5 AMP-activated protein kinase (AMPK) is an energy sensor activated by perturbed cellular energy status such as during muscle contraction. Activated AMPK is thought to regulate several key metabolic pathways. We used sex comparison to investigate whether AMPK signalling in skeletal muscle regulates fat oxidation during exercise. Moderately trained women and men completed 90 min bicycle exercise at 60%V O 2 peak . Both AMPK Thr 172 phosphorylation and α 2 AMPK activity were increased by exercise in men (∼200%, P < 0.001) but not significantly in women. The sex difference in muscle AMPK activation with exercise was accompanied by an increase in muscle free AMP (∼164%, P < 0.01), free AMP/ATP ratio (159%, P < 0.05), and creatine (∼42%, P < 0.001) in men but not in women (NS), suggesting that lack of AMPK activation in women was due to better maintenance of muscle cellular energy balance compared with men. During exercise, fat oxidation per kg lean body mass was higher in women than in men (P < 0.05). Regression analysis revealed that a higher proportion of type 1 muscle fibres (∼23%, P < 0.01) and a higher capillarization (∼23%, P < 0.05) in women than in men could partly explain the sex difference in α 2 AMPK activity (r = −0.54, P < 0.05) and fat oxidation (r = 0.64, P < 0.05) during exercise. On the other hand, fat oxidation appeared not to be regulated via AMPK. In conclusion, during prolonged submaximal exercise at 60%V O 2 peak , higher fat oxidation in women cannot be explained by higher AMPK signalling but is accompanied by improved muscle cellular energy balance in women probably due to sex specific muscle morphology.

. Malonyl-CoA and carnitine in regulation of fat oxidation in human skeletal muscle during exercise.

The collagens of the extracellular matrix are the most abundant structural proteins in the mammalian body. In tissue remodeling and in the invasive growth of malignant tumors, collagens constitute an important barrier, and consequently, the turnover of collagen is a rate-limiting process in these events. A recently discovered turnover route with importance for tumor growth involves intracellular collagen degradation and is governed by the collagen receptor, urokinase plasminogen activator receptor-associated protein (uPARAP or Endo180). The interplay between this mechanism and extracellular collagenolysis is not known. In this report, we demonstrate the existence of a new, composite collagen breakdown pathway. Thus, fibroblastmediated collagen degradation proceeds preferentially as a sequential mechanism in which extracellular collagenolysis is followed by uPARAP/Endo180-mediated endocytosis of large collagen fragments. First, we show that collagen that has been pre-cleaved by a mammalian collagenase is taken up much more efficiently than intact, native collagen by uPARAP/Endo180-positive cells. Second, we demonstrate that this preference is governed by the acquisition of a gelatin-like structure by the collagen, occurring upon collagenase-mediated cleavage under native conditions. Third, we demonstrate that the growth of uPARAP/Endo180-deficient fibroblasts on a native collagen matrix leads to substantial extracellular accumulation of well defined collagen fragments, whereas, wild-type fibroblasts possess the ability to direct an organized and complete degradation sequence comprising both the initial cleavage, the endocytic uptake, and the intracellular breakdown of collagen.Collagens are the most abundant protein constituents of the extracellular matrix. The sheet-like collagens of the basement membrane and the fibrillar matrix collagens all incorporate into dense, insoluble protein networks that form a critical barrier against processes of cell migration such as those connected to tissue remodeling, including the invasive growth of cancer. Consequently, the degradation of these matrices is one of the rate-limiting steps in cancer invasion (1).The physiological mechanisms responsible for collagen degradation have long been subject to investigation. Due to their unique structural features, collagens can only be degraded by a minority of mammalian extracellular proteases, but certain matrix metalloproteases (MMPs), 3 such as MMP-1, MMP-2, MMP-8, MMP-13, and the membrane-bound MMP-14 and -15, are indeed active against native collagens (2-10). The initial attack of these proteases leads to the generation of well defined collagen fragments, which, while still in the extracellular environment, may be subject to further degradation by gelatinases, MMP-2 or MMP-9, or other types of proteases (11-13).Importantly, however, collagen may also be degraded through an intracellular turnover pathway (11,14). Recent studies have shown that an endocytic route of collagen breakdown, mediated by the collagen internalization recep...

Despite a substantial effort to identify locally formed vasoactive compounds, responsible for the large increases in blood flow to skeletal muscle that occur with exercise, this issue remains largely unsolved. In many tissues, including skeletal muscle, endothelial-derived hyperpolarizing factors (EDHF) have been identified as factors that, independently of NO and prostanoids, can hyperpolarize smooth muscle, and thereby elicit vasodilatation, in response to substances that enhance endothelial calcium levels, e.g. acetylcholine and bradykinin Fisslthaler et al. 2000). Several different EDHFs appear to exist in different tissues and vessels (Quilley et al. 1997), although in coronary arteries of several species (Pinto et al. 1987;Hecker et al. 1994) the EDHF phenomenon involves a cytochrome P450 (CYP) epoxygenase-derived product. In porcine coronary arteries, a CYP 2C isoform homologous to CYP 2C9 plays a crucial role in the generation of the bradykinin-induced, endothelium-dependent hyperpolarization of vascular smooth muscle cells and the subsequent NO synthase-and cyclooxygenase-independent vasodilatation .In human skeletal muscle, bradykinin has been reported to induce hyperaemia independently of NO and prostanoids, suggesting an EDHF-mediated mechanism (Halcox et al. 2000) Recent in vitro studies have, moreover, demonstrated that a product of a CYP 2C enzyme homologous to CYP 2C8 and CYP 2C9 regulates EDHF-mediated responses in hamster skeletal muscle resistance arteries (Bolz et al. 2000). Thus, CYP 2C9 is clearly a potential candidate in the regulation of exercise hyperaemia in humans.We and others have previously shown that inhibition of NO synthesis by systemic infusion of either N v -monomethyl-L-arginine (L-NMMA; Shoemaker et al. 1997;Bradley et al. 1999;Rådegran & Saltin, 1999) Previous studies show that exercise-induced hyperaemia is unaffected by systemic inhibition of nitric oxide synthase (NOS) and it has been proposed that this may be due to compensation by other vasodilators. We studied the involvement of cytochrome P450 2C9 (CYP 2C9) in the regulation of skeletal muscle blood flow in humans and the interaction between CYP 2C9 and NOS. Seven males performed knee extensor exercise. Blood flow was measured by thermodilution and blood samples were drawn frequently from the femoral artery and vein at rest, during exercise and in recovery. ; P < 0.05). Oxygen uptake during exercise was 12 ± 3 % lower (9 min: 525 ± 46 vs. 594 ± 24 ml min _1 ; P < 0.05) with co-infusion of sulfaphenazole and L-NMMA, whereas oxygen uptake during sulfaphenazole infusion alone was not different from that of control (P > 0.05). The results demonstrate that CYP 2C9 plays an important role in the regulation of hyperaemia and oxygen uptake during exercise. Since inhibition of neither NOS nor CYP 2C9 alone affect skeletal muscle blood flow, an interaction between CYP 2C9 and NOS appears to exist so that a CYP-dependent vasodilator mechanism takes over when NO production is compromised.