Protective effects of silymarin-loaded chitosan nanoparticles in the diet-induced hyperlipidemia rat model.

Objectives: Obesity is a metabolic syndrome that leads to many chronic diseases worldwide. In this study, we investigate the antihyperlipidemic activities of chitosan nanoparticles (CH NPs) on silymarin (SIL) as a carrier in the drug delivery system that can improve some biochemical parameters and hormones in the model of hyperlipidemic rats receiving a high-fat diet (HFD). Materials and Methods: Physicochemical characterization of silymarin-loaded chitosannanoparticles (CH-SIL NPs) was done by Fourier-transform infrared (FTIR) spectroscopy, dynamic light scattering (DLS), and drug loading efficiency (LE). Diet-induced hyperlipidemic rats were treated with SIL (15 mg/kg/day) and CH-SIL NPs(15 mg/kg/day) for twelve weeks orally daily. The body weight loss (BW), food consumption, serum total cholesterol (TC), triglycerides (TG), high-density lipoprotein (HDL), levels of fasting blood glucose (FBG) in serum, serum insulin, cortisol, testosterone, and brain neuropeptide Y (NPY), Y1 and Y5 receptor mRNA expression were analyzed. Results: A significant reduction in BW and food consumption from 417 ± 16 g and 33 ± 1.03 in group HFD to 338 ± 10 g and 17.33 ± 1.02 in group CHS+HFD was observed, respectively. This data revealed that CH-SIL NPs improved hyperlipidemia, hyperinsulinemia, and hyperglycemia, reduced serum cortisol, and down-regulated NPY and Y1R with a significant increase in HDL and testosterone hormones compared to the control group. Conclusion: The developed Sil-loaded CH NPs were good agents for improving efficacy. It is the first report of the proposed weight loss mechanism of SIL CH NPs, thereby providing information about the anti-hyperlipidemic and antihyperglycemic effects of silymarin-loaded chitosan nanoparticles, a natural food with proper effects against metabolic disorders in case of hyperlipidemia that may lead to obesity and up-regulation of brain NPY.

Synergistic cytotoxicity effect of the combination of chitosan nanoencapsulated imatinib mesylate and quercetin in BCR-ABL positive K562 cells.

Objectives: Intolerable side effects and resistance to chemotherapeutic drugs have encouraged scientists to develop new methods of drug combinations with fewer complications. This study aimed to investigate the synergistic effects of quercetin and imatinib encapsulated in chitosan nanoparticles on cytotoxicity, apoptosis, and cell growth of the K562 cell line. Materials and Methods: Imatinib and quercetin were encapsulated in chitosan nanoparticles and their physical properties were determined using standard methods and SEM microscope images. BCR-ABL positive K562 cells were cultured in a cell culture medium, cytotoxicity of drugs was determined by MTT assay and the effects of nano drugs on apoptosis in cells were investigated by Annexin V-FITC staining. The expression level of genes associated with apoptosis in cells was measured by real-time PCR. Results: The IC50 for the combination of the nano drugs at 24 and 48 hr was 9.324 and 10.86 µg/ml, respectively. The data indicated that the encapsulated form of drugs induced apoptosis more effectively than the free form (P<0.05). Moreover, the synergistic effect of nano drugs in statistical analysis was proved (P=0.001). The combination of nano drugs resulted in the caspase 3, 8, and TP53 genes upregulation (P=0.001). Conclusion: The results of the present study showed that the encapsulated form of imatinib and quercetin nano drugs with chitosan has more cytotoxicity than the free form of the drugs. In addition, the combination of imatinib and quercetin as a nano-drug complex has a synergistic effect on the induction of apoptosis in imatinib-resistant K562 cells.

Protective Effects of Phoenixin-14 Peptide in the Indomethacin-Induced Duodenal Ulcer: An Experimental Study.

