It has long been recognized that the high lipophilicity of PCBs and related toxic compounds favors their localization to adipose tissue. However, their effects on adipocyte function have not been established. Our results suggest that lower concentrations of coplanar PCBs, acting as ligands of the AhR, promote adipocyte differentiation and increase the expression of proinflammatory adipokines. In contrast, higher concentrations, similar to TCDD, inhibit adipocyte differentiation. Importantly, when administered to WT, but not AhR-deficient mice, PCB-77 caused an increase in body weight gain. These results confirm in vitro findings and suggest that effects of PCB-77 are AhR mediated. In hypercholes-terolemic ApoE−/− mice, PCB-77 increased body weight associated with adipocyte hypertrophy, expanded adipose mass, and increased serum cholesterol concentrations and ectopic lipid deposition. These effects of PCB-77 were associated with increased atherosclerosis. These results suggest that exposure to PCB-77, a dioxin-like PCB, at relatively low levels may promote the development of obesity and obesity-associated atherosclerosis.
PCBs are highly lipophilic, with octanol:water partition coefficients of ≥ 104
, making their accumulation in nonlipid material negligible. In studies aimed at determining the congener-specific distribution of PCBs with chronic exposure, adult rats were treated five times per week for 4 weeks by gavage with Aroclor 1254 in corn oil (Kodavanti et al. 1998
). Total PCB (parts per milllion) accumulation in fat was 551 μg/g; the second-highest tissue accumulation of PCBs was in liver (38 μg/g). The mean blood:liver:fat tissue ratios were 1:22:359, similar to previously observed results for PCB-153 (Wyss et al. 1986
) or 2,2′3′,4,4′,5,5′-heptachlorobiphenyl (PCB-180) (Koss et al. 1993
). In the present study, adipocyte differentiation and proinflammatory adipokine expression were induced in 3T3-L1 adipocytes by PCB-77 but not by PCB-153. These PCB congeners differ slightly in their oil:water partition coefficients, favoring greater lipophilicity of PCB-153; however, both PCBs would be anticipated to accumulate in adipose tissue. A lack of effect of PCB-153 may have resulted from greater sequestration of this more-lipophilic PCB in the triacylglycerol droplet of the adipocyte, leaving less PCB-153 available to act at adipocyte target proteins. However, given that PCB-77 and TCDD exhibited similar effects and that TCDD has a high octanol:water partition coefficient comparable to PCB-153 (6.7 vs. 6.8, respectively), other mechanisms most likely mediated differences in effects of PCB-77 and PCB-153. Specifically, results from this study demonstrate that interactions with the AhR, for which both PCB-77 and TCDD possess affinity (but PCB-153 has low affinity), contributed to differences in the effects of these PCBs.
Phillips et al. (1995)
demonstrated that treatment of 3T3-L1 preadipocytes with 10 nM TCDD during the first 2 days of induction of differentiation resulted in a reduction in the number of fat cell colonies. The effect of TCDD to inhibit adipocyte differentiation was blocked by treatment with an AhR antagonist. To extend these findings to the in vivo
situation, Brodie et al. (1996)
demonstrated that a single high dose (175 μg/kg) of TCDD administered to rats resulted in inhibition of adipocyte differentiation. Additional studies by this group demonstrated that high-dose TCDD treatment in rats resulted in a reduction in preadipocyte differentiation to mature adipocytes, which was associated with decreases in transcription factor mRNAs (PPARγ, aP2, C/EBPβ) normally elevated during adipocyte differentiation (Brodie et al. 1997
). Using the 3T3-L1 adipocyte differentiation system, Shimba et al. (1998)
demonstrated that the level of AhR protein decreased with ongoing adipocyte differentiation. Using mouse embryo fibroblasts, Alexander et al. (1998)
demonstrated that treatment with 10 nM TCDD inhibited differentiation in cells from WT but not AhR−/−
mice. At high concentrations (approximately 10-fold greater than the dissociation constant), the literature and data from this study demonstrate that TCDD decreases adipocyte differentiation, commensurate with the wasting syndrome demonstrated from high-dose toxicity.
Several epidemiologic studies have suggested a link between exposure to dioxin, or PCBs, and diabetes (Bertazzi et al. 1997
; Calvert et al. 1999
; Codru et al. 2007
; Cranmer et al. 2000
; Lee et al. 2006
; Vasiliu et al. 2006
). Most of these studies represent findings from type 2 diabetics with obesity; however, increased relative risk for exposure to toxic compounds and the development of diabetes remains when data were adjusted for differences in body mass index across study populations. Interestingly, the association between persistent organic pollutants and diabetes was much stronger in obese subjects compared with lean subjects (Lee et al. 2006
). Moreover, in nondiabetic adults, results from the National Health and Nutrition Examination Survey (1999–2002) (Lee et al. 2007b
) demonstrated a linear positive relationship between serum concentrations of PCBs, including dioxin-like PCBs, and waist circumference. Unfortunately, similar epidemiologic studies have not been performed to define whether exposure to PCBs increases the risk for development of obesity or obesity-associated cardiovascular disease, including atherosclerosis.
