Leishmania infantum: prime boost vaccination with C-terminal extension of cysteine proteinase type I displays both type 1 and 2 immune signatures in BALB/c mice

The aims of current study are to describe the immunogenicity and protective efficacy of prime boost vaccine using C-terminal extension (CTE) of cysteine proteinase type I of Leishmania infantum in BALB/c mice. Group I as vaccinated group primed with 100 microg of pcDNA-CTE and 3 weeks later boosted with combination of 30 microg rCTE, 50 microg of CpG and Montanide 720. Groups II and III were served as control groups. Although, this vaccination regimen did not protect mice against the infectious challenge but it was highly immunogenic. IgG2a has been raised strongly against rCTE in contrast to IgG1 and remains high at every time point under study. By analysis of CTE synthetic peptides (CTE100) before challenge, both IgG1 and IgG2a were produced and for all overlapping synthetic peptides (CTE 1-8) IgG1 raised significantly. This statue is changed at 7 weeks after challenge and only CTE2 and CTE3 have shown to induce considerable amount of IgG1. In all groups, the level of IL-5 started to increase with high concentration shortly passing only 3 weeks after infectious challenge. In compare with two control groups, the vaccinated group produced significantly higher level of IL-5 at 7 weeks post-infection. The parasite burden of all groups is similar at 4 weeks post-challenge in both liver and spleen. In contrast, at 8 weeks post challenge, the spleen of the vaccinated group showed significantly higher level of parasite load in compare with two control groups. This study demonstrated that immunization with CTE display both type 1 and 2 immune signatures in experimental murine model of L. infantum infection.     

Is urine a reliable clinical sample for the diagnosis of human visceral leishmaniasis? A systematic review and meta-analysis

Visualization of amastigotes in lymph nodes, bone marrow, and other tissues samples remains the gold standard method for the diagnosis of visceral leishmaniasis (VL) in humans. This gold standard diagnostic method uses a technically challenging microscopy procedure that is often not accessible in many places in the world where VL is endemic. Here, we report the current systematic review and meta-analysis to evaluate whether urine is a reliable clinical sample for diagnosis of human VL. Data were extracted from ten available databases during the period from 2002 to 2017. Overall, 29 articles fulfilled the inclusion criteria and were used for data extraction in this systematic review. Most studies (72.4%) using urine specimens were reported from five countries: India 6 (20.7%), Iran 5 (17.2%), Bangladesh 4 (13.8%), Japan 3 (10.3%) and Spain 3 (10.3%), respectively. The most common diagnostic tests performed on urine were Katex (62.1%), ELISA (24.1%), and the rK39 (17.2%) assays. In meta-analysis the sensitivity and specificity of the three most commonly used diagnostic assays were rK39 (97%; CI: 91–99; 98%;76–100), ELISA (91%; 82–95; 99%; CI: 94–100), and Katex (83%; 73–90; 98%; 98–100), suggesting that the rK39 assay provided the highest sensitivity and the ELISA assay provided the highest specificity for diagnosis of VL from urine samples. Our findings suggest that urine is a valuable clinical sample for the diagnosis of human VL, particularly in areas where the gold standard test for VL is not available.     

Effects of Self‐Conditioning Techniques (Self‐Hypnosis) in Promoting Weight Loss in Patients with Severe Obesity: A Randomized Controlled Trial

Objective

The usefulness of the rapid‐induction techniques of hypnosis as an adjunctive weight‐loss treatment has not been defined. This randomized controlled trial evaluated whether self‐conditioning techniques (self‐hypnosis) added to lifestyle interventions contributed to weight loss (primary outcome), changes in metabolic and inflammatory variables, and quality of life (QoL) improvement (secondary outcomes) in severe obesity.

