Insulin activation of phosphatidylinositol 3-kinase in human skeletal muscle in vivo

Hickey, Matthew S., Charles J. Tanner, D. Sean O'Neill,Lydia J. Morgan, G. Lynis Dohm, and Joseph A. Houmard. Insulin activation of phosphatidylinositol 3-kinase in human skeletal muscle invivo. J. Appl. Physiol. 83(3):718-722, 1997.The purpose of this investigation was to determinewhether insulin-stimulated phosphatidylinositol 3-kinase (PI3-kinase)activity is detectable in needle biopsies of human skeletal muscle.Sixteen healthy nonobese males matched for age, percent fat, fastinginsulin, and fasting glucose participated in one of two experimentalprotocols. During an intravenous glucose tolerance test (IVGTT)protocol, insulin-stimulated PI3-kinase activity was determined frompercutaneous needle biopsies at 2, 5, and 15 min post-insulinadministration (0.025 U/kg). In the second group, a 2-h, 100 mU · m² · min¹euglycemic hyperinsulinemic clamp was performed, and biopsies wereobtained at 15, 60, and 120 min after insulin infusion was begun.Insulin stimulated PI3-kinase activity by 1.6 ± 0.2-, 2.2 ± 0.3-, and 2.2 ± 0.4-fold at 2, 5, and 15 min, respectively, duringthe IVGTT. During the clamp protocol, PI3-kinase was elevated by 5.3 ± 1.3-, 8.0 ± 2.6-, and 2.7 ± 1.4-fold abovebasal at 15, 60, and 120 min, respectively. Insulin-stimulatedPI3-kinase activity at 15 min post-insulin administration wassignificantly greater during the clamp protocol vs. the IVGTT(P < 0.05). These observations suggest that insulin-stimulated PI3-kinase activity is detectable inneedle biopsies of human skeletal muscle, and furthermore, that theeuglycemic, hyperinsulinemic clamp protocol may be a useful tool toassess insulin signaling in vivo.

     

A Modified Self-thinning Equation to Describe Size/Density Relationships for Defoliated Swards

Use of the self-thinning rule to describe size/density compensation(SDC) in defoliated swards is examined. It is shown that defoliationrelated variation in leaf area and associated morphogeneticchanges in plant structure necessitate slope corrections, designatedC_a and C_r , respectively. The theory predicts that reduced leafarea in more heavily defoliated swards will result in SDC atslopes more negative than -3/2 (variable leaf area SDC), andthat there will be a transition to -3/2 (constant leaf area)SDC at higher herbage mass. Empirical data from previous experiments with Lolium perenneL. and Medicago sativa L. are examined, and appear to confirmthe theoretical predictions, including the slope change at thepoint of transition from variable to constant leaf area SDC.This transition point, designated d_i , is subject to interspecificvariation and is related to the mature shoot size of a particularspecies. Some applications of this theory are discussed, andin particular a sward productivity index is proposed.Copyright1995, 1999 Academic Press Variable leaf area self-thinning, size/density compensation, Lolium perenne, Medicago sativa, sward productivity index     

Evidence that neutral spingomyelinase of cultured murine neuroblastoma cells is oriented externally on the plasma membrane

The activity of the neutral, Mg²⁺-stimulated sphingomyelinase of cultured neuroblastoma cells (N1E-115) is enriched in the plasma membrane fraction and is reduced following treatment of intact or broken cells with trypsin, α-chymotrypsin, papain, and protease. Two protease-sensitive enzymes of the cell interior (lactate dehydrogenase and NADPH-cytochrome c reductase) are not affected by protease treatment of intact cells. These results indicate that the neutral, Mg²⁺-stimulated sphingomyelinase is oriented externally on the plasma membrane of the cultured neuroblastoma cell.     

Serum indole-3-acetic acid in control subjects and newly abstinent alcoholics after an oral loading with L-tryptophan: a preliminary study using liquid chromatography with amperometric detection

Described in this paper is a rapid, isocratic assay for serum indole-3-acetic acid (IAA). The sample preparation involves only protein precipitation using sulfosalicylic acid, and the sensitivity of amperometric detection is in the picogram range. The chromatographic analysis time is approximately 4 min. The devised method was used for a longitudinal study of IAA levels in serum samples from control subjects and newly abstinent alcoholics. Dietary variations were eliminated by administering a 2.0-g loading dose of L-Trp to all subjects investigated. The results are presented in the form of cumulative frequency polygons. Preliminary data indicate no differences in IAA levels between newly abstinent alcoholics and control subjects.     