Phoenixin-14 (PNX -14 ) is a newly identified neuropeptide with potential anti-inflammatory effects in the gastrointestinal tract. In this study, we evaluated the protective effect of PNX-14 against the formation of experimental indomethacin (IND)-induced duodenal ulcer. Thirty-two male Sprague-Dawley rats were randomly assigned to the four following study groups: (1) negative control (2) IND (7.5 mg/kg subcutaneous IND), (3) famotidine (FA) (7.5 mg/kg subcutaneous IND followed by 40 mg/kg intraperitoneal FA), and (4) PNX-14 (7.5 mg/kg subcutaneous IND followed by 50 µ/kg intraperitoneal PNX-14). Outcome measures included macroscopic evaluation of duodenal lesion, serum levels of IL-1ß, TNF-α, IL-6, and IL-12, and tissue biochemical parameters of oxidative stress, including malondialdehyde (MDA) , myeloperoxidase (MPO) activity, superoxide dismutase (SOD) activity, and catalase activity. Results The macroscopic grade of duodenal lesions were significantly smaller in the PNX-14 group than in the IND group (p < 0.001). Serum inflammatory cytokines were significantly increased in the IND group. PNX-14 treatment significantly decreased the serum levels of inflammatory cytokines (p < 0.0001). Oxidative contents (MDA and MPO activity) were significantly smaller in the PNX-14 group compared with the IND group (p < 0.0001), while anti-oxidative contents (SOD and catalase activity) were significantly more (p < 0.0001). PNX-14 was superior to FA in several anti-inflammatory properties, such as inhibiting the release of inflammatory cytokines and increasing the catalase activity. PNX-14 showed significant protective effects against the formation of IND-induced duodenal ulcers. These results suggest a promising therapeutic implication for PNX-14 in the treatment of gastrointestinal inflammatory disorders.

Effect of ß-asarone in normal and ß-amyloid-induced Alzheimeric rats.

INTRODUCTION: ß-Asarone is a major component of Acorus tatarinowii Schott. It has pharmacological effects that include antihyperlipidemic, anti-inflammatory, and antioxidant activity. In the present study, the effect of ß-asarone on neurodegeneration induced by intrahippocampal administration of ß-amyloid was investigated in adult male Wistar rats. MATERIAL AND METHODS: The rats were randomly divided into 9 groups: normal control, sham-operated control, ß-asarone (12.5, 25, and 50 mg/kg intragastrically, daily) alone, Alzheimeric control rats (ß-amyloid, intrahippocampal), ß-asarone (12.5, 25, and 50 mg/kg intragastrically, daily) together with ß-amyloid, and treatment was performed accordingly. Animals were injected with ß-amyloid bilaterally. Animals received ß-asarone daily using an intragastric tube for 50 days, starting from 30 days before administration of the ß-amyloid. The rats were sacrificed and parameters of oxidative stress, superoxide dismutase (SOD) and glutathione peroxidase (GPX) activity were measured in hippocampus homogenate. Histopathological changes were examined by Bielschowsky staining. RESULTS: Our results showed that administration of ß-asarone (25 and 50 mg/kg) significantly increased the levels of antioxidant enzymes, including SOD (1.09 ±0.02, 1.21 ±0.02, p < 0.001, respectively) and GPX (58.94 ±0.78, 68.92 ±3.64, p < 0.001, respectively) in comparison with Alzheimeric control rats (SOD and GPX level for Alzheimeric control group: 0.44 ±0.01, 35.09 ±1.15, respectively). Histopathological examination showed that ß-asarone decreased cell loss in the cerebral cortex and hippocampus in Alzheimeric rats. CONCLUSIONS: These results indicate that ß-asarone is effective in providing protection against oxidative stress and neuronal damage induced by ß-amyloid.

Effect of troxerutin on oxidative stress and expression of genes regulating mitochondrial biogenesis in doxorubicin-induced myocardial injury in rats.

