Cerebellum Susceptibility to Neonatal Asphyxia: Possible Protective Effects of N-Acetylcysteine Amide.

BACKGROUND: After perinatal asphyxia, the cerebellum presents more damage than previously suggested. OBJECTIVES: To explore if the antioxidant N-acetylcysteine amide (NACA) could reduce cerebellar injury after hypoxia-reoxygenation in a neonatal pig model. METHODS: Twenty-four newborn pigs in two intervention groups were exposed to 8% oxygen and hypercapnia, until base excess fell to -20 mmol/l or the mean arterial blood pressure declined to <20 mmHg. After hypoxia, they received either NACA (NACA group, n = 12) or saline (vehicle-treated group, n = 12). One sham-operated group (n = 5) served as a control and was not subjected to hypoxia. Observation time after the end of hypoxia was 9.5 hours. RESULTS: The intranuclear proteolytic activity in Purkinje cells of asphyxiated vehicle-treated pigs was significantly higher than that in sham controls (p = 0.03). Treatment with NACA was associated with a trend to decreased intranuclear proteolytic activity (p = 0.08), There were significantly less mutations in the mtDNA of the NACA group compared with the vehicle-treated group, 2.0 × 10-4 (±2.0 × 10-4) versus 4.8 × 10-5(±3.6 × 10-4, p < 0.05). CONCLUSION: We found a trend to lower proteolytic activity in the core of Purkinje cells and significantly reduced mutation rate of mtDNA in the NACA group, which may indicate a positive effect of NACA after neonatal hypoxia. Measuring the proteolytic activity in the nucleus of Purkinje cells could be used to assess the effect of different neuroprotective substances after perinatal asphyxia.

Glucose metabolism and body composition in young adults treated with TBI during childhood.

After SCT in childhood, survivors may develop disorders of glucose metabolism. The role of obesity is controversial. We measured insulin sensitivity using the homeostasis model assessment (HOMA) and the frequently sampled i.v. glucose tolerance test (FSIVGTT), as well as body composition using dual-energy X-ray absorptiometry in 18 young adults median 18.2 years after SCT and compared them with matched controls. We also measured growth hormone (GH) secretion, and levels of leptin and adiponectin. HOMA showed insulin resistance in eight patients (44%), as opposed to none of the controls (P=0.008) and FSIVGTT showed a decreased sensitivity index in the patients (2.98 vs 4.54 mU/L/min, P=0.042). Dual energy X-ray absorptiometry showed a higher percentage fat mass in the patients (34.9 vs 24.3%, P=0.011), which correlated inversely with the sensitivity index (r=-0.52, P=0.032). The patients had a lower peak value of GH (GH(max) 9 vs 20.7 mU/L, P=0.002). Time post SCT correlated with percentage fat mass and inversely with GH(max). The patients had higher levels of leptin and lower levels of adiponectin, even after adjustment for fat mass. We propose that the decreased insulin sensitivity may primarily be explained by the adverse body composition, which may owe to long-standing GH deficiency.

Type 1 diabetic serum interferes with pancreatic beta-cell Ca2+-handling.

The aim of this study was to clarify the frequency of patients with type 1 diabetes that have serum that increases pancreatic beta-cell cytoplasmic free Ca(2+) concentration, [Ca(2+)](i), and if such an effect is also present in serum from first-degree relatives. We also studied a possible link between the serum effect and ethnic background as well as presence of autoantibodies. Sera obtained from three different countries were investigated as follows: 82 Swedish Caucasians with newly diagnosed type 1 diabetes, 56 Americans with different duration of type 1 diabetes, 117 American first-degree relatives of type 1 diabetic patients with a mixed ethnic background and 31 Caucasian Finnish children with newly diagnosed type 1 diabetes. Changes in [Ca(2+)](i) , upon depolarization, were measured in beta-cells incubated overnight with sera from type 1 diabetic patients, first-degree relatives or healthy controls. Our data show that there is a group constituting approximately 30% of type 1 diabetic patients of different gender, age, ethnic background and duration of the disease, as well as first-degree relatives of type 1 diabetic patients, that have sera that interfere with pancreatic beta-cell Ca(2+)-handling. This effect on beta-cell [Ca(2+)](i) could not be correlated to the presence of autoantibodies. In a defined subgroup of patients with type 1 diabetes and first-degree relatives a defect Ca(2+)-handling may aggravate development of beta-cell destruction.

