A) Adipocyte cell area in epididymal (EWAT), subcutaneous (SWAT) and mesenteric (MWAT) adipose tissue on high-fat diet (HFD). The areas of adipocytes were determined histologically. The values are based upon 200 measurements from each animal. B) Mass of the adipose depots on HFD. 14–15 animals were used for depot weight determination and 6–7 animals for adipocyte size determination. Cell size and depot mass differed over time in all three depots (p<0.05, Kruskall-Wallis analysis, 4 degrees of freedom). *Difference in cell size or depot mass between consecutive time points determined by Mann-Whitney analysis (p<0.05 adjusted to 0.0125 by Bonferroni correction). Error bars indicate standard deviation. Cell size - EWAT: time-point 0–1 weeks U(7) = 6, p = 0.02; 1–6 weeks U(7) = 24, p = 0.94; 6–9 weeks U(7,6) = 16, p = 0.47; 9–12 weeks U(6,7) = 11, p = 0.15; SWAT: time-point 0–1 weeks U(7) = 7, p = 0.02; 1–6 weeks U(7) = 16, p = 0.27; 6–9 weeks U(7,6) = 17, p = 0.56; 9–12 weeks U(6,7) = 9, p = 0.09; MWAT: time-point 0–1 weeks U(7) = 6, p = 0.02; 1–6 weeks U(7,6) = 11, p = 0.15; 6–9 weeks U(6,6) = 10, p = 0.2; 9–12 weeks U(6,7) = 3, p = 0.01; Depot mass - EWAT: time-point 0–1 weeks U(15) = 41, p = 0.003; 1–6 weeks U(15) = 92, p = 0.39; 6–9 weeks U(15,14) = 86.5, p = 0.42; 9–12 weeks U(14,15) = 51, p = 0.02; SWAT: time-point 0–1 weeks U(15) = 26, p = 0.0003; 1–6 weeks U(15) = 108, p = 0.85; 6–9 weeks U(15,14) = 92, p = 0.57; 9–12 weeks U(14,15) = 31, p = 0.001; MWAT: time-point 0–1 weeks U(15) = 29, p = 0.0005; 1–6 weeks U(15,14) = 83, p = 0.33; 6–9 weeks U(14,14) = 60, p = 0.08; 9–12 weeks U(14,15) = 71, p = 0.13.

Proportions of total fatty acids in epididymal adipose tissue at 0 and 12 weeks on high-fat diet: A) Saturated fatty acids (SFA) monounsaturated fatty acids (MUFA), linoleic acid (18∶2 ω-6) and α-linolenic acid (18∶3 ω-3). B) Low abundance omega-6 (ω-6) and omega-3 (ω-3) fatty acids. Error bars indicate standard deviation. *p<0.05 (Mann–Whitney analysis). SFA U(8) = 23, p = 0.34; MUFA U(8) = 14, p = 0.06; 18∶2 ω-6 U(8) = 0, p = 0.0007; 18∶3 ω-3 U(8) = 0, p = 0.0007; 18∶3 ω-6 U(8) = 0, p = 0.0007; 20∶3 ω-6 U(8) = 0, p = 0.0007; 20∶4 ω-6 U(8) = 0, p = 0.0007; 22∶4 ω-6 U(8) = 0, p = 0.0007; 22∶5 ω-6 U(8) = 3, p = 0.0023; 18∶4 ω-3 U(8) = 0, p = 0.0007; 20∶3 ω-3 U(8) = 0, p = 0.0007; 20∶4 ω-3 U(8) = 0, p = 0.0007; 22∶5 ω-3 U(8) = 0, p = 0.0007; 22∶6 ω-3 U(8) = 0, p = 0.0007.

Cell-associated incorporation of [¹⁴C]radiolabeled docosahexaenoic acid (DHA) and oleic acid (OA) in cellular triacylglycerols in the proximal and distal zone of epididymal (EWAT), subcutaneous (SWAT) and mesenteric (MWAT) adipose tissues. Adipose tissue explants from animals on chow diet were incubated in medium containing fatty acids bound to BSA, lipids were extracted and triacylglycerols were isolated by thin layer chromatography. The experiments were performed with 4–5 animals. DHA uptake but not OA uptake differed between depots (p<0.05, Kruskall-Wallis analysis, 3 degrees of freedom). *Difference determined by Mann-Whitney analysis (p<0.05 adjusted to 0.017 by Bonferroni correction). Error bars indicate standard deviation. DHA uptake: MWAT-EWAT proximal U(4,5) = 0, p = 0.014; SWAT-EWAT proximal U(5) = 3, 0.05; EWAT proximal-EWAT distal U(5,4) = 5, p = 0.22; OA uptake: MWAT-EWAT proximal U(4,5) = 3, p = 0.08; SWAT-EWAT proximal U(4,5) = 9, 0.80; EWAT proximal-EWAT distal U(5) = 9, p = 0.46.