Because of limitation of doxorubicin (DOX) clinical application in chemotherapy due to its cardiotoxicity, finding new strategies to reduce DOX challenge and improve patients' outcomes is crucial. Due to positive cardiovascular impacts of troxerutin (TXR), here we have investigated the effect of TXR on DOX-induced cardiotoxicity by evaluating the myocardial oxidative stress and expression of genes regulating mitochondrial biogenesis. Male Wistar rats (250-300 g) were randomly allocated into four groups: control, TXR, DOX, and TXR + DOX. Troxerutin (150 mg/kg) was orally administrated once a day through a gavage tube for 4 weeks before DOX challenge. The TXR-treated and time-matched control rats received intraperitoneal injection of DOX (20 mg/kg). Three days after DOX challenge, the left ventricular samples were obtained to determine the expression of genes regulating mitochondrial biogenesis via real-time PCR. Myocardial creatine kinase (CK-mB), oxidative stress markers, and mitochondrial function (generation of reactive oxygen species or ROS and ATP levels) were also evaluated using commercial kits and spectrophotometric and fluorometric methods. DOX administration significantly increased the levels of CK-mB, malondialdehyde (MDA), and mitochondrial ROS levels, while reduced the cellular ATP production and expression levels of SIRT-1, PGC-1α, and NRF-2 as well as superoxide dismutase, glutathione peroxidase, and catalase activity in comparison to control group (P < 0.05 to P < 0.01). Pretreatment of DOX-received rats with TXR significantly upregulated the expression of all biogenesis genes and antioxidant enzymes with non-significant effect on catalase activity, and significantly reduced CK-mB and MDA levels toward control values (P < 0.05 to P < 0.01). Mitochondrial ROS and ATP levels were also restored significantly by pretreatment with TXR (P < 0.05). The data suggested that preconditioning of rats with TXR had protective effect on DOX-induced cardiotoxicity through inducing antioxidative properties and restoring the mitochondrial function and the expression profiles of myocardial SIRT-1/PGC-1α/NRF-2 network.

Interaction between leptin and glutamatergic system on food intake regulation in neonatal chicken: role of NMDA and AMPA receptors.

Objective: The aim of the current study was to determine the possible interaction of the central leptin and Glutamatergic systems on feeding behavior in neonatal 3-hours food deprived (FD3) broilers chickens.Methods: In experiment 1, FD3 chicken received intracerebroventricular (ICV) injection of control solution (group i) and 2.5, 5 and 10 µg of Leptin (groups ii-iv). In experiment 2, FD3 chicken were ICV injected with (group i) control solution and groups ii-iv with 2.5, 5 and 10 nmol of AG-490 (JAK2 antagonist). In experiment 3, injections were (i) control solution, (ii) Leptin (10 µg), (iii) AG-490 (2.5 nmol) and (iv) Leptin + AG-490. In experiment 4, broiler chickens were ICV injected with (i) control solution, (ii) Leptin (10 µg), (iii) MK-801 (NMDA glutamate receptors antagonist; 15 nmol) and (iv) Leptin + MK-801. Experiments 5-9 were similar to experiment 1, except chicken were ICV injected with CNQX (AMPA receptor antagonist, 390 nmol), UBP-302 (Kainate receptor antagonist, 390 nmol), AIDA (mGluR1 antagonist, 2 nmol), LY341495 (mGluR2 antagonist, 150 nmol) and UBP1112 (mGluR3 antagonist, 2 nmol) instead of MK-801. Then, food intake was measured until 120 min after injection.Results: ICV injection of leptin (2.5, 5 and 10 µg) significantly decreased food intake in a dose dependent manner (p < 0.05). Also, ICV injection of the JAK2 antagonist (2.5, 5 and 10 nmol) had hyperphagic effect in chicken (p < 0.05). Co-administration of leptin + AG-490, partially decreased leptin-induced hypophagia in broiler chicken (p < 0.05). In addition, co-injection of leptin + MK-801 significalty inhibited leptin-induced hypophagia in neonatal chicken (p < 0.05). Also, co-administration of leptin + CNQX partially attenuated hypophagic effect of leptin in chicken (p < 0.05).Conclusion: The results of present study suggest that leptin has hypophagic effect in neonatal chicken and this effect is probably mediated via NMDA and AMPA glutamatergic receptors.

Pretreatment with vildagliptin boosts ischemic-postconditioning effects on cardioprotection and expression profile of genes regulating autophagy and mitochondrial fission/fusion in diabetic heart with reperfusion injury.

