Potential hypoglycemic effects of Chlorella in streptozotocin-induced diabetic mice

Chlorella, a type of unicellular fresh water algae, has been a popular foodstuff in Japan and Taiwan. Chlorella has been shown to produce hypoglycemic effects in alloxan-induced diabetic animals. However, there are no other reports of the effects of this substance in other diabetic animal models. Here we have used streptozocin (STZ)-induced diabetic mice to study the thypoglycemic effects of Chlorella. Diabetes was induced in ICR strain mice by the i.p. injection of STZ. Vehicle-treated ICR mice were used as normal control animals and glibenclamide was used as a positive drug control. The effects of Chlorella on basal blood glucose, exogenous insulin sensitivity test and plasma insulin levels were measured. In normal mice Chlorella produced a transient hypoglycemic effect at 90 min after acute administration; whereas glibenclamide produced a more sustained hypoglycemic effect between 90 min and 180 min after acute administration. Chlorella did not affect the basal blood glucose level in STZ mice. However, Chlorella enhanced and prolonged the hypoglycemic effects of injected insulin in STZ mice for a further 60 min compared to the normal vehicle-treated group. Plasma insulin levels were increased in normal mice after treatment with glibenclamide, whereas Chlorella had no such effect. The current results indicate that Chlorella enhances the hypoglycemic effects of exogenous insulin at a dose which does not produce hypoglycemia in STZ mice, suggesting that insulin sensitivity is increased in these mice.     

A case of autoimmune insulin antibody syndrome associated with polymyositis, empty sella and apparent high urinary output of immunoreactive insulin.

Patients with autoimmune insulin antibody are characterized by hypoglycemic attacks and antibodies to insulin in serum without prior insulin administration. In the present report, a patient with hypoglycemia due to autoimmune insulin antibody associated with primary empty sella syndrome and polymyositis appeared to have high urinary immunoreactive insulin (IRI) in the face of normal urinary C peptide. Consequently, the urinary IRI/C peptide ratio was apparently high. The amelioration of hypoglycemic attacks and polymyositis by prednisolone treatment was accompanied by the disappearance of the antibodies and complete normalization of the urinary IRI and IRI/C peptide ratio. No comparable rise in the urinary IRI and IRI/C peptide ratio was observed in the patients with other disorders studied. Glucose clamp and glucose tolerance study showed decreased sensitivity to exogenous or newly secreted insulin, prolonged half disappearance time of serum insulin, and normal disappearance of blood glucose. These results were consistent with the idea that autoantibodies buffered the effect of exogenous or newly secreted insulin and maintained a relatively constant level of serum free insulin which was not high enough when a large amount of glucose was loaded, but was too high after prolonged fasting, which eventually caused hypoglycemic attacks.     

Electrical stimulation alters fatty acid metabolism in isolated skeletal muscle

Little is known about the contribution of plasma free fatty acid (FFA) and intramuscular triacylglycerol (TG) as substrates for energy production during prolonged electrical stimulation of skeletal muscle. The purpose of this study was to investigate the effects of continuous and intermittent electrical stimulation protocols of different intensities on exogenous FFA oxidation, exogenous FFA incorporation into intracellular TG, and intracellular TG content in the isolated in vitro rat flexor digitorum brevis muscle preparation. Muscles were electrically stimulated for 0.5 h continuously at 0.2 Hz or intermittently (30 s on, 60 s off) at 0.2, 0.4, 0.8, and 5.0 Hz while incubated at 37 degrees C in 0.5 mM palmitate-3% bovine serum albumin medium (pH 7.4) in the presence of insulin (100 microU/ml) and glucose (11 mM). Control muscles were frozen immediately after excision or incubated for 0.5 h. At similar frequencies, less exogenous FFA esterification and more exogenous FFA oxidation occurred during continuous than during intermittent stimulation. As the frequency of intermittent stimulation increased, the amount of exogenous FFA esterified decreased and the amount of exogenous FFA oxidized increased. The data also indicate that at least a portion of TG was constantly being hydrolyzed during electrical stimulation. Under stimulation conditions in which exogenous FFA esterification was below the control (resting muscle) level, intramuscular TG content was significantly decreased compared with control TG content values. Thus both plasma FFA and intramuscular TG are substrates for energy production during electrical stimulation. However, the stimulation parameters employed affect the quantities utilized.     

