Tumor necrosis factor alpha is a key component in the obesity-linked elevation of plasminogen activator inhibitor 1

Abstract

Obesity is associated with a cluster of abnormalities, including hypertension, insulin resistance, hyperinsulinemia, and elevated levels of both plasminogen activator inhibitor 1 (PAI-1) and transforming growth factor beta (TGF-beta). Although these changes may increase the risk for accelerated atherosclerosis and fatal myocardial infarction, the underlying molecular mechanisms remain to be defined. Although tumor necrosis factor alpha (TNF-alpha) has been implicated in the insulin resistance associated with obesity, its role in other disorders of obesity is largely unknown. In this report, we show that in obese (ob/ob) mice, neutralization of TNF-alpha or deletion of both TNF receptors (TNFRs) results in significantly reduced levels of plasma PAI-1 antigen, plasma insulin, and adipose tissue PAI-1 and TGF-beta mRNAs. Studies in which exogenous TNF-alpha was infused into lean mice lacking individual TNFRs indicate that TNF-alpha signaling of PAI-1 in adipose tissue can be mediated by either the p55 or the p75 TNFR. However, TNF-alpha signaling of TGF-beta mRNA expression in adipose tissue is mediated exclusively via the p55 TNFR. Our results suggest that TNF-alpha is a common link between the insulin resistance and elevated PAI-1 and TGF-beta in obesity. The chronic elevation of TNF-alpha in obesity thus may directly promote the development of the complex cardiovascular risk profile associated with this condition.

Figure 1

Changes in PAI-1 expression in ob/ob mice after injection of a neutralizing antibody to TNF-α. Obese (ob/ob) male mice (four each, 14 weeks old) were injected i.p. with a control hamster IgG (black bars) or with a neutralizing hamster mAb against murine TNF-α (gray bars). Six hours later, the plasma (A) or adipose tissue (B) were collected and analyzed for PAI-1 antigen (A) or mRNA (B) as described in Materials and Methods. The level of PAI-1 antigen in plasma (A) or PAI-1 mRNA (B) in age-matched lean mice (n = 4) also is shown (empty bars). Error bars represent mean ± SD.

Figure 2

Effect of deletion of both TNFRs on PAI-1 expression in ob/ob mice. Plasma and adipose tissue were collected from 20- to 24-week-old male and female ob/ob mice and ob/ob mice lacking both TNFRs (p55^−/−, p75^−/−). Plasma PAI-1 antigen (A) or adipose tissue PAI-1 mRNA (B) were analyzed as described in Materials and Methods. (A) n = 6 for each group; bars represent mean ± SD. Comparison of ob/ob vs. ob/ob p55^−/−, p75^−/−: P < 0.01. (B) Error bars represent the mean for each group. Comparison of ob/ob vs. ob/ob p55^−/−, p75^−/−: P < 0.0009.

Figure 3

Effect of exogenous TNF-α on PAI-1 expression in lean wild-type and TNFR-deficient lean mice. Lean mice or TNFR-deficient lean mice (four each, 20–24 weeks old) were injected i.p. with 4 μg of murine recombinant TNF-α (open bars) or saline (filled bars). Three hours later, the plasma (A) or adipose tissue (B) were collected and analyzed for PAI-1 antigen (A) or mRNA (B) as described in Materials and Methods. Error bars represent mean ± SD.

Figure 4

Effect of TNF-α on the cellular localization of PAI-1 mRNA in wild-type and TNFR-deficient mice. In situ hybridization was performed on paraffin sections of adipose tissues from wild-type untreated mice (A) and TNF-α-treated wild-type (B), p55,p75 TNF-deficient (C), p55 TNFR-deficient (D), and p75 TNFR-deficient (E) mice. Slides were exposed for 4 weeks at 4°C and then stained with hematoxylin and eosin. a, adipocytes. Arrowheads indicate positive signals for PAI-1 mRNA in the fat (B, D, and E). Magnification: ×400 for all sections.

Figure 5

Effect of exogenous TNF-α treatment on TGF-β mRNA expression in lean wild-type and lean TNFR-deficient mice. Lean wild-type mice or TNFR-deficient lean mice (20–24 weeks old) were injected i.p. with 4 μg of murine recombinant TNF-α (open bars) or saline (filled bars). Three hours later, adipose tissues were collected and analyzed for TGF-β mRNA levels as described in Materials and Methods. n = 3, error bars represent ± SD.

Figure 6

Effect of TNF-α on the cellular localization of TGF-β mRNA in wild-type and TNFR-deficient mice. In situ hybridization was performed on paraffin sections of adipose tissues from wild-type untreated mice (A and B) and TNF-α-treated wild-type (C), p55,p75 TNFR-deficient (D), p55 TNFR-deficient (E), and p75 TNFR-deficient (E) mice. Slides were exposed for 8 weeks at 4°C and stained with hematoxylin and eosin. a, adipocytes. Arrowheads indicate positive signals for TGF-β mRNA in the fat. Magnification: ×400 for all sections.

Figure 7

TNF-α and cardiovascular disease in obesity. TNF-α is chronically elevated in adipose tissue in obesity and contributes to the insulin resistance/hyperinsulinemia and elevated TGF-β associated with this condition. TNF-α, insulin, and TGF-β may, in turn, lead to elevated PAI-1, tissue factor, and possibly other hemostatic genes that may promote cardiovascular risk.