Hypoinsulinemic Diabetes Research Articles

The spontaneous-diabetes rat: a model of noninsulin dependent diabetes mellitus by Yoshio Goto and Masaei Kakizaki (1981)

Y. Goto and M. Kakizaki produced a rat model of non-insulin dependent diabetes mellitus (NIDDM) by repetitive selective breeding of rats with slightly impaired glucose tolerance. In contrast to most obese diabetes models, which were genetically modified animals created by inducing a gene mutation, this rat was a unique model because it was a spontaneous diabetes model created by selective breeding. Furthermore, when it became clear that this rat was a non-obese diabetic model that exhibited hyperglycemia due to a decrease in insulin secretion capacity, it was recognized as a valuable model for elucidating non-obese, hypoinsulinemic diabetes commonly seen in East Asians. Diabetes is becoming an important health problem worldwide, especially in the East Asian region, thus Goto-Kakizaki (GK) rats are becoming increasingly important as a model of non-insulin dependent diabetes mellitus.

P2Y1 purinergic receptor identified as a diabetes target in a small-molecule screen to reverse circadian β-cell failure.

The mammalian circadian clock drives daily oscillations in physiology and behavior through an autoregulatory transcription feedback loop present in central and peripheral cells. Ablation of the core clock within the endocrine pancreas of adult animals impairs the transcription and splicing of genes involved in hormone exocytosis and causes hypoinsulinemic diabetes. Here, we developed a genetically sensitized small-molecule screen to identify druggable proteins and mechanistic pathways involved in circadian β-cell failure. Our approach was to generate β-cells expressing a nanoluciferase reporter within the proinsulin polypeptide to screen 2640 pharmacologically active compounds and identify insulinotropic molecules that bypass the secretory defect in CRISPR-Cas9-targeted clock mutant β-cells. We validated hit compounds in primary mouse islets and identified known modulators of ligand-gated ion channels and G-protein-coupled receptors, including the antihelmintic ivermectin. Single-cell electrophysiology in circadian mutant mouse and human cadaveric islets revealed ivermectin as a glucose-dependent secretagogue. Genetic, genomic, and pharmacological analyses established the P2Y1 receptor as a clock-controlled mediator of the insulinotropic activity of ivermectin. These findings identify the P2Y1 purinergic receptor as a diabetes target based upon a genetically sensitized phenotypic screen.

OR14-02 Circadian Regulation of Chromatin State Mediates Pancreatic Islet Incretin Response

The circadian clock is programmed by an autoregulatory transcription feedback loop present in brain and peripheral tissues that coordinates metabolism with nutritional state and the sleep-wake cycle. Epidemiologic and genetic studies indicate circadian disruption as a risk factor in the development of diabetes. We have demonstrated that conditional ablation of the β cell clock in adult life leads to hypoinsulinemic diabetes, and through mRNA-sequencing in mouse and human islets we revealed clock control of gene networks involved in insulin secretion, nutrient sensing, and exocytosis. A remaining question is: How does the core molecular clock modulate time-of-day dependent chromatin state to regulate pancreatic islet response to glucose and insulin secretagogues? Here we report that loss of the pancreatic β cell molecular clock results in closed chromatin at cAMP-responsive gene regulatory elements and dysregulated cAMP-dependent coregulator recruitment following cAMP agonism, consistent with a role for the molecular clock in mediating cell response to environmental stimuli. Further, tandem analyses of ATAC- and ChIP-sequencing in synchronized islets revealed dynamic chromatin accessibility across the 24-hour cycle at genes regulating insulin secretion and at genomic regions enriched for signal-inducible and circadian transcription factor motifs. Our genome-wide sequencing reveals a new role for the clock in global chromatin remodeling underlying the incretin response in pancreatic β cells.

Deficiency in prohormone convertase PC1 impairs prohormone processing in Prader-Willi syndrome.

