of antibody production in vitro. Inhibition of CYPlA-mediated metabolism within medaka immune cells also alleviates the immunotoxicity induced by benzo[a]pyrene-7,8-dihydrodiol, but not benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE). This suggests that BPDE may be an ultimate immunotoxicant. Results from this study in medaka are in agreement with previously conducted rodent studies that indicated a role for immunotoxic BaP metabolites in BaP-induced suppression of humoral immunity

Multiple indicators of sublethal stress (bioindicators) were used to assess the health and condition of two important estuarine fish species in the Pamlico Sound estuary following extensive flooding from three sequential hurricanes that occurred during early fall 1999. Bioindicators ranging from the biochemical to the reproductive and organism-level were used to assess the health of southern flounder and spot in Pamlico Sound compared to the health of these same species sampled from a relatively unaffected reference site in lower Core Sound. Many of the physiological, reproductive, immunological, histopathological, and general condition indices suggested that both species, and particularly spot, in Pamlico Sound were sublethally stressed and in poorer condition than fish sampled from Core Sound. The major environmental stressors causing these sublethal stress responses in Pamlico Sound fish appears to be those related to episodic hypoxic exposure or a combination of effects associated with hypoxic conditions such as alterations in preferred habitat and food availability. Although fish populations in Pamlico Sound do not appear to be severely damaged or impaired at this time, organisms that are sublethally stressed can incur increased vulnerability to additional or future stressors such as modified physicochemical regimes, changes in food and habitat availability, and increases in infectious pathogens. Because of the low flushing rate (similar to1 yr) of Pamlico Sound, recovery rate may be exceptionally slow, prolonging any adverse effects of altered nutrient regimes (such as hypoxia) on the health and fitness of resident fish populations. Flooding from the 1999 hurricanes may have contributed to the short-term health and condition of finfish species in Pamlico Sound and also influenced longer-term recovery and ecological status of this system. Longer-term manifestation of effects from flooding may be of particular concern as the frequency of hurricanes is expected to increase over the next few years and the accelerated uses of the coastal zone places further stress on estuarine resources

Smallmouth bass (Micropterus dolomieu) were collected to quantify the nature and prevalence of biomarker responses, including biochemical indices, toxicopathic lesions and general health indices, among fish collected from polychlorinated biphenyl (PCB)-contaminated and nearby uncontaminated reaches of the Kalamazoo River, Michigan, USA. Blood and tissue samples (gill, liver, spleen, head kidney, trunk kidney, thyroid and gonads) were collected and preserved at necropsy for biochemical and histological analyses. The body condition factor and liver somatic index were significantly lower in fish collected from the downstream, contaminated site. Plasma vitellogenin was not detected in male fish collected from either site. Liver ethoxyresorufin-O-deethylase activity and liver and spleen superoxide dismutase activity were significantly depressed in fish collected from the downstream site. Significant toxicopathic lesions such as glycogen depletion, enhanced macrophage aggregates, hepatic foci of cellular alteration (i.e. preneoplastic lesions) and neoplasia were also detected in the liver of fish collected from the downstream site. This study indicates that many of the biochemical and histopathological biomarker responses were associated with liver and body tissue PCB concentrations. Taken together, the biomarkers of exposure and effect strongly suggest that fish within the downstream site are adversely affected by PCBs and other chemical stressors