Rhodomyrtus tomentosa as a new anticancer molecular strategy in breast histology via Her2, IL33, EGFR, and MUC1.

The prevalence of breast cancer among patients in Indonesia is significant. Indonesian individuals maintain the belief that cancer cannot be cured alone by pharmaceuticals and treatment; herbal remedies must be used in conjunction. Rhodomyrtus tomentosa, also known as Haramonting, is an indigenous Indonesian medicinal plant renowned for its copious antioxidant properties. The objective of study was to assess the impact of haramonting on breast cancer by examining the expression of various biomarker proteins associated with breast cancer. Haramonting was administered to breast cancer model mice at different doses over a period of 30 days. Subsequently, blood and breast samples were obtained for immunohistochemistry and enzyme-linked immunosorbent assays (ELISA). Authors have discovered that there has been a notable rise in the proliferation of epithelial cells in the duct lobes, resulting in the formation of ducts and lobules. Additionally, the researchers discovered that the breasts exhibited distinct clinical and histological alterations. Haramonting possesses the capacity to restore the concentrations of malondialdehyde (MDA) and superoxide dismutase (SOD) to normal levels in the blood serum of rats afflicted with cancer. The histopathological analysis of the breast tissue revealed elevated levels of Her2, IL33, EGFR, and MUC1. The authors also discovered a notable increase in the growth of epithelial cells, with two or more layers of cells reaching towards the centre of the duct. The size of the epithelial cells exhibits variability; however, this state ameliorates with the administration of a dosage of 300 mg/kgBW of this botanical specimen. This study proposes that Haramonting may be effective in treating breast cancer.

Beneficial cardiac effects of cicletanine in conscious rabbits with metabolic syndrome.

BACKGROUND AND PURPOSE: High-fat diet and consequent metabolic syndrome (MS) can lead to elevated risk for cardiac arrhythmias. This preclinical study was to investigate if cicletanine (CIC) could produce cardioprotective effects in conscious rabbits exhibiting the main symptoms of MS. METHODS: NZW rabbits that had undergone an 8-week-long cholesterol-enriched diet (1.5%) were instrumented with a pacemaker electrode and randomly assigned into 3 groups according to the oral treatment of either CIC (50 mg·kg) or sotalol (25 mg·kg) and their placebo b.i.d. over 5 days. Study groups were subjected to either "arrhythmia challenge" by programmed electrical stimulation in the "Arrhythmogenesis" study (N = 54) or global myocardial ischemia by rapid pacing in the "Ventricular Overdrive Pacing-induced Myocardial Ischemia" study (N = 18). The antiarrhythmic effect was evaluated by the establishment of the incidence of programmed electrical stimulation-induced arrhythmias. Proarrhythmia indicators (eg, QTc, Tpeak-Tend) were also measured to assess the cardiac safety profile of CIC. To evaluate the background of antiarrhythmic effect, cardiac cyclic nucleotide (cyclic 3',5'-guanosine monophosphate [cGMP], cyclic 3',5'-adenosine monophosphate [cAMP]) and nitric oxide content were determined. The antiischemic effect was characterized by change of intracavital ST segment. RESULTS: Cicletanine treatment significantly decreased the incidence of ventricular arrhythmias, increased cardiac cGMP and nitric oxide content and reduced cardiac cAMP level. Cicletanine did not modify significantly QTc and Tpeak-Tend interval. The ST-segment change in response to rapid pacing was reduced significantly by CIC. (P < 0.05). CONCLUSIONS: Cicletanine exerts beneficial cardiac effects in rabbits with symptoms of MS, which may be of influence with regard to the clinical application of the drug.

Terpenes and terpenoids as main bioactive compounds of essential oils, their roles in human health and potential application as natural food preservatives.

Essential oils (EOs) are volatile and concentrated liquids extracted from different parts of plants. Bioactive compounds found in EOs, especially terpenes and terpenoids possess a wide range of biological activities including anticancer, antimicrobial, anti-inflammatory, antioxidant, and antiallergic. Available literature confirms that EOs exhibit antimicrobial and food preservative properties that are considered as a real potential application in food industry. Hence, the purpose of this review is to present an overview of current knowledge of EOs for application in pharmaceutical and medical industries as well as their potential as food preservatives in food industry.

