Treatment with the oral antidiabetic agent troglitazone improves beta cell responses to glucose in subjects with impaired glucose tolerance

Impaired glucose tolerance (IGT) is associated with defects in both insulin secretion and action and carries a high risk for conversion to non-insulin-dependent diabetes mellitus (NIDDM). Troglitazone, an insulin sensitizing agent, reduces glucose concentrations in subjects with NIDDM and IGT but is not known to affect insulin secretion. We sought to determine the role of beta cell function in mediating improved glucose tolerance. Obese subjects with IGT received 12 wk of either 400 mg daily of troglitazone (n = 14) or placebo (n = 7) in a randomized, double-blind design. Study measures at baseline and after treatment were glucose and insulin responses to a 75-g oral glucose tolerance test, insulin sensitivity index (SI) assessed by a frequently sampled intravenous glucose tolerance test, insulin secretion rates during a graded glucose infusion, and beta cell glucose-sensing ability during an oscillatory glucose infusion. Troglitazone reduced integrated glucose and insulin responses to oral glucose by 10% (P = 0.03) and 39% (P = 0.003), respectively. SI increased from 1.3+/-0.3 to 2.6+/-0.4 x 10(-)5min-1pM-1 (P = 0.005). Average insulin secretion rates adjusted for SI over the glucose interval 5-11 mmol/liter were increased by 52% (P = 0.02), and the ability of the beta cell to entrain to an exogenous oscillatory glucose infusion, as evaluated by analysis of spectral power, was improved by 49% (P = 0.04). No significant changes in these parameters were demonstrated in the placebo group. In addition to increasing insulin sensitivity, we demonstrate that troglitazone improves the reduced beta cell response to glucose characteristic of subjects with IGT. This appears to be an important factor in the observed improvement in glucose tolerance.     

The use of multiple antigenic peptide (MAP) in the immunodiagnosis of human immunodeficiency virus infection

In order to develop an ELISA system for the antibody detection of HIV-1 or HIV-2 infections, MAPs for HIV-1 gp41(584-618) and HIV-2 gp36(574-602) corresponding to the immunodominant regions of HIV-1 gp41 and HIV-2 gp36 were used as coating antigens in the ELISA. The MAPs were synthesized by the solid phase method using Fmoc-Lys(Fmoc)-OH and their molecular weights were confirmed by tricine gel electrophoresis. The MAPs reacted with all HIV positive sera (64 samples), but did not react with HIV negative sera (48 samples) obtained from healthy blood donors. The MAPs showed high sensitivity and specificity in anti-HIV 1/2 combo panel and anti-HIV-1 seroconversion panels. The results indicated that the ELISA system using synthetic MAPs of gp41(584-618) and gp36(574-602) as coating antigens can be used as an analytical system for the immunodiagnosis of HIV-1 or HIV-2 infections.     

Leptin is present in human milk and is related to maternal plasma leptin concentration and adiposity

Leptin is elevated during pregnancy and may be involved in the regulation of milk production in women. Immunoreactive leptin was quantified in human milk by modified radioimmunoassay. Leptin concentration was higher in whole vs. skim milk fractions; however, leptin concentration was not correlated with percentage milk fat. Leptin concentrations in whole and skim milk were correlated with maternal plasma leptin concentrations, maternal body weight, body mass index, and tricep skinfold thickness, but not with plasma insulin concentration. These data provide the first evidence for the presence of leptin in human milk in the range of concentrations found in human plasma and indicate that the concentration of leptin in milk reflects maternal adiposity. Determining the biological role(s) of milk-borne leptin could add to our understanding of neonatal metabolism and the mechanisms underlying the development of body fat and obesity in humans.     

