Novel halo- and thermo-tolerant Cohnella sp. A01 L-glutaminase: heterologous expression and biochemical characterization.

L-glutaminase importance to use in the food industry and medicine has attracted much attention. Enzymes stability has always been a challenge while working with them. We heterologously expressed and characterized a novel stable L-glutaminase from an extremophile bacterium (Cohnella sp. A01, PTCC No: 1921). Km, Vmax, catalytic efficiency and specific activity of rSAM were respectively 1.8 mM, 49 µmol/min, 1851 1/(S.mM) and 9.2 IU/mg. Activation energy for substrate to product conversion and irreversible thermo-inactivation were respectively 4 kJ/mol and 105 kJ/mol from the linear Arrhenius plot. rSAM had the highest activity at temperature 50 °C, pH 8 and was resistant to a wide range of temperature and pH. In compare to the other characterized glutaminases, rSAM was the most resistant to NaCl. Mg2+, glycerol, DTT, and BME enhanced the enzyme activity and iodoacetate and iodoacetamide inhibited it. rSAM had only been partially digested by some proteases. According to the Fluorimetry and Circular dichroism analysis, rSAM in pH range from 4 to 11 and temperatures up to 60 °C had structural stability. A cysteine residue in the enzyme active site and a thiol bond were predicted upon the modeled tertiary structure of rSAM. Present structural studies also confirmed the presence of a thiol bond in its structure.

Synergistic effects of deuterium depleted water and Mentha longifolia L. essential oils on sepsis-induced liver injuries through regulation of cyclooxygenase-2.

CONTEXT: Mentha longifolia L. (Lamiaceae), a traditional medicinal herb, has been highly valued for exhibiting antimicrobial, antioxidant and antispasmodic properties. OBJECTIVE: For the first time, the synergetic anti-inflammatory effects of deuterium depleted water (DDW) and M. longifolia essential oils (ML) were investigated in experimental sepsis. MATERIALS AND METHODS: Fifty Wistar rats were divided into 5 groups (n = 10): negative control (laparotomy), CLP, treatment groups including the combination of DDWs (15 and 30 ppm) and ML (100 mg/kg b.w) and indomethacin. At 24 h after CLP induction, lipid peroxidation (LP), glutathione (GSH), glutathione in S-transferases (GST), ferric reducing ability of plasma (FRAP), myeloperoxidase (MPO), prostaglandin E2 (PGE2), and COX-2 expression were determined in the plasma and liver tissues. RESULTS: Compared with the CLP group, the administration of DDWs and ML significantly (p < 0.05) prevented the increase of LP, COX-2 and PGE2 levels and liver enzymes. Additionally, the decreased levels of FRAP and GSH induced by sepsis were remarkably (p < 0.05) risen by the administration of DDWs and ML in comparison to the CLP group. However, no significant (p > 0.05) differences were observed regarding GST, ALP and bilirubin levels. Our results also proved the synergistic anti-inflammatory activities of the DDWs and ML. The anti-inflammatory effects of the DDWs and ML were confirmed by histopathological studies. DISCUSSION AND CONCLUSIONS: The combination of DDWs and ML exerted synergistic anti-inflammatory activity against CLP-induced sepsis possibly through modulating oxidative stress/antioxidant parameters.

Antibacterial Activity of Isolated Immunodominant Proteins of Naja Naja (Oxiana) Venom.

