Chitosan-LeoA-DNA Nanoparticles Promoted the Efficacy of Novel LeoA-DNA Vaccination on Mice Against Helicobacter pylori.

More than half of the world's population is infected with Helicobacter pylori (H. pylori), which may lead to chronic gastritis, peptic ulcers, and stomach cancer. LeoA, a conserved antigen of H. pylori, aids in preventing this infection by triggering specific CD3+ T-cell responses. In this study, recombinant plasmids containing the LeoA gene of H. pylori are created and conjugated with chitosan nanoparticle (CSNP) to immunize BALB/c mice against the H. pylori infection. We used the online Vaxign tool to analyze the genomes of five distinct strains of H. pylori, and we chose the outer membrane as a prospective vaccine candidate. Afterward, the proteins' immunogenicity was evaluated. The DNA vaccine was constructed and then encapsulated in CSNPs. The effectiveness of the vaccine's immunoprotective effects was evaluated in BALB/c mice. Purified activated splenic CD3+ T cells are used to test the anticancer effects in vitro. Nanovaccines had apparent spherical forms, were small (mean size, 150-250 nm), and positively charged (41.3 ± 3.11 mV). A consistently delayed release pattern and an entrapment efficiency (73.35 ± 3.48%) could be established. Compared to the non-encapsulated DNA vaccine, vaccinated BALB/c mice produced higher amounts of LeoA-specific IgG in plasma and TNF-α in splenocyte lysate. Moreover, BALB/c mice inoculated with nanovaccine demonstrated considerable immunity (87.5%) against the H. pylori challenge and reduced stomach injury and bacterial burdens in the stomach. The immunological state in individuals with GC with chronic infection with H. pylori is mimicked by the H. pylori DNA nanovaccines by inducing a shift from Th1 to Th2 in the response. In vitro human GC cell development is inhibited by activated CD3+ T lymphocytes. According to our findings, the H. pylori vaccine-activated CD3+ has potential immunotherapeutic benefits.

Optimization of xylanase production by Pichia kudriavzevii and Candida tropicalis isolated from the wood product workshop.

Enzymatic compounds can be found abundantly and provide numerous advantages in microbial organisms. Xylanases are used in various pharmaceutical, food, livestock, poultry, and paper industries. This study aimed to investigate xylanase-producing yeasts, xylose concentration curve and their enzymatic activity under various factors including carbon and nitrogen sources, temperature, and pH. Enzyme activity was evaluated under different conditions before, during, and after purification. The yeast strains were obtained from the wood product workshop and were subsequently cultivated on YPD (yeast extract peptone dextrose) medium. Additionally, the growth curve of the yeast and its molecular identification were conducted. The optimization and design process of xylan isolated from corn wood involved the use of Taguchi software to test different parameters like carbon and nitrogen sources, temperature, and pH, with the goal of determining the most optimal conditions for enzyme production. In addition, the Taguchi method was utilized to conduct a multifactorial optimization of xylanase enzyme activity. The isolated species were partially purified using ammonium sulfate precipitation and dialysis bag techniques. The results indicated that 3 species (8S, 18S, and 16W) after molecular identification based on 18S rRNA gene sequencing were identified as Candida tropicalis SBN-IAUF-1, Candida tropicalis SBN-IAUF-3, and Pichia kudriavzevii SBN-IAUF-2, respectively. The optimal parameters for wheat carbon source and peptone nitrogen source were found at 50 °C and pH 9.0 through single-factor optimization. By using the Taguchi approach, the best combination for highest activity was rice-derived carbon source and peptone nitrogen source at 50 °C and pH 6.0. The best conditions for xylanase enzyme production in single-factor optimization of wheat bran were 2135.6 U/mL, peptone 4475.25 U/mL, temperature 50 °C 1868 U/mL, and pH 9.0 2002.4 U/mL. Among the tested yeast, Candida tropicalis strain SBN-IAUF-1 to the access number MZ816946.1 in NCBI was found to be the best xylanase product. The highest ratio of enzyme production at the end of the delayed phase and the beginning of the logarithmic phase was concluded by comparing the growth ratio of 8S, 16W, and 18S yeasts with the level of enzymatic activity. This is the first report on the production of xylan polymer with a relative purity of 80% in Iran. The extracellular xylanases purified from the yeast species of C. tropicalis were introduced as a desirable biocatalyst due to their high enzymatic activity for the degradation of xylan polymers.