Prader-Willi syndrome (PWS) is caused by a loss of paternally expressed genes in an imprinted region of chromosome 15q. Among the canonical PWS phenotypes are hyperphagic obesity, central hypogonadism, and low growth hormone (GH). Rare microdeletions in PWS patients define a 91-kb minimum critical deletion region encompassing 3 genes, including the noncoding RNA gene SNORD116. Here, we found that protein and transcript levels of nescient helix loop helix 2 (NHLH2) and the prohormone convertase PC1 (encoded by PCSK1) were reduced in PWS patient induced pluripotent stem cell-derived (iPSC-derived) neurons. Moreover, Nhlh2 and Pcsk1 expression were reduced in hypothalami of fasted Snord116 paternal knockout (Snord116p-/m+) mice. Hypothalamic Agrp and Npy remained elevated following refeeding in association with relative hyperphagia in Snord116p-/m+ mice. Nhlh2-deficient mice display growth deficiencies as adolescents and hypogonadism, hyperphagia, and obesity as adults. Nhlh2 has also been shown to promote Pcsk1 expression. Humans and mice deficient in PC1 display hyperphagic obesity, hypogonadism, decreased GH, and hypoinsulinemic diabetes due to impaired prohormone processing. Here, we found that Snord116p-/m+ mice displayed in vivo functional defects in prohormone processing of proinsulin, pro-GH-releasing hormone, and proghrelin in association with reductions in islet, hypothalamic, and stomach PC1 content. Our findings suggest that the major neuroendocrine features of PWS are due to PC1 deficiency.

Increased Spontaneous Central Bleeding and Cognition Impairment in APP/PS1 Mice with Poorly Controlled Diabetes Mellitus.

Alzheimer’s disease (AD) and vascular dementia (VaD) are the most common causes of dementia, and borderlines are blurred in many cases. Aging remains the main risk factor to suffer dementia; however, epidemiological studies reveal that diabetes may also predispose to suffer AD. In order to further study this relationship, we have induced hypoinsulinemic diabetes to APPswe/PS1dE9 (APP/PS1) mice, a classical model of AD. APP/PS1 mice received streptozotocin (STZ) ip at 18 weeks of age, when AD pathology is not yet established in this animal model. Cognition was evaluated at 26 weeks of age in the Morris water maze and the new object discrimination tests. We observed that STZ-induced episodic and working memory impairment was significantly worsened in APP/PS1 mice. Postmortem assessment included brain atrophy, amyloid-beta and tau pathology, spontaneous bleeding, and increased central inflammation. Interestingly, in APP/PS1-STZ diabetic mice, we detected a shift in Aβ soluble/insoluble levels, towards more toxic soluble species. Phospho-tau levels were also increased in APP/PS1-STZ mice, accompanied by an exacerbated inflammatory process, both in the close proximity to senile plaque (SP) and in SP-free areas. The presence of hemorrhages was significantly higher in APP/PS1-STZ mice, and although pericytes and endothelium were only partially affected, it remains possible that blood-brain barrier alterations underlie observed pathological features. Our data support the implication of the diabetic process in AD and VaD, and it is feasible that improving metabolic control could delay observed central pathology.

Diabetic liver injury from streptozotocin is regulated through the caspase-8 homolog cFLIP involving activation of JNK2 and intrahepatic immunocompetent cells

The endemic occurrence of obesity and the associated risk factors that constitute the metabolic syndrome have been predicted to lead to a dramatic increase in chronic liver disease. Non-alcoholic steatohepatitis (NASH) has become the most frequent liver disease in countries with a high prevalence of obesity. In addition, hepatic steatosis and insulin resistance have been implicated in disease progression of other liver diseases, including chronic viral hepatitis and hepatocellular carcinoma. The molecular mechanisms underlying the link between insulin signaling and hepatocellular injury are only partly understood. We have explored the role of the antiapoptotic caspase-8 homolog cellular FLICE-inhibitory protein (cFLIP) on liver cell survival in a diabetic model with hypoinsulinemic diabetes in order to delineate the role of insulin signaling on hepatocellular survival. cFLIP regulates cellular injury from apoptosis signaling pathways, and loss of cFLIP was previously shown to promote injury from activated TNF and CD95/Apo-1 receptors. In mice lacking cFLIP in hepatocytes (flip−/−), loss of insulin following streptozotocin treatment resulted in caspase- and c-Jun N-terminal kinase (JNK)-dependent liver injury after 21 days. Substitution of insulin, inhibition of JNK using the SP600125 compound in vivo or genetic deletion of the mitogen-activated protein kinase (MAPK)9 (JNK2) in all tissues abolished the injurious effect. Strikingly, the difference in injury between wild-type and cFLIP-deficient mice occurred only in vivo and was accompanied by liver-infiltrating inflammatory cells with a trend toward increased amounts of NK1.1-positive cells and secretion of proinflammatory cytokines. Transfer of bone marrow from rag-1-deficient mice that are depleted from B and T lymphocytes prevented liver injury in flip−/− mice. These findings support a direct role of insulin on cellular survival by alternating the activation of injurious MAPK, caspases and the recruitment of inflammatory cells to the liver. Thus, increasing resistance to insulin signaling pathways in hepatocytes appears to be an important factor in the initiation and progression of chronic liver disease.