Proportions of saturated fatty acids (SFA), monounsaturated fatty acids (MUFA), linoleic acid (18∶2 ω-6) and α-linolenic acid (18∶3 ω-3) in proximal epididymal and distal epididymal (EWAT) adipose tissue, in mesenteric (MWAT), and in subcutaneous (SWAT) adipose tissue on chow feeding. SFA, linoleic acid and α-linolenic acid but not MUFA differed between depots (p<0.05, Kruskall-Wallis analysis, 3 degrees of freedom). *Difference between proximal EWAT and other adipose tissues determined by Mann-Whitney analysis (p<0.05 adjusted to 0.017 by Bonferroni correction). Error bars indicate standard deviation. SFA: MWAT-EWAT proximal U(5) = 0, p = 0.009; SWAT-EWAT proximal U(5) = 1, 0.02; EWAT proximal-EWAT distal U(5) = 12, p = 0.9; MUFA: MWAT-EWAT proximal U(5) = 7, p = 0.25; SWAT-EWAT proximal U(5) = 9, 0.46; EWAT proximal-EWAT distal U(5) = 11, p = 0.75; linoleic acid: MWAT-EWAT proximal U(5) = 0, p = 0.009; SWAT-EWAT proximal U(5) = 0, 0.009; EWAT proximal-EWAT distal U(5) = 10, p = 0.6; α-linolenic acid: MWAT-EWAT proximal U(5) = 0, p = 0.009; SWAT-EWAT proximal U(5) = 2, 0.03; EWAT proximal-EWAT distal U(5) = 7, p = 0.25.

Adipose tissue at the distal end of the depot, limited by the upper dashed line, is denoted distal adipose tissue. Adipose tissue associated with the head and the tail of epididymis, indicated by the lower dashed line, is denoted proximal adipose tissue.

Expression of acetyl-CoA synthetase short-chain family member 2 (Acss2), pyruvate dehydrogenase α1 (Pdha1), pyruvate dehydrogenase β (Pdhb) dihydrolipoamide S-acetyltransferase (Dlat), ATP citrate lyase (Acly), fatty acid synthase (Fasn), long chain elongation enzymes Elovl5 and Elovl6 and stearoyl-CoA desaturase 1 and 2 (Scd1/2) in subcutaneous (SWAT), mesenteric (MWAT) and epididymal (EWAT) adipose tissue. The y-axis scale is the same for all three depots. Gene expression is given as relative expression. N at time-point 0, 1, 6, 9 and 12 weeks were 4, 5, 7, 3, 6 for epididymal adipose tissue (EWAT); 5, 6, 4, 6, 5 for subcutaneous adipose tissue (SWAT); and 3, 5, 4, 5, 7 for mesenteric adipose tissue (MWAT), respectively. Error bars indicate standard deviation. Statistically significant changes in expression over time are indicated by *(p<0.05 (one way ANOVA)). SWAT: Acss2 F(4,21) = 10.1, p = 9.8×10⁻⁵; Pdha1 F(4,21) = 17.9, p = 1.5×10⁻⁶; Pdhb F(4,21) = 15.3, p = 5.3×10⁻⁶; Dlat F(4,21) = 8.8, p = 0.0002; Acly F(4,21) = 30.0, p = 2.0×10⁻⁸; Fasn F(4,21) = 8.8, p = 0.0002; Elovl5 F(4,21) = 2.9, p = 0.04; Elovl6 F(4,21) = 27.5, p = 4.4×10⁻⁸; Scd1 F(4,21) = 0.9, p = 0.47; Scd2 F(4,21) = 2.2, p = 0.10; MWAT: Acss2 F(4,18) = 4.3, p = 0.013; Pdha1 F(4,18) = 9.9, p = 0.0002; Pdhb F(4,18) = 7.2, p = 0.001; Dlat F(4,18) = 4.6, p = 0.009; Acly F(4,18) = 29.9, p = 9.9×10⁻⁸; Fasn F(4,18) = 5.3, p = 0.0.005; Elovl5 F(4,18) = 9.7, p = 0.0002; Elovl6 F(4,18) = 14.4, p = 1.9×10⁻⁵; Scd1 F(4,18) = 2.0, p = 0.14; Scd2 F(4,18) = 0.42, p = 0.79; EWAT: Acss2 F(4,20) = 1.1, p = 0.37; Pdha1 F(4,20) = 1.9, p = 0.19; Pdhb F(4,20) = 2.3, p = 0.09; Dlat F(4,20) = 2.0, p = 0.12; Acly F(4,20) = 1.4, p = 0.25; Fasn F(4,20) = 0.82, p = 0.53; Elovl5 F(4,20) = 1.7, p = 0.28; Elovl6 F(4,20) = 1.2, p = 0.32; Scd1 F(4,20) = 5.1, p = 0.005; Scd2 F(4,20) = 3.4, p = 0.027.