Use of immunoinformatics and the simulation approach to identify Helicobacter pylori epitopes to design a multi-epitope subunit vaccine for B- and T-cells.

BACKGROUND: Helicobacter pylori cause a variety of gastric malignancies, gastric ulcers, and cause erosive diseases. The extreme nature of the bacterium and the implantation of this bacterium protects it against designing a potent drug against it. Therefore, employing a precise and effective design for a more safe and stable antigenic vaccine against this pathogen can effectively control its associated infections. This study, aimed at improving the design of multiple subunit vaccines against H. pylori, adopts multiple immunoinformatics approaches in combination with other computational approaches. RESULTS: In this regard, 10 HTL, and 11 CTL epitopes were employed based on appropriate adopted MHC binding scores and c-terminal cut-off scores of 4 main selected proteins (APO, LeoA, IceA1, and IceA2). An adjuvant was added to the N end of the vaccine to achieve higher stability. For validation, immunogenicity and sensitization of physicochemical analyses were performed. The vaccine could be antigenic with significantly strong interactions with TOLK-2, 4, 5, and 9 receptors. The designed vaccine was subjected to Gromacs simulation and immune response prediction modelling that confirmed expression and immune-stimulating response efficiency. Besides, the designed vaccine showed better interactions with TLK-9. CONCLUSIONS: Based on our analyses, although the suggested vaccine could induce a clear response against H. pylori, precise laboratory validation is required to confirm its immunogenicity and safety status.

The effect of homocysteine thiolactone on paraoxonase and aryl esterase activity of human serum purified paraoxonase 1 in vitro experiments.

BACKGROUND: The important role of lipoproteins, particularly low-density lipoprotein (LDL) and high-density lipoprotein (HDL), has been highly regarded among the known causes of cardiovascular disease (CVD). A wide range of risk factors may cause structural and functional changes in lipoprotein particles, resulting in deposition and formation of atherosclerotic plaques. Homocysteine is one of the most important risk factors in heart disease, and its atherosclerotic properties appear to be related to its intermediate metabolite called homocysteine thiolactone (HCTL). The major aim of the present investigation was to study the effect of HCTL in different concentrations (10, 50, and 100 µM) on paraoxonase and aryl esterase activities of purified human serum paraoxonase 1 (PON1) antioxidant enzyme related to HDL, as an extracellular hydrolyzing enzyme of HCTL. METHODS: In order to purify PON1 enzyme from human serum, three-step chromatographic methods including DEAE Sephadex A50, Sephadex G100, and DEAE Sephadex A50 were used. Protein concentration and paraoxonase and aryl esterase activities of each fraction were measured separately and the highest activities fractions were collected and subsequently pooled together for the next steps. Ultimately, both activities of PON1 in the presence of different concentrations of HCTL were measured in triplicate by spectrophotometry technique. RESULTS: HCTL at concentrations of 50 and 100 µM decreased both paraoxonase and aryl esterase activities (P < 0.05) in comparison with the control group, which is directly related to the increase in HCTL concentration. However, at a concentration of 10 µM HCTL, no significant difference was observed in both paraoxonase and aryl esterase activities compared to the control group. CONCLUSION: HCTL is a highly toxic and reactive compound that is produced in all cells. Extracellular enzyme PON1 causes its hydrolysis with high efficiency. The results obtained from the present study showed that paraoxonase and aryl esterase activities decreased in vitro in the presence of HCTL and therefore, HCTL may cause changing in the protein structure of this enzyme. Previous in vivo studies have also shown decrease of PON1 activity in patients with hyperhomocysteinemia.

Association of BsmI variant of vitamin D receptor gene with polycystic ovary syndrome: A case-control study.