The aim of this study is to investigate antibacterial effects of immunodominant proteins isolated from the venom of Naja Naja Oxiana snake against Staphylococcus aureus, Escherichia coli, Bacillus subtilis and Pseudomonas aeruginosa. The innate immune system is an important line of defense against bacterial diseases. Antibacterial peptides and proteins produced by snake venoms have recently attracted significant attention due to their relevance to bacterial diseases and the potential of being converted into new therapeutic agents. Identification of immunodominant proteins of the venom of Naja Naja Oxiana snake was performed by SDS-PAGE and western blot analysis. Identified proteins were isolated directly from preparative gel electrophoresis by Electro-elution. In the next step, antibacterial effects of immunodominant proteins were tested against several strains of clinical isolates, including S.aureus, B.subtilis (Gram-positive bacteria) P.aeruginosa and E.coli (Gram-negative bacteria) using broth microdilution and disc-diffusion assays. In order to compare the results of the disc-diffusion assay, antibacterial effects of several antibiotics (Gentamicin, Ampicillin, Penicillin, Amoxicillin and Ciprofloxacin) were also examined using the same conditions. Results showed that immunodominant proteins of (14, and 65kDa) with high immunogenicity were very effective in inhibiting the growth of two Gram-positive bacteria (S.aureus, B.sub) that were tested. However, they were only moderately effective in inhibiting the growth of the two tested Gram-negative bacteria (P.aeruginosa and E.coli). However, immunodominant proteins of 22 kDa and 32kDa with high immunogenicity, showed slight effectiveness in inhibiting the growth of two; the Gram-positive and Gram-negative bacteria that were tested. To the best of our knowledge, these immunodominant proteins are novel antigens for potent antimicrobial effects against two gram-positive bacteria (S.aureus, B.subtilis ) and less antimicrobial effect against two gram-negative bacteria (E.coli, P.aeruginosa) that were prepared .

Fas gene variants in childhood acute lymphoblastic leukemia and association with prognosis.

Fas molecule is one of the main important molecules involved in apoptotic cell death. Single nucleotide polymorphisms in the promoter of Fas gene at positions -1377G/A and -670 A/G may affect its expression and play an important role in the pathology of leukemia. In the present study the association between these polymorphisms and risk of the development of acute lymphoblastic leukemia (ALL) in children with ALL compared to cancer-free control subjects was examined by polymerase chain reaction- based restriction fragment length polymorphism. The relationship between the polymorphisms and clinical and laboratory features of the patients and response to therapy were determined. No significant differences in genotype and allele frequencies between the patients and the control subjects at positions -670 and -1377 were detected. Evaluation of the prognostic factors revealed an association between the GG genotype at position -670 and liver involvement in ALL patients (p < 0.04). Although patients with -1377 AA genotype showed shorter mean complete remission duration, the result of survival analysis did not reach to be significant. In conclusion, results of this study showed no contribution of Fas genotypes at positions -670 and -1377 to risk of ALL in children. The association of Fas GG genotype at position -670 with liver involvement in the patients may show its important role in prognosis of ALL.

Lipid profile improvement after four group psychological interventions in combination to nutritional and physical activity instructing among overweight and obese individuals.

BACKGROUND: Research background approved weight loss would result in improvement of lipid profile. Nowadays, weight management is attained by modifying psychological mediators, as well as dieting and making individuals do more physical activity. Hence, This research was performed to compare effectiveness of different psychological interventions on lipid profile improvement among individuals with overweight and obesity. METHODS: Using quasi-experimental design, 68 participants with mean age 26 years and 9 months (ranged from 20 to 43) were randomly assigned into 4 groups (including LEARN, CBT, CT, and MCT), and eventually 61 participants completed the treatment protocols. BMI was calculated by Omron digital body composition instrument, and blood lipids and glucose concentrations were measured prior to and after all interventions. RESULTS: Data analysis by using t-test for dependent groups analysis revealed lipid profile, FBS, and BMI were significantly modified after applying 4 interventions comparing baseline and post treatment mean differences(P<0.05). Furthermore, repeated measure design analysis showed there were significant mean differences (P<0.05) in HDL cholesterol, TG levels and FBS levels among 4 groups. Scheffe test revealed more detailed significant differences among four groups in HDL cholesterol, and FBS concentration varieties (P<0.05). CONCLUSIONS: Adding psychological interventions to dietary plans and regular physical activities in overweight management would consider as a valuable step in optimizing physiological outcomes and that's because of correcting mind-body interactional pathways among obese and overweight individuals.

