Mitochondrial respiration in subcutaneous and visceral adipose tissue from patients with morbid obesity

Abstract

Adipose tissue exerts important endocrine and metabolic functions in health and disease. Yet the bioenergetics of this tissue is not characterized in humans and possible regional differences are not elucidated. Using high resolution respirometry, mitochondrial respiration was quantified in human abdominal subcutaneous and intra-abdominal visceral (omentum majus) adipose tissue from biopsies obtained in 20 obese patients undergoing bariatric surgery. Mitochondrial DNA (mtDNA) and genomic DNA (gDNA) were determined by the PCR technique for estimation of mitochondrial density. Adipose tissue samples were permeabilized and respirometric measurements were performed in duplicate at 37 degrees C. Substrates (glutamate (G) + malate (M) + octanoyl carnitine (O) + succinate (S)) were added sequentially to provide electrons to complex I + II. ADP ((D)) for state 3 respiration was added after GM. Uncoupled respiration was measured after addition of FCCP. Visceral fat contained more mitochondria per milligram of tissue than subcutaneous fat, but the cells were smaller. Robust, stable oxygen fluxes were found in both tissues, and coupled state 3 (GMOS(D)) and uncoupled respiration were significantly (P < 0.05) higher in visceral (0.95 +/- 0.05 and 1.15 +/- 0.06 pmol O(2) s(1) mg(1), respectively) compared with subcutaneous (0.76 +/- 0.04 and 0.98 +/- 0.05 pmol O(2) s(1) mg(1), respectively) adipose tissue. Expressed per mtDNA, visceral adipose tissue had significantly (P < 0.05) lower mitochondrial respiration. Substrate control ratios were higher and uncoupling control ratio lower (P < 0.05) in visceral compared with subcutaneous adipose tissue. We conclude that visceral fat is bioenergetically more active and more sensitive to mitochondrial substrate supply than subcutaneous fat. Oxidative phosphorylation has a higher relative activity in visceral compared with subcutaneous adipose tissue.

Figure 1. Typical original trace from the oxygraph

Left Y-axis (blue line): oxygen concentration in the chamber. Right Y-axis (red line): oxygen flux. X-axis: time in hours and minutes. Dig: digitonin; Mal+Glu: malate + glutamate; Oct: octanoyl carnitine; Suc: succinate; CytC: cytochrome c.

Figure 2. Respiratory flux rates in visceral (omentum) and abdominal subcutaneous adipose tissue obtained from obese patients during bariatric surgery (laparoscopic Roux-en-Y gastric bypass)

A, respiratory rates expressed per milligram of tissue. B, respiratory rates normalized to mtDNA content (left Y-axis) or normalized to number of cells (right Y-axis). Measurements were done in duplicate and by sequential addition of substrates to the respiratory chambers. Baseline: buffer; GM: +glutamate and malate for feeding into complex I (state 2 respiration); GM_D: +ADP (coupled state 3 respiration); GMO_D: +octanoyl carnitine (medium chain fatty acid); GMOS_D+succinate (for dual input of electrons into complex I + II); FCCP: uncoupled respiration. Data are mean ±s.e.m. *Significant difference (P < 0.05) between regions of fat.

Figure 3

A, respiratory control ratio (RCR) calculated as state 3/state 2 respiration. B, effect of addition of octanoyl carnitine, i.e. substrate control ratio (SCR), calculated as GMO_D/GM_D. C, effect of addition of succinate, i.e. SCR calculated as GMOS_D/GMO_D. D, effect of both octanoyl carnitine and succinate, i.e. SCR calculated as GMOS_D/GM_D. E, uncoupling control ratio (UCR), calculated as maximal uncoupled respiration by FCCP (state 3u) relative to ADP-activated state at maximal oxygen flux (coupled respiration with dual electron input to complex I + II (GMOS_D)). Values increase with limitation of the phosphorylation system. Individual data are shown. Horizontal bar shows mean value. *Significant difference (P < 0.05) between regions of adipose tissue. SCR for succinate (C) tended (P= 0.058) to be different between regions of adipose tissue.

Figure 4

A, detail (magnified frame in inset) of adipocyte (BIOPS protocol, after mechanical dissection) displaying the intracellular content of lipid (L) and a peripherally placed mitochondrion (M). Scale bar: 500 nm. Inset: view of adipocyte with part of an intercellular capillary (C). Lipid (L). Scale bar: 2 μm. B, detail (magnified frame in inset) of adipocyte (BIOPS protocol, after mechanical dissection) displaying the intracellular content of lipid (L) and a peripherally placed mitochondrion (M). In the extracellular space a few collagen fibres (arrow) can be seen. Scale bar: 500 nm. Inset: view of adipocyte. Lipid (L). Scale bar: 10 μm.