BACKGROUND: Polycystic ovarian syndrome (PCOS) is an endocrine disorder that affects women's fertility and causes alterations such as obesity, insulin resistance, menstrual irregularities, and polycystic ovaries. The results of the studies show that the issue of vitamin D and vitamin D receptor (VDR) is controversial for PCOS susceptibility. OBJECTIVE: To investigate the association of BsmI polymorphism in the VDR gene with metabolic parameters in obese PCOS women. MATERIALS AND METHODS: In this case-control study, 38 obese subjects with PCOS and 40 unrelated obese individuals were evaluated to determine the allelic and genotypic frequency of BsmI variant by Polymerase Chain Reaction Restriction Fragment Length Polymorphism method. Body Mass Index, parathyroid hormone, phosphorus, and calcium were evaluated in all participants. RESULTS: BsmI (rs1544410), (A/G) AA, AG, GG, A, and G percentage of genotypic/allelic frequencies were 65.8, 26.3, 7.9, 78.9, and 21.1 in cases and 57.5, 40, 2.5, 77.5, and 22.5 in controls, respectively. Statistical analysis revealed that the differences in genotypic (p = 0.31)/allelic (p = 0.83) frequencies and dominant (p = 0.45)/recessive (p = 0.35) models between the cases and controls were not significant. This study indicates no association between the BsmI genotypes and metabolic parameters. CONCLUSION: It can be concluded that VDR BsmI (rs1544410) Intron 8 (A > G) was not associated with obesity along with PCOS susceptibility in the studied groups.

The effect of endurance training with crocin consumption on the levels of MFN2 and DRP1 gene expression and glucose and insulin indices in the muscle tissue of diabetic rats.

This study aimed to investigate the effect of crocin consumption, high-intensity interval training (HIIT), and low-intensity continuous training (LICT) and their interactive effect on the gene expression of Mfn2 and Drp1 in the skeletal muscle and serum glucose and insulin indices in high-fat diet (HFD) and streptozotocin (STZ)-induced diabetic rats. Fifty-six adult rats were divided into eight groups of seven subjects: crocin consumption, HIIT, LICT, HIIT with crocin, LICT with crocin, diabetic control, healthy control, and sham (placebo). At the end of the course (5 months), metabolic indices were measured. Moreover, the Mfn2 and Drp1 gene expression levels in all groups were measured using RT-PCR. The statistical analysis showed that in the exercise training (HIIT and LICT) and the crocin consumption groups, the glucose and insulin indices significantly improved (p = .005). Moreover, in these groups, the levels of gene expression of Mfn2 and Drp1 significantly increased and decreased, respectively (p = .001). Exercise training and crocin consumption appear to, either in combination or individually, have a beneficial effect on mitochondrial dynamics and diabetes by improving the mitochondrial fusion and fission indices (Mfn2 and Drp1), and by modifying the insulin resistance index and glucose homeostasis. PRACTICAL APPLICATIONS: Mfn2 and Drp1, as the main regulators of the mitochondrial fusion and fission, play an important role in maintaining mitochondrial dynamics and type 2 diabetes. Thus, the regulation of mitochondrial dynamics is an intricate process that retains the balance between mitochondrial fission and fusion, and any disturbance in this balance can lead to mitochondrial-associated diseases including insulin resistance and T2D. There is evidence that herbal antioxidants Including crocin and exercise training help improve the mitochondrial activity and insulin sensitivity in T2D. Considering the importance of the two Drp1 and Mfn1 genes in the mitochondrial dynamic pathway and coding the proteins that play a key role in relation to T2D, this study primarily examined the interactive effects of endurance training (HIIT and LICT) along with crocin consumption on the expression the genes mentioned above; the results obtained in this study can provide a new approach to the treatment of HFD + STZ-induced diabetic rats.

Effect of Metformin Treatment on Insulin Resistance Markers, and Circulating Irisin in Women with Polycystic Ovarian Syndrome (PCOS).

Polycystic ovarian syndrome (PCOS) is considered as a common endocrinal dysfunction among adult women characterized by polycystic ovaries, anovulation, and hyperandrogenism. Irisin is associated with metabolic parameters and insulin resistance. However, the association of irisin with PCOS remains poorly delineated. This study was aimed to examine circulating irisin levels and effects of metformin on this parameter in women with PCOS. Moreover, the association of irisin with insulin resistance markers was determined. Thirty-nine females with PCOS, aged 20-40 years, participated in this study and received 500 mg of metformin once daily for 3 months. Serum creatinine, blood urea nitrogen, high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein (LDL), cholesterol, triglyceride, fasting blood sugar, testosterone, 17-hydroxyprogesterone, and irisin were assayed in the studied groups. Circulating irisin was significantly higher in PCOS women. Circulating irisin levels correlated with 17-hydroxyprogesterone, testosterone, and insulin. Three months metformin treatment decreased circulating irisin in PCOS women and improved IR. Circulating irisin is directly associated with insulin resistance in PCOS women and may be used as a biomarker for IR in these patients. Moreover, metformin as a confounding therapy in metabolic diseases can be used to regulate circulating irisin levels in PCOS women.