Association between Hypoadiponectinemia and Low Serum Concentrations of Calcium and Vitamin D in Women with Polycystic Ovary Syndrome.

Objective. To investigate the possible association of calcium and vitamin D deficiency with hypoadiponectinemia in women with PCOS. Subjects and Methods. In this case-control study, 103 PCOS cases and 103 controls included. The concentrations of calcium, 25-OH-vitamin D (25OHD), adiponectin, insulin, glucose, total cholesterol, HDL-cholesterol, triglyceride (TG), and androgens were measured in fasting blood samples. Results. Adiponectin (8.4 ± 2.7 ng/mL versus 13.6 ± 5 ng/mL in control group, P : 0.00), calcium (2 ± 0.1 mmol/L versus 2.55 ± 0.17 mmol/L in controls, P : 0.00), and 25-OH-Vit D (30 ± 2.99 nmol/L versus 43.7 ± 5.2 nmol/L in control group, P : 0.00) levels were decreased in women with PCOS. Subjects with PCOS had higher concentrations of TG (1.4 ± 0.77 mmol/L versus 1.18 ± 0.75 mmol/L in control group, P : 0.019) and dehydroepiandrosterone sulfate (DHEA-S) (10.7 ± 11 mmol/L versus 9.7 ± 10.4, P : 0.02 in control group). There were significant correlations between adiponectin concentrations with calcium (r : 0.78, P : 0.00) and 25OHD levels (r : 0.82, P : 0.00). The association of hypoadiponectinemia and PCOS was not significant considering 25OHD as a confounding factor. Conclusion. The present findings indicate that the association of hypoadiponectinemia with PCOS is dependent on vitamin D. A possible beneficiary effect of vitamin D on the metabolic parameters in PCOS may be suggested.

Serum magnesium concentrations in polycystic ovary syndrome and its association with insulin resistance.

OBJECTIVE: It has been revealed that low serum magnesium (Mg) is often associated with insulin resistance (IR), cardiovascular problems, diabetes mellitus, and hypertension. Patients with polycystic ovary syndrome (PCOS) are known to have a high incidence of insulin resistance. This study was designed to determine whether women with PCOS exhibit serum magnesium deficiency and its potential association with IR. SUBJECTS AND METHODS: In this cross-sectional study, 103 cases with PCOS and 103 normal women who were matched for their age and body mass index (BMI) were included. Blood samples were collected after an overnight fast and concentrations of calcium, magnesium, testosterone, dehydroepianderosterone sulfate, insulin, fasting plasma glucose (FPG), triglyceride, total cholesterol, and HDL-cholesterol were measured. RESULTS: The risk of PCOS for subjects with Mg deficiency was 19 times greater than those who had normal serum Mg concentrations (p ≤ 0.0001). No correlation was found between Mg and insulin sensitivity or secretion, FPG, dyslipidemias, and also androgen concentrations. After adjustment for calcium concentration the role of magnesium to predict PCOS attenuated and became non-significant (ß:-1.9, p: 0.7). CONCLUSION: The present study provides the first evidence showing that magnesium deficiency is not associated with IR in PCOS. According the evidences of this study, serum calcium concentration is more potent predictor of PCOS than serum Mg and only calcium, not Mg, is related to insulin resistance in PCOS.

Decreased adiponectin levels in polycystic ovary syndrome, independent of body mass index.