Neurogenic insulin resistance in guinea-pigs with cisplatin-induced neuropathy.

The aim of the present work was to study whether neurotoxicity produced by cisplatin modified tissue insulin sensitivity in guinea-pigs. One week after selective sensory denervation of the anterior hepatic plexus by means of perineurial 2% capsaicin treatment, hyperinsulinaemic euglycaemic glucose clamp were performed to estimate insulin sensitivity in male guinea-pigs. The guinea-pigs underwent regional sensory denervation of the anterior hepatic plexus exhibited insulin resistance, whereas systemic capsaicin desensitization increased insulin sensitivity. Intraportal administration of L-nitro-arginine methyl ester (L-NAME decreased, whereas capsaicin increased insulin sensitivity. Neither atropine nor acetylcholine produced any significant effect. In animals with preceding regional capsaicin desensitization, none of the pharmacological maneuvers modified the resulting insulin resistant state. Cisplatin pretreatment induced sensory neuropathy and decreased insulin sensitivity. Insulin sensitivity did not change after either regional or systemic capsaicin desensitization in the cisplatin-treated animals. CGRP(8-37), a nonselective calcitonin gene-related peptide (CGRP) antagonist (50 microg/kg i.v.), significantly increased insulin sensitivity in normal animals but only a tendency to insulin sensitization was seen after cisplatin treatment. Cisplatin treatment, similar to regional capsaicin desensitization of the anterior hepatic plexus, produced a significant decrease in insulin-stimulated uptake of 2-deoxy-D [L-14C] glucose in cardiac and gastrocnemius muscle with no effect on percentage suppression of endogenous glucose production by hyperinsulinaemia. We conclude that the majority of cisplatin-induced insulin resistance is related to functional deterioration of the hepatic insulin sensitizing substance (HISS) mechanism.

Beneficial effect of resveratrol on cholecystokinin-induced experimental pancreatitis.

Resveratrol is a phytoalexin with strong antioxidant and anti-inflammatory effects reaching high concentrations in red wine. The aim of our study was to test the effects of resveratrol pretreatment on cholecystokinin-octapeptide (CCK-8)-induced acute pancreatitis in rats. Animals were divided into a control group, a group treated with CCK-8 and a group receiving 10 mg/kg resveratrol prior to CCK-8 administration. Resveratrol ameliorated the CCK-8-induced changes in the laboratory parameters, and reduced the histological damage in the pancreas. The drug failed to improve the pancreatic antioxidant state, but increased the amount of hepatic reduced glutathione and prevented the reduction of hepatic catalase activity. Resveratrol-induced inhibition of nuclear factor kappa B (NF-kappaB) activation or reduction of the pancreatic tumor necrosis factor-alpha (TNF-alpha) concentration could not be demonstrated. In conclusion, the beneficial effects of resveratrol on acute pancreatitis seem to be mediated by the antioxidant effect of resveratrol or by an NF-kappaB-independent anti-inflammatory mechanism.

Effect of melatonin on the severity of L-arginine-induced experimental acute pancreatitis in rats.

AIM: To determine the effect of melatonin pre- and post-treatment on the severity of L-arginine (L-Arg) -induced experimental pancreatitis in rats. METHODS: Male Wistar rats (25) were divided into five groups. Those in group A received two injections of 3.2g/kg body weight L-Arg i.p. at an interval of 1h. In group MA, the rats were treated with 50 mg/kg body weight melatonin i.p. 30 min prior to L-Arg administration. In group AM, the rats received the same dose of melatonin 1h after L-Arg was given. In group M, a single dose of melatonin was administered as described previously. In group C the control animals received physiological saline injections i.p. All rats were exsanguinated 24 h after the second L-Arg injection. RESULTS: L-Arg administration caused severe necrotizing pancreatitis confirmed by the significant elevations in the serum amylase level, the pancreatic weight/body weight ratio (pw/bw), the pancreatic IL-6 content and the myeloperoxidase activity, relative to the control values. Elevation of the serum amylase level was significantly reduced in rats given melatonin following L-Arg compared to rats injected with L-Arg only. The activities of the pancreatic antioxidant enzymes (Cu/Zn-superoxide dismutase (Cu/Zn-SOD) and catalase (CAT)) were significantly increased 24 h after pancreatitis induction. Melatonin given in advance of L-Arg significantly reduced the pancreatic CAT activity relative to that in the rats treated with L-Arg alone. In the liver, L-Arg significantly increased the lipid peroxidation level, and the glutathione peroxidase and Cu/Zn-SOD activities, whereas the Mn-SOD activity was reduced as compared to the control rats. Melatonin pre-treatment prevented these changes. CONCLUSION: Melatonin is an antioxidant that is able to counteract some of the L-Arg-induced changes during acute pancreatitis, and may therefore be helpful in the supportive therapy of patients with acute necrotizing pancreatitis.