The burden of myocardial ischemia/reperfusion (IR) injury is 2-3-folds higher in diabetic patients, so protecting diabetic hearts is clinically important. Here, we investigated the effect of combinational therapy with vildagliptin and ischemic postconditioning (IPostC) on cardioprotection and the expression of genes regulating autophagy and mitochondrial function in diabetic hearts with IR injury. Type 2 diabetes was induced through high-fat diet and streptozotocin protocol in Wistar rats. Vildagliptin was orally administered to diabetic rats 5 weeks before IR injury. Myocardial-IR injury was modeled by ligation of left the coronary artery for 30 min followed by 60-min reperfusion, on a Langendorff-perfusion system. IPostC was applied at early reperfusion as 6 alternative cycles of 10-s reperfusion/ischemia. Creatine-kinase levels were measured spectrometrically, and infarct size was evaluated by TTC staining method. Left ventricles were harvested for assessing the expression levels of autophagy and mitochondrial-related genes using real-time PCR. Induction of diabetes significantly increased creatine-kinase release in comparison to healthy rats, and all treatments significantly reduced the release of enzyme toward control levels (P < 0.05). Only the combination therapy (IPostC + vildagliptin) could significantly reduce the infarct size of diabetic hearts as compared to untreated diabetic-IR group (P < 0.01). The levels of autophagy genes LC3 and p62 were significantly higher in diabetic groups than healthy ones. Induction of IR injury in diabetic hearts enhanced mitochondrial fission (drp-1) and reduced mitochondrial fusion (mfn1 and mfn2) genes. IPostC alone had no significant effect on the gene expression and vildagliptin alone could only affect LC3-II and mfn2 expressions. Nevertheless, administration of combination therapy significantly reduced the expression of both autophagy genes and increased both LC3-II/I and mfn2/1 ratios as compared with diabetic-IR hearts (P < 0.01-0.05). Application of this combination therapy could overcome the diabetes-induced failure of cardioprotection by individual treatments and improve mitochondrial dynamic and autophagy flux.

The Lentiviral Vector Pseudotyped by Modified Rabies Glycoprotein Does Not Cause Reactive Gliosis and Neurodegeneration in Rat Hippocampus

Background: A human immunodeficiency virus type 1 (HIV-1)-based lentiviral vector (LV) pseudotyped by a variant of rabies envelope glycoprotein, FUG-B2, has previously been prepared and used in transfection of hippocampal CA1 ("Cornu Ammonis" area 1) neurons. This study aimed to verify reactive gliosis and neuronal damage after injection of the vector into the rat hippocampus. Methods: HEK 293T cells were transfected with transfer (fck-Jaws-GFP-ER2), envelope (FUG-B2), and packaging (pMDLg/pRRE, pRSV-Rev) plasmids, and the vector was injected into CA1 of the rat hippocampus. After one week, transduction efficiency, and the number of neuronal and astroglial cells were determined in CA1 and CA3 by double staining of the brain slices. Results: Hippocampal cells were successfully transfected as 92.7% of CA1 and 95.8% of CA3 neuronal cells expressed GFP. The frequency of neuronal and astroglial cells in CA1 and CA3 of the vector-injected rats remained unchanged compared to those in the control and the saline-injected rats. Furthermore, no morphological change was found in hippocampal astrocytes and neuronal cells. Conclusion: The HIV-1-based LV pseudotyped by FUG-B2 is safe and does not cause neuroinflammation and neuronal loss once directly delivered into the rat hippocampus.

Effect of sinapic acid on memory deficits and neuronal degeneration induced by intracerebroventricular administration of streptozotocin in rats.

The present study aimed to elucidate the neuroprotective effect of sinapic acid on intracerebroventricular streptozotocin (ICV-STZ) induced neuronal loss and memory impairment. To test this hypothesis, male Wistar rats were randomly divided into 11 groups: normal control, sham-operated control, sinapic acid (2.5, 5, 10, and 20 mg/kg bw intragastrically, daily) alone, Alzheimer control rats (ICV-STZ, 3 mg/kg bw), sinapic acid (2.5, 5, 10, and 20 mg/kg bw intragastrically, daily) together with STZ, and the treatment was performed accordingly. After 28 days of ICV-STZ administration, the animals were assessed for cognitive performance using passive avoidance test and then sacrificed for biochemical and histopathological examinations. Sinapic acid was found to be effective in improving antioxidant status and preventing memory loss in Alzheimer rats. Moreover, TNF-α level in the hippocampus was significantly decreased by sinapic acid. Also, administration of sinapic acid significantly increased the levels of antioxidant enzymes and decreased malondialdehyde level in the hippocampus. Histopathological examination showed that sinapic acid reduced cell loss in the cerebral cortex and hippocampus in Alzheimer's rats. The present study suggests that sinapic acid is effective in the prevention of memory loss and improvement of oxidative stress and might be beneficial in the treatment of Alzheimer's disease.