Effect of transcutaneous transcerebral electrostimulation as electroanesthesia on the beta-endorphin content of the cerebrospinal fluid and blood plasma

The beta-endorphin content was measured in the cerebrospinal fluid (CSF) and blood plasma of patients before and after 30 minutes of transcutaneous transcerebral electric stimulation in the electric anesthesia mode. The output current was biphasic and rectangular. It was composed of high-frequency pulse trains (peak-to-peak intensity 250-300 mA, frequency 167 kHz) modulated by low frequency (77 Hz). Electrical stimulation resulted in an appreciable increase in the beta-endorphin content in the CSF and blood plasma of patients. The data obtained attest to the intensification of the neuromodulator release to the CSF and blood plasma and to the involvement of the endorphinergic brain systems in the realization of the analgetic effect of transcutaneous transcerebral electric stimulation.     

Role of heparin in the realization of the hypoglycemic action of insulin

The presence of reactive heparin in blood circulation is necessary for realization of the hypoglycemic action of insulin. This is confirmed by the fact that after heparin binding by protamin sulfate both endogenous and exogenous insulin do not exhibit any hypoglycemic activity. In animals with different basal concentration of heparin in the blood, the blockade of insulin action is attained by application of different doses of protamin sulfate, respectively. Based on the data obtained one can determine an approximate blood heparin concentration necessary for realization of the hypoglycemic action of insulin.     

Lysosomal proteolysis: effects of aging and insulin

Age-related characteristics of the effect of insulin on the activity of lysosomal proteolytic enzymes were studied. The relationship between the insulin effect on protein degradation and insulin degradation was analyzed. The effect of insulin on the activities of lysosomal enzymes was opposite in young and old rats (inhibitory in 3-month-old and stimulatory in 24-month-old animals). The activities of proteolytic enzymes were regulated by insulin in a glucose-independent manner: similar hypoglycemic effects of insulin in animals of different ages were accompanied by opposite changes in the activities of lysosomal enzymes. The inhibition of lysosomal enzymes by insulin in 3-month-old rats is consistent with a notion on the inhibitory effect of insulin on protein degradation. An opposite insulin effect in 24-month-old rats (i.e., stimulation of proteolytic activity by insulin) may be partly associated with attenuation of the degradation of insulin, resulting in disturbances in signaling that mediates the regulatory effects of insulin on protein degradation.     

Effect of ethanolic extracts of Ananas comosus L. leaves on insulin sensitivity in rats and HepG2

Ethanolic extracts of Ananas comosus L. leaves (AC) enriched with phenols have hypoglycemic activity in diabetic rats. Here, we investigated the effect of AC on insulin sensitivity in rats and HepG2. In high-fat diet-fed and low-dose streptozotozin-treated diabetic Wistar rats subjected to challenge with exogenous human insulin, AC treatment at an oral dose of 0.40 g/kg could significantly improve sensitivity to exogenous insulin. After a sub-acute treatment, AC also could inhibit the development of insulin resistance in high-fat diet-fed and low-dose streptozotozin-treated diabetic rats following the test of loss of tolbutamide-induced blood glucose lowering action. For intravenous insulin/glucose infusion test, high-fat diet-fed and low-dose alloxan-treated Wistar rats were associated with insulin resistance, which was improved after AC or fenofibrate treatment. AC application inhibited the development of insulin resistance in HepG2 cells. The above animal models were well developed to simulate type 2 diabetes. Taken together, our results suggest that AC may improve insulin sensitivity in type 2 diabetes and could be developed into a new potential natural product for handling of insulin resistance in diabetic patients.     