An Overview of FGF19 and FGF21: The Therapeutic Role in the Treatment of the Metabolic Disorders and Obesity.

Fibroblast growth factors (FGFs) are responsible for the regulation of a wide range of biological functions, among which cellular proliferation, survival, migration, and differentiation could be pointed out. FGF19 controls the enterohepatic bile acid/cholesterol system, and FGF21 modulates fatty acid/glucose metabolism. Obesity, type 2 diabetes, coronary artery disease, and cancer, all can alter FGF21 circulating concentrations. In contrast to FGF21, metabolic diseases exhibit reduced serum FGF19 levels. Accordingly, FGF19 and FGF21 play important roles in regulating glucose and lipid metabolism. Hence, we present here a timely review on the relationship between FGF19/21 and metabolic diseases, especially obesity, and their probable role in development and treatment of obesity seems necessary.

Stabilization of bovine lactoperoxidase in the presence of ectoine.

Lactoperoxidase (LPO) is a heme peroxidase with various applications in industry and medicine. In this study, the effects of ectoine, as a compatible solute, on the structure, thermal stability, thermodynamic parameters, activity, and stability of LPO have been investigated. The results showed that the catalytic activity of LPO was improved by increasing ectoine concentration. The UV-visible absorption spectroscopy and FTIR spectra studies indicated that ectoine could bind to the LPO spontaneously. Moreover, ectoine increased the enzyme Tm and Gibbs free energy. The fluorescence measurements showed that LPO fluorescence was quenched in the presence of ectoine. The quenching mechanism was probably a static quenching by forming a ground state complex. The thermodynamic parameters indicated that hydrogen bonding and Vander Waals forces played a key role in the LPO-ectoine interaction process. The findings suggest that ectoine could be used as a lactoperoxidase stabilizing agent for industrial or medical purposes.

Cadmium Testicular Toxicity in Male Wistar Rats: Protective Roles of Zinc and Magnesium.

Cadmium (Cd) is a highly toxic element, which may cause toxicity to most organs in the body. Zinc (Zn) and magnesium (Mg) are essential minerals with probable benefits on Cd harmful effects. Finding an efficient and non-pathological treatment against Cd toxicity seems promising. Fifty adult rats were divided into ten experimental groups of five rats each. The Cd group was treated with 1 mg Cd/kg and the control group received 0.5 cm3 normal saline. The other eight groups received Zn (0.5 and 1.5 mg/kg) and Mg (0.5 and 1.5 mg/kg) either alone or in combination with 1 mg Cd/kg through IP injection for 3 weeks. Testis malondialdehyde (MDA), sperm parameters, and testis histopathology were investigated. Cd reduced sperm parameters and increased testis MDA. Moreover, Cd exposure caused a significant histological damage in testis of male rats. However, Zn or Mg treatment prevented and reversed Cd toxic alterations in testis. These findings suggest that co-administration of Zn or Mg could improve cadmium testicular toxicity in male Wistar rats.

Protective Role of Zinc and Magnesium against Cadmium Nephrotoxicity in Male Wistar Rats.

Cd is a toxic metal that has a destructive impact on most organ systems. This work aims to determine Zn or Mg protective effects against Cd renal toxicity. In this study, rats were divided into six groups. The Cd group was treated with 1 mg Cd/kg, and the control group received 0.5 cm3 normal saline, intraperitoneally. The other four groups received one of the following dosages of 1 mg/kg Cd + 0.5 mg/kg Zn, 1 mg/kg Cd + 1.5 mg/kg Zn, 1 mg/kg Cd + 0.5 mg/kg Mg, or 1 mg/kg Cd + 1.5 mg/kg Mg through IP injection for 3 weeks. Kidney malondialdehyde (MDA) and serum sodium, potassium, urea, creatinine, and protein were measured. Light microscopic examination was used for histological studies. Cd reduced serum creatinine and protein, and increased urea, sodium, and potassium. Moreover, Cd exposure caused a significant enhancement in MDA levels as well as histological damage in kidneys. Zn or Mg treatment prevented and reversed toxic alterations induced by Cd. These results suggest that Zn and Mg may have protective effects against Cd renal toxicity.

Lymphocyte cytotoxicity of oxLDL in patients with atherosclerosis.