Lack of mutations in the TSHr and Gsalpha genes in TSHr antibody negative Graves' disease.

The aim of this study was to investigate whether TSHr antibody negative Graves' disease is associated with somatic mutations in the TSHr or Gsalpha genes and whether histopathologically defined thyroid lesions, i.e., hyperfunctioning adenoma, non-functioning follicular adenomas, or nodules in toxic and non-toxic multinodular goiters are associated with such mutations. No mutations but three germ-line polymorphisms were found in patients with TSHr antibody negative Graves' disease. The three polymorphisms are expected to have no or only minor effects on the signaling properties, and is not associated with altered antigenecity imposed by such mutations. Two heterozygous somatic TSHr mutations were found in two hyperfunctioning adenomas and in two toxic multinodular goiters. The lack of TSHr and Gsalpha mutations in TSHr antibody negative Graves' disease patients indicates that such mutations are neither primary nor secondary events in this disease. The results also confirm that somatic gain-of-function TSHr mutations are present in hyperfunctioning follicular adenomas and goiters, but not in non-functioning thyroid lesions.

Clinical, histopathologic, and genetic investigation in two large families with dentinogenesis imperfecta type II.

Dentinogenesis imperfecta (DI) type II, an inherited disorder affecting dentin, has been linked to mutations in the dentin sialophosphoprotein ( DSPP) gene on chromosome 4q21. The gene product is cleaved into two dentin-specific matrix proteins, dentin sialoprotein (DSP) and dentin phosphoprotein. The aim of this investigation was to study genotypes and phenotypes in two affected families with special reference to clinical, radiographic, and histopathologic manifestations. Seven affected members of Family A and five of Family B were documented clinically and radiographically; 14 and 10 teeth, respectively, were available for histopathologic investigation and prepared for ground sections, which were assessed semiquantitatively for dysplastic manifestations in the dentin according to the scoring system, dysplastic dentin score (DDS). Venous blood samples were collected from six affected and ten unaffected members of Family A, and from eight affected and six unaffected members of Family B. Genomic DNA was extracted and used for sequence analyses. The two families presented with different missense mutations. An Arg68Trp missense mutation in the DSP part of the gene was revealed in all six analyzed affected individuals in Family A. This mutation was not present in any of the ten healthy members. In Family B, an Ala15Val missense mutation involving the last residue of the signal peptide was found in all eight affected but in none of the six healthy members. The clinical and radiographic disturbances and DDS were more severe in Family B. The data indicate the presence of a genotype-phenotype correlation in DI type II.

Orlistat treatment in obese prepubertal children: a pilot study.

AIM: This study investigated orlistat treatment in obese prepubertal children with regard to tolerance, safety and psychological well-being. METHODS: 11 healthy, severely obese prepubertal children (age 8.3-12.3 y, body mass index standard deviation score 5.3-9.2) were recruited for a 12 wk open treatment. Before, during and after treatment, the participants were investigated by psychological evaluation, blood chemistry, and parameters reflecting obesity and fat mass. RESULTS: The participants were able to comply with the treatment, as indicated by pill counts and self reports, and expressed a desire to continue the treatment after the study period. Gastrointestinal side effects were mild and tolerable. No negative effects on psychological or physical well-being were detected, and the psychological evaluation demonstrated increased avoidance of fattening food, body shape preoccupation and oral control (p = 0.011). The median weight loss was 4.0 kg (range -12.7 to +2.5 kg, p = 0.016) and was highly correlated to decreased fat mass (regression coefficient 0.953, p < 0.01). CONCLUSION: This pilot study indicates that obese prepubertal children were able to reduce their fat intake to avoid gastrointestinal side effects. Thus, orlistat may be suitable as a component in behaviour-modification programmes for obese children, and the results prompt a placebo-controlled investigation of its effectiveness in promoting weight loss.

