Expression and regulation of soluble epoxide hydrolase in adipose tissue

Abstract

Obesity is an increasingly important public health issue reaching epidemic proportions. Visceral obesity has been defined as an important element of the metabolic syndrome and expansion of the visceral fat mass has been shown to contribute to the development of insulin resistance and cardiovascular disease. To identify novel contributors to cardiovascular and metabolic abnormalities in obesity, we analyzed the adipose proteome and identified soluble epoxide hydrolase (sEH) in the epididymal fat pad from C57BL/6J mice that received either a regular diet or a "western diet." sEH was synthesized in adipocytes and expression levels increased upon differentiation of 3T3-L1 preadipocytes. Although normalized sEH mRNA and protein levels did not differ in the fat pads from mice receiving a regular or a "western diet," total adipose sEH activity was higher in the obese mice, even after normalization for body weight. Furthermore, peroxisome proliferator-activated receptor gamma (PPARgamma) agonists increased the expression of sEH in mature 3T3-L1 adipocytes in vitro and in adipose tissue in vivo. Considering the established role for sEH in inflammation, cardiovascular diseases, and lipid metabolism, and the suggested involvement of sEH in the development of type 2 diabetes, our study has identified adipose sEH as a potential novel therapeutic target that might affect the development of metabolic and cardiovascular abnormalities in obesity.

Figure 1

Tissue soluble epoxide hydrolase expression. (a) Soluble epoxide hydrolase (sEH) mRNA levels and (b,c) activity for liver, kidney, and epididymal fat pad. (a) All data are expressed relative to liver standard fat diet (SFD). Diet had no effect on sEH mRNA levels (P > 0.05 for each organ). (b,c) sEH activity is expressed per mg protein (b) and for the total organ after normalization for body weight (c). * P< 0.001 vs. liver for the same diet; **P< 0.01 vs. kidney for the same diet; ^†P< 0.01 vs. SFD. n = 5.

Figure 2

Soluble epoxide hydrolase (sEH) in 3T3-L1 cells. (a) Differentiation of 3T3-L1 preadipocytes was analyzed by Oil Red O staining at days 0, 2, 4, and 7 (left to right). Bar = 50 µm. Western blotting for perilipin A and B and sEH indicated that sEH protein levels increased during preadipocyte differentiation ((b), densitometric analysis (rdu, relative densitometric units); (c) representative bands for perilipin A, perilipin B, sEH, and control actin). n = 3. (d) aP2, sEH, and Pref-1 mRNA levels determined by real-time quantitative PCR indicated that sEH mRNA levels increased during preadipocyte differentiation. n = 6.

Figure 3

Troglitazone regulates soluble epoxide hydrolase (sEH) expression in mature 3T3-L1 adipocytes. (a,b) sEH mRNA levels in 3T3-L1 adipocytes determined by real-time quantitative PCR during incubation with troglitazone (a, 10 µmol/l; b, 48 h). n = 6. (c–f) Western blotting of sEH and control actin in 3T3-L1 adipocytes during incubation with troglitazone (c,e, 10 µmol/l; d,f, 48 h) (rdu, relative densitometric units). (e,f) Western blots used for quantitation in c and d, respectively. n = 3. ^†P< 0.05; *P< 0.01; **P< 0.001 vs. baseline.

Figure 4

Rosiglitazone increases soluble epoxide hydrolase (sEH) expression in adipose tissue. (a) sEH mRNA levels and (b) activity were increased in the epididymal fat pad of obese C57BL/6J mice by rosiglitazone. (c) Rosiglitazone also increased sEH expression in subcutaneous fat whereas levels of kidney and liver sEH mRNA were not influenced. n = 5. *P< 0.05 vs. vehicle; **P< 0.05 vs. epididymal fat.