Prostaglandin Endoperoxide H Synthase-2 (PGHS-2) Variants and Risk of Obesity and Microvascular Dysfunction Among Tunisians: Relevance of rs5277 (306G/C) and rs5275 (8473T/C) Genetic Markers

The purpose of this study was to determine the impact of six PGHS-2 genetic variants on obesity development and microvascular dysfunction. The study included 305 Tunisian subjects (186 normal weights, 35 overweights and 84 obeses). PCR analyses were used for allelic discrimination between polymorphisms. Prostaglandin (PGE2, PGI2), leptin, and matrix metalloproteinase (MMP1, 2, 3, 9) levels were evaluated by ELISA. Fatty acid composition was performed by gas chromatography–mass spectrometry. Our results revealed that subjects carrying the PGHS-2 306CC (rs5277) and 8473CC (rs5275) genotypes present higher anthropometric values compared to wild-type genotypes (306GG, BMI (Kg/m²): 27.11?±?0.58; WC (cm): 93.09?±?1.58; 306CC, BMI: 33.83?±?2.46; WC: 109.93?±?5.41; 8473TT, BMI: 27.75?±?0.68; WC: 93.96?±?1.75; 8473CC, BMI: 33.72?±?2.2; WC: 117.89?±?2.94). A reduced microvascular reactivity and a higher PGE2 level were also found in individuals with the 306CC and 8473CC genotypes in comparison to 306GG and 8473TT carriers (306GG, Peak Ach-CVC (PU/mmHg): 0.46?±?0.03; PGE2 (pg/ml): 7933.1?±?702; 306CC, Peak Ach-CVC: 0.24?±?0.01; PGE2: 13,380.3?±?966.2; 8473TT, Peak Ach-CVC: 0.48?±?0.05; PGE2: 7086.41?±?700.31; 8473CC, Peak Ach-CVC: 0.23?±?0.01; PGE2: 13,175.7?±?1165.8). Fatty acid analysis showed a significant increase of palmitic acid (PA) (34.2?±?2.09 vs. 16.82%?±?1.76, P?<?0.001), stearic acid (SA) (25.76?±?3.29 vs. 9.05%?±?2.53, P?<?0.001), and linoleic acid (LA) (5.25?±?1.18 vs. 0.5%?±?0.09, P?<?0.001) levels in individuals carrying the PGHS-2 306CC genotype when compared to GG genotype individuals. Subjects with the 8473CC genotype showed also a significant increase of PA, SA ,and LA levels when compared to TT genotype carriers (PA: 38.02?±?1.51 vs. 12.65%?±?1.54, P?<?0.001; SA: 32.96?±?1.87 vs. 1.38%?±?0.56, P?<?0.001; LA: 26.84?±?2.09 vs. 3.7%?±?1.54, P?<?0.001). Logistic regression analysis revealed that PGHS-2 306CC and 8473CC variants are significantly associated with obesity status (OR 6.25, CI (1.8–21.6), P?=?0.004; OR 3.01, CI (1.13–8.52), P?=?0.03, respectively). Haplotypes containing the C₃₀₆:T₈₄₇₃ (OR 2.91; P?=?0.01) and G_306:C₈₄₇₃ (OR 5.25; P?=?0.002) combinations were associated with an enhanced risk for obesity development in the studied population. In conclusion, our results highlight that PGHS-2 306G/C and 8473T/C variants could be useful indicators of obesity development, inflammation, and microvascular dysfunction among Tunisians.

     

Human cellular immune response to the saliva of Phlebotomus papatasi is mediated by IL-10-producing CD8+ T cells and Th1-polarized CD4+ lymphocytes

Background

The saliva of sand flies strongly enhances the infectivity of Leishmania in mice. Additionally, pre-exposure to saliva can protect mice from disease progression probably through the induction of a cellular immune response.

Methodology/Principal Findings

We analysed the cellular immune response against the saliva of Phlebotomus papatasi in humans and defined the phenotypic characteristics and cytokine production pattern of specific lymphocytes by flow cytometry. Additionally, proliferation and IFN-γ production of activated cells were analysed in magnetically separated CD4+ and CD8+ T cells. A proliferative response of peripheral blood mononuclear cells against the saliva of Phlebotomus papatasi was demonstrated in nearly 30% of naturally exposed individuals. Salivary extracts did not induce any secretion of IFN-γ but triggered the production of IL-10 primarily by CD8+ lymphocytes. In magnetically separated lymphocytes, the saliva induced the proliferation of both CD4+ and CD8+ T cells which was further enhanced after IL-10 blockage. Interestingly, when activated CD4+ lymphocytes were separated from CD8+ cells, they produced high amounts of IFN-γ.

