Long-acting insulin analogues elicit atypical signalling events mediated by the insulin receptor and insulin-like growth factor-I receptor.

AIMS/HYPOTHESIS: Insulin analogues were developed to improve the pharmacological properties of injected insulin and to better mimic endogenous insulin output. However, certain insulin analogues have been suggested to display IGF-I-like biological activities. Furthermore, several recent epidemiological studies have suggested a potential increase in cancer risk for treatment of diabetes patients with long-acting analogue insulin glargine (A21Gly,B31Arg,B32Arg human insulin). Additional studies, however, reported no increased cancer risk. The purpose of the present study was to identify the receptor(s) and signal transduction pathways responsible for the biological actions of insulin glargine and insulin detemir (B29Lys[ε-tetradecanoyl],desB30 human insulin). METHODS: The colon cancer-derived cell line HCT116 was treated with increasing doses of insulin glargine, insulin detemir, regular insulin or IGF-I, and receptor activation was evaluated by immunoprecipitation assays. IGF-I receptor (IGF-IR) internalisation following insulin glargine treatment was assessed by confocal microscopy. Activation of the Akt and extracellular signal-regulated kinase pathways was evaluated by western blots. The anti-apoptotic effect of the analogues was measured by poly-(ADP ribose) polymerase antibody and annexin assays. RESULTS: We found evidence for dual activation of the insulin receptor and IGF-IR by the analogues. Dose-dependency experiments showed that insulin glargine was able to phosphorylate the IGF-IR at fivefold lower doses than those required to activate the insulin receptor. We also showed that insulin glargine can lead to prolonged activation of the receptors and therefore promote abnormal signalling. Confocal imaging experiments showed that insulin glargine, but not regular insulin induced IGF-IR internalisation similarly to IGF-I. Finally, both analogues displayed IGF-I-like anti-apoptotic activities and stimulated cell cycle progression. CONCLUSIONS/INTERPRETATION: Our data indicate that insulin glargine and insulin detemir display atypical signalling activities that differ from those elicited by regular insulin and involve activation of the anti-apoptotic IGF-IR.

Differential effect of insulin-like growth factor-1 and growth hormone on hypothalamic regulation of growth hormone secretion in the rat.

Pharmacological administration of either growth hormone (GH) or insulin-like growth factor 1 (IGF-1) were reported to inhibit endogenous GH release in humans and in the laboratory animal. We have evaluated the short-term differential mechanisms whereby the two hormones affect hypothalamic regulation of GH secretion. Wistar male rats (90 days old) were injected i.p. with either GH (recombinant GH NIAMDD, Baltimore, MD, USA), rIGF-1 (Fujisawa Pharmaceutical Co. Ltd., Osaka, Japan) or saline. Animals were sacrificed at 15, 30, 60 and 120 minutes following injection. Hypothalami were dissected and extracted immediately and the levels of growth hormone-releasing hormone (GHRH) and somatostatin were determined using specific antisera. Trunk blood was collected for GH and IGF-1 determination by RIA. Administration of IGF-1 or GH markedly decreased hypothalamic somatostatin stores by 77% and 54% respectively, within 15 minutes. Concomitantly, the wide range of GH levels found in the control group was reduced in the IGF-1 treated group suggesting that the pulsatile pattern of GH secretion was suppressed. Growth hormone administration induced an increase in hypothalamic GHRH stores (60% at 120 minutes). During this period serum IGF-1 levels were not altered. It is suggested that short term modulation of hypothalamic neurohormones by GH and IGF-1 is mediated by rapid stimulation of somatostatin release by both hormones, and inhibition of GHRH release is induced only by GH.

Renal function in Laron syndrome patients treated by insulin-like growth factor-I.

Eight children with Laron syndrome (5 males, 3 females) aged 3-14.5 years received daily subcutaneous injections of 150 micrograms/kg recombinant insulin-like growth factor-I (IGF-I) for 5 months. The children were examined weekly for the 1st month and then once monthly. At each visit, overnight fasting blood was drawn for serum IGF-I and blood chemistry measurements and a 24-h urine collection was performed for the determination of calcium, phosphorus, creatinine and nitrogen. The main effects related to kidney function were: an initial weight gain with a mild transitory reduction in the urinary volume, an increase in serum electrolyte concentrations and a decrease in urinary electrolyte excretion. The lower than normal mean (+/- SEM) basal creatinine clearance (76.7 +/- 15.8 ml/min per 1.73 m2) increased towards the normal range during treatment to 124.9 +/- 13 ml/min per 1.73 m2, with a mean increment of 73.4 +/- 28% (P < 0.02) from basal values after 2 months of treatment, without changes in the serum creatinine. Initially an increase in blood urea nitrogen was observed together with a reduction in urinary nitrogen excretion. During the IGF-I therapy the urinary calcium excretion increased from 0.7 +/- 0.2 nmol/day to 1.5 +/- 0.3 nmol/day and the tubular reabsorption of phosphate increased from 1.24 +/- 0.06 to more than 1.38 +/- 0.04 nmol/l (P < 0.002), resulting in a significant increase in serum phosphate levels from 1.51 +/- 0.06 to more than 1.63 +/- 0.04 nmol/l (P < 0.005).(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of acute and chronic administration of clonidine on hypothalamic content of growth hormone-releasing hormone and somatostatin in the rat.