Biological Timekeeping in Obesity and Diabetes

Circadian systems are programmed by a transcription‐translation feedback loop that generates daily cycles of energy storage and utilization in synchrony with the 24 hr rotation of the Earth. In mammals, master pacemaker neurons located within the suprachiasmatic nucleus are entrained to light, and generate 24 hr oscillations in extra‐SCN neurons and most peripheral tissues, including those involved in glucose and lipid metabolism. During wakefulness and feeding, the molecular clock promotes glucose‐stimulated insulin secretion within b‐cells of the Islets of Langerhans. Clock ablation limited to pancreas results in hypoinsulinemic diabetes, indicating a primary function of peripheral clocks in glucose homeostasis. In contrast, ablation of the liver clock impairs oxidative metabolism, indicating that peripheral tissue clocks exert opposing effects in response to feeding and fasting. At the molecular level, clock disruption causes reduced NAD+ biosynthesis and leads to alterations in function of NAD+‐dependent ribosylases and de‐acetylases involved in oxidative metabolism. Circadian systems thus contribute to long‐term energy and glucose homeostasis across alternating behavioral and nutrient states.

Altered dipeptidyl peptidase-4 activity during the progression of hyperinsulinemic obesity and islet atrophy in spontaneously late-stage type 2 diabetic rats

Altered dipeptidyl peptidase-4 (DPP4) activity during the progression of late-stage type 2 diabetes was measured in Otsuka Long-Evans Tokushima fatty (OLETF) rats. Compared with OLETF rats subjected to 30% food restriction, food-satiated OLETF rats exhibited spontaneous hyperphagic obesity, insulin resistance, hyperglycemia, hyperinsulinemia, and increased plasma DPP4 activity during the early phase of the experiment (up to ∼30 wk). Subsequently, their plasma DPP4 activity as well as their body weight, body fat, and plasma insulin concentration declined to control levels during the late phase, resulting in excessive polyuria, proteinuria, dyslipidemia, pancreatic islet atrophy, hypoinsulinemia, and diabetes, which changed from insulin-resistant diabetes to hypoinsulinemic diabetes secondary to progressive islet insufficiency, and their fasting blood glucose level remained high. Since plasma DPP4 activity demonstrated significant positive correlations with body weight and the fasting plasma insulin level but not with the fasting blood glucose level during the late stage of diabetes, body fat and fasting plasma insulin levels may be useful factors for predicting the control of plasma DPP4 activity. In contrast, pancreatic DPP4 activity was significantly increased, and the expression of pancreatic insulin was significantly reduced in late-stage diabetic OLETF rats, suggesting that a relationship exists between the activation of pancreatic DPP4 and insulin depletion in pancreatic islet atrophy. In conclusion, it is suggested that plasma DPP4 activity changes in accordance with the progression of hyperinsulinemic obesity and pancreatic islet atrophy. DPP4 activity may play an important role in insulin homeostasis.

