Phylogenetic analysis of mtCR-1 sequences of Tunisian and Egyptian hares (Lepus sp. or spp., Lagomorpha) with different coat colours

North African hares are currently considered belonging to cape hares (Lepus capensis), except for an isolated occurrence of L. victoriae in NW Algeria. However, the few existing molecular data are not unequivocal. Here, we study sequence variation (415 bp) in the hypervariable domain-1 of the mitochondrial (mt) control region, of hares with different coat colour from north-central Tunisia and NW Egypt, to test Petter's [(1959): Eléments d’une révision des Lièvres africains du sous-genre Lepus. Mammalia 23, 41–67] hypothesis that North African hares belong to L. capensis. Seven Tunisian and one Egyptian haplotypes were revealed from 28 hares and compared phylogenetically to 245 haplotypes of various Lepus species downloaded from GenBank. Neighbour joining (NJ) and principal coordinate (PCO) analyses based on a Tamura-Nei 93 distance matrix, as well as maximum parsimony (MP) analysis concordantly grouped all currently obtained haplotypes together into one monophyletic clade, and revealed relatively close relationships to the clades of African scrub hares (L. saxatilis) and brown hares (L. europaeus). The three distinguished coat colour types of Tunisian hares were paralleled only to a small extent by sequence differentiation. Haplotypes of L. capensis from the nominal Cape province of South Africa, North Africa, and China clustered into different major clades, respectively, with Chinese L. capensis haplotypes forming only a subclade within a major clade that encompassed predominantly “mountain/arctic hare-type sequences” in addition to sequences of several other palaearctic and nearctic species. One further Chinese L. capensis haplotype clustered into the L. comus clade. These results indicated occurrence of introgression and/or shared ancestral polymorphism. Such an evolutionary scenario implies using nucelar markers in addition to mtDNA for phylogenetic inferences in the genus Lepus; nevertheless, mtDNA is still useful for inferring phylogenetic history and biogeography of hares.     

Date seed oil limit oxidative injuries induced by hydrogen peroxide in human skin organ culture

The skin is chronically exposed to pro-oxidant agents, leading to the generation of reactive oxygen species (ROS). To protect the skin against an over-load of oxidant species, we studied the chemoprotective effect of one new natural product: "date seed oil: DSO". This oil may serve as a potential source of natural antioxidants such as phenols and tocopherols. Here, the antioxidative potential of DSO was compared that of to extra virgin olive oil. Adult human skin was maintained in organ culture in the presence of the DSO and extra virgin olive oil before the addition of hydrogen peroxide (H2O2), in order to prevent the tissue from its oxidizing effects. Skin specimens were collected for histology and for melanin studies. In the investigated model system, DSO protects skin against oxidative injuries. It has a significant chemoprotective effect, by inhibition of damage caused by H_{2}O_{2} compared with specimens without such addition endowing with a radical scavenging ability. The various components from DSO were much more potent antioxidant and more free radical scavengers of the H2O2 than those of olive oil. Our study shows that topical DSO treatment of the skin stimulates events in the epidermis leading to repair skin damage possibly due to antioxidant synergisms.     

Residues in the alternative reading frame tumor suppressor that influence its stability and p53-independent activities

The Alternative Reading Frame (ARF) protein suppresses tumorigenesis through p53-dependent and p53-independent pathways. Most of ARF's anti-proliferative activity is conferred by sequences in its first exon. Previous work showed specific amino acid changes occurred in that region during primate evolution, so we programmed those changes into human p14ARF to assay their functional impact. Two human p14ARF residues (Ala¹⁴ and Thr³¹) were found to destabilize the protein while two others (Val²⁴ and Ala⁴¹) promoted more efficient p53 stabilization and activation. Despite those effects, all modified p14ARF forms displayed robust p53-dependent anti-proliferative activity demonstrating there are no significant biological differences in p53-mediated growth suppression associated with simian versus human p14ARF residues. In contrast, p53-independent p14ARF function was considerably altered by several residue changes. Val²⁴ was required for p53-independent growth suppression whereas multiple residues (Val²⁴, Thr³¹, Ala⁴¹ and His⁶⁰) enabled p14ARF to block or reverse the inherent chromosomal instability of p53-null MEFs. Together, these data pinpoint specific residues outside of established p14ARF functional domains that influence its expression and signaling activities. Most intriguingly, this work reveals a novel and direct role for p14ARF in the p53-independent maintenance of genomic stability.     

Association of GSTM1 and GSTT1 Polymorphisms with Chronic Obstructive Pulmonary Disease in a Tunisian Population

GSTM1 and GSTT1 polymorphisms have been proposed in relationship with chronic obstructive pulmonary disease (COPD). We investigated the association between these polymorphisms and COPD (as well as its subtypes emphysema and chronic bronchitis) in 234 COPD patients and 182 healthy controls in the Tunisian population. Genotyping was performed using multiplex PCR. GSTM1-null genotype frequency was significantly higher in COPD patients than in controls (P = 0.02); however, multivariate analysis of cofounding variables showed no independent association with this genotype (P = 0.073). In contrast, the association of the GSTM1-null genotype with emphysema was significant, even after adjustment for risk factors (P = 0.011). There were no significant differences in GSTT1 genotypes between patients and controls. The GSTM1 null allele is likely not an independent risk factor for COPD but is related to emphysema, whereas the GSTT1 gene is not associated with the disease.     