Glutathione synthetase deficiency: is gamma-glutamylcysteine accumulation a way to cope with oxidative stress in cells with insufficient levels of glutathione?

Glutathione (GSH) plays a major role in the cellular defence against oxidative stress and other vital cellular functions. It therefore seems inevitable that patients with severe depletion of GSH will not survive. However, at least some with glutathione synthetase (GS) deficiency do. This study was done to determine whether these patients have a mechanism to compensate for their GSH deficiency. Cell-free extracts of cultured fibroblasts from 9 patients with GS deficiency and 9 control subjects were analysed by HPLC for low-molecular-weight thiol compounds. The patients' cells contained 7.4 nmol of GSH per mg of protein (median; range 2.8-25.2) compared to 33.0 nmol in control fibroblasts (range 26.7-51.4) (p < 0.01). On the other hand, the patients' cells accumulated 18.1 nmol of gamma-glutamylcysteine (gamma-GC) per mg of protein (median; range 6.9-71.7), whereas the control cells contained 0.1 nmol (range 0.05-0.16) (p < 0.01). The cysteine concentrations in the patients' cells were 20.7 nmol/mg protein (median; range 9.4-52.9) compared to 8.9 nmol in control cells (range 3.0-12.4) (p < 0.01). Cultured fibroblasts from patients with GS deficiency have low levels of GSH, but instead accumulate gamma-GC. We suggest that gamma-GC, which contains both reactive groups of GSH (i.e. the sulphydryl and gamma-glutamyl groups), can compensate for GSH in the cellular defence against oxidative stress. Thus, gamma-GC may alleviate, but only partly prevent, serious consequences of insufficient GSH levels in affected patients. Since the sum of the levels of GSH and gamma-GC in GS-deficient cells (median 31.5 nmol/mg protein, range 16.2-79.0) was similar to the level of GSH alone in control cells (33.0 nmol/mg protein, range 26.7-51.4), we propose that the cultured fibroblasts may have a mechanism to regulate in a coordinated way the levels of GSH and gamma-GC; for instance, by both compounds acting as feedback inhibitors of gamma-GC synthetase.

Cooperative binding of gamma-glutamyl substrate to human glutathione synthetase.

Human glutathione synthetase is responsible for catalyzing the final step in glutathione biosynthesis. It is a homodimer with a monomer subunit MW of 52 kDa. Kinetic analysis reveals a departure from linearity of the Lineweaver-Burk double reciprocal plot for the binding of gamma-glutamyl substrate, indicating cooperative binding. The measured apparent K(m) values for gamma-glutamyl-alpha-aminobutyrate (an analog of gamma-glutamyl-alpha-aminobutyrate) are 63 and 164 microM, respectively. Neither ATP (K(m) of 248 microM) nor glycine (K(m) of 452 microM) exhibits such cooperative binding behavior. Although ATP is proposed to play a key role in the sequential binding of gamma-glutamyl substrate to the enzyme, the cooperative binding of the gamma-glutamyl substrate is not affected by alterations of ATP concentration. Quantitative analysis of the kinetic results for gamma-glutamyl substrate binding gives a Hill coefficient (h) of 0.75, indicating negative cooperativity. Our studies, for the first time, show that human glutathione synthetase is an allosteric enzyme with cooperative binding for gamma-glutamyl substrate.

Effects of growth hormone treatment on the leptin system and body composition in obese prepubertal boys.