A) De novo lipogenesis determined by incorporation of [¹⁴C] from radiolabeled glucose in triacylglycerols in explants from the proximal and distal zone of epididymal (EWAT), subcutaneous (SWAT) and mesenteric (MWAT) adipose tissue. B) Expression of genes encoding fatty acid synthase (Fasn) and ATP citrate lyase (Acly) determined by RT-PCR. De novo lipogenesis and Fasn expression but not Acly expression differed between depots (p<0.05, Kruskall-Wallis analysis, 3 degrees of freedom). *Difference between proximal EWAT and other adipose tissues determined by Mann-Whitney analysis (p<0.05 adjusted to 0.017 by Bonferroni correction). Experiments were performed with 4–5 animals. Error bars indicate standard deviation. de novo lipogenesis: MWAT-EWAT proximal U(4,5) = 0, p = 0.014; SWAT-EWAT proximal U(5) = 5, p = 0.12; EWAT proximal-EWAT distal U(5) = 0, p = 0.009; Fasn: MWAT-EWAT proximal U(4) = 2, p = 0.08; SWAT-EWAT proximal U(4) = 0, 0.02; EWAT proximal-EWAT distal U(4) = 0, p = 0.02; Acly: MWAT-EWAT proximal U(4) = 2, p = 0.08; SWAT-EWAT proximal U(4) = 2, 0.08; EWAT proximal-EWAT distal U(4,3) = 5, p = 0.72.

A) Gene expression levels of the androgen receptor (Ar) in proximal and distal epididymal (EWAT) adipose tissue, in mesenteric (MWAT), and in subcutaneous (SWAT) adipose tissue determined by RT-PCR. B) Expression of the gene encoding the androgen receptor (Ar) determined by RT-PCR plotted against rate of lipogenesis measured by incorporation of [¹⁴C]glucose in TAG in adipose tissue explants. C) Expression of carbohydrate responsive element binding protein (ChREBP), ATP citrate lyase (Acly) and fatty acid synthase (Fasn) on 12 weeks of high-fat diet determined by microarray analysis. The expression level at baseline was set to 1 for all genes. (A) Ar expression differed between depots (p<0.05, Kruskall-Wallis analysis, 3 degrees of freedom), *Difference between proximal EWAT and other adipose tissues determined by Mann-Whitney analysis (p<0.05 adjusted to 0.017 by Bonferroni correction). MWAT-EWAT proximal U(5) = 0, p = 0.009; SWAT-EWAT proximal U(5) = 1, 0.02; EWAT proximal-EWAT distal U(5) = 3, p = 0.05 (B) the relation between lipogenesis and Ar expression was determined by Pearson correlation coefficient with one-sided significance calculated by Cronbach's Alpha, (C) *p<0.05 (one way ANOVA with Tukey post hoc analysis, 4 degrees of freedom. Error bars indicate standard deviation. SWAT: ChREBP F(4,21) = 10, p = 0.0001; Acly F(4,21) = 30.0, p = 2.0×10⁻⁸; Fasn F(4,21) = 8.8, p = 0.0002; MWAT: ChREBP F(4,18) = 8.6, p = 0.0004; Acly F(4,18) = 29.9, p = 9.9×10⁻⁸; Fasn F(4,18) = 5.3, p = 0.0.005; EWAT: ChREBP F(4,20) = 2.8, p = 0.055; Acly F(4,20) = 1.4, p = 0.25; Fasn F(4,20) = 0.82, p = 0.53.

Androgen suppression of adipose tissue de novo lipogenesis is regulated by androgen receptor (Ar). Ar is highly expressed in proximal epididymal adipose tissue (EWATp). Low rate of de novo lipogenesis causes high content of dietary fatty acids and on chow diet the essential fatty acids linoleic acid and α-linolenic acid accumulate. Delta-6 and delta-5 desaturase (D6D and D5D) in epididymis convert linoleic acid and α-linolenic acid into polyunsaturated fatty acids (PUFA) with 20 and 22 carbon atoms required for spermatogenesis. When animals are fed high-fat diet (HFD) rich in saturated fatty acids adipose tissue is depleted of PUFA, potentially resulting in impaired sperm quality and decreased fertility.