Obesity, a condition that would predictably increase the body burden of lipophilic PCBs, is associated with an elevation in the systemic concentrations of a variety of factors that are produced and released from adipocytes (Lago et al. 2007
; Lau et al. 2005
). Many of these factors have been linked to diseases clustering around an obesity phenotype, including coronary artery disease, the primary cause of death in the obese population. Thus, factors that regulate adipokine secretion from adipocytes may influence not only obesity but also obesity-associated atherosclerosis. The present results demonstrate that PCB-77 promotes the expression and secretion of a variety of proinflammatory adipokines from 3T3-L1 adipocytes and decreases the expression of adiponectin, an anti-inflammatory adipokine. Although previous studies have demonstrated proinflammatory effects of PCBs in various cell types (Choi et al. 2003
; Eum et al. 2006
; Hennig et al. 1999
; Ramadass et al. 2003
), to our knowledge, this is the first report demonstrating that PCBs can promote the production and elaboration of these factors from adipocytes. Interestingly, Vogel et al. (2007)
recently reported that a single injection of TCDD to C57BL/6 mice resulted in an increase in MCP-1 and KC-1 in liver and adipose tissue. However, enhanced expression of these adipokines in adipose tissue was associated with increased expression of the macrophage marker F4/80, suggesting that this effect may have resulted from enhanced macrophage infiltration into adipose tissue. Our results extend these findings by demonstrating that PCB-77 can act directly on 3T3-L1 adipocytes to promote the mRNA abundance and secretion of several pro-inflammatory adipokines.
To extend results from in vitro
experiments to an in vivo
model, we administered PCB-77 to WT or AhR-deficient mice or to hyper-cholesterolemic ApoE−/−
mice. In previous studies, C57BL/6 mice that received 30 mg/kg/day of PCB-77 consumed in food for a total of 16 weeks exhibited a reduction in body weight (Goodwill et al. 2007
). Thus, chronic dosing with high doses of PCB-77 appears to mimic the wasting syndrome that results from toxic TCDD exposure. In the present study, we injected mice four times (49 mg/kg per dose) with PCB-77 over a 6-week period; although this was a higher daily dose than used by Goodwill et al. (2007)
, it was far less cumulative. Our choice of PCB-77 dose was based on previous studies demonstrating that this dose exhibits proinflammatory effects on endothelial cells when injected into mice (Hennig et al. 2002b
) and is classified as a moderate exposure dose in experimental animals (Brouwer et al. 1999
; Jensen et al. 2001
; Wassermann and Wassermann 1979
). In agreement, PCB-77 levels in adipose tissue of mice from this study were far lower than those reported in adipose tissue from rats administered Arochlor [55 vs. 551 μg/g; (Kodavanti et al. 1998
)]. Our results demonstrate that in contrast to high-dose PCB-77 dosing in vivo
, lower doses exhibit an opposite effect to increase body weight. Moreover, these effects were AhR mediated. The ability of PCB-77 to increase adipose mass with associated adipocyte hypertrophy may relate to the in vitro
effects of PCB-77 to promote adipocyte differentiation. Alternatively, the ability of PCB-77 to increase CD36 mRNA abundance in adipocytes may have promoted lipid uptake and contributed to adipocyte hypertrophy.
Previous investigators demonstrated that dietary exposure to PCBs results in fatty liver and hypercholesterolemia in rats (Kato et al. 1980
; Nagaoka et al. 1990
; Quazi et al. 1983
). These effects were primarily attributed to an increase in hepatic cholesterol synthesis. In the present study, a low dose of PCB-77 resulted in a marked increase in serum cholesterol concentrations in ApoE−/−
mice, with predominant increases in VLDL cholesterol. These results extend previous findings by demonstrating that in a mouse model exhibiting hyper-cholesterolemia and atherosclerosis, marked elevations in serum cholesterol concentrations are induced by PCB-77. Moreover, elevations in VLDL cholesterol by PCB-77 in this study were associated with increased atherosclerosis. To our knowledge, this is the first study that has directly examined the effects of PCB exposure on experimental atherosclerosis.
In conclusion, at low exposure levels, coplanar PCB-77 promoted adipocyte differentiation and proinflammatory adipokine expression. In contrast, both PCB-77 and TCDD inhibited adipocyte differentiation at higher concentrations. Effects of PCB-77 to promote adipocyte differentiation and regulate body weight were AhR mediated. Importantly, when administered in vivo to ApoE−/− mice at a moderate dose, PCB-77 resulted in an increase in body weight, adipose mass and adipocyte area, serum cholesterol concentrations, and atherosclerosis. These results suggest that low-level exposure to coplanar PCBs may contribute to the development of obesity and to obesity-associated atherosclerosis.