UNLABELLED: Leptin correlates with measures of body fat stores. Growth hormone (GH) treatment may affect leptin levels either directly or indirectly by influencing body composition and circulating insulin level. Here, the effects of GH treatment on the leptin axis and body composition of six severely obese, but otherwise healthy, prepubertal boys were studied. Fasting serum leptin was significantly reduced after only 3 wk of GH treatment. Body fat percentage, but not BMI, decreased significantly (p < 0.05) after 3 mo. The serum leptin concentration per unit fat mass decreased significantly during GH treatment (p < 0.05), suggesting that such treatment might have a direct effect on serum leptin independently of the effects on body composition. Leptin RNA expression in abdominal subcutaneous tissue was not significantly changed by treatment. CONCLUSIONS: The data indicate that GH has an early downregulatory effect on the circulating leptin level independently of the concomitant changes in body composition. Whether GH affects leptin production or metabolism needs further study.

Total parenteral nutrition after surgery rapidly increases serum leptin levels.

OBJECTIVE: In humans, leptin is regulated by long-term changes in energy intake. However, short-term regulation of serum leptin by nutrients has been difficult to show. The aim of this study was to investigate whether short periods of fasting and stress sensitise the leptin response to nutrients. SUBJECTS AND EXPERIMENTAL PROTOCOL: Fourteen patients of normal weight undergoing elective open cholecystectomy were randomised into two groups. One group received saline infusion during surgery and for 24 h postoperatively. The other group also received saline during the surgical procedure, but total parenteral nutrition (TPN) was started immediately after surgery. Blood samples were drawn before as well as 2, 4, 8, 16, and 24 h after the start of surgery to determine the serum levels of leptin and other hormones. RESULTS: Postoperative TPN induced a significant rise in serum leptin within 6 h, reaching a more than fourfold increase within 14 h (P<0.001). Serum glucose and insulin levels increased within 2 h. Growth hormone and IGF-1 serum levels also increased significantly in the group receiving TPN. Serum cortisol levels increased postoperatively in both groups, which may explain why no significant reduction in serum leptin was observed in the group receiving saline. Free tri-iodothyronine (T3) decreased in both groups, while catecholamines were similar in the groups. CONCLUSION: During fasting and surgical stress, nutrients rapidly increased the serum leptin levels in humans in a manner similar to that previously reported in rodents. This may be mediated by increases in serum glucose, insulin and cortisol.

Insulin-degrading enzyme identified as a candidate diabetes susceptibility gene in GK rats.

Genetic analysis of the diabetic GK rat has revealed several diabetes susceptibility loci. Congenic strains have been established for the major diabetes locus, Niddm1, by transfer of GK alleles onto the genome of the normoglycemic F344 rat. Niddm1 was dissected into two subloci, physically separated in the congenic strains Niddm1b and Niddm1i, each with at least one disease susceptibility gene. Here we have mapped Niddm1b to 1 cM by genetic and pathophysiological characterization of new congenic substrains for the locus. The gene encoding insulin-degrading enzyme (IDE:) was located to this 1 cM region, and the two amino acid substitutions (H18R and A890V) identified in the GK allele reduced insulin-degrading activity by 31% in transfected cells. However, when the H18R and A890V variants were studied separately, no effects were observed, demonstrating a synergistic effect of the two variants on insulin degradation. No effect on insulin degradation was observed in cell lysates, indicating that the effect is coupled to receptor-mediated internalization of insulin. Congenic rats with the IDE: GK allele displayed post-prandial hyperglycemia, reduced lipogenesis in fat cells, blunted insulin-stimulated glucose transmembrane uptake and reduced insulin degradation in isolated muscle. Analysis of additional rat strains demonstrated that the dysfunctional IDE: allele was unique to GK. These data point to an important role for IDE: in the diabetic phenotype in GK.

Kinetic properties of missense mutations in patients with glutathione synthetase deficiency.

Patients with hereditary glutathione synthetase (GS) (EC 6.3.2.3) deficiency present with variable clinical pictures, presumably related to the nature of the mutations involved. In order to elucidate the relationship between genotype, enzyme function and clinical phenotype, we have characterized enzyme kinetic parameters of missense mutations R125C, R267W, R330C and G464V from patients with GS deficiency. One of the mutations predominantly affected the K(m) value, with decreased affinity for glycine, two mutations influenced both K(m) and V(max) values, and one mutation reduced the stability of the enzyme. This characterization agrees well with predictions based on the recently reported crystal structure of human GS. Thus our data indicate that different mutations can affect the catalytic capacity of GS by decreasing substrate affinity, maximal velocity or enzyme stability.

