Modulation of Immune-Related Gene Expressions in Zebrafish (Danio rerio) by Dietary Purslane (Portulaca oleracea) Extract.

To promote fish's immunity against pathogens in the aquaculture industry, fish dietary fortification with additives or compounds has increasingly attracted attention. In the present study, zebrafish (Danio rerio) was used as an animal model to investigate the effects of purslane, Portulaca oleracea, extract (PE) on the relative expression level of some immune-related genes. A total of 300 zebrafish were randomly divided into four treatment groups and fed for 8 weeks with the basal diets supplemented with 0.5, 1, 1.5, and 2% of PE. The control group was fed with a basal diet without PE. At the end of 8 weeks, the mRNA expression levels of interleukin 1-beta (IL-1ß), interleukin 10 (IL-10), transforming growth factor-beta (TGF-ß), tumor necrosis factor-alpha (TNF-α), superoxide dismutase (SOD), and lysozyme (LYZ) in the fish were evaluated. The results showed that the mRNA expression level of IL-1ß was significantly upregulated in the fish fed with 1 and 2% PE compared to the control group (p < 0.05). Moreover, the evaluation of the mRNA expression level of TGF-ß was significantly increased in a dose-dependent manner in the 1.5 and 2% fed groups compared to the control group (p < 0.05). However, the IL-10 was significantly downregulated in all treated groups compared to the control group (p < 0.05). The expression of the TNF-α gene was not affected amongst all groups by the inclusion of PE in the zebrafish diet (p > 0.05). Based on the results, the diet supplemented with 1.5 and 2% PE significantly upregulated the mRNA expression levels of LYZ and SOD, respectively, compared to the control group (p < 0.05). In conclusion, dietary inclusion of PE may result in beneficial effects on some immune responses via upregulation of some immune genes in zebrafish.

The effects of Sorafenib and Natural killer cell co-injection in combinational treatment of hepatocellular carcinoma; an in vivo approach.

BACKGROUND: Natural killer cells (NKC) and Sorafenib (Sor) are two important agents for the treatment of hepatocellular carcinoma (HCC). Over the past decade, the interaction of Sor and NKC against HCC has been widely challenging. This study aimed to assess the efficacy of NKC & Sor for the treatment of HCC in vivo. METHODS: Subcutaneous xenograft models of HCC were established in nude mice. For safety assessment of treatment, the kidney and liver functions were analyzed. Paraffin embedded tumor sections were histopathologically studied and immunohistochemistry (IHC) tests were done to evaluate the angiogenesis (CD34) and proliferation (Ki67) indexes. The terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay was performed to identify the tumor cells undergoing apoptosis. The serum levels of tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ) were measured by enzyme-linked immunosorbent assay (ELISA) and expression levels of major inflammatory cytokines and cytoplasmic granules in xenograft HCC were quantified using real-time PCR. RESULTS: NKC & Sor significantly inhibited necrosis and apoptosis in tumor cells and increased angiogenesis and proliferation of HCC compared to the monotherapy of NKC or Sor alone. The serum levels of TNF-α, IFN-γ as well as the expression levels of TNF-α, IFN-γ, interleukins (ILs)-1, 6, 10, granzyme-B and perforin in the xenograft HCC tissues of the treated mice with NKC & Sor were significantly lower than those of treated with NKC or Sor alone. CONCLUSION: Therapy with the specific dosage of NKC & Sor could not inhibit the HCC xenograft growth rate through a synergistic effect in a mouse model of HCC.

The Role of Forced and Voluntary Training on Accumulation of Neural Cell Adhesion Molecule and Polysialic Acid in Muscle of Mice with Experimental Autoimmune Encephalomyelitis.

It has been suggested that depletion of adhesion molecules is one of the factors associated with or possibly responsible for multiple sclerosis (MS) progression. The aim of this study was to investigate the effect of forced and voluntary training before and after induction of experimental autoimmune encephalomyelitis (EAE) on accumulation of neural cell adhesion molecule (NCAM) and polysialic acid (PSA) in neuromuscular junction denervation in plantaris and soleus muscles in C57BL/6 female mice. A total of 40 female C57BL/6 mice, 10-week-old, were randomly divided into four groups, including induced control groups without EAE induction, induced EAE without training, and forced and voluntary training groups. Myelin oligodendrocyte glycoprotein peptide 35-55 (300 µg in saline; MOG 35-55; KJ Ross-Petersen ApS, Denmark) was injected subcutaneously at the base of the tail of each mouse. Clinical assessment of EAE was performed daily using a 15-point scoring system following immunization. Training groups performed the swimming program for 30 min/day, 5 times/week, for 4 weeks. Mice had access to a treadmill for one hour per day, 5times/week, for 4 weeks in individual cage. The mice were scarified, and the plantaris and soleus muscles were then isolated for investigation of proteins expression using IHC. An analysis of the preventive exercise (before) and recovery exercise (after) of the EAE was performed. Images of the stained sections were taken using a fluorescent microscope. Quantitative image analysis was performed using ImageJ software package. The obtained data from the mean percentage expression of PSA and NCAM in pre- and post-soleus and plantaris muscles showed that the highest and lowest expression levels of PSA and NCAM belonged to control and swim EAE (SE) groups, respectively. The low expression levels of PSA and NCAM were detected in rat with MS without intervention. In conclusion, the relationship between increasing levels of NCAM and PSA protein expression and voluntary and compulsory activity were detectable both in pre and post-soleus and plantaris. However, voluntary activity resulted in more expression levels of NCAM and PSA than that of compulsory. In conclusion, since it has been suggested that depletion of NCAM is one of the factors associated with or possibly responsible for MS progression, these findings show exercise MS progression may be reduced by increasing expression of exercise-related adhesion molecule such as NCAM and PSA (a glycan modification of the NCAM).