Interplay between nitric oxide and VIP in CCK-8-induced phasic contractile activity in the rabbit sphincter of Oddi.

AIM: The sphincter of Oddi (SO) plays an important role in delivery of bile into the duodenum. To establish whether vasoactive intestinal polypeptide (VIP) and nitric oxide (NO) were involved in phasic contractile activity of the rabbit SO stimulated by cholecystokinin-octapeptide (CCK-8). METHODS: Isolated SO muscle rings were cleaned of fat and mounted horizontally on two small L-shaped hooks one of which was connected to a force transducer for the measurement of isometric tension. The experiments were carried out in a thermostatically controlled (37+/-0.2 degrees) organ bath (5 mL) containing Krebs solution. The organ fluid was gassed with 95% O(2) and 50 mL/L CO(2) to keep the pH at 7.40+/-0.05. Contractile responses to CCK-8 (1 micromol/L) were evaluated in the presence and absence of N(G)-nitro-L-arginine (LNNA), an inhibitor of NO synthase (100 micromol/L), and (p-chloro-D-Phe(6)-Leu(17))-VIP (VIPa, 30 micromol/L), a VIP receptor antagonist. RESULTS: CCK-8 stimulated the phasic activity of the SO. NO synthase inhibition increased the frequency and amplitude of contractions with a slight increase in developed tension. Pre-incubation with VIPa also attenuated this CCK-8 effect. The combined application of LNNA and VIPa abolished the phasic activity of the muscle rings with a marked increase in tension in response to CCK-8. CONCLUSION: VIP and NO together contribute to an increase in phasic activity of SO.

Effect of herpesvirus infection on pancreatic duct cell secretion.

AIM: To examine the effect of acute infection caused by herpesvirus (pseudorabies virus, PRV) on pancreatic ductal secretion. METHODS: The virulent Ba-DupGreen (BDG) and non-virulent Ka-RREp0lacgfp (KEG) genetically modified strains of PRV were used in this study and both of them contain the gene for green fluorescent protein (GFP). Small intra/interlobular ducts were infected with BDG virus (10(7) PFU/mL for 6 h) or with KEG virus (10(10) PFU/mL for 6 h), while non-infected ducts were incubated only with the culture media. The ducts were then cultured for a further 18 h. The rate of HCO(3)(-) secretion (base efflux -J(B-)) was determined from the buffering capacity of the cells and the initial rate of intracellular acidification (1) after sudden blockage of basolateral base loaders with dihydro-4,4-diisothiocyanatostilbene-2,2-disulfonic acid (500 micromol/L) and amiloride (200 micromol/L), and (2) after alkali loading the ducts by exposure to NH(4)Cl. All the experiments were performed in HCO(3)(-)-buffered Ringer solution at 37 degrees (n = 5 ducts for each experimental condition). Viral structural proteins were visualized by immunohistochemistry. Virally-encoded GFP and immunofluorescence signals were recorded by a confocal laser scanning microscope. RESULTS: The BDG virus infected the majority of accessible cells of the duct as judged by the appearance of GFP and viral antigens in the ductal cells. KEG virus caused a similarly high efficiency of infection. After blockage of basolateral base loaders, BDG infection significantly elevated -J(B-) 24 h after the infection, compared to the non-infected group. However, KEG infection did not modify -J(B-). After alkali loading the ducts, -J(B-) was significantly elevated in the BDG group compared to the control group 24 h after the infection. As we found with the inhibitor stop method, no change was observed in the group KEG compared to the non-infected group. CONCLUSION: Incubation with the BDG or KEG strains of PRV results in an effective infection of ductal epithelial cells. The BDG strain of PRV, which is able to initiate a lytic viral cycle, stimulates HCO(3)(-) secretion in guinea pig pancreatic duct by about four- to fivefold, 24 h after the infection. However, the KEG strain of PRV, which can infect, but fails to replicate, has no effect on HCO(3)(-) secretion. We suggest that this response of pancreatic ducts to virulent PRV infection may represent a defense mechanism against invasive pathogens to avoid pancreatic injury.

