Blockade of the Naloxone-induced Aversion in Morphine-conditioned Wistar Rats by L-Arginine Intra-central Amygdala

AbstractObjective(s)Single injection of naloxone, a selective antagonist of morphine, prior to the drug conditioning testing was used to investigate on morphine dependence.Materials and MethodsConditioning to morphine (2.5-10 mg/kg, s.c.) was established in adult male Wistar rats (weighing 200-250 g) using an unbiased procedure. Nitric oxide agents were microinjected into the central amygdala prior to naloxone-paired place conditioning testing.ResultsThe results showed that morphine produced a significant dose-dependent place preference in animals. Naloxone (0.1-0.4 mg/kg, i.p.) injections pre-testing of the response to morphine (7.5 mg/kg, s.c.) caused a significant aversion at the higher doses (0.4 mg/kg, i.p.). This response was reversed by microinjection of L-arginine (0.3-3 µg/rat, intra-central amygdala) prior to naloxone on the day of the testing. The response to L-arginine was blocked by pre-injection of NG-nitro-L-arginine methyl ester (L-NAME) (intra-central amygdala).ConclusionA single injection of naloxone on the test day of morphine place conditioning may simply reveal the occurrence of morphine dependence in rats, and that the nitric oxide in the central amygdala most likely plays a key role in this phenomenon

Identification, Cloning and Structural Analysis of Major Genes from Portulaca oleracea L. Hairy Roots that Involved in the Biosynthesis of Dopamine

Dopamine is one of the important medications of Portulaca oleracea L. To optimize the production of dopamine, one of the methods is the identification and engineering of metabolite pathways. To investigate the tyrosine decarboxylase (TDC) and tyrosinase, which seem to be the most important genes in dopamine synthesis pathway, hairy roots were produced from Portulaca oleracea using Agrobacterium rhizogenes and total RNA was extracted from hairy roots. A cDNA library was synthesized using RT-PCR. Then, the twogenes were amplified, isolated and cloned in a pTG 19-T vector. Bioinformatics' databases were used to predict the details of the structural, functional and biological characteristic of these genes. Nucleotide sequence analysis revealed that the cloned cDNAs expressed TDC and tyrosinase, and contained a single open reading frame of 1800 bp and 1750 bp, respectively. TDC has the most similarity with TDC of Arabidopsis thaliana (L.) Heynh.,but tyrosinase has 98% similarity withtyrosinase of Agaricus bisporus. Because of More negatively charged amino acids the TDC has hydrophobic properties, therefore affinity and hydrophilic chromatography can be used for purification of TDC. But tyrosinase has hydrophilic properties and hydrophobicity chromatography can be used for its purification. There were two peroxisomal signal peptide (KLAKEFEQL) and (KIEGRPLHL) in the TDC and tyrosinase, respectively. Therefore, they are biologically active in the peroxisomes, and included in biosynthesis dopamine through the transformation of L-lysine to L-dopa and finally to the dopamine. In conclusion, increasing the expression of TDC and tyrosinase through the genetic engineering can increase dopamine production in the Portolaca

The effect of Migri-Heal® on nitric oxide production in an in vitro inflammatory model of primary microglial cells

Background: Recently, much attention has been directed towards considering activated microgelial cells as putative targets for treatment of neurological disorders. MigriHeal® as a novel herbal remedy was introduced for the treatment of migraine headaches. The previous researches has shown that MigriHeal® extracts can decrease NO in an in vitro inflammatory model. The aim of this study was to investigate the effect of MigriHeal® on NO generation from LPS- stimulated microglia cells.Materials and Methods: Neonatal rat primary microglial cells were isolated from the mixed glial cultures and the purity of the cultures was determined by immunocytochemistry. Microglial cells were pretreated with Migri-Heal® and activated by 1μg/ml LPS. Subsequently, NO levels in the culture supernatants were measured by a griess reaction. Our results showed that Migri-Heal® 50μg/ml significantly reduced NO level in inflamed microglia in a dose-dependent manner. Results: The results showed that different concentrations of Migri-Heal® had no prominent effect on cell viability in presence of LPS as compared with the control group. In addition, the pretreatment of microglia cells with Migri-Heal® can prevent from a morphological changes of the cells into the round and phagocytic shape. Conclusion: Our study demonstrated that MigriHeal® might have NO scavenging properties. Integrative studies are warranted to uncover the novel pharmacological insights of this herbal remedy as an putative therapeutic approach against diseases - associated with inflammation

Computational study of SENP1 in cancer by novel natural compounds and ZINC database screening