The effects of cooperative and individualistic reward on intrinsic motivation

The effects of cooperative versus individualistic reward on students' intrinsic motivation were investigated. The controlling aspects of extrinsic reward may be heightened or produce greater ego threat in the individualistic situation when compared with a group situation. We predicted that students in the cooperative social situation would show higher levels of intrinsic motivation. Fifth-grade students from existing cooperative groups were assigned randomly to receive a tangible reward based on either cooperative or individualistic achievement for completing pattern block designs. Cooperation affected intrinsic motivation positively. Students in the cooperative dyad solved the block designs more quickly, interacted positively, and viewed the task as easier than did those in the individualistic situation, and they reported that their peers were helpful. There was little evidence that the controlling functions of reward or ego-threat were factors in producing the outcome. Some evidence supporting the importance of the social nature of cooperation was provided.     

Development of a high-volume in situ mRNA hybridization assay for the quantification of gene expression utilizing scintillating microplates

The 94-kDa glucose-regulated protein (GRP94) is a member of the 90-kDa heat-shock protein (HSP90) family. In this study, we expressed the barley (Hordeum vulgare L.) GRP94 and the alpha isoform of human HSP90 (HSP90 alpha) in Escherichia coli and compared their dimer-forming abilities. Native polyacrylamide gel electrophoresis revealed that GRP94 (amino acids 69-809) and the full-length form of HSP90 alpha existed in the dimeric state. The C-terminal 326 amino acids of GRP94 or the C-terminal 200 amino acids of HSP90 alpha were sufficient for the dimerization. Limited proteolysis of the C-terminal half of GRP94 with thrombin revealed a 16-kDa fragment, which was derived from the C-terminus of GRP94 through the cleavage of either the Arg710-His711 or the Arg735-Leu736 bond. These cleavage sites were nearly, if not completely, equivalent to the proteolyzed region of HSP90 alpha. Their structural similarity prompted us to investigate, by use of a coexpression system, the possibility that the two proteins form a heterodimeric complex. A two-step affinity chromatography that specifically trapped only the complex revealed that the C-terminal 200 amino acids of HSP90 alpha and the C-terminal 326 amino acids of GRP94 associated with HSP90 alpha and GRP94, respectively. However, the C-terminal 326 amino acids of GRP94 failed to form a complex with HSP90 alpha. In conclusion, these results indicate the similarity of the general dimeric conformation of the two HSP90 family member proteins, but show that the similarity is not sufficient to allow heterodimer formation.     

Generation and characterization of gp100 peptide-specific NK-T cell clones

MHC-restricted cytotoxic T lymphocytes (CTLs) specific for antigens expressed by malignant cells are important components of immune responses against human cancer. Peripheral blood monocytes of HLA-A2+ healthy donors were used to induce dendritic cells (DCs) by granulocyte-macrophage colony-stimulating factor and interleukin-4 and loaded with a gp100 peptide (YLEPGPVTA). By applying these peptide-loaded DCs, a CTL line that displayed high cytotoxic reactivity with peptide-loaded target cells was generated. A total of 11 gp100 peptide-specific CTL clones were generated from this cell line. Several of these CTL clones were studied in detail. Of particular interest was clone CTL-45, which, contrary to the parental cell line, displayed strong NK activity and, by flow-cytometric analysis, revealed a CD3+, TCR BV17, CD8+ and CD56+ phenotype. This clone was strictly peptide-specific and effectively killed a panel of melanoma cells expressing HLA-A2 and gp100. Tumor-specific T cells with this kind of dual function are potentially of great clinical importance as they have a backup mechanism that may go into action when tumor cells escape specific killing by losing their HLA-class I molecules.     

Effect on normal vaginal flora of three intravaginal microbicidal agents potentially active against human immunodeficiency virus type 1

The effect on normal vaginal flora of three intravaginal microbicides potentially active against human immunodeficiency virus type 1 was examined. Volunteers received dextrin sulfate (D2S), nonoxynol-9 (N-9), or docusate sodium in separate placebo-controlled studies. High vaginal swabs were obtained for bacterial culture before and after microbicide application. D2S did not affect the vaginal flora. However, lactobacilli decreased by > or = 10(2) cfu/mL in 9 (56%) of 16 women given N-9 and in 5 (63%) of 8 women given docusate sodium. Women using N-9 were also significantly more likely to become colonized abnormally (usually with aerobic gram-negative rods) than were those using placebo, as were women using docusate sodium. Women with reduced lactobacilli were less likely to regain normal flora than were those whose lactobacilli were unaffected. However, coliform colonization occurred whether lactobacilli produced H2O2 or not. Continuous use of N-9 could induce susceptibility to urinary and gynecological infection. It is essential that potential microbicides are examined for activity against normal vaginal flora.     