Effect of long-term olanzapine treatment on meal-induced insulin sensitization and on gastrointestinal peptides in female Sprague-Dawley rats.

Meal-induced insulin sensitization (MIS), an endogenous adaptive mechanism is activated post-prandially. Reduced MIS leads to diabetes, but its activation improves insulin sensitivity. MIS is preserved to single olanzapine administration, therefore we aimed to investigate the chronic effect of olanzapine on fasted-state insulin sensitivity and on MIS in female Sprague-Dawley rats. Daily food and water intake, stool and urine production and body weight were determined. The MIS was characterized by a rapid insulin sensitivity test. Fasting hepatic and peripheral insulin sensitivity were determined by a hyperinsulinaemic euglycaemic glucose clamping supplemented with radiotracer technique. Fasted and post-prandial blood samples were obtained for plasma insulin, leptin, ghrelin, amylin, GLP-1, GIP, PYY and PP determination. Adiposity was characterized by weighing intra-abdominal and inguinal fat pads. Olanzapine caused hepatic insulin resistance and a reduced metabolic clearance rate of insulin, but the MIS retained its function. Body weight and adiposity were enhanced, but olanzapine failed to increase food intake. Fasting insulin and leptin were elevated and the post-prandial reduction in ghrelin level was inhibited by olanzapine.The MIS remained functionally intact after long-term olanzapine treatment. Altered insulin, leptin and ghrelin levels indicate olanzapine-induced metabolic derangements. Pharmacological activation of MIS could potentially be exploited to treat or prevent olanzapine-induced insulin resistance.

Meal-induced insulin sensitization is preserved after acute olanzapine administration in female Sprague-Dawley rats.

Olanzapine, an atypical antipsychotic, can acutely induce fasting insulin resistance, but we do not know whether it is able to modulate the meal-induced insulin sensitization (MIS). Two main experimental groups (control and olanzapine-treated) were created with two subgroups (fasted and re-fed) within each. After oral vehicle/olanzapine administration, the first meal size and duration and the total amount of consumed food was recorded in conscious rats. Then, under anaesthesia, the carotid artery and jugular vein was prepared and cannulated to obtain samples for blood glucose and hormone determination as well as for insulin/glucose infusion, respectively. Basal insulin sensitivity and MIS was determined by homeostasis model assessment (HOMA) calculation and by rapid insulin sensitivity test, respectively. In fasted animals, olanzapine increased blood glucose and plasma insulin and reduced basal insulin sensitivity, but it failed to modify other hormone levels. Postprandial leptin and glucose-dependent insulinotropic polypeptide (GIP) levels increased, and ghrelin level decreased significantly (p < 0.05) both in vehicle- and olanzapine-treated groups, but plasma insulin increased only in vehicle-treated animals. Furthermore, decrement in ghrelin level was attenuated in olanzapine-treated animals compared to controls. There was no significant change in the first meal size and duration or in the total amount of food consumed. Olanzapine had no effect on the MIS. We demonstrated that olanzapine can induce insulin resistance without weight gain in healthy rats. Furthermore, the MIS was preserved after acute olanzapine treatment. The blunted postprandial ghrelin and insulin response could contribute to the effect of olanzapine on feeding behaviour. Pharmacological induction of MIS may improve the olanzapine-induced insulin resistance.

Ethanol inhibits the motility of rabbit sphincter of Oddi in vitro.

