Cell Therapy and Investigation of the Angiogenesis of Fibroblasts with Collagen Hydrogel on the Healing of Diabetic Wounds.

Objectives: A diabetic ulcer is a common disease in patients with diabetes. Because of antibiotic resistance, new therapeutic alternatives are being considered in diabetic foot patients to reduce complications and mortality. This study aimed to evaluate the effect of collagen hydrogel on the wound-healing process in diabetic rats. Materials and Methods: Diabetic wounds were induced with streptozotocin in all 42 male Wistar rats. The rats were divided into four groups: (a) treated with fibroblast cells, (b) collagen hydrogel, (c) collagen cultured with fibroblast cells, and (d) a control group. Microscopic and histological (hematoxylin and eosin staining and Mason trichrome staining), measurement of wound surface with image J, skin density and thickness by the ultrasound probe, and skin elasticity with cytometer tool were used to evaluate wound healing at days 14 and 21 after the treatment. Results: The results showed that treating diabetic wounds with fibroblasts cultured in collagen hydrogel greatly reduces inflammatory responses in the skin tissue and significantly accelerates the healing process. In addition, 21 days after the start of treatment, skin elasticity, thickness, and density were higher in the collagen + fibroblast group than in the control group. Conclusion: In addition, the results of the present study show that diabetic wound dressing can significantly reduce the inflammatory phase in the wound healing process by increasing the speed of collagen synthesis, skin density and elasticity, and angiogenesis.

Expression and characterization of hemagglutinin-neuraminidase protein from Newcastle disease virus in Bacillus subtilis WB800.

BACKGROUND: Newcastle disease virus (NDV) belongs to the genus Avaluvirus and Paramyxoviridae family, and it can cause acute, highly contagious Newcastle disease in poultry. The two proteins, haemagglutinin neuraminidase (HN) and Fusion (F), are the main virulence factor of the virus and play an essential role in immunogenicity against the virus. In most paramyxoviruses, the F protein requires HN protein to fuse the membrane, and HN proteins substantially enhance the viruses' fusion activity. RESULTS: The present study describes the successful cloning and expression of HN protein from NDV in Bacillus subtilis WB800 using the modified shuttle vector pHT43. HN coding sequence was cloned into the pGet II vector. It was then subcloned into the PHT43 shuttle vector and transferred to Escherichia coli for replication. The recombinant plasmid was extracted from E. coli and used to transform B. subtilis by electroporation. After induction of recombinant B. subtilis by IPTG, total cell protein and the protein secreted into the media were analysed through a time course using SDS-PAGE. The expressed HN protein was purified using cation exchange chromatography followed by metal affinity chromatography, using the 6× His epitope introduced at the carboxyl terminus of the recombinant protein. The accuracy of the PHT43-HN construct was confirmed by sequencing and enzymatic digestion. SDS-PAGE results showed that the recombinant HN protein was successfully expressed and secreted into the medium. Moreover, the purified HN protein showed neuraminidase activity with characteristics similar to the indigenous HN NDV protein. B. subtilis is a free endotoxin host that could be a favourite prokaryotic platform for producing the recombinant HN protein. CONCLUSION: The establishment of this expression and purification system has allowed us to explore further the biochemical characteristics of HN protein and obtain material that could be suitable for a new production of NDV candidate vaccine with high immunogenicity.

The effect of opine on matrix metalloproteinase expression in mice with breast cancer.

Regarding the anti-inflammatory and anti-tumour effects of arginine and its derivatives, this study evaluates matrix metalloproteinase (MMPs) expression in an animal model of breast cancer following administration of octopine. In this study, 40 animals of Balb/C mice were divided into 5 groups: the healthy control, the cancer control, the cancer group receiving 50 mg of octopine, the cancer group receiving 100 mg of octopine and the cancer group receiving 150 mg of octopine for 3 weeks. 4T1 cell line was used to induce cancer. Biopsy specimens were enrolled from mice and MMP-1, MMP-3 and MMP-9 gene expression evaluated using real-time PCR, while these protein amounts were measured using immunohistochemistry and ELISA methods. Data were analysed using one-way ANOVA, Kruskal-Wallis and Mann-Whitney U tests (p < .05). The results showed that 100 mg octopine consumption had significant decreasing effect on MMP-9 expression (p = .02) in the treatment group compared with cancerous non-treated mice. Furthermore, results from immunohistochemistry and ELISA confirmed this effect, the protein amount of MMP-9 was significantly decreased in group treating with 100 mg octopine (.005). The use of octopine has a beneficial effect on reducing MMP-9 in mice breast cancer.