BACKGROUND: Atherosclerosis, a chronic inflammatory disease of the vessel wall, is characterized by local and systemic immune responses to a variety of antigens. Oxidized low-density lipoprotein (oxLDL) is considered as an important determining factor in the pathogenesis of atherosclerosis. OBJECTIVE: The purpose of this study was to investigate the degree of peripheral blood mononuclear cells (PBMC) vulnerability to in vitro oxLDL-induced cytotoxicity from atherosclerotic patients in comparison to healthy individuals. METHODS: Thirty patients with atherosclerotic lesions, confirmed by angiography, and 30 matched healthy individuals were investigated. PBMC was prepared from individuals' blood samples which were further stimulated with low dose (1 µg/mL) and high dose (50 µg/mL) of extensively oxidized LDL. MTT assay was utilized to measure cell viability and proliferation. Stimulation index (SI) was calculated as mean ratio of optical density (OD) of the stimulated cells divided by OD of untreated cells. RESULTS: Low dose oxLDL treatment caused no significant proliferative or cytotoxic effect in the control group; however, similar treatment caused significant cytotoxic effect in the patients compared to the controls (p=0.026). High dose oxLDL treatment induced more significant cytotoxicity in the patients compared to the controls (p=0.006). Comparison of the SI between the two groups of patients and controls showed significantly lower index by either the low (p=0.03) or the high dose (p<0.001) oxLDL in the patients compared to the controls. CONCLUSIONS: PBMC from patients with atherosclerosis showed increased susceptibility to oxLDL-induced cytotoxicity. Our results imply that prolonged exposure to elevated levels of circulating oxLDL could weaken the cellular defense mechanisms by progressive depletion of the pool of antiapoptotic proteins, rendering the cells more vulnerable to oxLDL-induced cell death.

Antioxidant effects of Citrus aurantifolia (Christm) juice and peel extract on LDL oxidation.

BACKGROUND: We studied the antioxidant effects of fresh juice and peel extract of Citrus aurantifolia (Christm). METHODS: Low density lipoprotein (LDL) was separated from one hypercholesterolemic human serum by modified Bronzert and Brewer procedure. Oxidation of LDL was measured at 234 nm against 0, 5, 10, 20, 25, 30 and 40 µl of fresh lime juice and 0, 5, 10, 15 and 20 µl of peel polyphenolic extract solution in DMSO. RESULTS: 5 µl of lime juice didn't change LDL oxidation. 10 µl of juice inhibited LDL oxidation, and with increasing the juice concentration, LDL was oxidized faster. The higher concentrations of peel extract prevented LDL oxidation better than the lower ones. CONCLUSIONS: Both juice and peel demonstrated antioxidant properties, but the excessive consumption of lime juice seems not to be beneficial. Regarding the intensity and type of flavonoids, lime juice and peel may show different effects.

Effect of CRP on Some of the in vitro Physicochemical Properties of LDL.

BACKGROUND: Atherosclerosis is the most important underlying cause of cardiovascular diseases (CVD) which recently has been classified as an inflammatory disorder. Accumulation of large amounts of oxidized LDL in the intima during local inflammation reaction led to increase several factors such as C -reactive protein (CRP). It has also been reported that CRP is able to bind with modified forms of LDL as well as oxidized LDL. These findings suggest possible positive or negative involvement of this protein in atherogenesis. The main objective of the present study was to assess the influence of CRP on LDL oxidation and the possible physical \changes of LDL in the presence of CRP in vitro. METHODS: In this study, the susceptibility of purified LDL to oxidation was assayed by monitoring of formation of conjugated dienes in different physiological concentrations of CRP (0 - 0.5 -2 µg/ml) using a shimadzu spectrophotometer. Electrophoresis was used to determine the electrophoretic mobility of LDL in those conditions. RESULTS: CRP significantly reduced the susceptibility of Cu(++) -induced LDL oxidation through increasing the lag timeand there was positive relationship between these findings and CRP concentration (P < 0.05). CRP caused a significant reduction in the electrophotretic mobility of LDL compared to native LDL (n-LDL) (P < 0.05). CONCLUSION: A considerable reduction was shown in LDL oxidation, in higher concentration of CRP, via an unknown mechanism. The electrophoretic mobility of LDL, in the oxidative condition, decreases in the presence of CRP compared to n-LDL, which can be indicative of the effect of this protein on the physical and chemical properties of LDL. It seems that, other pathway than LDL oxidation is responsible for the effect of CRP on the atherogenesis processes.