AIM: The role of the sphincter of Oddi (SO) in ethanol (ETOH)-induced pancreatitis is controversial. Our aim was to characterise the effect of ETOH on basal and stimulated SO motility. METHODS: SOs removed from white rabbits were placed in an organ bath (Krebs solution, pH7.4, 37 degrees). The effects of 2 mL/L, 4 mL/L, 6 mL/L and 8 mL/L of ETOH on the contractile responses of the sphincter were determined. SOs were stimulated with either 0.1 mumol/L carbachol, 1 mumol/L erythromycin or 0.1 mumol/L cholecystokinin (CCK). RESULTS: ETOH at a dose of 4 mL/L significantly decreased the baseline contractile amplitude from 11.98+/-0.05 mN to 11.19+/-0.07 mN. However, no significant changes in the contractile frequency were observed. ETOH (0.6%) significantly decreased both the baseline amplitude and the frequency compared to the control group (10.50+/-0.01 mN, 12.13+/-0.10 mN and 3.53+/-0.13 c/min, 5.5+/-0.13 cycles(c)/min, respectively). Moreover, 0.8% of ETOH resulted in complete relaxation of the SO. Carbachol (0.1 micromol/L) or erythromycin (1 micromol/L) stimulated the baseline amplitudes (by 82% and 75%, respectively) and the contractile frequencies (by 150% and 106%, respectively). In the carbachol or erythromycin-stimulated groups 2-6 mL/L of ETOH significantly inhibited both the amplitude and the frequency. Interestingly, a 4-5 min administration of 0.6% ETOH suddenly and completely relaxed the SO. CCK (0.1 micromol/L) stimulated the baseline amplitude from 12.37+/-0.05 mN to 27.40+/-1.82 mN within 1.60+/-0.24 min. After this peak, the amplitude decreased to 17.17+/-0.22 mN and remained constant during the experiment. The frequency peaked at 12.8+/-0.2 c/min, after which the constant frequency was 9.43+/-0.24 c/min throughout the rest of the experiment. ETOH at a dose of 4 mL/L significantly decreased the amplitude from 16.13+/-0.23 mN to 14.93+/-0.19 mN. However, no significant changes in the contractile frequency were observed. ETOH at a dose of 6 mL/L inhibited both the amplitudes and the frequencies in the CCK-stimulated group, while 8 mL/L of ETOH completely relaxed the SO. CONCLUSION: ETOH strongly inhibits the basal, carbachol, erythromycin, and CCK-stimulated rabbit SO motility. Therefore, it is possible that during alcohol-intake the relaxed SO opens the way for pancreatic fluid to flow out into the duodenum in rabbits. This relaxation of the SO may protect the pancreas against alcohol-induced damage.

L-arginine-induced experimental pancreatitis.

Despite medical treatment, the lethality of severe acute pancreatitis is still high (20-30%). Therefore, it is very important to find good animal models to characterise the events of this severe disease. In 1984, Mizunuma et al. developed a new type of experimental necrotizing pancreatitis by intraperitoneal administration of a high dose of L-arginine in rats. This non-invasive model is highly reproducible and produces selective, dose-dependent acinar cell necrosis. Not only is this a good model to study the pathomechanisms of acute necrotizing pancreatitis, but it is also excellent to observe and influence the time course changes of the disease. By writing this review we illuminate some new aspects of cell physiology and pathology of acute necrotizing pancreatitis. Unfortunately, the reviews about acute experimental pancreatitis usually did not discuss this model. Therefore, the aim of this manuscript was to summarise the observations and address some challenges for the future in L-arginine-induced pancreatitis.

Insulin is necessary for the hypertrophic effect of cholecystokinin-octapeptide following acute necrotizing experimental pancreatitis.