Biphasic insulin secretion during intravenous glucose tolerance test promotes optimal interstitial insulin profile

We examined the hindlimb lymph insulin profile during simulated intravenous glucose tolerance tests (IVGTTs) in anesthetized dogs to test the following hypotheses: 1) the biphasic insulin response to intravenous glucose can be seen as a priming bolus and a secondary infusion that effect a rapid stepwise increase in the interstitial insulin concentration and 2) the activation of glucose utilization (rate of glucose uptake [Rd]) during an IVGTT is more similar to the dynamics of the interstitial insulin profile than that of the arterial plasma. Three insulin profiles were infused: a normal biphasic pattern, a second phase infusion only, and a biphasic pattern with a fourfold greater first phase and a normal second phase. During the normal biphasic infusion, lymph insulin quickly reached and maintained a steady-state concentration (10 min, 26.42 +/- 0.86 microU/ml). With second phase only, it took lymph insulin 35 min to reach a steady state of lower concentration (13.13 +/- 0.46 microU/ml) than the normal. And with a fourfold greater first phase, lymph insulin plateaued quickly (16 min, 140.87 +/- 1.68 microU/ml), but for a shorter duration than the normal. For each profile, the time course of activation of Rd did not follow the time course of insulin in the plasma, but was more similar to that of insulin in the interstitial fluid. These results show that the biphasic response allows interstitial insulin to rapidly reach and maintain a steady state beneficial to activation and maintenance of glucose utilization.     

Validation of the in vivo intravascular ultrasound measurement of in-stent neointimal hyperplasia volumes

OBJECTIVES: This study was undertaken to validate the in vivo intravascular ultrasound (IVUS) measurement of in-stent neointimal hyperplasia (IH) volumes. BACKGROUND: Because stents reduce restenosis compared to balloon angioplasty, stent use has increased significantly. As a result, in-stent restenosis is now an important clinical problem. Serial IVUS studies have shown that in-stent restenosis is secondary to intimal hyperplasia. To evaluate strategies to reduce in-stent restenosis, accurate measurement of in-stent neointimal tissue is important. METHODS: Using a porcine coronary artery model of in-stent restenosis, single Palmaz-Schatz stents were implanted into 16 animals with a stent:artery ratio of 1.3:1. Intravascular ultrasound imaging was performed at 1 month, immediately prior to animal sacrifice. In vivo IVUS and ex vivo histomorphometric measurements included stent, lumen and IH areas; IH volumes were calculated with Simpson's rule. RESULTS: Intravascular ultrasound measurements of IH (30.4+/-11.0 mm3) volumes correlated strongly with histomorphometric measurements (26.7+/-8.5 mm3, r=0.965, p < 0.0001). The difference between the IVUS and the histomorphometric measurements of IVUS volume was 4.1+/-2.7 mm3 or 15.8+/-11% (standard error of the estimate=0.7). Both histomorphometry and IVUS showed that IH was concentric and uniformly distributed over the length of the stent. Intravascular ultrasound detected neointimal thickening of < or =0.2 mm in 5 of 16 stents. Sample size calculations based on the IVUS measurement of IH volumes showed that 12 stented lesions/arm would be required to show a 50% reduction in IVUS-measured IH volume and 44 stented lesions/arm would be required to show a 25% reduction in IH volume. CONCLUSION: In vivo IVUS measurement of IH volumes correlated strongly with ex vivo histomorphometry. Using volumetric IVUS end points, small sample sizes should be necessary to demonstrate effectiveness of strategies to reduce in-stent restenosis.