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The opioid agonists [leucine]enkephalin, [D-Ala2,D-Leu5]enkephalin and dynorphin-(1-13)-peptide, but not morphine, stimulated the conversion of [2-14C]pyruvate into glucose and glycogenolysis when added directly to isolated hepatocytes. Naloxone produced a small but significant inhibition of both the basal and stimulated rate of incorporation of label into glucose but had no effect on the total glucose output by the cells. The effects of the opioid peptides were mediated by a cyclic AMP-independent mechanism.

Alterations in the long-chain acyl-CoA binding to albumin in the carnitine palmitoyltransferase (CPT) assay appreciably affect the reaction at commonly used substrate concentrations. Since in the CPT assay the latter are typically well below saturation or Vmax. values, the measured enzyme activity depends on both the absolute quantity of albumin in the CPT assay and any biochemical modification of its binding. The present study verifies the striking dependence of the K0.5 for palmitoyl-CoA on albumin and the misleading ‘activation’ of the enzyme by compounds that also avidly bind to albumin. In assessing the intracellular physiological relevance of any modifier of CPT, the effects of protein binding in the assay assume particular importance. Indeed, any compound that alters CPT activity may do so, not directly, but as an assay artifact changing the free or unbound substrate concentrations.

Albumin elimination rates were determined in 3-, 12-, 24- and 36-month-old female WAG/Rij rats. No change in elimination half-life was found with age. However, as there was an increase in the whole-body albumin pool, a concomitant increase in albumin clearance was observed at between 12 and 36 months of age. It was concluded that the increase in clearance between 12 and 24 months of age was only due to a change in the animal's physiology, whereas between 24 and 36 months of age it was also due to changes in the albumin molecule. The age-related changes in albumin clearance were thought not to be caused by changes in the albumin excretion via the urine or via the gastrointestinal tract.

Metabolic acidosis stimulates the rate of glutamine release from muscle, and this in turn is used by the kidney in acid-base balance. NH4Cl, HCl or diabetic ketoacidosis increases the maximum activity of glutamine synthetase in skeletal muscle. Starvation and administration of adrenal steroids also increase the activity of the enzyme in muscle.

Pig synovial fibroblasts were shown to produce a protein that caused live cartilage to resorb its proteoglycan matrix in vitro. Fibroblasts were obtained either from synovial tissue digest or by allowing them to grow out of explants. The population derived from the digests was homogeneous and free of macrophage-like cells after two passages, but was still producing the cartilage-resorbing protein after seven passages. The active protein was found to have Mr 20,000 on gell filtration, and pI 4.8 on isoelectric focussing in polyacrylamide gel. It was indistinguishable from a protein with the same activity from pig mononuclear leucocytes, which has been called catabolin. Production of the protein was increased if the synovial fibroblasts were cultured with the tumour promoter phorbol 12-myristate 13-acetate. Fibroblasts from other sources (joint capsule and peritoneum) also apparently made the protein. The possibility that catabolin is the same as interleukin-1 is discussed: if they are, then the results suggest that fibroblasts can make an interleukin-1-life protein.

At micromolar concentrations, acetyl-CoA inhibited hepatic carnitine acyltransferase activity and mitochondrial fatty acid oxidation. The inhibitory effects were not nearly as potent on a molar basis as those of malonyl-CoA; nevertheless, the cytosolic concentrations of acetyl-CoA, as yet unknown, may be sufficient (greater than 30 microM) to curtail appreciably the mitochondrial transfer of long-chain acyl-CoA units and fatty acid oxidation. Hence acetyl-CoA may also partially regulate hepatic ketogenesis.

Bacitracin is a proteolytic inhibitor which interacts with the intracellular processing of insulin. Its effects on pyruvate, fatty acid and amino acid metabolism were examined in rat hepatocyte suspensions. Bacitracin (0.25-1.0 mM) increased the oxidation of [1-14C]pyruvate by 50-70% and presumably therefore increased the flux through pyruvate dehydrogenase. This was found both in the presence of extracellular Ca2+ and in its absence, but not in the presence of 2 mM-2-chloropropionate, which inhibits pyruvate dehydrogenase kinase. Insulin did not further stimulate [1-14C]pyruvate oxidation in the presence of 1 mM-bacitracin. Bacitracin decreased 14CO2 formation from [2-14C]pyruvate (20-40%) and [U-14C]palmitate (30-70%), suggesting a decreased flux through the tricarboxylic acid cycle. Fatty acid oxidation before acetyl-CoA formation was also decreased. Bacitracin decreased the incorporation of label from [3H]leucine into protein in the absence of insulin, but not in its presence. Bacitracin is commonly used in studies on insulin action. Our results suggest that in such studies the effects noted may be related not only to an interaction of bacitracin with the intracellular processing of insulin but also to direct metabolic effects of bacitracin independent of insulin.

Monoclonal IgG1 (immunoglobulin G1) PA2.1 and MA2.1 antibodies recognize polymorphic sites of the human transplantation antigen HLA-A2. They are distinguishable because MA2.1 binds HLA-A2 and HLA-B17, whereas PA2.1 binds HLA-A2 and HLA-A28. The affinities of PA2.1-Fab for HLA-A2, three HLA-A2 variants and HLA-A28 are similar and relatively low (1.9 × 10(7) M-1). The affinities of MA2.1-Fab for HLA-A2, three HLA-A2 variants and HLA-B17 are similar and high (1.2 × 10(9) M-1). The difference in affinity is due to the rates of dissociation, which give half-times of dissociation of 290 min for MA2.1-Fab and 4 min for PA2.1-Fab. For both Fab, equilibrium measurements and kinetic determinations gave consistent estimates for affinity. When PA2.1-F(ab)2 or IgG is incubated with cells it reaches equilibrium within 3 h, with most molecules bound bivalently to the cell. Under similar conditions, MA2.1-F(ab)2 does not reach equilibrium and a significant proportion of molecules bound with one and two sites are found. For the lower-affinity antibody (PA2.1), estimates of the binding constants for one- and two-site interactions could be made. By simple Scatchard analysis the avidity of F(ab)2 or IgG is 1.3 × 10(9) M-1, giving an enhancement factor of 68 between bivalent and univalent binding. This is a measure of the equilibrium constant for the interchange between bivalent and univalent binding. Analysis of the results with more realistic models indicates that the actual value is larger (10(3)-10(4) M-1) than 68 M-1. The avidities of F(ab)2 and IgG for HLA-A2 are identical, showing the Fc does not interfere with bivalent binding to cells.