AIM: In previous experiments we have demonstrated that by administering low doses of cholecystokinin-octapeptide (CCK-8), the process of regeneration following L-arginine (Arg)-induced pancreatitis is accelerated. In rats that were also diabetic (induced by streptozotocin, STZ), pancreatic regeneration was not observed. The aim of this study was to deduce whether the administration of exogenous insulin could in fact restore the hypertrophic effect of CCK-8 in diabetic-pancreatitic rats. METHODS: Male Wistar rats were used for the experiments. Diabetes mellitus was induced by administering 60 mg/kg body mass of STZ intraperitoneally (i.p.), then, on d 8, pancreatitis was induced by 200 mg/100 g body mass Arg i.p. twice at an interval of 1 h. The animals were injected subcutaneously twice daily (at 7 a.m. and 7 p.m.) with 1 ?g/kg of CCK-8 and/or 2 IU mixed insulin (300 g/L short-action and 700 g/L intermediate-action insulin) for 14 d after pancreatitis induction. Following this the animals were killed and the serum amylase, glucose and insulin levels as well as the plasma glucagon levels, the pancreatic mass/body mass ratio (pm/bm), the pancreatic contents of DNA, protein, amylase, lipase and trypsinogen were measured. Pancreatic tissue samples were examined by light microscopy on paraffin-embedded sections. RESULTS: In the diabetic-pancreatitic rats treatment with insulin and CCK-8 significantly elevated pw/bm and the pancreatic contents of protein, amylase and lipase vs the rats receiving only CCK-8 treatment. CCK-8 administered in combination with insulin also elevated the number of acinar cells with mitotic activities, whereas CCK-8 alone had no effect on laboratory parameters or the mitotic activities in diabetic-pancreatitic rats. CONCLUSION: Despite the hypertrophic effect of CCK-8 being absent following acute pancreatitis in diabetic-rats, the simultaneous administration of exogenous insulin restored this effect. Our results clearly demonstrate that insulin is necessary for the hypertrophic effect of low-doses of CCK-8 following acute pancreatitis.

The role of acute hyperinsulinemia in the development of cardiac arrhythmias.

Patients with perturbed metabolic control are more prone to develop cardiac rhythm disturbances. The main purpose of the present preclinical study was to investigate the possible role of euglycemic hyperinsulinemia in development of cardiac arrhythmias. Euglycemic hyperinsulinemia was induced in conscious rabbits equipped with a right ventricular pacemaker electrode catheter by hyperinsulinemic euglycemic glucose clamp (HEGC) applying two different rates of insulin infusion (5 and 10 mIU/kg/min) and variable rate of glucose infusion to maintain euglycemia (5.5 ± 0.5 mmol/l). The effect of hyperinsulinemia on cardiac electrophysiological parameters was continuously monitored by means of 12-lead surface ECG recording. Arrhythmia incidence was determined by means of programmed electrical stimulation (PES). The possible role of adrenergic activation was investigated by determination of plasma catecholamine levels and intravenous administration of a beta adrenergic blocking agent, metoprolol. All of the measurements were performed during the steady-state period of HEGC and subsequent to metoprolol administration. Both 5 and 10 mIU/kg/min insulin infusion prolonged significantly QTend, QTc, and Tpeak-Tend intervals. The incidence of ventricular arrhythmias generated by PES was increased significantly by euglycemic hyperinsulinemia and exhibited linear relationship to plasma levels of insulin. No alteration on plasma catecholamine levels could be observed; however, metoprolol treatment restored the prolonged QTend, QTc, and Tpeak-Tend intervals and significantly reduced the hyperinsulinemia-induced increase of arrhythmia incidence. Euglycemic hyperinsulinemia can exert proarrhythmic effect presumably due to the enhancement of transmural dispersion of repolarization. Metoprolol treatment may be of benefit in hyperinsulinemia associated with increased incidence of cardiac arrhythmias.

Identification of PPARγ ligands with One-dimensional Drug Profile Matching.

INTRODUCTION: Computational molecular database screening helps to decrease the time and resources needed for drug development. Reintroduction of generic drugs by second medical use patents also contributes to cheaper and faster drug development processes. We screened, in silico, the Food and Drug Administration-approved generic drug database by means of the One-dimensional Drug Profile Matching (oDPM) method in order to find potential peroxisome proliferator-activated receptor gamma (PPARγ) agonists. The PPARγ action of the selected generics was also investigated by in vitro and in vivo experiments. MATERIALS AND METHODS: The in silico oDPM method was used to determine the binding potency of 1,255 generics to 149 proteins collected. In vitro PPARγ activation was determined by measuring fatty acid-binding protein 4/adipocyte protein gene expression in a Mono Mac 6 cell line. The in vivo insulin sensitizing effect of the selected compound (nitazoxanide; 50-200 mg/kg/day over 8 days; n = 8) was established in type 2 diabetic rats by hyperinsulinemic euglycemic glucose clamping. RESULTS: After examining the closest neighbors of each of the reference set's members and counting their most abundant neighbors, ten generic drugs were selected with oDPM. Among them, four enhanced fatty acid-binding protein/adipocyte protein gene expression in the Mono Mac 6 cell line, but only bromfenac and nitazoxanide showed dose-dependent actions. Induction by nitazoxanide was higher than by bromfenac. Nitazoxanide lowered fasting blood glucose levels and improved insulin sensitivity in type 2 diabetic rats. CONCLUSION: We demonstrated that the oDPM method can predict previously unknown therapeutic effects of generic drugs. Nitazoxanide can be the prototype chemical structure of the new generation of insulin sensitizers.