TLR2 and TLR4 Signaling Pathways and Gastric Cancer: Insights from Transcriptomics and Sample Validation

Background: Pattern recognition receptors, especially toll-like receptors (TLRs), as the first line of defense for pathogen detection, were found to be associated with H.¬ pylori infection and gastric cancer (GC). However, the expression levels of TLRs, i.e. TLR2 and TLR4, as the main receptors sensed by H.¬ pylori, still remain largely ambiguous. We aimed to investigate the patterns of key transcripts of TLR2 and TLR4 in 100 GC transcriptome data. Additionally, we evaluated TLR2 and TLR4 gene expressions in gastric biopsies of Iranian GC patients, in order to validate RNA-seq outputs. Methods: For this study, 100 runs of GC samples and controls were processed and analyzed using map read to reference. Differential gene expression method was used to distinguish between GC and normal samples in the expression of TLRs and other innate immune molecules. Also, using qRT-PCR assay, transcripts of TLRs molecules for 15 GC and 15 control samples were analyzed based on the analysis of variance and least significant differences. Results: The results clearly showed that all signaling pathways molecules of TLR4, especially TLR4 (p = 0.019), NF-κB (p ¬= 0.047), IL-1ß (p = 0.0096), and TNF-α (p = 0.048), were upregulated in a cancerous condition in different parts and at various stages of GC. Conclusion: Our findings suggested that molecules involved in inflammation, including TLR4 and its related pro-inflammatory cytokines, may be responsible for the development and progression of GC. Accordingly, the control of H. pylori infection reduces inflammation in the gastric system and can play an important role in preventing gastrointestinal disorders.

Biological Characteristics and Optical Reflectance Spectroscopy of Human Placenta Derived Mesenchymal Stem Cells for Application in Regenerative Medicine.

Introduction: The efficiency of stem cell isolation, culture, and biological characterization techniques for treatment is facing serious challenges. The purpose of this study was to provide a protocol for isolation and culture of three types of mesenchymal stem cells (MSCs) derived from the human placenta, amniotic membrane, and umbilical cord with high efficiency used for cell therapy. Methods: During this experimental laboratory study, 10 complete placenta samples were prepared from cesarean section mothers. The protocol for isolation and culture of mesenchymal cells from the placenta tissue, umbilical cord, and amniotic membrane was enzymatically optimized. The morphological features of mesenchymal cells were investigated using an inverted microscope and their biological features were measured using flow cytometry. The differentiation potential of the cells was evaluated by measuring their differentiation capacity into osteocytes and adipocytes. The absorption and reflectance features of the cells were recorded by optical spectroscopy. Finally, the data were statistically analyzed. Results: The expression of CD44, CD73, CD90 and CD29 markers in human placenta tissue-derived cells was significant. CD14, CD34 and CD45 markers were not expressed or were slightly expressed. These cells were highly viable and successfully differentiated into osteocytes and adipocytes. MSCs absorbed more light than visible light by showing light absorption peaks at wavelengths of about 435 and 550 nm. Conclusion: The protocol used in this study for isolation and culture of human placenta tissue-derived MSCs had significant efficiency for the production of MSCs for use in cell therapy and tissue engineering.

Supplementation of freezing media with alpha lipoic acid preserves the structural and functional characteristics of sperm against cryodamage in infertile men with asthenoteratozoospermia.

