Assessment of Hepatic Mitochondrial Oxidation and Pyruvate Cycling in NAFLD by (13)C Magnetic Resonance Spectroscopy

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease and there is great interest in understanding the potential role of alterations in mitochondrial metabolism in its pathogenesis. To address this question we assessed rates of hepatic mitochondrial oxidation in subjects with and without NAFLD by monitoring the rate of (13)C labeling in hepatic [5-(13)C]glutamate and [1-(13)C]glutamate by (13)C MRS during an infusion of [1-(13)C]acetate. We found that rates of hepatic mitochondrial oxidation were similar between NAFLD and Control subjects. We also assessed rates of hepatic pyruvate cycling during an infusion of [3-(13)C]lactate by monitoring the (13)C label in hepatic [2-(13)C]alanine and [2-(13)C]glutamate and found that this flux also was similar between groups and more than 10-fold lower than previously reported. Contrary to previous studies we show that hepatic mitochondrial oxidation and pyruvate cycling are not altered in NAFLD and do not account for the hepatic fat accumulation

Reversal of nonalcoholic hepatic steatosis, hepatic insulin resistance, and hyperglycemia by moderate weight reduction in patients with type 2 diabetes

To examine the mechanism by which moderate weight reduction improves basal and insulin-stimulated rates of glucose metabolism in patients with type 2 diabetes, we used (1)H magnetic resonance spectroscopy to assess intrahepatic lipid (IHL) and intramyocellular lipid (IMCL) content in conjunction with hyperinsulinemic-euglycemic clamps using [6,6-(2)H(2)]glucose to assess rates of glucose production and insulin-stimulated peripheral glucose uptake. Eight obese patients with type 2 diabetes were studied before and after weight stabilization on a moderately hypocaloric very-low-fat diet (3%). The diabetic patients were markedly insulin resistant in both liver and muscle compared with the lean control subjects. These changes were associated with marked increases in IHL (12.2 ± 3.4 vs. 0.6 ± 0.1%; P = 0.02) and IMCL (2.0 ± 0.3 vs. 1.2 ± 0.1%; P = 0.02) compared with the control subjects. A weight loss of only ~8 kg resulted in normalization of fasting plasma glucose concentrations (8.8 ± 0.5 vs. 6.4 ± 0.3 mmol/l; P < 0.0005), rates of basal glucose production (193 ± 7 vs. 153 ± 10 mg/min; P < 0.0005), and the percentage suppression of hepatic glucose production during the clamp (29 ± 22 vs. 99 ± 3%; P = 0.003). These improvements in basal and insulin-stimulated hepatic glucose metabolism were associated with an 81 ± 4% reduction in IHL (P = 0.0009) but no significant change in insulin-stimulated peripheral glucose uptake or IMCL (2.0 ± 0.3 vs. 1.9 ± 0.3%; P = 0.21). In conclusion, these data support the hypothesis that moderate weight loss normalizes fasting hyperglycemia in patients with poorly controlled type 2 diabetes by mobilizing a relatively small pool of IHL, which reverses hepatic insulin resistance and normalizes rates of basal glucose production, independent of any changes in insulin-stimulated peripheral glucose metabolism

Direct assessment of hepatic mitochondrial oxidative and anaplerotic fluxes in humans using dynamic ¹³C magnetic resonance spectroscopy

Despite the central role of the liver in the regulation of glucose and lipid metabolism there are currently no methods to directly assess hepatic oxidative metabolism in humans in vivo. By utilizing a novel (13)C-labeling strategy in combination with (13)C magnetic resonance spectroscopy we show that rates of mitochondrial oxidation and anaplerosis in human liver can be directly determined noninvasively. Using this approach we found the mean rates of hepatic TCA cycle flux (V(TCA)) and anaplerotic flux (V(ANA)) to be 0.43 ± 0.04 μmol (g-liver-min)(−1) and 0.60 ± 0.11 μmol (g-liver-min)(−1), respectively, in fourteen healthy, lean, individuals. We also found the ratio V(ANA)/V(TCA) to be 1.39 ± 0.22, which is several fold lower than recently published estimates using an indirect approach. This method will be useful for understanding the pathogenesis of non-alcoholic fatty liver disease and type 2 diabetes as well as assessing the effectiveness of new therapies targeting these pathways in man

Intracellular energetics and critical Po2 in resting ischemic human skeletal muscle in vivo

During ischemia and some types of muscular contractions, oxygen tension (Po2) declines to the point that mitochondrial ATP synthesis becomes limited by oxygen availability. Although this critical Po2 has been determined in animal tissue in vitro and in situ, there remains controversy concerning potential disparities between values measured in vivo and ex vivo. To address this issue, we used concurrent heteronuclear magnetic resonance spectroscopy (MRS) to determine the critical intracellular Po2 in resting human skeletal muscle in vivo. We interleaved measurements of deoxymyoglobin using 1H-MRS with measures of high-energy phosphates and pH using 31P-MRS, during 15 min of ischemia in the tibialis anterior muscles of 6 young men. ATP production and intramyocellular Po2 were quantified throughout ischemia. Critical Po2, determined as the Po2 corresponding to the point where PCr begins to decline (PCrip) in resting muscle during ischemia, was 0.35 ± 0.20 Torr, means ± SD. This in vivo value is consistent with reported values ex vivo and does not support the notion that critical Po2 in resting muscle is higher when measured in vivo. Furthermore, we observed a 4.5-fold range of critical Po2 values among the individuals studied. Regression analyses revealed that time to PCrip was associated with critical Po2 and the rate of myoglobin desaturation (r = 0.83, P = 0.04) but not the rate of ATP consumption during ischemia. The apparent dissociation between ATP demand and myoglobin deoxygenation during ischemia suggests that some degree of uncoupling between intracellular energetics and oxygenation is a potentially important factor that influences critical Po2 in vivo