Enhanced tumorigenesis of forestomach tumors induced by N‐Methyl‐N′‐nitro‐N‐nitrosoguanidine in rats with hypoinsulinemic diabetes

Hyperinsulinemia and hyperglycemia in prediabetic and diabetic patients are thought to increase the risk of developing neoplasms because insulin is a growth factor with pre-eminent metabolic but also mitogenic effects. To determine the effect of hypoinsulinemic diabetic conditions on carcinogenesis, we examined N-Methyl-N'-nitro-N-nitrosoguanidine (MNNG)-induced forestomach carcinogenesis in hypoinsulinemic diabetic WBN/Kob rats aged about 45 weeks (DM) compared with non-diabetic younger WBN/Kob rats (C1), non-diabetic Wistar rats age-matched to DM (C2), and non-diabetic Wistar rats age-matched to C1 (C3). All rats were treated with MNNG by gavage and were killed at 40 weeks after dosing. Various-sized tumors were disseminated throughout the forestomach of all rats, and the ratio of the area of tumors to the whole forestomach area was 23.3% in the DM group and was higher than in the C1-3 (4.2-14.3%) groups. The incidence of carcinoma was much higher in the DM group (36.8%) than in the C1-3 (7.1-16.7%) groups, and the incidence of papilloma was also significantly higher in the DM group (84.2%) than in the C1-3 (28.5-50.0%) groups. The average thickness of the squamous epithelium in the non-neoplastic mucosa was significantly greater in the DM group (50.8 mum) than in the C1-3 (29.6-37.9 microm) groups. Immunohistochemically, the Ki-67-positive index in the non-tumorous mucosa of the DM group (42.0%) was significantly higher than that of the C1-3 groups (18.8-33.3%). These results suggest that prolonged hyperglycemic conditions without hyperinsulinemia enhance tumorigenesis of MNNG-induced tumors by enhanced proliferative activity of the squamous epithelium in the rat forestomach.

Characterization of diabetic nephropathy in a transgenic model of hypoinsulinemic diabetes

Genetic mouse models provide a unique opportunity to investigate gene function in the natural course of the disease. Although diabetic nephropathy (DN) in models of type II diabetes has been well characterized, diabetic renal disease in hypoinsulinemic diabetic mice is still incompletely understood. Here, we characterized renal changes in the pdx1(PB)-HNF6 transgenic mouse that exhibits beta-cell dysfunction and nonobese hypoinsulinemic diabetes. Male transgenic mice developed hyperglycemia by the age of 7 wk and survived for over 1 yr without insulin treatment. Diabetes ensued earlier and progressed more severely in the HNF6 males than the females. The HNF6 males exhibited albuminuria as early as 10 wk of age, and the urinary albumin excretion increased with age, exceeding 150 microg/24 h at 11 mo of age. Diabetic males developed renal hypertrophy after 7 wk of age, whereas glomerular hyperfiltration was not observed in the mice. Hypertension and hyperlipidemia were not observed in the diabetic mice. Histological analysis of the HNF6 kidneys displayed diabetic glomerular changes, including glomerular enlargement, diffuse mesangial proliferation and matrix expansion, thickened glomerular basement membrane, and arteriolar hyalinosis. Mesangial matrix accumulation increased with age, resulting in nodular lesions by 44 wk of age. Immunohistochemistry showed accumulation of type IV collagen and TGF-beta1 in the mesangial area. No significant immune complex deposition was observed in the HNF6 glomeruli. Thus the HNF6 mouse exhibits diabetic renal changes that parallel the early phase of human DN. The model should facilitate studies of genetic and environmental factors that may affect DN in hypoinsulinemic diabetes.

Skeletal muscle basal AMP-activated protein kinase activity is chronically elevated in alloxan-diabetic dogs: impact of exercise.