Effects of growth hormone treatment in obese prepubertal boys.

Childhood obesity is associated with several abnormalities of the GH axis, including decreased spontaneous secretion, decreased response to exogenous secretagogues, and altered pulsatile pattern of secretion. In adults, GH treatment reduces abdominal obesity and improves insulin sensitivity, as well as blood lipid profiles. Whether GH has similar effects in obese children has not been investigated previously. In this study, seven prepubertal severely obese boys aged 10-12 yr were treated with GH for 6 months and followed for an additional 6 months. No diet or exercise modifications were initiated. Body fat percentage decreased from 51.3% to 46.1% after treatment (P = 0.03). Frequently sampled iv glucose tolerance tests revealed an increased responsivity of the acute insulin secretion (P = 0.04) and a nonsignificant trend toward improved insulin sensitivity. In isolated adipocytes, the maximum isoprenaline- and terbutaline-induced lipolysis were increased approximately 2.5-fold (P = 0.02). The sensitivity of the adipocytes to isoprenaline was unchanged, whereas the sensitivity to terbutaline was increased (P = 0.04). No effect was observed on basal or insulin-stimulated lipogenesis. In conclusion, GH treatment for 6 months of obese prepubertal boys reduces body fat, possibly, via stimulation of catecholamine-induced lipolysis, without negative effects on glucose homeostasis.

Pathophysiological and genetic characterization of the major diabetes locus in GK rats.

Genetic studies of the type 2 diabetes-like GK rat have revealed several susceptibility loci for the compound diabetes phenotype. Congenic strains were established for Niddm1, the major quantitative trait locus (QTL) for postprandial glucose levels, by transfer of GK alleles onto the genome of the normoglycemic F344 rat. Despite the polygenic nature of diabetes in GK, the locus-specific diabetes phenotype was retained in the congenic strain Niddmla, containing a GK-derived genomic fragment of 52 cM from the Niddm1 locus. Furthermore, Niddm1 was divided into two non-overlapping loci, physically separated in the two congenic strains Niddmlb and Niddm1i with distinct metabolic phenotypes. Both strains displayed postprandial hyperglycemia and reduced insulin action in isolated adipose cells. Furthermore, Niddm1i already exhibits a pronounced in vivo insulin secretion defect at 65 days, while Niddm1b develops a relative insulin secretory defect at 95 days. This suggests that Niddm1i impairs mechanisms common to insulin secretion in pancreatic B-cells and insulin action in adipocytes. Niddm1b rats show signs of increasing insulin resistance with age associated with obesity, hyperinsulinemia, and dyslipidemia. Moreover, the data indicated nonallelic interaction (epistasis) between Niddm1b and Niddm1i on the postprandial glucose levels. These data emphasize the pathophysiological complexity of diabetes, even within an apparently single QTL, and demonstrate the potential of the GK model in transforming the multifactorial diabetes phenotype into single traits, suitable for positional cloning.

Insulin induced hypoglycaemia: comparison of glucose and glycerol concentrations in plasma and microdialysate from subcutaneous adipose tissue.