Investigation of the metabolic effects of chronic clozapine treatment on CCK-1 receptor deficient Otsuka Long Evans Tokushima Fatty (OLETF) rats.

Clozapine increases meal size and meal duration, effects similar to the pharmacological blockade or congenital deficiency of CCK-1 receptor. We aimed to investigate the role of CCK-1 receptor in clozapine-induced weight gain and insulin sensitivity in CCK-1 receptor deficient, male Otsuka Long Evans Tokushima Fatty rats (OLETF). Long Evans Tokushima Otsuka (LETO) rats served as healthy control. Animals were orally treated with either clozapine (10mg/kg) or its vehicle over 25 days. Daily metabolic parameters were measured by metabolic cages. The insulin sensitivity was determined by hyperinsulinaemic euglycaemic glucose clamping (HEGC). Adiposity was determined by measuring the perirenal, intraabdominal and epididymal white adipose tissue fat pads. Hypothalamic mRNA expression of CCK-1 and CCK-2 receptor was measured by real-time PCR, plasma insulin by radioimmunoassay. Clozapine failed to increase weight gain or daily food intake, but it increased adiposity, 1st meal size and duration and decreased insulin sensitivity both in OLETF or LETO rats. The glucose infusion rate during the steady state of the HEGC was unaltered, but the metabolic clearance rate of insulin was reduced by the clozapine treatment. Hypothalamic mRNA of CCK-1 and CCK-2 receptor was elevated in LETO rats, but the mRNA of CCK-2 receptor was reduced by clozapine in OLETF rats. Our results suggest that the CCK-1 receptor has no direct role in the clozapine-induced adiposity and insulin resistance. We also demonstrated that atypical antipsychotic treatment can induce insulin resistance in the absence of manifest obesity in male rats.

Insulin resistance occurs in parallel with sensory neuropathy in streptozotocin-induced diabetes in rats: differential response to early vs late insulin supplementation.

We investigated whether progressive sensory neuropathy was accompanied by changes in whole-body insulin sensitivity (WBIS) in rats made diabetic by streptozotocin (STZ). The effects of early and late insulin supplementation were also studied. The STZ-treated rats failed to gain weight and exhibited stable hyperglycemia and low plasma insulin levels with a decrease in nerve conduction velocity (NCV) measured in A and C fibers of the saphenous nerve. A decreased sensory neuropeptide (SNP) release such as that of substance P, somatostatin, and calcitonin gene-related peptide determined from organ fluid of tracheal preparations subjected to electrical field stimulation also occurred in diabetic animals. These features were accompanied by a decrease in WBIS measured by hyperinsulinemic-euglycemic glucose clamping and a decrease in insulin-stimulated glucose uptake in cardiac and gastrocnemius muscle. When insulin supplementation with slow-release implants (2 IU/d) was started 4 weeks after STZ injection, blood glucose level normalized. Both insulin sensitivity and sensory nerve function reflected in either NCV or SNP release completely recovered by the 12th post-STZ week. When the insulin implants were applied from the eighth post-STZ week, both WBIS and glucose uptake remained significantly decreased, with a seriously impaired NCV and SNP release with strong hyperglycemia. Late insulin supplementation, however, even by using double implantation from the 10th post-STZ week, was unable to restore blood glucose, WBIS, NCV, and SNP release by the 12th week. Insulin resistance occurs in parallel with sensory neuropathy in STZ-diabetic rats. Both can be improved by early but not late insulin supplementation.

Involvement of cholecystokinin in baseline and post-prandial whole body insulin sensitivity in rats.