The effect of diabetes and exercise on skeletal muscle (SkM) AMP-activated protein kinase (AMPK)alpha1 and -alpha2 activities and site-specific phosphorylation of acetyl-CoA carboxylase was examined in the same six dogs before alloxan (35 mg/kg)-induced diabetes (C) and after 4-5 wk of suboptimally controlled hyperglycemic and hypoinsulinemic diabetes (DHG) in the presence and absence of 300-min phlorizin (50 microg.kg-1.min-1)-induced "normoglycemia" (DNG). In each study, the dog underwent a 150-min [3-3H]glucose infusion period, followed by a 30-min treadmill exercise test (60-70% maximal oxygen capacity) to measure the rate of glucose disposal into peripheral tissues (Rdtissue). SkM biopsies were taken from the thigh (vastus lateralis) before and immediately after exercise. In the C and DHG states, the rise in plasma free fatty acids (FFA) with exercise ( approximately 40%) was similar. In the DNG group, preexercise FFA were significantly higher, but the absolute rise in FFA with exercise was similar. However, the exercise-induced increment in Rdtissue was significantly blunted (by approximately 40-50%) in the DNG group compared with the other states. In SkM, preexercise AMPKalpha1 and -alpha2 activities were significantly elevated (by approximately 60-125%) in both diabetic states, but unlike the C group these activities did not rise further with exercise. Additionally, preexercise acetyl-CoA carboxylase phosphorylation in both diabetic states was elevated by approximately 70-80%, but the increases with exercise were similar to the C group. Preexercise AMPKalpha1 and -alpha2 activities were negatively correlated with Rdtissue during exercise for the combined groups (both P < 0.02). In conclusion, the elevated preexercise SkM AMPKalpha1 and -alpha2 activities contribute to the ongoing basal supply of glucose and fatty acid metabolism in suboptimally controlled hypoinsulinemic diabetic dogs; but whether they also play a permissive role in the metabolic stress response to exercise remains uncertain.

Peripheral neuropathy in transgenic diabetic mice: restoration of C-fiber function with human recombinant nerve growth factor.

Mice (Ins.Dd1) with hypoinsulinemic diabetes were created by increased expression of syngeneic major histocompatibility complex (MHC) class I protein in pancreatic beta-cells. The diabetic state was characterized in these mice by high glucose concentrations and islet pathology. To determine whether a neuropathy would develop, motor and sensory conduction velocities (CV) were determined in the sciatic nerves of 2-, 4-, and 7-month-old control and diabetic littermate male mice. Recording bipolar electrodes were placed in the plantar muscles of the hind foot of anesthetized (ketamine/xylazine) mice. Bipolar stimulating electrodes were positioned near the sciatic nerve at the sciatic notch or near the tibial nerve at the ankle. Motor CV from alpha-motor fibers and sensory CV from proprioceptive Aalpha nerves were measured and expressed as meters per second (m/s). Group data are reported as mean +/- SE and compared by analysis of variance. The CVs from nondiabetic mice (controls) were not different across the three ages and averaged 41.3 +/- 1.7 m/s for motor and 38.7 +/- 1.7 m/s for sensory. The motor CVs from diabetic mice at 2 and 4 months were similar to controls. Sensory CVs were unchanged at 2 months but were lower at 4 months (18.9 +/- 2.4 m/s). Both sensory (23.9 +/- 2.1 m/s) and motor (18.9 +/- 1.8 m/s) CVs were significantly reduced at 7 months, which is indicative of a polyneuropathy. NGF has well-known trophic effects on sympathetic and small sensory neurons. To determine whether NGF could influence this neuropathy, 6-month-old control and diabetic mice were divided into the following groups: 1) control + vehicle, 2) diabetic + vehicle, and 3) diabetic + NGF (1 mg/kg, 3x week, s.c.). After 1 month of treatment, motor and sensory CVs were determined. In some mice, the branches of the sciatic nerve were exposed and in situ recordings from the sural nerve were performed to determine compound C-fiber CV, integral, and amplitude. Sensory CV, determined via Hoffmann's reflex (H-reflex) (A-fiber), was decreased in diabetic compared with control animals as expected (P < 0.05), and NGF did not alter this parameter. Continuing diabetes reduced the amplitude (0.9 +/- 0.2 vs. 3.2 +/- 0.7 mV x 10(-2); P < 0.05) and integral (6.9 +/- 1.9 mV/ms vs. 18.8 +/- 4.4 mV/ms; P < 0.05) of the C-fiber response versus control, suggesting fiber loss. NGF treatment normalized C-fiber amplitude (2.9 +/- 0.8 mV x 10(-2)) and integral (21.2 +/- 6.5 mV/ms) in animals with established diabetes, with no effect on blood glucose. The C-fiber CV was similar in all groups, indicating that the animals had some normally conducting small fiber sensory nerves. These studies characterized a motor and sensory polyneuropathy in transgenic diabetic mice and are the first to demonstrate directly that NGF treatment can protect or restore abnormal sensory C-fiber function.