AIMS: To investigate the dynamics between plasma and dialysate glucose during hypoglycaemia in children. STUDY DESIGN: Six children in prepuberty or early puberty were investigated by multiple blood sampling and microdialysis of subcutaneous adipose tissue during a standard arginine-insulin tolerance test. Glucose and glycerol, as an index of lipolysis, were measured in samples from both compartments. Plasma concentrations of insulin and the main counterregulatory hormones were also measured. RESULTS: Plasma and dialysate glucose concentrations were very similar at baseline and increased in concert after infusion of arginine, probably in response to glucagon release. After insulin injection, glucose in both plasma and dialysate fell in parallel. The subsequent hypoglycaemic stress response induced a rapid rebound in the plasma concentration with a mean (SD) delay in the dialysate of 16 (3) minutes. Plasma glycerol was approximately fivefold lower than in the dialysate and did not fluctuate significantly. Dialysate glycerol decreased with arginine infusion and reached a nadir immediately following insulin administration. Subsequently, the antilipolytic effect of insulin was overcome by the hypoglycaemic stress response, and lipolysis prevailed in spite of hyperinsulinaemia. CONCLUSION: After rapidly induced hypoglycaemia, rebound of interstitial glucose concentrations is significantly delayed compared with plasma concentrations, and the antilipolytic effect of hyperinsulinaemia is opposed possibly by the hypoglycaemic stress response.

Growth hormone treatment downregulates serum leptin levels in children independent of changes in body mass index.

The changes in serum leptin levels during growth hormone (GH) treatment were studied in 27 children, 17 with GH deficiency (GHD), 10 with idiopathic short stature (ISS), and 9 with Prader-Willi syndrome (PWS). Within 1 month of GH treatment, serum leptin levels decreased by 40% in the GHD children (p < 0.01). There was no significant change in serum leptin level in the children with ISS. In children with PWS, the mean serum leptin level decreased by almost 60% after 3 months of treatment (p < 0.001). Thereafter, no further decline was observed in any of the 3 groups. Changes in body composition became evident first after the 3 months of treatment. In the GHD children, the BMI was unchanged while the mean body fat percentage was 2.7% lower after 1 year of GH treatment (p < 0.05). In the ISS children, neither BMI nor body fat percentage were significantly changed during treatment. The PWS children exhibited a significant decrease in BMI after 6 months of GH treatment without any further change during the remaining period of treatment. In this group, the mean body fat percentage decreased from 42 +/- 2.4 to 28 +/- 2.2% after treatment (p < 0.001). The finding that the fall in leptin occurs before changes in body composition become detectable suggests a direct effect of GH on leptin production, metabolism, or clearance.

Effect of growth hormone treatment on insulin action in adipocytes from children with Prader-Willi syndrome.

OBJECTIVE: To study the effect of growth hormone (GH) treatment (2-4 months) on insulin action in adipocytes isolated from children with Prader-Willi syndrome (PWS), in whom GH deficiency appears to be a primary defect. We investigated the complex effects of GH on carbohydrate metabolism, as part of a current clinical trial of GH treatment in children with PWS. METHODS: Biopsies of subcutaneous abdominal adipose tissue were obtained from 12 children with PWS before and after 2-4 months of GH treatment. Lipogenesis was determined by the incorporation of radiolabelled glucose into lipids in isolated adipocytes, and glycerol release to the incubation medium was used as an index of lipolysis. GLUT4 RNA was measured by solution hybridization. RESULTS: With low glucose concentrations, at which glucose transport is rate-limiting, maximal insulin-induced lipogenesis was increased by 120% after GH treatment (P < 0.05), but the sensitivity to insulin (half-maximum effective hormone concentration) was unchanged. This was not accompanied by a significant change in the RNA expression of GLUT4. Neither responsiveness (maximum effect) nor sensitivity of insulin-induced inhibition of lipolysis was affected by GH treatment. CONCLUSIONS: GH treatment of children with PWS results in an upregulation of insulin-induced lipogenesis in isolated adipocytes, with no effect on insulin-induced inhibition of lipolysis. The data suggest that the site of the effect of GH on lipogenesis is distal to the insulin hormone-receptor interaction, but does not involve altered GLUT4 expression.

Changes in cytoplasmic ATP concentration parallels changes in ATP-regulated K+-channel activity in insulin-secreting cells.

