A functional SNP in miR-146a and genetic susceptibility to drug-resistant epilepsy.

OBJECTIVES: The study aimed to assess the relationship between the functional single nucleotide polymorphism (SNP) (rs57095329) of miR-146a, the progression of drug-resistant epilepsy (DRE), and the severity of the disease (seizure frequency) in a group of Egyptian children epilepsy patients. SUBJECTS AND METHODS: 110 Egyptian children were recruited and divided into two groups, the epilepsy patients (n = 60) and the healthy control children (n = 50). The patient's group was equally subdivided into two subgroups: drug-resistant and drug-responsive epilepsy patients. Genomic DNA samples from all participants were screened for the incidence of the rs57095329 SNP of the miR-146a gene by the Real-Time PCR. RESULTS: There was no statistical significance between epilepsy patients compared to controls regarding the rs57095329 SNP genotypes and alleles. Contrarily, there was significant difference between the drug-resistant epilepsy and the drug-responsive cases (P < 0.05). The genotypes AG (P < 0.007, OR: 0.118, 95% CI (0.022-0.636)) and GG (P = 0.016, OR: 0.123, 95% CI (0.023-0.769)) were higher among the drug-resistant, while AA was higher among the drug-responsive patients. The alleles A and G were higher among all cases, with a statistically significant difference (P = 0.028, OR: 0.441, 95% CI (0.211-0.919)). A significant difference was reported in the dominant model (AA versus AG+GG) (P = 0.005, OR: 0.12395% CI (0.025-0.621)). CONCLUSION: Therefore, miR-146a might be a potential therapeutic target for epilepsy treatment. The study was limited by the low number of young epileptic patients, the refusal of some parents to participate, and the incomplete medical history of some cases in the study, which forced their exclusion. More studies might be necessary to investigate other effective drugs to overcome the resistance issues induced by miR-146a rs57095329 polymorphisms.

Purification and Characterization of Ornithine Decarboxylase from Aspergillus terreus; Kinetics of Inhibition by Various Inhibitors.

l-Ornithine decarboxylase (ODC) is the rate-limiting enzyme of de novo polyamine synthesis in humans and fungi. Elevated levels of polyamine by over-induction of ODC activity in response to tumor-promoting factors has been frequently reported. Since ODC from fungi and human have the same molecular properties and regulatory mechanisms, thus, fungal ODC has been used as model enzyme in the preliminary studies. Thus, the aim of this work was to purify ODC from fungi, and assess its kinetics of inhibition towards various compounds. Forty fungal isolates were screened for ODC production, twenty fungal isolates have the higher potency to grow on L-ornithine as sole nitrogen source. Aspergillus terreus was the most potent ODC producer (2.1 µmol/mg/min), followed by Penicillium crustosum and Fusarium fujikuori. These isolates were molecularly identified based on their ITS sequences, which have been deposited in the NCBI database under accession numbers MH156195, MH155304 and MH152411, respectively. ODC was purified and characterized from A. terreus using SDS-PAGE, showing a whole molecule mass of ~110 kDa and a 50 kDa subunit structure revealing its homodimeric identity. The enzyme had a maximum activity at 37 °C, pH 7.4-7.8 and thermal stability for 20 h at 37 °C, and 90 days storage stability at 4 °C. A. terreus ODC had a maximum affinity (Km) for l-ornithine, l-lysine and l-arginine (0.95, 1.34 and 1.4 mM) and catalytic efficiency (kcat/Km) (4.6, 2.83, 2.46 × 10-5 mM-1·s-1). The enzyme activity was strongly inhibited by DFMO (0.02 µg/mL), curcumin (IC50 0.04 µg/mL), propargylglycine (20.9 µg/mL) and hydroxylamine (32.9 µg/mL). These results emphasize the strong inhibitory effect of curcumin on ODC activity and subsequent polyamine synthesis. Further molecular dynamic studies to elucidate the mechanistics of ODC inhibition by curcumin are ongoing.