Conclusion

Herein, we demonstrated that the overall effect of Phlebotomus papatasi saliva was dominated by the activation of IL-10-producing CD8+ cells suggesting a possible detrimental effect of pre-exposure to saliva on human leishmaniasis outcome. However, the activation of Th1 lymphocytes by the saliva provides the rationale to better define the nature of the salivary antigens that could be used for vaccine development.     

Influence of static magnetic field on cadmium toxicity: Study of oxidative stress and DNA damage in rat tissues

In the present study, we investigated the effect of co-exposure to static magnetic field (SMF) and cadmium (Cd) on the biochemical parameters, antioxidant enzymes activity and DNA damage in rat tissues. Animals were treated with cadmium (CdCl₂, 40 mg/L, per os) in drinking water during 4 weeks. Cd treatment induced an increase of plasma lactate dehydrogenase (LDH) and transaminases levels. Moreover, Cd treatment increased malondialdehyde (MDA) and 8-oxodGuo levels in rat tissues. However, the antioxidant enzymes activity such as the glutathione peroxidase (GPx), catalase (CAT) and the superoxide dismutase (SOD) were decreased in liver and kidney, while we noted a huge increase of hepatic and renal cadmium content. Interestingly, the combined effect of SMF (128mT, 1 h/day during 30 consecutive days) and Cd (40 mg/L, per os) decreased the GPx and CAT activities in liver compared to cadmium treated group. However, the association between SMF and Cd failed to alter transaminases, MDA and 8-oxodGuo concentration.

Cd treatment altered antioxidant enzymes and DNA in liver and kidney of rats. Moreover, SMF associated to Cd disrupt this antioxidant response in liver compared to Cd-treated rats.     

Mutation spectrum of primary hyperoxaluria type 1 in Tunisia: Implication for diagnosis in North Africa

Background

Primary hyperoxaluria type 1 (PH1) is an autosomal recessive inherited metabolic disease, characterized by progressive kidney failure due to renal deposition of calcium oxalate. Mutations in the AGXT gene, encoding the liver-specific enzyme alanine glyoxylate aminotransferase, are responsible for the disease. We aimed to determine the mutational spectrum causing PH1 and to provide an accurate tool for diagnosis as well as for prenatal diagnosis in the affected families.

Methods

Direct sequencing was used to detect mutations in the AGXT gene in DNA samples from 13 patients belonging to 12 Tunisian families.

Results

Molecular analysis revealed five mutations causing PH1 in Tunisia. The mutations were identified along exons 1, 2, 4, 5 and 7. The most predominant mutations were the Maghrebian “p.I244T” and the Arabic “p.G190R”. Furthermore, three other mutations characteristic of different ethnic groups were found in our study population. These results confirm the mutational heterogeneity related to PH1 in Tunisian population. All the mutations are in a homozygous state, reflecting the high impact of endogamy in our population.

Conclusion

Mutation analysis through DNA sequencing can provide a useful first line investigation for PH1. This identification could provide an accurate tool for prenatal diagnosis, genetic counseling and screen for potential presymptomatic individuals.     

Genetic polymorphism in the manganese superoxide dismutase gene is associated with an increased risk for hepatocellular carcinoma in HCV-infected Moroccan patients

Hepatocellular carcinoma (HCC) ranks among the 10 most common cancers worldwide. The main risk factors for its development are hepatitis B and C virus infections. Hepatitis B and C viruses induce chronic inflammation and oxidative stress that could predispose a cell to mutagenesis and proliferation. Manganese superoxide dismutase (MnSOD) catalyses the detoxification of free radicals, thus playing a crucial role in the protection against damage. A valine (Val) to alanine (Ala) substitution at amino acid 9, mapping within the mitochondrion-targeting sequence of the MnSOD gene, has been associated with an increased cancer risk. The aim of our study was to investigate a possible association of the Val/Ala-MnSOD polymorphism and HCC development in Moroccan patients. Genotypes were determined by means of PCR and RFLP analysis in 96 patients with HCC and 222 control subjects matched for age, sex, and ethnicity. Homozygous Ala/Ala carriers were 31% in the cases and 18% in the controls, which corresponds to an odds ratio (OR) of 2.89, with a 95% confidence interval (CI) of 1.47-5.68. Stratification into subgroups based on HCV infection status revealed an even more increased risk for homozygous Ala/Ala carriers with hepatitis C infection (38.2% in the cases versus 14.8% in the control subjects OR, 5.09; 95% CI, 1.76-14.66). Our findings provide further evidence of an association between the Ala-9Val MnSOD polymorphism and HCC occurrence in hepatitis C virus-infected Moroccan patients.     