Methods

Individuals (with BMI = 35‐50 kg/m²) without organic or psychiatric comorbidity were randomly assigned to the intervention (n = 60) or control arm (n = 60). All received exercise and behavioral recommendations and individualized diets. The intervention consisted of three hypnosis sessions, during which self‐hypnosis was taught to increase self‐control before eating. Diet, exercise, satiety, QoL, anthropometric measurements, and blood variables were collected and measured at enrollment and at 1 year (trial end).

Results

A similar weight loss was observed in the intervention (?6.5 kg) and control (?5.6 kg) arms (β = ?0.45; 95% CI: ?3.78 to 2.88; P = 0.79). However, habitual hypnosis users lost more weight (?9.6 kg; β = ?10.2; 95% CI: ?14.2 to ?6.18; P < 0.001) and greatly reduced their caloric intake (?682.5 kcal; β = ?643.6; 95% CI: ?1064.0 to ?223.2; P = 0.005) in linear regression models. At trial end, the intervention arm showed lower C‐reactive protein values (β = ?2.55; 95% CI: ?3.80 to ?1.31; P < 0.001), higher satiety (β = 19.2; 95% CI: 7.71‐30.6; P = 0.001), and better QoL (β = 0.09; 95% CI: 0.02‐0.16; P = 0.01).

Conclusions

Self‐hypnosis was not associated with differences in weight change but was associated with improved satiety, QoL, and inflammation. Indeed, habitual hypnosis users showed a greater weight loss.

     

The anticancer impacts of free and liposomal caffeic acid phenethyl ester (CAPE) on melanoma cell line (A375)

The deadliest type of skin cancer, malignant melanoma, is also the reason for the majority of skin cancer-related deaths. The objective of this article was to investigate the efficiency of free caffeic acid phenethyl ester (CAPE) and liposomal CAPE in inducing apoptosis in melanoma cells (A375) in in vitro. CAPE was loaded into liposomes made up of hydrogenated soybean phosphatidylcholine, cholesterol, and 1,2-distearoyl-sn-glycero-3 phosphoethanolamine-N-[methoxy (polyethylene glycol)-2000], and their physicochemical properties were assessed. (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test was performed for comparing the cytotoxicity of free CAPE and liposomal CAPE at dosages of 10, 15, 25, 50, 75 and the highest dose of 100 μg/mL for period of 24 and 48 h on A375 cell line to calculate IC50. Apoptosis and necrosis were evaluated in A375 melanoma cancer cells using flow cytometry. Atomic force microscopy was utilized to determine the nanomechanical attributes of the membrane structure of A375 cells. To determine whether there were any effects on apoptosis, the expression of PI3K/AKT1 and BAX/BCL2 genes was analyzed using the real-time polymerase chain reaction technique. According to our results, the maximum amount of drug release from nanoliposomes was determined to be 91% and the encapsulation efficiency of CAPE in liposomes was 85.24%. Also, the release of free CAPE was assessed to be 97%. Compared with liposomal CAPE, free CAPE showed a greater effect on reducing the cancer cell survival after 24 and 48 h. Therefore, IC50 values of A375 cells treated with free and liposomal CAPE were calculated as 47.34 and 63.39 μg/mL for 24 h. After 48 h of incubation of A375 cells with free and liposomal CAPE, IC50 values were determined as 30.55 and 44.83 μg/mL, respectively. The flow cytometry analysis revealed that the apoptosis induced in A375 cancer cells was greater when treated with free CAPE than when treated with liposomal CAPE. The highest nanomechanical changes in the amount of cell adhesion forces, and elastic modulus value were seen in free CAPE. Subsequently, the greatest decrease in PI3K/AKT1 gene expression ratio occurred in free CAPE.     