Enhancing the Therapeutic Efficacy of Daunorubicin and Mitoxantrone with Bavachinin, Candidone, and Tephrosin.

The capability of flavonoids in sensitizing cancer cells was demonstrated in numerous works to chemotherapy and converse multidrug resistance by modulating efflux pumps and apoptosis mechanisms. Three flavonoids, namely, bavachinin, tephrosin, and candidone, have been recently introduced to cancer treatment research presenting various activities, such as antibacterial, immunomodulatory, cell death, and anticancer. Less information exists regarding the therapeutic significance of these flavonoids in cancer treatment, especially in overcoming multidrug resistance (MDR). Here, we tempted to investigate the potency of these agents in reversing MDR by analyzing their effects as chemosensitizers on cell cytotoxicity, P-gp and ABCG2 protein expression levels, and their function on two multidrug-resistant cell lines, P-gp-overexpressing human gastric adenocarcinoma cell line (EPG85.257RDB) and ABCG2-overexpressing human epithelial breast cancer cell line (MCF7/MX). The inhibitory concentration of 10% (IC10) of bavachinin, tephrosin, and candidone in EPG85.257RDB cells was 1588.7 ± 202.2, 264.8 ± 86.15, and 1338.6 ± 114.11 nM, respectively. Moreover, these values in MCF7/MX cell were 2406.4 ± 257.63, 38.8 ± 4.28, and 27.9 ± 5.59 nM, respectively. Expression levels of ABCG2 and P-gp were not significantly downregulated by these flavonoids. Maximum levels of daunorubicin and mitoxantrone accumulations and minimum rates of drug efflux in both cell lines were detected 48 hrs posttreatment with tephrosin and bavachinin, respectively. Chemosensitization to mitoxantrone and daunorubicin treatments was, respectively, achieved in MCF7/MX and EPG85.257RDB cells in response to IC10 of bavachinin and tephrosin, independently. These effects did not follow time-dependent manner, and each flavonoid had its cell-dependent patterns. Overall, bavachinin, tephrosin, and candidone showed potency to sensitize MDR cells to daunorubicin and mitoxantrone and could be considered as attractive MDR modulators for cancer treatment. However, their action was time and cell specific.

Effect of Selenium Nanoparticle Supplementation on Tissue Inflammation, Blood Cell Count, and IGF-1 Levels in Spinal Cord Injury-Induced Rats.

Selenium is known to be a neuroprotective agent in respect to a number of neuronal diseases and pain. The aim of this study was to evaluate the neuroprotective effect of the oral administration of selenium nanoparticles in rats with spinal cord injury (SCI). Forty adult female rats were randomly assigned to two equal groups as experimental and control. Under general inhalation anesthesia, in both groups, SCI was created, at the T9-10 level of the column. On the third day after the operation, a supplement of selenium nanoparticle was administered to the experimental group at 0.2 mg/kg per day. The histology of the site of injury, IGF-1 serum concentrations, and changes in the white blood cells were examined in both groups at different pre-surgical and post-surgical times. The results of the current study showed a significant decrease in the total white blood cells, including lymphocyte, neutrophil, and monocyte in the experimental group compared to the control group. Histological evaluation showed that the inflammatory responses reduced significantly in the experimental group compared to the control group. In conclusion, we speculate that the decrease in the number of inflammatory cells after oral administration of the selenium nanoparticles is due to the neuroprotective effects of this nanoparticle.

Enhancement of therapeutic DNA vaccine potency by melatonin through inhibiting VEGF expression and induction of antitumor immunity mediated by CD8+ T cells.

To be effective, therapeutic cancer vaccines should stimulate both an effective cell-mediated and a robust cytotoxic CD8+ T-cell response against human papillomavirus (HPV)-infected cells to treat the pre-existing tumors and prevent potential future tumors. In this study, the therapeutic experiments were designed in order to evaluate antitumor effect against the syngeneic TC-1 tumor model. The anti-tumor efficacy of a HPV-16 E7 DNA vaccine adjuvanted with melatonin (MLT) was evaluated in a C57BL/6 mouse tumor model by measuring tumor growth post vaccination and the survival rate of tumor-bearing mice, analyzing the specific lymphocyte proliferation responses in control and vaccinated mice by MTT assay. The E7-specific cytotoxic T cells (CTL) were analyzed by lymphocyte proliferation and lactate dehydrogenates (LDH) release assays. IFN-γ, IL-4 and TNF-α secretion in splenocyte cultures as well as vascular endothelial growth factor (VEGF) and IL-10 in the tumor microenvironment were assayed by ELISA. Our results demonstrated that subcutaneous administration of C57BL/6 mice with a DNA vaccine adjuvanted with MLT dose-dependently and significantly induced strong HPV16 E7-specific CD8+ cytotoxicity and IFN-γ and TNF-α responses capable of reducing HPV-16 E7-expressing tumor volume. A significantly higher level of E7-specific T-cell proliferation was also found in the adjuvanted vaccine group. Furthermore, tumor growth was significantly inhibited when the DNA vaccine was combined with MLT and the survival time of TC-1 tumor bearing mice was also significantly prolonged. In vivo studies further demonstrated that MLT decreased the accumulation of IL-10 and VEGF in the tumor microenvironment of vaccinated mice. These data indicate that melatonin as an adjuvant augmented the cancer vaccine efficiency against HPV-associated tumors in a dose dependent manner.