The gene expression of caspasses is up-regulated during the signaling response of Aedes caspius against larvicidal bacteria

Our current knowledge on the key molecular mechanisms and cognate signaling transduction, by the δ-endotoxin-mediated mosquitoticidal effects, associated with exposure to Bacillus thuringiensis (Bt) and Bacillus sphaericus (Bs), is limited. Moreover, this observed mosquitocidal activity that is related to program cell death is largely unknown. Therefore, in an attempt to answer this question, the current study was primarily sought to provide evidence as to the molecular mechanism of mortality in Bt/Bs infected Aedes caspius mosquito larvae. Thus, the impact of Bt and Bs treatment on the expression of some selected apoptosis related caspase genes in A. caspius mosquito larvae was investigated, via quantitative reverse-transcriptase PCR (qRT-PCR). Mosquito larvae were collected from natural water niches. Larvae were grown to adult stage and were subsequently identified as A. caspius at Natural History Museum, London, UK. Remarkably, light and transmission electron microscopy studies of the midgut epithelial tissues revealed that both Bt and Bs brought about significant histopathological effects. Moreover, this treatment resulted in severe destruction at the sub-cellular organelle level for the mitochondria. Interestingly, qRT-PCR studies revealed that the treatment of A. caspius mosquito larvae with both Bt and Bs caused a significant up-regulation in the transcription level of all caspase genes under study, namely: CASPS17, CASPS18, CASPS19, CASPS20 and CASPS21. The results are discussed in the light of our current understanding of the signaling transduction pathway of apoptosis in insects and mosquitoes and the putative role of caspases gene expression in response to the treatment of A. caspius mosquito larvae with larvicidal bacteria.Keywords: Aedes caspius, Bacillus thuringenesis, Bacillus sphaericus, apoptosis, caspase, larvicidal bacteri

Thymoquinone inhibits growth of human medulloblastoma cells by inducing oxidative stress and caspase-dependent apoptosis while suppressing NF-jB signaling and IL-8 expression

Medulloblastoma (MB) is the most common malignant brain tumor of childhood. The transcription factor NF-κB is overexpressed in human MB and is a critical factor for MB tumor growth. NF-κB is known to regulate the expression of interleukin-8 (IL-8), the chemokine that enhances cancer cell growth and resistance to chemotherapy. We have recently shown that thymoquinone (TQ) suppresses growth of hepatocellular carcinoma cells in part by inhibiting NF-κB signaling. Here we sought to extend these studies in MB cells and show that TQ suppresses growth of MB cells in a dose- and time-dependent manner, causes G2M cell cycle arrest, and induces apoptosis. TQ significantly increased generation of reactive oxygen species (ROS), while pretreatment of MB cells with the ROS scavenger N-acetylcysteine (NAC) abrogated TQ-induced cell death and apoptosis, suggesting that TQ-induced cell death and apoptosis are oxidative stress-mediated. TQ inhibitory effects were associated with inhibition of NF-κB and altered expression of its downstream effectors IL-8 and its receptors, the anti-apoptotic Bcl-2, Bcl-xL, X-IAP, and FLIP, as well as the pro-apoptotic TRAIL-R1, caspase-8, caspase-9, Bcl-xS, and cytochrome c. TQ-triggered apoptosis was substantiated by up-regulation of the executioner caspase-3 and caspase-7, as well as cleavage of the death substrate poly(ADP-ribose)polymerase. Interestingly, pretreatment of MB cells with NAC or the pan-caspase inhibitor zVAD-fmk abrogated TQ-induced apoptosis, loss of cyclin B1 and NF-κB activity, suggesting that these TQ-mediated effects are oxidative stress- and caspase-dependent. These findings reveal that TQ induces both extrinsic and intrinsic pathways of apoptosis in MB cells, and suggest its potential usefulness in the treatment of MB

Origanum majorana L. polyphenols: in vivo antiepileptic effect, in silico evaluation of their bioavailability, and interaction with the NMDA receptor

Introduction: Epilepsy is a chronic brain disease characterized by repeated seizures and caused by excessive glutamate receptor activation. Many plants are traditionally used in the treatment of this disease. This study aimed to evaluate the bioavailability of a polyphenolic extract obtained from Origanum majorana L. (OMP) leaves, as well as its antiepileptic activity and its potential mechanism of action.Methods: We have developed and validated a simple, rapid, and accurate stability-indicating reversed-phase liquid chromatographic method for the simultaneous determination of caffeine and quercetin in rat plasma. The OMP antiepileptic effect was evaluated with pilocarpine-induced seizures, and a docking method was used to determine the possible interaction between caffeic acid and quercetin with the N-methyl-D-aspartate (NMDA) receptor.Results and Discussion: Both compounds tested showed low bioavailability in unchanged form. However, the tested extract showed an anticonvulsant effect due to the considerably delayed onset of seizures in the pilocarpine model at a dose of 100 mg/kg. The molecular docking proved a high-affinity interaction between the caffeic acid and quercetin with the NMDA receptor. Taken together, OLP polyphenols demonstrated good antiepileptic activity, probably due to the interaction of quercetin, caffeic acid, or their metabolites with the NMDA receptor

Regulation of the Fruit-Specific PEP Carboxylase SlPPC2 Promoter at Early Stages of Tomato Fruit Development