Chitosan magnetic nanoparticles for drug delivery systems

The potential of magnetic nanoparticles (MNPs) in drug delivery systems (DDSs) is mainly related to its magnetic core and surface coating. These coatings can eliminate or minimize their aggregation under physiological conditions. Also, they can provide functional groups for bioconjugation to anticancer drugs and/or targeted ligands. Chitosan, as a derivative of chitin, is an attractive natural biopolymer from renewable resources with the presence of reactive amino and hydroxyl functional groups in its structure. Chitosan nanoparticles (NPs), due to their huge surface to volume ratio as compared to the chitosan in its bulk form, have outstanding physico-chemical, antimicrobial and biological properties. These unique properties make chitosan NPs a promising biopolymer for the application of DDSs. In this review, the current state and challenges for the application magnetic chitosan NPs in drug delivery systems were investigated. The present review also revisits the limitations and commercial impediments to provide insight for future works.     

Cold-induced physiological and biochemical responses of three grapevine cultivars differing in cold tolerance

In this study the cold tolerance potential of three Vitis vinifera cultivars including ‘Red Sultana’, ‘White Sultana,’ and ‘Flame Seedless’ was evaluated under greenhouse condition. After 15 leaves stage in average, the grapevine plants were subjected to cold stress regimes (4, 0 and ? 4 °C) and compared with control plants (24 °C). A clear increase in leaf electrolyte leakage (EL), thiobarbituric acid reactive substances (TBARS), and H₂O₂ concentrations was observed with decreasing temperature from 4 to ? 4 °C in all grapevine cultivars. Chilled plants showed marked increases in their abscisic acid (ABA), soluble sugars, and proline contents in compared to control vines. Upon exposure to cold stress, the EL, TBARS, H₂O₂, and relative water content of ‘Red Sultana’ were found to be lower compared to ‘White Sultana’ and ‘Flame Seedless’. Under 0 °C condition, ‘Red Sultana’ had the highest superoxide dismutase, guaiacol peroxidase and catalase activities, which was approximately twofold higher than those of all other cultivars. Soluble sugars such as glucose, fructose, and sucrose increased from 4 to ? 4 °C. These increments were higher in ‘Red Sultana’ compared to other cultivars which was concomitant with higher accumulation of endogenous ABA concentration in this cultivar. Higher accumulation of ABA and soluble sugars in ‘Red Sultana’ confirmed the key roles of these compounds in cold tolerance which could be applied as a cold tolerance marker for early selection of grapevine cultivars with the aim to establish vineyards in cold winter regions.     

Delivery of HIV-1 Nef Protein in Mammalian Cells Using Cell Penetrating Peptides as a Candidate Therapeutic Vaccine

The HIV-1 Nef protein expressed early in viral life cycle has been known as a potent candidate for therapeutic vaccine development. Due to different cell barriers, various cell penetrating peptides (CPPs) such as Pep-1 and CADY-2 have been known to deliver biologically active proteins to cytoplasmic compartments via the plasma membrane. In current study, we firstly evaluated the efficiency of lentiviral vector (pCDH-CMV-MCS-EF1-cGFP-T2A-puro) and eukaryotic expression vector (pEGFP-N1) for expression of HIV-1 Nef protein in HEK-293T cells using TurboFect transfection reagent. Our results showed that both vectors can effectively express the Nef proteins within the target cell. The pEGFP-N1 was more effective than pCDH-GFP for protein expression. Furthermore, Nef protein was expressed in E. coli as GST-Nef fusion and transfected by the amphipathic CPPs including Pep-1 and CADY-2 into HEK-293T cells. The size and morphology of the GST-Nef/CPP complexes were evaluated by scanning electron microscopy, and Zetasizer. Our data indicated that the recombinant GST-Nef protein generated in BL21 strain migrated as a clear band of ~50 kDa in SDS-PAGE. The CPP/GST-Nef nanoparticles were formed with a diameter of below 200 nm and notably delivered into HEK-293T cells. Generally, the Nef protein was expressed in prokaryotic and eukaryotic expression systems using different vectors and efficiently transfected in mammalian cells using various delivery systems. The in vitro efficient delivery of HIV-1 Nef gene and also its protein supports the potential of Nef DNA constructs and CPPs as potent carriers of Nef protein for HIV vaccine design in Future.