The aim of this study was to examine the effects of alpha lipoic acid (ALA) supplementation during semen cryopreservation on the sperm quality, chromatin integrity, oxidative stress, and expression level of BAX, BCL2, HSP70 and iNOS genes in semen samples obtained from infertile men with asthenoteratozoospermia. METHODS: Twenty freshly ejaculated semen samples were cryopreserved with sperm freezing medium supplemented with 0.00, 0.02, 0.05, 0.1, 0.5, and 1 mmol/mL of ALA. The samples were analyzed according to the WHO guidelines before and after freezing. Sperm ROS production level, DNA fragmentation and cryo-capacitation were assessed using flow cytometry, TUNEL assay and chlortetracycline (CTC) test, respectively. Expression level of stress protein (HSP70), pro-apoptotic Bax, anti-apoptotic Bcl-2, and iNOS genes was assessed by real-time PCR assay. RESULTS: The effective concentrations of ALA (0.02 and 0.5 mM) significantly improved the motility, viability and morphology of the frozen-thawed sperms compared to the control group treated with 0.00 mM of ALA. During cryopreservation, treatment of semen with 0.02 mM of ALA, as the optimal concentration, significantly decreased DNA fragmentation and oxidative stress level (P < 0.05), protected the acrosome integrity, and led to insignificant reduction in BAX gene expression level and significant increase in expression level of BCL2, HSP70, and iNOS genes compared with control group. CONCLUSION: Our findings revealed that the adding ALA to semen samples obtained from infertile men with asthenoteratozoospermia plays a significant protective role against cryodamage by preserving the sperm functional parameters.

Effect of Vitex Agnus-Castus Ethanolic Extract on Sex Hormones in Streptozotocin-Induced Diabetic Rats.

Objective: Diabetes mellitus is recognized as one of the serious global health problems. There are evidences regarding the high prevalence of sexual dysfunction in diabetic patients. Experimental studies revealed a positive effect of Vitex agnus-castus (Vitex), on sexual function and behaviors. In this research, the effect of Vitex on sexual hormones in streptozotocin-(STZ) induced diabetic rats was investigated. Materials and methods: A Thirty adult female Wistar rats were divided into five groups. 1-control group (non-diabetic), 2- diabetic group (received normal saline) and three induced diabetic groups treated with different doses (400, 200 and 100 mg/kg) of Vitex. Treatment groups received Vitex fruit extract by gavage for 7 days. The levels of luteinizing hormone (LH), follicle-stimulating hormone (FSH), progesterone and estrogen in serum were measured. Results: Levels of LH, FSH, estrogen and progesterone and average body weight was lower in diabetic group compared to control group (p <0.010). Animals received high dose of Vitex fruit extract (400mg/kg) had significantly higher levels of serum LH, FSH, estrogen and progesterone compared to diabetic group (p < 0.010). In animals receiving minimum dose (100mg/kg) of Vitex, no difference was observed compared to diabetic group (p > 0.010). Conclusion: It can be concluded that Vitex fruit extract probably has regulatory effect on diabetes-induced change in the levels of sex hormones in female rats. Vitex fruit extract can improve serum levels of sex hormones in an animal model of STZ-induced diabetes.

Effects of phosphodieastrase type 5 inhibitions on morphine withdrawal symptoms in mice / Efectos comportamentales por inhibición de la fosfodiesterasa tipo 5 sobre los síntomas de abstinencia de morfina en ratones

BACKGROUND: Chronic morphine exposure creates dependence and, upon cessation, withdrawal symptoms. Studies indicate the phosphodiesterase type 5 (PDE5) inhibitor sildenafil may provide centrally mediated benefits against withdrawal, and therefore, this study evaluated morphine withdrawal signs in dependent mice with and without sildenafil treatment. METHOD: Dependence was induced by repeated treatments with morphine over 5 consecutive days. The morphine-dependent mice received sildenafil (1, 5, 10, or 20 mg/kg, i.p.) 15 min prior to the precipitation of morphine withdrawal. On the last day, naloxone was injected 2 hours after the last morphine injection, and withdrawal signs were evaluated for 30 min after naloxone injection. RESULTS: The administration of sildenafil reduced all of the morphine withdrawal symptoms. CONCLUSIONS: The administration of sildenafil diminished morphine withdrawal signs in morphine-dependent mice. We hypothesize that the mechanism involves enhanced cyclic guanosine monophosphate (cGMP) activity, but further studies are recommended for a better understanding