Transgenic Mice Overexpressing Type 2 Nitric-oxide Synthase in Pancreatic β Cells Develop Insulin-dependent Diabetes without Insulitis

We generated transgenic mice carrying the mouse type 2 nitric-oxide synthase (NOS2) cDNA under the control of the insulin promoter. Western and immunohistochemical analyses revealed that NOS2 was expressed abundantly in transgenic islets but not in control islets. When islets were isolated and cultured, high levels of nitrite were released from the transgenic islets. In transgenic mice, the beta cell mass was markedly reduced without the infiltration of macrophages or lymphocytes, and extensive DNA strand breaks were detected in the islets by in situ nick translation. All the transgenic mice developed hypoinsulinemic diabetes by 4 weeks of age, and treatment with an inhibitor of NOS2, aminoguanidine (200 mg/kg body weight every 12 h), prevented or delayed the development of diabetes. The present study shows that the production of nitric oxide by beta cell NOS2 plays an essential role in the beta cell degeneration.

Diabetes causes differential changes in CNS noradrenergic and dopaminergic neurons in the rat: a molecular study

We have previously reported that chronic elevation of insulin in the CNS of rats results in opposing changes of the mRNA expression for the norepinephrine transporter (NET; decreased) and the dopamine transporter (DAT; increased). In the present study we tested the hypothesis that a chronic depletion of insulin would result in opposite changes of NET and DAT mRNA expression, from those observed with chronic elevation of insulin. Rats were treated with streptozotocin to produce hypoinsulinemic diabetes. One week later, steady state levels of mRNA were measured by in situ hybridization for NET in the locus coeruleus (LC) and for DAT in the ventral tegmental area/substantia nigra compacta (VTA/SNc). The mRNA for tyrosine hydroxylase (TH), the rate-limiting enzyme for NE and DA synthesis, was measured in these same brain regions. In the diabetic animals, NET mRNA was significantly elevated (159 ± 22% of average control level) while DAT mRNA was non-significantly decreased (78 ± 9% of average control level). Additionally, TH mRNA was significantly altered in both the LC (131 ± 11 % of average control level) and VTA/SNc (79 ± 5% of average control level). We conclude that endogenous insulin is one physiological regulator of the synthesis and re-uptake of NE and DA in the CNS.

Endocytosis is inhibited in hepatocytes from diabetic rats.

The consequences of type I diabetes on cellular endocytosis were investigated by comparing [125I]insulin, [125I]alpha 2-macroglobulin, and Lucifer yellow uptake in hepatocytes freshly isolated from control and STZ-induced diabetic rats. In addition to the previously described reversible inhibition of ligand-induced internalization of the insulin receptor, we report a decrease in the constitutive receptor-mediated endocytosis of alpha 2-macroglobulin and a near abolition of fluid-phase endocytosis of Lucifer yellow in cells from diabetic animals. Despite decreased receptor autophosphorylation and internalization, the ligand-induced surface redistribution of the insulin receptor was normal in the diabetic cell population. By contrast, the insulin receptor association with clathrin-coated pits was impaired in diabetic cells as a result of a decreased concentration of these specialized invaginations on the nonvillous cell surface. The morphology and diameter of clathrin-coated pits were similar in both conditions under study. These results demonstrate a general impairment of endocytosis in hypoinsulinemic diabetes: receptor-mediated endocytosis was less affected than fluid-phase endocytosis. Impaired endocytosis of specific ligands or other macromolecules could be an important mechanism underlying the accumulation of extracellular matrix or even blood cholesterol removal in diabetes.

Induction of type I diabetes by interferon-alpha in transgenic mice.

Type I diabetes is an autoimmune disease involving an interaction between an epigenetic event (possibly a viral infection), the pancreatic beta cells, and the immune system in a genetically susceptible host. The possibility that the type I interferons could mediate this interaction was tested with transgenic mice in which the insulin-producing beta cells expressed an interferon-alpha. These mice developed a hypoinsulinemic diabetes associated with a mixed inflammation centered on the islets. The inflammation and the diabetes were prevented with a neutralizing antibody to the interferon-alpha. Thus, the expression of interferon-alpha by the beta cells could be causal in the development of type I diabetes, which suggests a therapeutic approach to this disease.