The objective of the study was to investigate the role of cholecystokinin (CCK) on the food-induced insulin sensitization phenomenon in healthy Long Evans Tokushima Otsuka (LETO) and Otsuka Long Evans Tokushima Fatty (OLETF) rats. Whole body insulin sensitivity determined by hyperinsulinaemic euglycaemic glucose clamping and the rapid insulin sensitivity test served as endpoints. Determinations were done in both fasted and re-fed animals. The involvement of CCK in post-prandial insulin sensitization was assessed by using proglumide, a CCK receptor blocker, by assessment of hypothalamic CCK-1/CCK-2 receptor expression by rt-PCR technique and by plasma insulin immunoreactivity determinations by means of radioimmunoassay as pharmacological, genetic and analytical approaches, respectively. The body weight of the OLETF rats and the amount of food consumed much exceeded those seen with LETO rats. The post-prandial increase in insulin sensitivity was marked in LETO, but not in OLETF rats. Intravenous proglumide attenuated post-prandial insulin sensitivity in LETO rats, with no effect in OLETF rats. Nevertheless, baseline insulin sensitivity was much lower in OLETF than in LETO rats. Treatment with rosiglitazone increased baseline insulin sensitivity of OLETF rats and evoked an increase in CCK-1 receptor gene expression in LETO rats. The results provide evidence for the involvement of CCK receptors in adjustment of both fasting and post-prandial insulin sensitivity. The data obtained with OLETF rats strongly suggest the predominant role of CCK-1 receptors.

Meal-induced enhancement in insulin sensitivity is not triggered by hyperinsulinemia in rats.

Several reports confirmed the phenomenon of postprandial increase in whole-body insulin sensitivity. Although the initial step of this process is unknown, the pivotal role of postprandial hyperinsulinemia has strongly been suggested. The aim of the present study was to determine whether hyperinsulinemia per se induces insulin sensitization in healthy male Wistar rats. Rapid insulin sensitivity test (RIST) were performed in fasted, anesthetized rats before and during stable hyperinsulinemia achieved by hyperinsulinemic euglycemic glucose clamping (HEGC) with insulin infused either through the jugular vein (systemic HEGC) or into the portal circulation (portal HEGC) at a rate of 3 mU/(kg min). Insulin sensitivity expressed by the rapid insulin sensitivity (RIST) index (in milligrams per kilogram) was characterized by the total amount of glucose needed to maintain prestudy blood glucose level succeeding an intravenous bolus infusion of 50 mU/kg insulin over 5 minutes. In fasted animals, the RIST index was 37.4 +/- 3.1 mg/kg. When hyperinsulinemia mimicking the postprandial state was achieved by systemic HEGC, the RIST index (39.7 +/- 10.6 mg/kg) showed no significant changes as compared with the pre-HEGC values. Hyperinsulinemia achieved by portal insulin infusion also failed to modify the RIST index (35.7 +/- 4.3 mg/kg). The results demonstrate that acute hyperinsulinemia, no matter how induced, does not yield any sensitization to the hypoglycemic effect of insulin.

Cyclic GMP-mediated activation of a glibenclamide-sensitive mechanism in the rabbit sphincter of Oddi.

We investigated whether glibenclamide-sensitive potassium channels are involved in cyclic GMP (cGMP)-mediated relaxation of the rabbit Oddi's sphincter. Changes in isometric tension were measured in the presence of atropine (1 microM) and guanethidine (4 microM). Concentration-response curves for nitroglycerin, vasoactive intestinal polypeptide (VIP), and sodium nitroprusside (SNP) were shifted to the right in the presence of (p-chloro-D-Phe6, Leu17)-VIP (VIPa), a VIP receptor antagonist. Glibenclamide (1 microM) attenuated the relaxations to VIP, nitroglycerin, or 8-bromo cGMP. In the presence of tetrodotoxin (TTX), glibenclamide attenuated relaxations to VIP without effect on those to nitroglycerin. Furthermore, nitroglycerin increased both cAMP and cGMP concentrations, however, it failed to increase the tissue cAMP concentration in the presence of TTX. VIPa also blocked the increase in content of either cyclic nucleotide. VIP increased cAMP with a TTX-sensitive increase in cGMP content. 8-Bromo cGMP (1 microM) significantly increased the tissue cAMP content. This was blocked by either TTX or VIPa (both 1 microM). We conclude that ATP-sensitive potassium channel (KATP) activation contributes to cGMP-mediated relaxation of the Oddi's sphincter of the rabbit. Activation of KATP results from a cyclic AMP-mediated process due to cGMP-dependent VIP release from neurons.