Diabetogenic activity of 20 kDa human growth hormone (20K-hGH) and 22K-hGH in rats.

To compare the diabetogenic activity of 20 kDa human growth hormone (20K-hGH) with that of 22K-hGH, we evaluated insulin sensitivity with a euglycaemic clamp in rats. The glucose infusion rate (GIR) in euglycaemic clamp studies was measured as an indicator of insulin sensitivity. [(14)C]glucose and 2-[(3)H] deoxy- D -glucose injection were used to calculate the rate of glucose utilization (R(d)), the hepatic glucose output (HGO), and the glucose metabolic index (R(g)'). Both 20K- and 22K-hGH were infused at equivalent rates (1.0 (mg/kg)/day). A 24 h infusion of 20K-hGH had no significant effects on the GIR, R(d), HGO and R(g)(')except for in gastrocnemius muscle. In contrast, 22K-hGH significantly lowered the GIR compared with the control (P< 0.001) and 20K-hGH groups (P< 0.01). The infusion of 22K-hGH also reduced R(d)compared with the controls and the 20K-hGH rats by 46.6% (P< 0.001) and 39.6% (P< 0.05) respectively, while no differences were observed in the HGO. Moreover, 22K-hGH inhibited glucose uptake, which was estimated from the insulin-stimulated R(g)' in some tissues. These results suggest that 22K-hGH inhibits the uptake and use of glucose in various tissues, which then leads to insulin resistance. In conclusion, the diabetogenicity of 20K-hGH is much weaker than that of 22K-hGH, and the reduced insulin-antagonizing action of 20K-hGH could have important clinical benefits.

Differences in the effects of 20 K- and 22 K-hGH on water retention in rats.

Antidiuretic actions induced by two growth hormone (GH) isoforms (20 K- and 22 K-hGH; 0.2 and 2.0 mg/kg) were evaluated in rats, as fluid retention may cause oedema, one of the adverse effects of GH. Both GH isoforms (2.0 mg/kg) suppressed urine excretion in hypophysectomized rats (P< 0.01), but only the 22 K-hGH isoform (2.0 mg/kg) suppressed urine excretion in intact rats (P< 0.01). In addition, prolactin (PRL) suppressed urine excretion in intact rats (P< 0.05). In conclusion, 20 K-hGH has less potency in causing urine retention than 22 K-hGH and since 20 K-hGH is missing 15 amino acids found in 22 K-hGH, these amino acids may be important for the antidiuretic action of GH. Since prolactin suppressed urine excretion, a part of the antidiuretic action of GH may be related to PRL-R activation.

Advanced glycation end products-driven angiogenesis in vitro. Induction of the growth and tube formation of human microvascular endothelial cells through autocrine vascular endothelial growth factor.

This study was undertaken to determine whether and how advanced glycation end products (AGE), senescent macroproteins accumulated in various tissues under hyperglycemic states, cause angiogenesis, the principal vascular derangement in diabetic microangiopathy. We first prepared AGE-bovine serum albumin (BSA) and anti-AGE antiserum using AGE-RNase A. Then AGE-BSA was administered to human skin microvascular endothelial cells in culture, and their growth was examined. The AGE-BSA, but not nonglycated BSA, was found to induce a statistically significant increase in the number of viable endothelial cells as well as their synthesis of DNA. The increase in DNA synthesis by AGE-BSA was abolished by anti-AGE antibodies. AGE-BSA also stimulated the tube formation of endothelial cells on Matrigel. We obtained the following evidence that it is vascular endothelial growth factor (VEGF) that mainly mediates the angiogenic activities of AGE. (1) Quantitative reverse transcription-polymerase chain reaction analysis of poly(A)+ RNA from microvascular endothelial cells revealed that AGE-BSA up-regulated the levels of mRNAs for the secretory forms of VEGF in time- and dose-dependent manners, while endothelial cell expression of the genes encoding the two VEGF receptors, kinase insert domain-containing receptor and fms-like tyrosine kinase 1, remained unchanged by the AGE treatment. Immunoprecipitation analysis revealed that AGE-BSA did increase de novo synthesis of VEGF. (2) Monoclonal antibody against human VEGF completely neutralized both the AGE-induced DNA synthesis and tube formation of the endothelial cells. The results suggest that AGE can elicit angiogenesis through the induction of autocrine vascular VEGF, thereby playing an active part in the development and progression of diabetic microangiopathies.

A nuclear factor containing the leucine-rich repeats expressed in murine cerebellar neurons.

A nuclear protein, termed leucine-rich acidic nuclear protein (LANP), has been isolated from among rat cerebellar proteins whose expression was transiently increased during an early stage of postnatal development. The amino acid sequence, deduced from its cDNA, showed that LANP contains 247 amino acids consisting of two distinct structural domains: the N-terminal domain characterized by "leucine-rich repeat," which is found in many eukaryotic proteins and which potentially functions in mediating protein-protein interactions, and the C-terminal domain characterized by a cluster of acidic amino acids with a putative nuclear localization signal. Immunohistochemical study using an antibody against LANP revealed that the protein is localized mainly in nuclei of Purkinje cells. In the rat cerebellum on postnatal day 7, LANP mRNA was expressed moderately in the external granule and Purkinje cells and weakly in the internal granule cells. The expression in these cells, especially in Purkinje cells, increased in the second postnatal week and thereafter decreased to an adult level. The structural characteristics, localization, and the stage- and cell type-specific expression suggest a potential role of LANP in a signal transduction pathway that directs differentiation of cerebellar neurons.