The failure of some obese subjects to lose weight while eating a diet they report as low in calories is due to an energy intake substantially higher than reported and an overestimation of physical activity, not to an abnormality in thermogenesis.

Glycogen synthase kinase 3β (GSK3β) is involved in metabolism, neurodegeneration, and cancer. Inhibition of GSK3β activity is the primary mechanism that regulates this widely expressed active kinase. Although the protein kinase Akt inhibits GSK3β by phosphorylation at the N terminus, preventing Akt-mediated phosphorylation does not affect the cell-survival pathway activated through the GSK3β substrate β-catenin. Here, we show that p38 mitogen-activated protein kinase (MAPK) also inactivates GSK3β by direct phosphorylation at its C terminus, and this inactivation can lead to an accumulation of β-catenin. p38 MAPK-mediated phosphorylation of GSK3β occurs primarily in the brain and thymocytes. Activation of β-catenin-mediated signaling through GSK3β inhibition provides a potential mechanism for p38 MAPK-mediated survival in specific tissues.The p38 mitogen-activated protein kinase (MAPK) is activated through phosphorylation primarily by MAPK kinase 3 (MKK3) and MKK6 in response to cellular stress and cytokines. The p38 MAPK pathway functions in the control of differentiation, the blockade of proliferation, and in the induction of apoptosis (1). It is also activated in response to DNA double-stranded breaks (DSBs) induced by ionizing irradiation or chemotherapeutic drugs, and it participates in the induction of a G 2 /M cell-cycle checkpoint (2,3). p38 MAPK can also promote survival (4-6) by unknown mechanisms. During T cell receptor β (TCRβ) rearrangement, V(D)J recombination-mediated DSBs also activate p38 MAPK in immature thymocytes at the double negative 3 (DN3) stage of development (7,8). The expression of a constitutively active mutant of MKK6 [MKK6(Glu)] in thymocytes of transgenic mice (MKK6 transgenic mice) activates a p53-mediated G 2 /M phase cell-cycle checkpoint (8). Like recombination-activating gene (Rag) deficiency, persistent activation of p38 MAPK interferes with the differentiation of thymocytes beyond the DN3 stage. However, MKK6 transgenic thymocytes (but not Rag -/-thymocytes) survive and accumulate in vivo (8), suggesting that

The transcription factor NF-B, a central regulator of immunity, is subject to regulation by redox changes. We now report that cysteine-179 of the inhibitory B kinase (IKK) ␤-subunit of the IKK signalosome is a central target for oxidative inactivation by means of S-glutathionylation. S-glutathionylation of IKK-␤ Cys-179 is reversed by glutaredoxin (GRX), which restores kinase activity.

To quantify proteins on a global level from mammalian tissue, a method was developed to metabolically introduce 15N stable isotopes into the proteins of Rattus norvegicus for use as internal standards. The long-term metabolic labeling of rats with a diet enriched in 15N did not result in adverse health consequences. The average 15N amino acid enrichments reflected the relative turnover rates in the different tissues and ranged from 74.3 mpe in brain to 92.2 mpe in plasma. Using the 15N-enriched liver as a quantitative internal standard, changes in individual protein levels in response to cycloheximide treatment were measured for 310 proteins. These measurements revealed 127 proteins with altered protein level (p < 0.05). Most proteins with altered level have previously reported functions involving xenobiotic metabolism and protein-folding machinery of the endoplasmic reticulum. This approach is a powerful tool for the global quantitation of proteins, is capable of measuring proteome-wide changes in response to a drug, and will be useful for studying animal models of disease.

Leucine metabolism in vivo can be determined from a primed, continuous infusion of L-[1-13C]leucine by measuring, at isotopic steady state, plasm [-13C]leucine enrichment, expired 13CO2 enrichment, and CO2 production rate. With an appropriate priming dose of L-[1-13C]leucine and NaH13CO3, isotopic steady state is reached in less than 2 h, and the infusion is completed in 4 h. The method can determine rates of leucine turnover, oxidation, and incorporation into protein with typical relative uncertainties of 2, 10, and 4%, respectively. The method requires no more than 1 ml of blood and uses stable isotope rather than radioisotope techniques. Thus, the method is applicable to studies of human beings of all ages. L-[1-13C]leucine may be infused with a second amino acid labeled with 15N for simultaneous determination of the kinetics of two amino acids.

Maintenance of reduced or elevated body weight results in respective decreases or increases in energy expended in physical activity, defined as 24-h energy expenditure excluding resting energy expenditure and the thermic effect of feeding, beyond those attributable to weight change. We examined skeletal muscle work efficiency by graded cycle ergometry and, in some subjects, rates of gastrocnemius muscle ATP flux during exercise by magnetic resonance spectroscopy (MRS), in 30 subjects (15 males, 15 females) at initial weight and 10% below initial weight and in 8 subjects (7 males, 1 female) at initial weight and 10% above initial weight to determine whether changes in skeletal muscle work efficiency at altered body weight were correlated with changes in the energy expended in physical activity. At reduced weight, muscle work efficiency was increased in both cycle ergometry [mean (SD) change = +26.5 (26.7)%, P < 0.001] and MRS [ATP flux change = -15.2 (23.2)%, P = 0.044] studies. Weight gain resulted in decreased muscle work efficiency by ergometry [mean (SD) change = -17.8 (20.5)%, P = 0.043]. Changes in muscle efficiency at altered body weight accounted for 35% of the change in daily energy expended in physical activity.

1. Measurements have been made of whole-body and skeletal muscle protein synthesis in fed and fasted adults with L-[1-13C]leucine. 2. The marked increase in whole-body synthesis on feeding largely reflects the changes in protein synthesis in muscle, which doubles on feeding, compared with a 40% increase in that of the rest of the body. 3. Skeletal muscle in fed man contributes more than half to total protein synthesis occurring in the whole body.

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