ANTECEDENTES: se sabe que la exposición crónica a la morfina conduce a la dependencia. El cese en el consumo de morfina conlleva al desarrollo del síndrome de abstinencia. Numerosos estudios indican los efectos beneficiosos del sildenafil en el sistema nervioso central. Es por ello que el presente estudio evaluó el efecto del sildenafil sobre el síndrome de abstinencia a la morfina. MÉTODO: los ratones dependientes de morfina recibieron dosis diferentes de sildenafil (1, 5, 10 o 20 mg/kg i.p.) 15 minutos antes de producir el síndrome de abstinencia a la morfina. En el último día, la naloxona se inyectó dos horas después de la última inyección de morfina. Los signos de abstinencia fueron evaluados durante 30 minutos después de la inyección de naloxona. RESULTADOS: la administración de sildenafil redujo todos los síntomas de abstinencia relacionados con la morfina. CONCLUSIONES: se puede concluir que la administración de sildenafil disminuyó los signos de abstinencia a la morfina. Estos resultados muestran cómo los incrementos en el nivel del GMP cíclico mediante la vía PDE5 reduce los síntomas de abstinencia a la morfina

Effects of phosphodieastrase type 5 inhibitions on morphine withdrawal symptoms in mice.

BACKGROUND: Chronic morphine exposure creates dependence and, upon cessation, withdrawal symptoms. Studies indicate the phosphodiesterase type 5 (PDE5) inhibitor sildenafil may provide centrally mediated benefits against withdrawal, and therefore, this study evaluated morphine withdrawal signs in dependent mice with and without sildenafil treatment. METHOD: Dependence was induced by repeated treatments with morphine over 5 consecutive days. The morphine-dependent mice received sildenafil (1, 5, 10, or 20 mg/kg, i.p.) 15 min prior to the precipitation of morphine withdrawal. On the last day, naloxone was injected 2 hours after the last morphine injection, and withdrawal signs were evaluated for 30 min after naloxone injection. RESULTS: The administration of sildenafil reduced all of the morphine withdrawal symptoms. CONCLUSIONS: The administration of sildenafil diminished morphine withdrawal signs in morphine-dependent mice. We hypothesize that the mechanism involves enhanced cyclic guanosine monophosphate (cGMP) activity, but further studies are recommended for a better understanding.

Effect of Prunus domestica L. (mirabelle) on learning and memory in mice.

BACKGROUND: Plums have been known to have various pharmacological activities. AIMS: The aim of the present study was to evaluate the effect of administration of hydro-alcoholic extract of plum, using passive avoidance task. SETTINGS AND DESIGN: Mice were divided into four groups (n = 7 each) one control and three plum (75, 100, 150 mg/kg) groups. MATERIALS AND METHODS: The control group received saline and plum groups received the extract by oral gavage for 7 days. The number of trials to acquisition, step through latency (STLr) in the retention test and the time spent in the dark compartment (TDC) during the retention test were measured. STATISTICAL ANALYSIS: Differences between groups were tested by one-way ANOVA with Tukey post-hoc test. RESULTS: A significant difference was found in the number of trials to acquisition between the groups. The results also indicated in the retention test, administration of 75 and 100 mg/kg plum caused an increased STLr (compared with the untreated control group). The results also showed that the total time spent in TDC by the animals of the extract groups was lower than that of the control group. CONCLUSIONS: Hydro-alcoholic extract of plum has a beneficial effect on learning and memory in passive avoidance task. It can be concluded that its antioxidant and antidyslipidemic activities may be involved in the obtained effects.

Involvement of the ventral tegmental area in the inhibitory avoidance memory in rats.