Opiate-prostaglandin interactions in the regulation of insulin secretion from rat islets of Langerhans in vitro

The inadequate insulin secretory response to glucose stimulation in non-insulin dependent diabetes has been attributed to many factors including high PGE2 levels blunting the secretory response, and to the existence of inhibitory opiate activity in vivo. The purpose of the present work was to see if there was a connection between these two independent theories. Radioimmunoassayable PGE2 in islets of Langerhans was found to be proportional to islet number and protein content and was typically 4 to 5pg/micrograms islet protein. Indomethacin (2.8 X 10(-5) M), sodium salicylate (1.25 X 10(-3) M) and chlorpropamide (7.2 X 10(-5) M) all lowered islet PGE2 levels and stimulated insulin release in vitro. Dynorphin (1-13), stimulated insulin release at a concentration of 6 X 10(-9) M, while lowering islet PGE2. Conversely, at a higher concentration, (6 X 10(-7) M), dynorphin had no stimulatory effect on insulin secretion and did not lower PGE2 levels in islets or in the incubation media. The stimulatory effects of dynorphin and sodium salicylate on insulin secretion were blocked by exogenous PGE2 (10(-5) M). PGE2 at a lower concentration (10(-9) M) did not exert any inhibitory effect on dynorphin- or sodium salicylate-induced insulin release. This concentration of exogenous PGE2 stimulated insulin release in the presence of 6mM glucose. Results from these experiments suggest that since an opioid peptide can lower endogenous PGE2 production in islets and since the stimulatory effects of the opioid peptide are reversed by exogenous PGE2 there may be interactions between these two modulators of insulin secretion.     

Effect of adenosine triphosphate (ATP) on the secretion of insulin induced by tolbutamide in the absence of glucose

Our experiments were carried out on the isolated perfused rat pancreas. The effect of ATP on insulin secretion induced by tolbutamide (a hypoglycemic sulfonylurea) was studied in the absence of glucose. The addition of ATP (0.165 mmol/l) to the perfusion medium did not significantly modify the first phase induced by tolbutamide (0.4 mmol/l) but potentiated the second phase.     

Transcranial electrical stimulation activates the reparative regeneration and insulin-producing function of beta-cells in alloxan-induced diabetic rats

In alloxan-induced diabetic rats, it was demonstrated that transcranial electrical stimulation of the brain endorphinergic structures activated the reparative regeneration of the damaged beta-cells of the Langerhans pancreatic islets. This was estimated on the histological sections of pancreas with hematoxylin-eosin staining. Several small newborn islets were found to originate from pancreatic progenitor cells. After transcranial electrical stimulation of insulin granules, beta-cells (Gomori's staining) were observed as an indication of the restoration of the insulin production. Correspondingly the increase of the blood insulin level was estimated by immune-enzyme method. The dynamics of the plasma insulin increase had a significant negative correlation with decrease of the blood glucose level. The glucose-lowering action of the transcranial electrical stimulation in alloxan-induced diabetic rats seems to be based on stimulation of the regeneration of damaged beta-cells with the restoration of their insulin production.     

Decreased sensitivity to the hypoglycemic action of insulin in animals with a lowered blood heparin concentration

It has been shown that in animals with reduced heparin blood concentration the hypoglycemic insulin action was considerably low. This was established for rats with alloxan diabetes and ageing rats with depressed anticoagulation system and atherosclerosis enhanced by prolonged atherogenic diet. With insulin injection (0.2-0.3 U/200 g), blood sugar concentration in such animals was 2-2.5 times lower than in normal ones. The compensation of endogenous heparin deficiency by an intravenous injection of heparin normalizes the reaction of animals to exogenous insulin.     

Lateral modulation of the hypoglycemic action of insulin

The results of the experimental investigation on 15 rabbits are presented here. Hypoglycemic action of the standard exogenic insulin dose strengthened after performing transcerebral lateral electrostimulation on the right side with the weak current impulse.     

The differential effect of insulin in vivo on the peripheral utilization of glucose and ketone bodies in the rat.

This in vivo study assessed the immediate effects of insulin on glucose and ketone body utilization in the fed, fasted, and diabetic ketoacidotic rat. The experimental design consisted of the functional removal of the liver (the site of glucose and ketone body production) and the pancreas from the anesthetized animals. This surgical procedure permitted the assessment of the effect of exogenously administered insulin on the rate of both glucose and ketone body utilization by peripheral tissues. Insulin exerted hypoglycemic activity in all three metabolic states studied. This hypoglycemic activity contrasted to the lack of demonstrable effect of this hormone on ketone body uptake by peripheral tissues. It was concluded that in the rat, the immediate effect of insulin, i.e. within 30 minutes, was to exert hypoglycemic activity without simultaneous hypoketoniemic activity.     