Hypothalamic GH-releasing hormone (GHRH) and somatostatin contents were determined in male rats at pre- and mid-pubertal stages following acute (0.15 mg/kg) or chronic (0.1 mg/kg twice daily for 7 days) administration of clonidine. Hypothalamic GHRH content increased markedly during transition from pre- to mid- and late-pubertal stages (2.05 +/- 0.17 vs 3.13 +/- 0.26 and 3.46 +/- 0.20 ng/hypothalamus respectively, means +/- S.E.M.). Hypothalamic GHRH content was markedly decreased 1 h after clonidine administration in mid-pubertal but not in prepubertal rats (3.67 +/- 0.23 vs 2.65 +/- 0.22 ng/hypothalamus). Hypothalamic somatostatin content, on the other hand, decreased in both age groups, although the decrease in the mid-pubertal group was more pronounced (51% and 38% respectively). In addition, the GH responsiveness to clonidine was higher at mid-puberty than at prepuberty. Determination of hypothalamic somatostatin and GHRH content in the chronic experiment 4 h after the last drug administration revealed a marked decrease in GHRH in the mid-pubertal rats and a slight decrease in the prepubertal rats (1.78 +/- 0.32 vs 3.15 +/- 0.31 and 2.01 +/- 0.30 vs 2.35 +/- 0.15 ng/hypothalamus; P less than 0.005 and P less than 0.05 respectively) whereas somatostatin levels were not altered. It is suggested that the clonidine-induced GH secretion is modulated by a chain of events which involves primary stimulation of GHRH release resulting in increased GH secretion which, via a negative-feedback mechanism, triggers an enhancement of somatostatin release which ultimately normalizes the system. This mechanism is age-dependent and reaches full maturity only at the onset of puberty.

Growth hormone, somatomedin and prolactin--relationship to brain function.

In recent years it has been found that the brain of several mammals including have receptors for somatomedin both IGF-I and II and measurable hGH has been identified in human brain tissue. IGF-II has been determined in CSF. The presence of these hormones in brain tissue seems to be of developmental and functional importance as experimental studies in frogs, tadpoles and rats showed that injection of growth hormone enhanced brain growth and increased the ratio of neurons to glia. In man early initiation of hGH therapy to children with hGH (who have a less than normal head circumference) induced a fast catch-up growth of the head and improved their IQ. The data available seems to indicate that growth hormones and/or the somatomedins play an important role in the early brain development, maturation and function. In case of hereditary or congenital GH-RH, hGH or somatomedin deficiency, the effectiveness of therapy seems age limited similar to hypothyroidism. The finding of prolactin receptors in human brain and the report of a child with congenital hypoprolactinemia who had mild mental retardation raises the possibility that also prolactin plays a role in brain function.

Long-term effect of D-Trp6-luteinizing hormone-releasing hormone on testicular size and luteinizing hormone, follicle-stimulating hormone, and testosterone levels in hypothalamic hypogonadotropic males.

Six men, ages 18 to 34 years, with hypothalamic hypogonadotropism were treated with D-Trp6-luteinizing hormone-releasing hormone (10 micrograms intramuscularly on alternate days) for a period of 6 months. They underwent an intravenous luteinizing hormone-releasing hormone (LH-RH) test (50 micrograms/sq m) before and after 1, 3, and 6 months of treatment. During the first 3 months of therapy, the mean (+/- standard deviation) testicular volume increased from 3.5 +/- 1.0 ml to 6.0 +/- 2.0 ml, but decreased to 5.0 +/- 1.0 ml after 6 months. A significant increase in the plasma LH response to LH-RH over pretreatment levels was noted after 1 month (10.2 +/- 4.2 mIU/ml versus 1.6 +/- 1.0 mIU/ml, P less than 0.001) and 3 months (3.0 +/- 1.6 mIU/ml, P less than 0.01) with a subsequent decline to pretreatment levels after 6 months of treatment. The follicle-stimulating hormone response to LH-RH was not significant. It is concluded that D-Trp6-LH-RH induced an initial stimulation in these patients but, probably because of the excessively high dose used, a paradoxical inhibitory response was obtained after 3 months of therapy.

Multiple pituitary hormone deficiencies in eight siblings of one Jewish Moroccan family.

In a Jewish Moroccan inbred family, 8 of 12 siblings were found to have multiple deficiencies of pituitary hormones, including GH, TSH and gonadotrophins. The parents showed no deficiency and are in good health, as are the other 4 siblings. The investigations carried out indicate that in this family the etiology is hereditary in nature, probably being autosomal recessive, with the defect located in the pituitary gland.

Pubertal development in the Prader-Labhart-Willi syndrome.

The sexual maturation in the Prader-Labhart-Willi (PLW) syndrome was investigated in 14 patients, 10 females and 4 males. A wide variability in the pattern of pubertal development was found including delayed puberty in 5 patients and normal puberty in 4 patients; sexual precocity was also observed in 5 patients, true precocious puberty in one patient and incomplete sexual precocity in the form of precocious pubarche in 4 patients. In 5 patients, 3 of them with precocious pubarche, the appearance of the pubertal signs was followed by a delay or arrest in their future development. An LH-RH stimulation test was performed in 11 patients. In the 6 patients who eventually developed normal puberty, the basal levels and the peak responses of both LH and FSH were within the range of those observed in normal controls of the same pubertal stage. In 4 patients showing marked delay or arrest of puberty, the basal levels were normal or low and the responses of LH and FSH to LH-RH were blunted. Priming with repeated LH-RH stimulation in one of the male patients led to an augmented LH response, suggesting a hypothalamic hypogonadotrophism. It is concluded that the lack of uniformity in the pattern of sexual maturation in the PLW syndrome is due to a variability in the location and extent of a hypothalamic lesion, which may comprise an active process continuing beyond the perinatal period.