The ventral tegmental area (VTA) is a neural structure that sends strong efferent projections to the hippocampus. Output from the VTA can affect hippocampal-dependent neural processes that are critical for learning and memory, including long term potentiation and theta activity. However, no study to date has elucidated what role the VTA plays in mediating the different stages of learning and memory. Therefore, the current study was designed to assess how reversible inactivation of the VTA may affect the acquisition, consolidation and retrieval of memory in rats using an inhibitory avoidance (IA) task. In this experiment, rats with chronically implanted cannulae aimed at the VTA were trained on an inhibitory avoidance task. They received intra-VTA infusions of lidocaine or saline immediately before training, after training or before a memory retention test. The results indicate that inactivation of the VTA prior to the first acquisition session increased the number of trials rats required to reach the acquisition criterion. Similarly, inactivation of the VTA after acquisition training decreased the step-through latency and increased the amount of time spent in the dark compartment relative to the saline-treated group. However, inactivation of the VTA immediately prior to the memory retention test failed to alter either step-through latency or the amount of time spent in the dark compartment. Overall, these results suggest the VTA facilitates the acquisition and consolidation of IA learning and memory.

Economic appraisal of urine opiates screening test: a study in kerman, iran.

BACKGROUND: Cost effectiveness, the ratio of relative costs of a program to its desired outcomes, is one of the basic issues in various screening programs performed to detect opium abuse. This study aimed to find the cost-effectiveness of opiates abuse screening through urine analysis. METHODS: A total number of 64698 individuals were selected and divided into to five distinct groups based on the reason for which they were tested. Cost-effectiveness of opiates abuse screening in each group was calculated by dividing the total cost, including personnel and overhead costs, to the number of detected cases. Finally, the results were compared. FINDINGS: The total number of positive cases based on rapid screening assay (RSA) and thin layer chromatography (TLC) was 3460 (5.3%). According to incremental cost-effectiveness analysis, screening program of the group referred by the police was the most cost-effective program with the breakeven point at 2%. CONCLUSION: According to the obtained results, continuation of drug abuse screening programs is recommended.

Ascorbic acid supplementation could affect passive avoidance learning and memory in rat.

Ascorbic acid (vitamin C) is required for health and, in particular, its supplementation has beneficial effects in some pathological conditions. There are conflicting reports regarding the usefulness of ascorbic acid in the treatment of dementia. In this study, we investigated the effects of acute, short- and long-term pre-training administration of ascorbic acid (60,120 mg/kg) on passive avoidance learning (PAL) and memory in rats. Retention test was done 24h after training. The results showed that acute injection of ascorbic acid had no significant effect on PAL. On the other hand, both in the short- and long-term ascorbic acid treated groups trials to acquisition were less than control group. Also, ascorbic acid prolonged the step-through latency (STL) and decreased the time spent in the dark compartment in retention test. Thus, it can be concluded that short- and long-term supplementation with ascorbic acid has facilitatory effects on acquisition and retrieval processes of passive avoidance learning and memory in rats.

The role of GABAergic transmission in the dentate gyrus on acquisition, consolidation and retrieval of an inhibitory avoidance learning and memory task in the rat.

The hippocampal GABAergic interneurons are responsible for controlling the input of large principal cell populations, and they thereby determine the oscillatory discharge patterns and synaptic plasticity in the hippocampus. Such oscillations within neuronal systems serve various complex functions, such as perception, cognition, plasticity and memory. The aim of this study is to define the function of GABAergic synaptic transmission in the dentate gyrus (DG) of the hippocampus in the different stages of inhibitory avoidance (IA) learning and memory in the rat. Two cannulae were implanted above the hippocampal DG. Then the rats were trained on a step-through IA learning task. Each rat received intra-DG injection of picrotoxin (PTX) or saline before training, after training or before the retrieval test. The results show that post-training injection of PTX impaired the IA memory. On the other hand, pre-training and pre-retrieval injection of PTX had no significant effect on the IA activity. Therefore, it seems that GABAergic transmission in the DG is involved in the consolidation step (but not in the acquisition and retrieval steps) of the IA task by controlling the input to the principal cells.