Transduction efficacy and retrograde movement of a lentiviral vector pseudotyped by modified rabies glycoprotein throughout the trisynaptic circuit of the rat hippocampus.

BACKGROUND: The trisynaptic circuit (entorhinal cortex-dentate gyrus-CA3-CA1) is a key unidirectional network in the hippocampus. Damage to the hippocampus interrupts this circuit and causes neurological disorders. Efficient delivery of therapeutic genes into this network is of great interest with respect to treating trisynaptic circuit pathologies. METHODS: We generated a lentivector system pseudotyped by a variant of rabies glycoprotein, FUG-B2. The efficiency of the vector in the retrograde transduction of the rat hippocampal neurons (i.e. the entorhinal cortex from the dentate gyrus, the dentate gyrus from CA3, and CA3 from CA1) was examined by direct injection of the vector into the dentate gyrus, CA3 and CA1. To distinguish transduction of the neuronal and glial cells, as well as selective retrograde gene transfer, double-staining of the green fluorescent protein (GFP) expressing cells with the specific neuron biomarker NeuN (neuronal nuclear protein) and the specific glia biomarker GFAP (glial fibrillary acidic protein) was performed across the network. RESULTS: The transgene was successfully introduced into the circuit. More than 80% of the neuronal and glial cells at the injection sites preserved GFP expression during the 2-month period after vector injection. Importantly, GFP was expressed selectively in almost 80.0% of the presynaptic neuronal cells by retrograde axonal transport of the vector. CONCLUSIONS: The FUG-B2-based vector system can efficiently introduce the transgene into the rat hippocampal neurons both directly and indirectly through retrograde monosynaptic movement. This efficient and long-lasting gene delivery might provide a tool for treating neurological disorders originating in hippocampal circuits.

Interaction between central GABAA receptor and dopaminergic system on food intake in neonatal chicks: role of D1 and GABAA receptors.

OBJECTIVE: The present study was designed to examine the role of central γ-Aminobutyric acidA receptors and dopaminergic system on feeding behaviour in neonatal layer-type chicken. METHODS: In this study, six experiments were designed, each with four treatment groups (n = 44 in each experiment). In experiment 1, four groups of 3-h food-deprived chicks received a dose of either the intracerebroventricular injection of (1) control solution, (2) Levo-dihydroxyphenylalanine as precursor of dopamine; 125 nmol, (3) Gaboxadol (γ-Aminobutyric acidA receptor agonist, 0.2 µg) and (4) Levo-dihydroxyphenylalanine (125 nmol) plus Gaboxadol (0.2 µg). Experiments 2-6 were similar to experiment 1, except that the chickens were intracerebroventricular-injected with 6-hydroxydopamine (is a neurotoxin; 2.5 nmol), SCH23390 (D1 receptor antagonist, 5 nmol), AMI-193 (D2 receptor antagonist, 5 nmol), NGB2904 (D3 receptor antagonist, 6.4 nmol) and L-741,742 (D4 receptor antagonist, 6 nmol) instead of levo-dihydroxyphenylalanine. Then, the cumulative food intake was measured until 120 min post-injection. RESULTS: According to the results, intracerebroventricular injection of Gaboxadol (0.2 µg) significantly increased the food intake (P < 0.05). Co-injection of the 6-hydroxydopamine + Gaboxadol significantly amplified the food intake (P < 0.05). Intracerebroventricular injection of SCH23390 (5 nmol) + Gaboxadol (0.2 µg) significantly amplified the Gaboxadol-induced hyperphagia (P < 0.05). No significant effect was observed by co-injection of the D2-D4 receptor antagonists + Gaboxadol (P > 0.05). CONCLUSION: These results suggested the interconnection between central Dopaminergic and γ-Aminobutyric acidA on the feeding behaviour mediates via D1 and γ-Aminobutyric acidA receptors in 3-h food-deprived neonatal layer-type chicken.