Albumin-induced plasma volume expansion: diurnal and temperature effects

To develop a reliable procedure for the acute expansion of plasma volume (PV), 26 male volunteers were randomly assigned to either a thermoneutral (25 degrees C and 40% relative humidity) or hot-dry (37 degrees C and 25% relative humidity) environment; subsequently each subject was seated for at least 1 h and then infused intravenously with either 100 or 200 ml of a 25% albumin solution or 0.9% saline. On the day before each infusion, PV was estimated by dye dilution using indocyanine green. Net percent change in PV (using hematocrit and hemoglobin values) was calculated at 1, 3, 6, 9, 12, and 24 h postinfusion. The PV of subjects residing in the heat after a 100-ml saline infusion increased significantly over 1-h values at 6, 9, and 12 h postinfusion but not at 24 h. The same trend, although not significant, was apparent at room temperature. The data suggest a slow isooncotic circadian pattern of PV expansion and contraction. The infusion of hyperoncotic albumin produced rapid expansion of plasma volume. With the low dose (25 g) at 1 h postinfusion, the expansion was 379 +/- 102 ml in the heat and 301 +/- 160 ml at room temperature. With the high dose (50 g) at 1 h postinfusion, the expansion was 479 +/- 84 ml in the heat and 427 +/- 147 ml at room temperature. The high dose produced an expansion that persisted for at least 9 h in subjects in either environment. The data suggest a mechanism for the retention of fluid during heat acclimatization and a useful procedure for plasma volume expansion in humans.     

Gas-chromatographic resolution of the enantiomers of 3-(3,4-dihydroxyphenyl)alanine and its α-methyl analog

Methylation of (R,S)-DOPA with diazomethane gave the trimethyl derivative in which the phenolic hydroxy groups and the carboxy group were methylated. N-Methylated side products were also formed. N-Acylation of the racemic trimethyl derivative with (S)-α-methoxy-α-trifluoromethylphenylacetyl chloride gave two diastereomeric amides which were resolved by gas chromatography, the diastereomer derived from (S)-(−)-DOPA cluting first. The procedure was also applied to α-methyl-DOPA.     

A note on the structure of system state spaces and its implications on the existence of non-repeatable experiments

The implication of state space structure on the existence of a repeatable experimentE designed to determine if a states∈L has propertyP or notP is investigated. It is shown that if a state spaceL is connected, then no experimentE is repeatable. This formalism is used to demonstrate that if a propertyP has an associated set of points inL which is dense with dense complement inL, then there exists no repeatable experimentE which can be used to test whethers has propertyP or notP. Other consequences of this formalization are discussed.     

Glucocorticoid modulation of lymphokine-induced macrophage proliferation

The ability of glucocorticoids to modify lymphokine-induced macrophage proliferation, an in vitro correlate of cellular immunity in the guinea pig, was investigated. Lymphocyte production of macrophage mitogenic factor (MMF) was decreased in the presence of physiological concentrations of glucocorticoids. Inhibition was concentration dependent (IC₅₀ of triamcinolone acetonide (TA): 2 × 10^?9M), glucocorticoid specific, and reversed by cortexolone. In contrast, pharmacological concentrations of glucocorticoids were necessary to inhibit macrophage proliferation induced by suboptimal dilutions of MMF. This inhibition was concentration dependent (IC₅₀ of TA: 4 × 10^?7M), glucocorticoid specific, and reversed by cortexolone. At supraoptimal dilutions of MMF, glucocorticoids caused a twofold potentiation of MMF-induced macrophage proliferation. Potentiation was concentration dependent (EC₅₀ of TA: 3 × 10^?8M), glucocorticoid specific, reversed by glucocorticoid antagonists, and occurred in the presence of indomethacin. Thus, glucocorticoids regulate both the initiation and effector phases of this in vitro model of delayed hypersensitivity. However, the results indicate that the major mechanism of glucocorticoid-mediated anti-inflammatory action occurs at the level of the MMF-producing lymphocyte rather than at the effector macrophage, as MMF-induced proliferation is likely controlled by opposing glucocorticoid-sensitive mechanisms.