Diacylglycerols modulate phosphorylation of the insulin receptor from human mononuclear cells

It has been found that 1,2- but not 1,3-diacylglycerols stimulated phosphorylation of the insulin receptor of cultured human monocyte-like (U-937) and lymphoblastoid (IM-9) cells both in the intact- and broken-cell systems. The stimulation of the receptor's beta-subunit phosphorylation was dose-dependent, with optimal effect at 100 micrograms/ml of diacylglycerol. The effects of insulin and 1,2-diacylglycerols on the phosphorylation of partially purified insulin receptors were additive. Phosphoamino acid analysis showed a major effect of diacylglycerols on phosphorylation of tyrosine residues. The diacylglycerols also stimulated tyrosine kinase activity of the partially purified U-937 and IM-9 insulin receptors 2.5-3.5-fold when measured by phosphorylation of an exogenous substrate, poly(Glu80Tyr20) in the absence of any added insulin, calcium or phospholipid. Since this diacylglycerol effect could not be reproduced under conditions optimal for protein kinase C activation and the purified protein kinase C did not stimulate phosphorylation of the beta-subunit of the insulin receptor in this system, it is unlikely that the diacylglycerol effect was mediated by protein kinase C. Since these exogenous 1,2-diacylglycerols at the same high concentration also inhibited 125I-insulin binding to the insulin receptor of the intact U-937 and IM-9 cells, diacylglycerols could modulate the function of the insulin receptor and insulin action in human mononuclear cells.     

Response of cortical evoked potentials to electric stimulation of the acoustic nerve in animals with damaged cochlea

Evoked potentials of the auditory cortex during the electrical stimulation of the cochlea were studied in acute experiments on cats. A series of electric pulses of short duration and different frequency delivered to the streptomycin-damaged cochlea were used as a stimulus. It has been shown that an amplitude and latency of electrical cortex responses depended on the number of pulses in series and on the interpulse intervals. Amplitudes of evoked responses increased with the growth of the number of stimuli. Latent periods changed in a narrower stimulation frequency band. Dependence of the induced potentials' amplitude growth on the increase in the number of electric pulses changed as a result of the two-fold enhancement of the stimulation amplitude.     

Insulin regulation of steroidogenic activity in rat culture luteal cells

The effect of insulin on the function of rat luteal cells in monolayer culture was examined. Cells were obtained from PMSG-hCG primed immature rats and further cultured in serum free medium with or without insulin. The hormone produced an increase of progesterone production and maximal stimulation was achieved at 0.2 nM of insulin (100% stimulation). This effect was enhanced by addition of methyl-isobutyl-xantine (MIX 0.1 mM) to the culture medium. However, the stimulation produced by LH was not augmented by the presence of insulin. The conversion of progesterone into 20 alpha-hydroxy-progesterone was also enhanced after insulin treatment. Luteal cells were also cultured in the presence of 25-hydroxy-cholesterol (10 micrograms/ml). In these conditions insulin produced a 2-fold increase in progesterone production. Aromatase activity was assessed by adding androstenedione (0.25 microM) as substrate. Insulin produced a 14-fold stimulation of estradiol production after 24 h of culture. Insulin action was tested in short time incubations of luteal cells in a glucose free medium, in these experiments the hormone was able to induce a significant increase in progesterone and 20 alpha-hydroxy-progesterone production. These data suggest that luteal cell function is regulated by insulin and that this hormone has a direct effect on the steroidogenic process.     

Synthesis and hypoglycemic effect of chrysin derivatives

A series of 18 chrysin derivatives, prepared by alkylation and condensation, were fully characterized by NMR and other techniques and tested in vivo against the diabetes mellitus. Several modified compounds especially those with propyl, butyl, octyl and tolyl groups were found to have hypoglycemic effect on diabetec mice in spite of the fact that chrysin itself had inhibited insulin release by 40-60%. None of the test animals died at the maximum dose 500mg/kg and did not cause any significant change in general feature, water and food consumption, body weight and organ weight when we examined the acute oral toxicity of those compounds having significant hypoglycemic effect.     