Modulatory function of NMDA glutamate receptor on MC3/MC4 receptors agonist-induced hypophagia in neonatal meat-type chicken.

Melanocortin 3 and 4 receptors (MC3R and MC4R) are known as the main receptors for melanocortin-induced hypophagia in mammalian and poultry. Also, central glutamatergic system has mediatory role on function of the melanocortin system in some brain areas. So, the aim of the current study was to determine the role of MC3/MC4 receptors agonist on food intake and its interaction with glutamatergic in 3-h food-deprived (FD3) neonatal broilers. In experiment 1, chickens were intracerebroventricular (ICV) injected with control solution, MTII (MC3/MC4 receptors agonist; 2.45, 4.8 and 9.8 pmol). In experiment 2, control solution, SHU9119 (MC3/MC4 receptors antagonist; 0.5, 1 and 2 nmol) were ICV injected. In experiment 3, birds ICV injected with control solution, SHU9119 (0.5 nmol), MTII (9.8 pmol) and co-injection of the SHU9119 + MTII. Experiments 4-8 were similar to experiment 3, except birds injected with MK-801 (NMDA glutamate receptors antagonist, 15 nmol), CNQX (AMPA glutamate receptors antagonist; 390 nmol), AIDA (mGLUR1 glutamate receptors antagonist; 2 nmol), LY341495 (mGLUR2 glutamate receptors antagonist; 150 nmol) and UBP1112 (mGLUR3 glutamate receptors antagonist; 2 nmol) instead of SHU9119. Then, cumulative food intake was recorded until 120 min after injection. According to the results, dose dependent hypophagia observed after ICV injection of the MTII (p < 0.05). ICV injection of SHU9119 significantly increased food intake in birds (p < 0.05). Co-injection of SHU9119 + MTII significantly inhibited MTII- induced hypophagia in neonatal chicks (p < 0.05). In addition, hypophagia- induced by MTII was significantly attenuated with co-injection of MTII + MK-801(p < 0.05). These results suggested MC3 and MC4 receptors have inhibitory role on food intake and this effect is probably mediated by NMDA glutamate receptors in neonatal chickens.

Effects of berberine on ß-secretase activity in a rabbit model of Alzheimer's disease.

INTRODUCTION: Relevant aspects of Alzheimer's disease (AD) can be modeled by aluminium-maltolate injection into specific regions of the brain. The possible role of berberine chloride (BC) as an anti-inflammatory agent in the brain has been previously addressed. MATERIAL AND METHODS: Rabbits were divided into control (C), untreated lesion (L) and BC-treated + lesion (L + BC) groups. Animals in L + BC received BC (50 mg/ kg) orally 1 day after surgery and daily for 2 weeks. The lesion was induced by injection of 100 µl of either vehicle or water containing 25 mM aluminium-maltol into intraventricular fissure. Weight loss, ataxia, paralysis and tremor were monitored. For histopathology, Bielschowsky silver and H&E staining were employed. ß-Secretase activity in hippocampus was finally assessed. RESULTS: All L animals died on days 12-15 after lesion. Seven to 10 days after lesion, abnormal symptoms as well as cachexia were seen in over 90% of cases. L rabbits lost an average of 0.5 kg which was significant on days 10 and 12 (p < 0.05); this was not completely prevented by BC. Up to day 15, all L animals had lost their lives (p < 0.001). BC treatment protected the hippocampus from degeneration, altered the behavior and decreased the activity of ß-site amyloid precursor protein cleaving enzyme-1 (BACE-1). CONCLUSIONS: Considering the findings in regard to physiological abilities, histological changes and BACE-1 activity in hippocampus changes, it is concluded that BC treatment could be an effective therapy in restoring Al maltol-induced behavioral derangements in the rabbit model of AD.