A molecular study of Tunisian populations of Dugesia sicula (Plathelminthes, Tricladida) through an identification of a set of genes.

Cell regeneration is a natural repair of different types of tissue after an injury or a lesion, and is associated with asexual reproduction in some animals such as planarians. Its understanding and improvement could have repercussions for tissue repair and regeneration as far as humans are concerned. In this context, we have proceeded to an essential step, which is the identification of the genes involved in planarian regeneration in the model species. Dugesia sicula Lepori (D. sicula) is distributed around the Mediterranean Sea, and this population is found in most of Tunisian dams. The collection of identified genes is already known in other species. DjFoxG, DjPC2, DjotxA, and Cathepsin-D were identified by the PCR technique and their expression was confirmed by RT-PCR and in situ hybridization. DjFoxG gene, the FoxG1 homolog, is expressed throughout the planarian body, abundantly on stem cells. Consecutively, we choose a central nervous system (CNS) marker; the prohormone convertase 2 (DjPC2) gene. DjotxA was observed in the brain and especially in the region surrounding the eyes (visual cells). The regenerative cells of the gut of D. sicula were scored by the Cathepsin-D gene expression, which belongs to the aspartyl protease family. We found significant results through RT-PCR and In Situ Hybridization (ISH) techniques, confirming the expression of DjFoxG, DjPC2, DjotxA and Cathepsin-D genes in our specimens.

Biochemical and Molecular Study of Carpobrotus edulis Bioactive Properties and Their Effects on Dugesia sicula (Turbellaria, Tricladida) Regeneration.

The traditional medicinal properties of Carpobrotus edulis are well recognized, particularly in Tunisia where it is used for wound healing. Thus, in this study, biochemical and molecular properties of its leaves' bioactive aqueous-acetone extract were investigated. The total phenolic content (TPC) of the extract was estimated to be 184 ± 5 mg/100 g of fresh matter (FM). The qualitative and quantitative polyphenolic profile was determined by ultra performance liquid chromatography with diode array detection (UPLC-DAD) and showed that chlorogenic acid was the major compound (43.7%). The extract exhibits potent antioxidant capacities with IC50 = 56.19 and 58.91 µg/ml, as accessed via the anionic DPPH and cationic ABTS radical scavenging assays, respectively. The extract has high antibacterial properties, especially against the Gram+ Staphylococcus aureus and Bacillus cereus strains. To investigate the extract effect on regeneration, the flatworm Dugesia sicula Lepori, 1948, was used as a model. The macroscopic analysis of planarian cultures in ordinary medium containing phenolic extract at non-toxic concentrations illustrated that the extract caused morphological changes. Additionally, the molecular study through the fluorescence-activated cell sorting (FACS) technique showed that C. edulis polyphenols can harm the stem cells' development. These results emphasize the ecotoxicological impact of phenolic rejections in the environment on flatworms' physiology.

Pomegranate and mint syrup addition to green tea beverage stabilized its polyphenolic content and biofunctional potentials during refrigerated storage.

The chemical stability of the green tea (GT) preparation during refrigerated storage was investigated following the addition of mint (MS) or pomegranate (PS) syrups, a common habit in the Mediterranean countries that improves the savor of this popular beverage. The supernatants recovered by centrifuging GT supplemented or not with mint (GTMS) or pomegranate (GTPS) syrup were examined for their polyphenolic profiles using the high performance liquid chromatography with diode array detection and electrospray ionization-mass spectrometry. Following storage at 4 °C for 15 days, not-supplemented GT showed a significant decrease (≈92 %) of its phenolic content. However, the decrease was relatively lesser in GTPS (≈36 %) and in GTMS (≈40 %). The observed slight increase of the extractable polyphenolics in PS and MS during the storage might explain in part the relatively limited decrease of GTPS and GTMS total phenolic content. However, chromatographic examination proved that some tea compounds, particularly caffeine, were preserved following PS and MS supplementation. Likewise, syrups'addition to GT significantly (P < 0.5) limited the reduction of its antioxidant capacity as revealed by the DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,2'-azino-bis-(3-ethylbenz-thialzoline-6-sulfonic acid)) assays. As expected, the antimicrobial trials showed that Gram (+) Staphylococcus aureus and Staphylococcus epidermidis were the most sensitive strains to tea polyphenols. The syrups supplementation noticeably preserved the tea bacteriostatic and bactericide activities during storage. The obtained analytical results demonstrate that MS or PS addition to green tea beverage stabilized its polyphenolic content and biofunctional properties during refrigerated storage, thus, scientifically supporting this popular practice in the Mediterranean countries.

Use of molecular and in silico bioinformatic tools to investigate pesticide binding to insect (Lepidoptera) phenoloxidases (PO): insights to toxicological aspects.

In the present work, the promising bioinformatic tools, based on structure-affinity analysis, allowed to screen several pesticides supposed to bind to the insect immune Phenoloxidases (PO). First, the binding of aminoparathion, a reference compound, to the PO was structurally (3D) validated in accordance with previous reports. Second, using the same docking conditions, a range of pesticides was checked for their binding ability to the crystal 3D structure (PDB 3HSS) of the insect Manduca sexta (Lepidoptera) PO. The obtained data showed that many of the tested pesticides were able to bind, in silico, to M. sexta PO. The combination of in vitro (chemical and biochemical) and in silico (automated docking) approaches was found advantageous to elucidate the behavior of phenolic pesticides as substrate-analogues when binding to the active site of insect POs. Our findings emphasize new ecotoxicological aspects of pesticide residues in the agro-chemical and environmental circles.

Cell viability effects and antioxidant and antimicrobial activities of Tunisian date syrup (Rub El Tamer) polyphenolic extracts.

The aqueous-acetone polyphenolic extract of the traditionally derived date syrup, known as "Rub El Tamer", was analyzed using RP-HPLC-DAD and ESI-MS. The phenolic content of the extract was 394.53 ± 1.13 mg per 100 g of syrup with caffeoylsinapylquinic acid as the most abundant compound (72.23%). The extract exhibited strong antioxidant activities as evaluated using the ABTS (2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)), DPPH (2,2-diphenyl-1-picrylhydrazyl) and FRAP (ferric reducing antioxidant power) methods. The extract antimicrobial potential against a range of microorganism strains showed that Staphylococcus aureus, Staphylococcus epidermidis, and Bacillus cereus were the most sensitive bacteria with MBC in the range of 0.5-0.05 mg/mL. Furthermore, in the presence of the syrup extract (8.18-131 µg/mL), the Human SH-SY5Y neuroblastoma and the 3T3 fibroblast cell lines showed dissimilar reduction of viability suggesting a higher cytotoxic effect against tumorigenic cells. Our results provide new insights into date syrup characterization which should stimulate further studies of this hot desert resource.