Defective ENaC processing and function in tissue kallikrein-deficient mice

An inverse relationship exists between urinary tissue kallikrein (TK) excretion and blood pressure in humans and rodents. In the kidney TK is synthesized in large amounts in the connecting tubule and is mainly released into the urinary fluid where its function remains unknown. In the present study mice with no functional gene coding for TK (TK-/-) were used to test whether the enzyme regulates apically expressed sodium transporters. Semiquantitative immunoblotting of the renal cortex revealed an absence of the 70-kDa form of gamma-ENaC in TK-/- mice. Urinary Na+ excretion after amiloride injection was blunted in TK-/- mice, consistent with reduced renal ENaC activity. Amiloride-sensitive transepithelial potential difference in the colon, where TK is also expressed, was decreased in TK-/- mice, whereas amiloride-sensitive alveolar fluid clearance in the lung, where TK is not expressed, was unchanged. In mice lacking the B2 receptor for kinins, the abundance of the 70-kDa form of gamma-ENaC was increased, indicating that its absence in TK-/- mice is not kinin-mediated. Incubation of membrane proteins from renal cortex of TK-/- mice with TK resulted in the appearance of the 70-kDa band of the gamma-ENaC, indicating that TK was able to promote gamma-ENaC cleavage in vitro. Finally, in mouse cortical collecting ducts isolated and microperfused in vitro, the addition of TK in the luminal fluid increased significantly intracellular Na+ concentration, consistent with an activation of the luminal entry of the cation. The results demonstrate that TK, like several other proteases, can activate ENaC in the kidney and the colon.     

Phenolic composition and biological activities of Tunisian Nigella sativa L. shoots and roots

In the present investigation, methanolic extracts from shoots and roots of Tunisian Nigella sativa were assayed for their antioxidant and antimutagenic activities. The phenolic composition of the methanolic extracts was determined by RP-HPLC. The predominant phenolic compound was vanillic acid with a mean concentration of 143.21 and 89.94 mg per 100 g dry weight of shoots and roots, respectively. Shoots and roots showed comparable and strong superoxide scavenger activity; however, shoots exhibited higher DPPH radical scavenging, reducing and chelating activities than roots. Mutagenic and antimutagenic activities were determined by using the Ames test. Shoots and roots demonstrated important antimutagenic effects. Roots exhibited stronger activity than shoots with an inhibition percentage of 71.32%.     

Identification of lactobacilli residing in chicken ceca with antagonism against Campylobacter

Bacteriocins produced by Lactobacillus salivarius have been recently recognized as a natural means to control Campylobacter and Salmonella in live poultry. This finding is of relevance since Campylobacter jejuni and Campylobacter coli are the predominant species isolated from poultry that are associated with human campylobacteriosis. In the present work, lactic acid bacteria (LAB) isolated from the cecum of twenty Tunisian chickens were identified and those isolates with antagonism against Campylobacter were further characterized. Following their preliminary confirmation as LAB, 150 strains were identified by combining morphological criteria, biochemical tests, and molecular methods, the latter inluding intergenic 16S- 23S PCR, specific lactobacilli PCR, and a biphasic approach. Most of the LAB isolated belonged to the genus Lactobacillus, among them Lb. sakei (33.3%), Lb. salivarius (19.4%), Lb. reuteri (8.6%), and Lb. curvatus (8.6%). The other LAB strains included those of the genus Weissella (16.7%), Enterococcus faecalis (5.3%), Leuconostoc mesenteroides (2.7%), Lactococcus graviae (2.7%), and Streptococcus sp. (2.7%). The Lactobacilli strains were tested for their antagonism against C. jejuni and C. coli. The activity of three of them, Lb. salivarius SMXD51, Lb. salivarius MMS122, and Lb. salivarius MMS151, against the aforementioned target strains could be ascribed to the production of bacteriocins.