BACKGROUND: Insulin resistance has been shown to have an association with polycystic ovary syndrome (PCOS). This study was designed to evaluate the potential role of adiponectin, which is linked with insulin resistance, in the etiology of PCOS and its relationship to obesity. METHODS: This case-control study consisted of 103 newly diagnosed PCOS cases and 73 female controls seen at a referral university hospital in Zanjan, Iran. Serum adiponectin, insulin, plasma fasting glucose, and lipid levels were measured. The homeostasis model assessment index was used to determine the level of insulin resistance. Women were classified as follows: Group I (normal nonlean women); group II (normal lean women); group III (nonlean women with PCOS); and group IV (lean women with PCOS) RESULTS: Adiponectin levels were decreased in women with PCOS (8.4 +/- 2.7 ng/mL vs. 13.6 +/- 5 ng/mL in the control group, P < 0.001). There was no significant difference between the adiponectin concentrations of women in group III and that in group IV (8.1 +/- 2.8 ng/mL vs. 9.2 +/- 2.6 ng/mL, respectively, P = 0.1). Adiponectin levels were significantly lower in group I compared with group II. A weak but significant negative correlation was found between adiponectin and insulin levels in all the subjects. Multiple regression analyses showed that the presence of PCOS was the only significant determinant of serum adiponectin levels. CONCLUSIONS: Adiponectin levels were reduced in all the women with PCOS. There seemed to be an interaction between adiponectin and PCOS pathogenesis that was independent of body mass index.

Anti-leishmanial effects of trinitroglycerin in BALB/C mice infected with Leishmania major via nitric oxide pathway.

This study investigated whether trinitroglycerine (TNG) as nitric oxide (NO) releasing agent had anti-leishmanial effects and mediated pathology in BALB/c mice infected with Leishmania major. Cutaneous leishmaniasis (CL), a zoonotic infection caused by leishmania protozoa is still one of the health problems in the world and in Iran. NO is involved in host immune responses against intracellular L. major, and leishmania killing by macrophages is mediated by this substance. Moreover, application of CL treatment with NO-donors has been recently indicated. In our study, TNG was used for its ability to increase NO and to modify CL infection in mice, in order to evaluate NO effects on lesion size and formation, parasite proliferation inside macrophages, amastigote visceralization in target organs, and NO induction in plasma and organ suspensions. Data obtained in this study indicated that TNG increased plasma and liver-NO, reduced lesion sizes, removed amastigotes from lesions, livers, spleens, and lymph nodes, declined proliferation of amastigotes, hepatomegaly, and increased survival rate. However, TNG reduced spleen-NO and had no significant effects on spelenomegaly. The results show that TNG therapy reduced leishmaniasis and pathology in association with raised NO levels. TNG had some antiparasitic activity by reduction of positive smears from lesions, livers, spleens, and lymph nodes, which could emphasize the role of TNG to inhibit visceralization of L. major in target organs.

Evaluation of anti-leishmanial activity by induction of nitric oxide and inhibition of prostaglandin in Balb/c mice infected with Leishmania major.

Cutaneous leishmaniasis is still one of the health problems in Iran and in the region. Nitric oxide (NO) has a key mechanism in the elimination of parasite from the body by its anti-leishmanial activity. Prostaglandin (PG) is a critical inhibitory factor of infected macrophage to decrease their anti-leishmanial activity. This study was designed to induce NO by L-arginine (L-Arg) precursor and inhibit PG production by anti-inflammatory Indomethacin (INDO) in Leishmania major infected Balb/c mice, in order to evaluate the effects of NO and PG on delay of lesion formation, size of lesion and proliferation of amastigotes inside macrophages. Liver, spleen and lymph nodes were also studied as target organs to detect amastigotes. Serum, liver and spleen suspensions were investigated for NO induction by using Griess microassay and serum PG was determined by ELISA. The results indicated that NO production was inhibited by Leishmania in infected Balb/c mice as compared with naive animals. Serum NO was inhibited by a combination therapy of L-Arg and INDO. Although NO was decreased in the liver by L-Arg, however it increased in the spleen after L-Arg and INDO application. A significant decline was observed in lesion size from Week 6 after infection by INDO. Both L-Arg and INDO had significant inhibitory effects on visceralization of leishmania in target organs. Only L-Arg decreased proliferation of promastigotes in macrophages. Pathophysiological signs including hepatomegaly, splenomegaly, survival rate and body weight all were affected in this experiment. Statistical analysis of data revealed an association between NO induction and PG inhibition in leishmaniasis. These data may indicate a possible candidatory for L-Arg and INDO as novel drugs for the treatment of leishmaniasis in mouse model.