Partial reversal of .alpha.2U globulin gene expression by thyroxine in the liver of diabetic rats

Synthesis of alpha 2u globulin and its mRNA has been used as an index to monitor the effect of thyroxine on specific gene expression in the liver of hypoinsulinemic male rats. Administration of a physiological dose of thyroxine can partially reverse (to approximately 30% of the normal control) the marked reduction (more than 90%) in the hepatic levels of alpha 2u globulin and its mRNA during streptozotocin-induced diabetes. Estimation of newly synthesized alpha 2u globulin RNA transcripts from the native chromatin of isolated liver nuclei by "nuclear runoff experiments" showed that thyroxine can elevate the rate of transcription of alpha 2u globulin gene in the diabetic rat. Hypoinsulinemic diabetes is also found to be associated with an approximately 35% reduction in the thyroid hormone receptor level as compared to the normal control. The stimulatory effect of thyroxine on the synthesis of alpha 2u globulin and its mRNA was also evident in spontaneous diabetic Wistar "BB" rats. From these studies it can be concluded that severe hypoinsulinemia can cause a decrease in thyroid hormone action at the level of specific gene expression.

Receptor-mediated endocytosis of polypeptide hormones is a regulated process: inhibition of [125I]iodoinsulin internalization in hypoinsulinemic diabetes of rat and man.

Much data suggest that receptor-mediated endocytosis is regulated in states of hormone excess. Thus, in hyperinsulinemic states there is an accelerated loss of cell surface insulin receptors. In the present experiments we addressed this question in hypoinsulinemic states, in which insulin binding to cell surface receptors is generally increased. In hepatocytes obtained from hypoinsulinemic streptozotocin-induced diabetic rats, [125I]iodoglucagon internalization was increased, while at the same time [125I]iodoinsulin internalization was decreased. The defect in [125I]iodoinsulin internalization was corrected by insulin treatment of the animal. In peripheral blood monocytes from patients with type I insulinopenic diabetes, internalization of [125I]iodoinsulin was impaired; this defect was not present in insulin-treated patients. These data in the hypoinsulinemic rat and human diabetes suggest that receptor-mediated endocytosis is regulated in states of insulin deficiency as well as insulin excess. Delayed or reduced internalization of the insulin-receptor complex could amplify the muted signal caused by deficient hormone secretion.

The mechanisms of up-regulation of the hepatic insulin receptor in hypoinsulinemic diabetes mellitus.

The mechanism underlying the increased insulin binding found in hepatic plasma membranes from streptozotocin-diabetic rats was evaluated by measuring insulin binding to intact and Triton X-100-soluble extracts of plasma membranes prepared from the livers of control rats and rats administered streptozotocin (85 mg/kg). In addition, to assess whether the cellular content of hepatic insulin receptors is also increased in diabetic animals, we measured insulin-binding activity in intact and soluble extracts of total hepatic cellular membrane preparations (100,000 X g cellular pellets). The data indicate that while insulin binding is increased (52 +/- 3%) in intact hepatic plasma membranes from diabetic rats compared to control rats, there is no comparable increase in insulin binding in intact total cellular membranes or in Triton X-100-soluble extracts of plasma membranes or total cellular membranes. We therefore conclude that the enhanced insulin binding found in the livers of diabetic rats is the result of a local redistribution of plasma membrane insulin receptors from cryptic to exposed sites. Finally, the data suggest the presence of a negative modulator of insulin-binding affinity in intact plasma and total cellular membranes.

Concentration and secretion of gastric somatostatin in streptozotocin-diabetic rats.

Both the release and the content of gastric somatostatin were investigated in streptozotocin-diabetic rats. Fundic and antral somatostatin contents were both increased in streptozotocin-diabetic rats compared with control animals. Basal somatostatin levels in the perfusates of streptozotocin-treated animals were not significantly different from those of the control animals. However, the peak values of somatostatin release induced by 5 x 10(-9) M glucagon in the diabetic animals were significantly higher than those of the controls. These results lead us to conclude that a hyperfunctioning state of the gastric D-Cells exists in hypoinsulinemic diabetes.