Introduction: Sentrin-specific protease 1 (SENP1) is a protein whose main function is deSUMOylation. SENP1 inhibits apoptosis, and increases angiogenesis, estrogen and androgen receptor transcription and c-Jun transcription factor, proliferation, growth, cell migration, and invasion of cancer. The in vivo and in vitro studies also demonstrated which natural compounds, especially phytochemicals, minerals, and vitamins, prevent cancer. More than 3,000 plant species have been reported in modern medicine. Natural compounds have many anti-cancerous andanti-turmeric properties such as antioxidative, antiangiogenic, antiproliferative, and pro-apoptotic properties.Methods: In this study, we investigated the interaction of some natural compounds with SENP1 to inhibit its activity. We also screened the ZINC database including natural compounds. Molecular docking was performed, and toxicity of compounds was determined; then, molecular dynamics simulation (MDS) and essential dynamics (ED) were performed on natural compounds with higher free binding energies and minimal side effects. By searching in a large library, virtual screening of the ZINC database was performed using LibDock and CDOCKER, and the final top 20 compounds were allowed for docking against SENP1. According to the docking study, the top three leading molecules were selected and further analyzed by MDS and ED.Results: The results suggest that resveratrol (from the selected compounds) and ZINC33916875 (from the ZINC database) could be more promising SENP1 inhibitory ligands.Discussion: Because these compounds can inhibit SENP1 activity, then they can be novel candidates for cancer treatment. However, wet laboratory experiments are needed to validate their efficacy as SENP1 inhibitors

Collagen as Adherent Substratum and Inducer of Dorsal Root Ganglia Outgrowth

ABSTRACT Neurite outgrowth from dorsal root ganglion (DRG) explants is a method of evaluating neurotrophic activity of growth factors. When complete medium containing collagen was supplemented with nerve growth factor (NGF) DRG outgrowth was observed after 18 h. In the absence of NGF and in the presence of collagen, the DRG outgrowth took place after 72 h. In wells not supplemented with collagen gel in substratum, no DRG outgrowth was observed. Partially, DRG differentiation was observed in the presence of NGF. In the absence of NGF and collagen, there was no DRG outgrowth detected. It seems that, in some circumstances, cells degenerated by DRG may be an indication of an apoptosis phenomenon. Therefore, we suggested that collagen as a substratum is more effective than NGF

Betanin purification from red beetroots and evaluation of its anti-oxidant and anti-inflammatory activity on LPS-activated microglial cells.

Microglial activation can release free radicals and various pro-inflammatory cytokines, which implicates the progress of a neurodegenerative disease. Therefore suppression of microglial activation can be an appropriate strategy for combating neurodegenerative diseases. Betanin is a red food dye that acts as free radical scavenger and can be a promising candidate for this purpose. In this study, purification of betanin from red beetroots was carried out by normal phase colum chromatography, yielding 500 mg of betanin from 100 g of red beetroot. The purified betanin was evaluated by TLC, UV-visible, HPLC, ESI-MASS, FT-IR spectroscopy. Investigation on the inhibitory effect of betanin on activated microglia was performed using primary microglial culture. The results showed that betanin significantly inhibited lipopolysaccharide induced microglial function including the production of nitric oxide free radicals, reactive oxygen species, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and interleukin-1 beta (IL-1β). Moreover, betanin modulated mitochondrial membrane potential, lysosomal membrane permeabilization and adenosine triphosphate. We further investigated the interaction of betanin with TNF-α, IL-6 and Nitric oxide synthase (iNOS or NOS2) using in silico molecular docking analysis. The docking results demonstrated that betanin have significant negative binding energy against active sites of TNF-α, IL-6 and iNOS

Detection of Interleukin-19 mRNA in C57BL/6 Mice Astroglial Cells and Brain Cortex

Introduction: Astrocytes are the most abundant glial cell type. In addition to their neurological roles, astrocytes also have immune functions. They have been involved in antigen presentation in the central nervous system (CNS). Activated astrocytes express adhesion molecules, chemokines and release several inflammatory mediators, pro-inflammatory cytokines, neurotrophic and neuroprotective factors, thus these cells have a dual role within the CNS: neuroinflammation and repair processes. IL-19, IL-20, IL-22, IL-24, IL-26, IL-28A, IL-28B, and IL-29 are members of the IL-10 family of cytokines. These cytokines have different biological functions in spite of partial amino acid sequences homology. Signal transduction of the IL-10 family of cytokines is through R1-type and R2-type receptors. Methods: No information has been available about the expression and regulation of IL-19 in mice astrocytes and brain. To investigate the expression of IL-19, we examined its expression in C57BL/6 mice astroglial cells in response to lipopolysaccharide (LPS), using reversetranscription polymerase chain reaction (RT-PCR) method.  Results: We provide for the first time, evidence that astrocytes can express IL-19 mRNA following LPS stimulation. Furthermore, we have found the expression of IL-19 mRNA in the cortex of adult C57BL/6 mice following intraperitoneal (i.p.) administration of LPS.  Discussion: This finding will contribute to current knowledge on the function and behavior of cells and mediators during inflammatory conditions in the brai