Distinct roles of beta1 integrins during angiogenesis

Integrins are transmembrane receptors which bind extracellular matrix proteins and enable not only cell adhesion and cytoskeleton organization but also transduction of critical signals into the cells to promote survival, proliferation, differentiation, or migration programs. Integrins participate in many aspects of vascular biology. The past few years have experienced a sustained interest in the implication of integrin receptors in tumor angiogenesis. We will focus our review on studies giving concrete evidence to a role of the beta1 class of integrins in angiogenesis, and we will provide an overview of the molecular mechanisms involved in their action.     

The epigenetic regulator BRD4 is required for myofibroblast differentiation of knee fibroblasts

Arthrofibrosis, which is characterized by excessive scar tissue and limited motion, can complicate the daily functioning of patients after total knee arthroplasty (TKA). Molecular hallmarks of arthrofibrosis include pathologic accumulation of myofibroblasts and disproportionate collagen deposition. Epigenetic mechanisms, including posttranslation modification of histones, control gene expression and may regulate fibrotic events. This study assessed the role of the bromodomain and extra-terminal (BET) proteins on myofibroblast differentiation. This group of epigenetic regulators recognize acetylated lysines and are targeted by a class of drugs known as BET inhibitors. RNA-seq analysis revealed robust mRNA expression of three BET members (BRD2, BRD3, and BRD4) while the fourth member (BRDT) is not expressed in primary TKA knee outgrowth fibroblasts. RT-qPCR and western blot analyses revealed that BET inhibition with the small molecule JQ1 impairs TGFβ1-induced expression of ACTA2, a key myofibroblast marker, in primary outgrowth knee fibroblasts. Similarly, JQ1 administration also reduced COL3A1 mRNA levels and collagen deposition as monitored by picrosirius red staining. Interestingly, the inhibitory effects of JQ1 on ACTA2 mRNA and protein expression, as well as COL3A1 expression and collagen deposition, were paralleled by siRNA-mediated depletion of BRD4. Together, these data reveal that BRD4-mediated epigenetic events support TGFβ1-mediated myofibroblast differentiation and collagen deposition as seen in arthrofibrosis. To our knowledge, these are the first studies that assess epigenetic regulators and their downstream events in the context of arthrofibrosis. Future studies may reveal clinical utility for drugs that target epigenetic pathways, specifically BET proteins, in the prevention and treatment of arthrofibrosis.     

Cis ε-viniferin: A New Antifungal Resveratrol Dehydrodimer from Cyphostemma crotalarioides Roots

Six antifungal constituents were isolated from the roots of Cyphostemma crotalarioides (Vitaceae): a new product, Cis‐ε‐viniferin 3, along with five known compounds: trans‐resveratrol I and its oligomers: trans ε‐viniferin 2, gnetin C 4, pallidol 5 and gnetin E 6. It is the first time such compounds have been reported in this plant species. The roots tissues were found to be rich in other resveratrol oligomers, however, the available amounts of the other purified products did not allow structure investigations. No fungicidal activity was detected in the leaves of this plant.     

Sequential events of apoptosis induced by zearalenone in cultured hepatocarcinoma cells

Zearalenone (ZEA) is a fungal metabolite that can contaminate feed and foodstuffs and can cause serious health problems for animals as well as for humans. The present investigation was conducted to determine the chronological succession of the main events that characterise ZEA-induced toxicity in human hepatocarcinoma cells. To this aim, we have monitored the effects of ZEA on (1) cell viability, (2) heat-shock protein expression, (3) oxidative damage, (4) DNA fragmentation, (5) the cell cycle and (6) the cell-death-signalling pathway. Our results demonstrated that ZEA reduced cell viability in a time- and dose-dependent manner. When we exposed HepG2 cells to 100 μM ZEA (80% of cells are viable) for different treatment times (2, 4, 8, 24, 30, 48 and 60 h), we demonstrated an induction of Hsp70 protein, an increase in reactive oxygen species (ROS) generation, DNA fragmentation and cell-cycle arrest. These events begin after only 2 h of mycotoxin exposure and are earlier than those implicated in the execution of apoptosis. However, significant apoptotic cell death was observed after at least 30 h of ZEA exposure as a consequence of increased Bax expression, decreased Bcl-2 expression and mitochondrial membrane potential (Δψm)-released cytochrome c and activated caspase-3 and caspase-9.     

Exploring the binding pocket for pyridopyrimidine ligands at the CCK1 receptor by molecular docking

Pyridopyrimidine-based analogues are among the most highly potent and selective antagonists of cholecystokinin receptor subtype-1 (CCK1R) described to date. To better understand the structural and chemical features responsible for the recognition mechanism, and to explore the binding pocket of these compounds, we performed automated molecular docking using GOLD2.2 software on some derivatives with structural diversity, and propose a putative binding conformation for each compound. The docking protocol was guided by the key role of the Asn333 residue, as revealed by site directed mutagenesis studies. The results suggest two putative binding modes located in the same pocket. Both are characterized by interaction with the main residues revealed by experiment, Asn333 and Arg336, and differ in the spatial position of the Boc-Trp moiety of these compounds. Hydrophobic contacts with residues Thr117, Phe107, Ile352 and Ile329 are also in agreement with experimental data. Despite the poor correlation obtained between the estimated binding energies and the experimental activity, the proposed models allow us to suggest a plausible explanation of the observed binding data in accordance with chemical characteristics of the compounds, and also to explain the observed diastereoselectivity of this family of antagonists towards CCK1R. The most reasonable selected binding conformations could be the starting point for future studies. Figure Superimposition of the two putative binding conformations revealed by molecular docking for pyridopyrimidine-based CCK1 antagonists