Changes in cytoplasmic ATP concentration were monitored in intact insulin-producing cells and correlated to changes in the activity of ATP-sensitive K+-channels (KATP channels). Luciferase was introduced into HIT M2.2 cells and whole pancreatic islets by transient expression of firefly (Photinus pyralis) luciferase cDNA. In transfected cells, extracellular addition of luciferin increased the luminescence signal to a maximum within 50-120 s. Addition of 1 microM of the mitochondrial uncoupler FCCP decreased the luminescence, an effect partly reversed upon withdrawal of the compound. High concentrations of glucose increased cytoplasmic free ATP concentration. Changes in the luminescence signal were accompanied by changes in activity of the ATP-sensitive K+-channel. Transfection per se did not deteriorate cell function, as verified by experiments showing similar changes in cytoplasmic free Ca2+-concentration, [Ca2+li, in both transfected and non-transfected cells. By measuring the cytoplasmic ATP concentration and KATP channel activity under similar experimental conditions, it was possible to establish, for the first time, a direct relationship between these two parameters. This indeed suggests that the cytoplasmic ATP concentration has a crucial role in the regulation of KATP channel activity under physiological conditions.

Antilipolytic effect of insulin and insulin receptor messenger RNA expression in adipocytes of infants, children, and adults.

Human adipose tissue exhibits marked age-dependent differences in morphology and regulation of lipolysis. The mechanism behind this phenomenon is poorly characterized. The aim of the present investigation was to evaluate the antilipolytic effect of insulin and the expression of insulin receptor mRNA in human adipose tissue during development. To study the antilipolytic effect of insulin, lipolysis was induced with submaximal effective concentrations of isoprenaline or forskolin in adipocytes from five infants below 2 mo of age, five infants above 2 mo of age, six children, and five adults. Isoprenaline was equally effective in stimulating lipolysis in all age groups, whereas forskolin was significantly more effective in adipocytes isolated from adults than from infants and children. Increasing the concentration of insulin during the incubation allowed construction of dose-response curves of insulin inhibition of lipolysis. Maximal inhibition was observed with 30 microU/mL of insulin in the presence of either isoprenaline or forskolin. No differences in the maximal antilipolytic effect of insulin or in the insulin sensitivity expressed as ED50 were observed. Furthermore, the expression of insulin receptor mRNA in adipocytes did not differ from six infants, five children, and six adults as determined with a solution hybridization RNase protection assay. Thus, age-dependent variations in the regulation of lipolysis do not appear to be accompanied by variations in insulin action and insulin receptor gene expression in isolated adipocytes.

B cells of aging rats: impaired stimulus-secretion coupling but normal susceptibility to adverse effects of a diabetic state.

A diabetic state impairs B-cell function and survival. We tested whether the negative effects are exacerbated by the aging process. Islets were isolated from old (63.3 +/- 2.3 weeks) and young (11.3 +/- 0.5 weeks) inbred Wistar rats. Age did not affect DNA and insulin content, yet both glucose-induced (27.8 mmol/L) and arginine-induced induced (10 mmol/L) insulin responses in old islets were significantly reduced. Islets were transplanted under the kidney capsule of recipients that were either nondiabetic or severely diabetic after streptozotocin (STZ) treatment (blood glucose > 20 mmol/L). Following 8 weeks' transplantation to nondiabetic recipients, perfused kidneys with grafts of old islets exhibited the same insulin responses to glucose as grafts of young islets. However, responses to arginine were reduced in grafts of old islets (28 +/- 4 microU/min) relative to grafts of young islets. (70 +/- 18 microU/min, P < .05). Insulin mRNA content was similar in grafts of old islets and grafts of young islets. Following 8 weeks' transplantation to diabetic recipients, 27.8 mmol/L glucose failed to induce insulin secretion in grafts of old islets and grafts of young islets alike, whereas arginine-induced insulin secretion was unaffected in grafts of old islets but reduced in grafts of young islets. Insulin mRNA content was reduced to a similar extent by the diabetic state (to 28% in grafts of old islets and to 27% in grafts of young islets grafts in nondiabetic recipients). We conclude that aging, although leading to impaired stimulus-secretion coupling, does not increase susceptibility to the negative effects of a diabetic state on B-cell function as presently tested.