Combined effects of troglitazone and muscle contraction on insulin sensitization in Balb-c mouse muscle

Thiazolidinediones, represented by troglitazone, are insulin-sensitizing agents with proven efficacy for the treatment of type 2 diabetes. Exercise is also recommended for patients with type 2 diabetes because it both stimulates glucose uptake directly and it increases insulin sensitivity following exercise. The purpose of this study was to investigate the effects of troglitazone combined with exercise on 2-deoxyglucose (2DG) uptake in both the epitrochlearis and soleus muscle of Balb-c mice. Acute, 1-h treatment with troglitazone (10 or 20 microM), in the presence or absence of insulin, had no effect on 2DG uptake in either muscle. Chronic treatment with troglitazone by feeding enhanced the insulin sensitivity and responsiveness of 2DG uptake primarily in the epitrochlearis. Direct electrical stimulation of in situ muscle was used to model exercise while the contralateral muscle was used as the unexercised control. This model mimicked exercise in that glycogen was depleted, immediate 2DG uptake was enhanced, and there was a post-exercise increase in insulin sensitivity. Troglitazone feeding had no effect on 2DG uptake in the soleus when measured immediately after electrical stimultion. However, 2DG uptake in the unstimulated epitrochlearis from troglitazone-fed mice was elevated when measured immediately after removal such that no additional effects of the electrical stimulation were measured. We found that the insulin-sensitizing effect of troglitazone was not additive to the insulin-sensitizing effect of exercise, which suggests that troglitazone and exercise share similar pathways. A unique finding in this study was the differential response to troglitazone between the epitrochlearis (fast twitch) and the soleus (slow twitch) muscle types. Possible mechanisms are discussed.     

Effect of morphine and acetylcholine on contractile activity and cyclic AMP in guinea-pig ileum

Neither acute nor prolonged exposure to morphine altered cAMP content or spontaneous movements of longitudinal muscle-myenteric plexus strips of the guinea-pig ileum. By contrast, exogenous acetylcholine or electrical stimulation of the strips elicited both a decrease of cAMP concentration and a twitch response. Atropine blocked the effects of stimulation on these parameters. Addition of morphine to electrically stimulated strips inhibited the twitch response but did not affect cAMP levels. Incubation with morphine led to the development of tolerance to the inhibitory effect on twitch activity and prevented the fall in cAMP normally elicited by electrical stimulation. These results suggest that muscarinic activation is associated with a reduction of cAMP content, an effect which would be impaired in opiate-tolerant tissues.     

Insulin stimulation of the insulin receptor kinase can occur in the complete absence of beta subunit autophosphorylation

The glutamic acid:tyrosine (Glu:Tyr) synthetic polymer was observed to inhibit the insulin receptor beta subunit autophosphorylation with an IC50 of 0.20 mg/ml in the absence and 0.15 mg/ml in the presence of insulin. Even though complete blockade of beta subunit autophosphorylation was observed at 4.0 mg/ml Glu:Tyr, insulin was still capable of stimulating the exogenous protein kinase activity of the insulin receptor toward Glu:Tyr. Histone H2B (1.3 mg/ml) was also observed to inhibit the beta subunit autophosphorylation by approximately 80% with an IC50 of 0.31 and 0.35 mg/ml in the absence and presence of insulin, respectively. Similar to the results with Glu:Tyr, insulin was found to stimulate histone H2B phosphorylation under these conditions. Comparisons between the time courses of beta subunit autophosphorylation with those of Glu:Tyr phosphorylation both in the presence and absence of insulin confirmed that insulin can stimulate the exogenous protein kinase activity of the insulin receptor in the complete absence of beta subunit autophosphorylation. Prephosphorylation of the insulin receptor (from 0 to 1.3 mol of phosphate/mol of insulin receptor) in the absence of insulin was found to have no significant effect on the exogenous protein kinase activity when assayed both in the presence and absence of insulin. Insulin was observed to stimulate the phosphorylation of Glu:Tyr approximately 3-fold independent of the extent of beta subunit autophosphorylation. In contrast, prephosphorylation of the insulin receptors in the presence of insulin was observed to enhance the exogenous protein kinase activity dependent on the extent of autophosphorylation, such that by 1.4 mol of phosphate incorporated per mol of insulin receptor, insulin was found to maximally stimulate the initial rate of Glu:Tyr phosphorylation (approximately 9-fold). These results demonstrate that the insulin-dependent autophosphorylation of the insulin receptor results in an amplification of the insulin stimulation of the exogenous protein kinase activity, whereas the insulin-independent autophosphorylation does not.