The SlPPC2 phosphoenolpyruvate carboxylase (PEPC; EC 4.1.1.31) gene from tomato (Solanum lycopersicum) is differentially and specifically expressed in expanding tissues of developing tomato fruit. We recently showed that a 1966 bp DNA fragment located upstream of the ATG codon of the SlPPC2 gene (GenBank AJ313434) confers appropriate fruit-specificity in transgenic tomato. In this study, we further investigated the regulation of the SlPPC2 promoter gene by analysing the SlPPC2 cis-regulating region fused to either the firefly luciferase (LUC) or the β-glucuronidase (GUS) reporter gene, using stable genetic transformation and biolistic transient expression assays in the fruit. Biolistic analyses of 5′ SlPPC2 promoter deletions fused to LUC in fruits at the 8th day after anthesis revealed that positive regulatory regions are mostly located in the distal region of the promoter. In addition, a 5′ UTR leader intron present in the 1966 bp fragment contributes to the proper temporal regulation of LUC activity during fruit development. Interestingly, the SlPPC2 promoter responds to hormones (ethylene) and metabolites (sugars) regulating fruit growth and metabolism. When tested by transient expression assays, the chimeric promoter:LUC fusion constructs allowed gene expression in both fruit and leaf, suggesting that integration into the chromatin is required for fruit-specificity. These results clearly demonstrate that SlPPC2 gene is under tight transcriptional regulation in the developing fruit and that its promoter can be employed to drive transgene expression specifically during the cell expansion stage of tomato fruit. Taken together, the SlPPC2 promoter offers great potential as a candidate for driving transgene expression specifically in developing tomato fruit from various tomato cultivars

In vivo and in silico studies of the effects of oil extracted from Cannabis sativa L. seeds on healing of burned skin wounds in rats

IntroductionThis study investigates the potential effects of cannabis seed oil (CSO) on the wound healing process. The aim was to assess the efficacy of CSO in treating skin wounds using an animal model and to explore its anti-inflammatory properties through in silico analysis.MethodsEighteen male albino Wistar rats, weighing between 200 and 250 g, were divided into three groups: an untreated negative control group, a group treated with the reference drug silver sulfadiazine (SSD) (0.01 g/mL), and a group treated topically with CSO (0.962 g/mL). The initial wound diameter for all groups was 1 cm. In silico studies were conducted using Maestro 11.5 to evaluate the anti-inflammatory effects of phytoconstituents against cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2).ResultsCSO and SSD treatments led to a significant reduction (p <0.05) in the size of burned skin wounds by day 5, with contraction rates of 53.95% and 45.94%, respectively, compared to the untreated negative control group. By day 15, wounds treated with CSO and SSD had nearly healed, showing contraction rates of 98.8% and 98.15%, respectively. By day 20, the wounds treated with CSO had fully healed (100%), while those treated with SSD had almost completely healed, with a contraction rate of 98.97%. Histological examination revealed granulated tissue, neo-blood vessels, fibroblasts, and collagen fibers in wounds treated with CSO. In silico studies identified arachidic acid, γ-linolenic acid, and linolenic acid as potent inhibitors of COX-1 and COX-2. Serum biochemical parameters indicated no significant changes (p > 0.05) in liver and kidney function in rats treated with CSO, whereas a significant increase (p < 0.01) in ALAT level was observed in rats treated with SSD.DiscussionThe findings demonstrate that CSO has a promising effect on wound healing. The CSO treatment resulted in significant wound contraction and histological improvements, with no adverse effects on liver and kidney function.However, the study's limitations, including the small sample size and the need for detailed elucidation of CSO's mechanism of action, suggest that further research is necessary. Future studies should focus on exploring the molecular pathways and signaling processes involved in CSO’s pharmacological effects

The Effect of Walterinnesia aegyptia Venom Proteins on TCA Cycle Activity and Mitochondrial NAD+-Redox State in Cultured Human Fibroblasts

Fibroblast cultures were used to study the effects of crude Walterinnesia aegyptia venom and its F1–F7 protein fractions on TCA cycle enzyme activities and mitochondrial NAD-redox state. Confluent cells were incubated with 10 μg of venom proteins for 4 hours at 37°C. The activities of all studied TCA enzymes and the non-TCA mitochondrial NADP+-dependent isocitrate dehydrogenase underwent significant reductions of similar magnitude (50–60% of control activity) upon incubation of cells with the crude venom and fractions F4, F5, and F7 and 60–70% for fractions F3 and F6. In addition, the crude and fractions F3–F7 venom proteins caused a drop in mitochondrial NAD+ and NADP+ levels equivalent to around 25% of control values. Whereas the crude and fractions F4, F5, and F7 venom proteins caused similar magnitude drops in NADH and NADPH (around 55% of control levels), fractions F3 and F6 caused a more drastic drop (60–70% of control levels) of both reduced coenzymes. Results indicate that the effects of venom proteins could be directed at the mitochondrial level and/or the rates of NAD+ and NADP+ biosynthesis

Expression profiles of <i>SlTIFY</i> genes in response to jasmonic and abscisic acid.

<p>Three-week-old tomato seedlings were subjected to exogenous JA (B) or ABA (B) treatments for different times; roots and leaves were collected and analyzed separately. Relative expression levels of the <i>SlTIFY</i> genes were analyzed by real-time quantitative RT-PCR (qPCR), and log10-transformed fold-change values were used for creating the heatmap (original data were shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0177381#pone.0177381.s009" target="_blank">S3 Table</a>). The colour-based gene expression scale is shown at the bottom of each panel.</p

Expression profiles of <i>SlTIFY</i> genes under abiotic stress conditions.

<p>Expression profile of <i>SlTIFY</i> genes in response to salinity (A) orosmotic stresses (B). Three-week-old tomato plants were subjected to salt or osmotic stresses; roots and leaves were collected at different times and analyzed separately. Relative expression levels of the <i>SlTIFY</i> genes were analyzed by real-time quantitative RT-PCR (qPCR), and log10-transformed fold-change values were used for creating the heatmap (original data in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0177381#pone.0177381.s009" target="_blank">S3 Table</a>). The colour-based gene expression scale is included.</p