Protective role of alpha-tocopherol-succinate (provitamin-E) in cyclophosphamide induced testicular gametogenic and steroidogenic disorders: a correlative approach to oxidative stress

The present study was undertaken to find out the adverse effects of cyclophosphamide on testicular activities along with testicular oxidative stress at its therapeutic dose and the protective effects of alpha-tocopherol succinate on testicular dysfunctions induced by cyclophosphamide in mature albino rats. A significant diminution in the activities of testicular delta 5, 3 beta-hydroxysteroid dehydrogenase (HSD) and 17 beta-hydroxysteroid dehydrogenase (HSD) along with significant reduction in the plasma level of testosterone and number of spermatogonia-A (ASg), preleptotene spermatocytes (pLSc), midpachytene spermatocytes (mPSc) and step 7 spermatids (7Sd) at stage VII of spermatogenic cycle were observed following cyclophosphamide treatment. Oxidative stress was also noted in testis, which was enlightened by significant elevation in the level of malondialdehyde (MDA) and conjugated dienes along with significant reduction in the activities of testicular peroxidase and catalase. Co-administration of alpha-tocopherol succinate in cyclophosphamide-treated rats resulted a significant restoration of all the above-mentioned parameters to the control level. The results of our experiment suggest that cyclophosphamide treatment at its clinical dose is associated with antigonadal activities as well as induction of oxidative stress in gonad that can be ameliorated significantly by alpha-tocopherol succinate co-administration. So, our data have some potential clinical implications.     

Smad3 and Snail show circadian expression in human gingival fibroblasts, human mesenchymal stem cell, and in mouse liver

Smads are intracellular signaling mediators. Complexes of Smad2 and Smad3 with Smad4 transmit transforming growth factor-beta (TGF-β) receptor-induced signaling. Snail plays important roles in mesoderm formation, gastrulation, neural crest development, and epithelial mesenchymal transition. However, it remains unknown whether Smad3 and Snail expression is circadian rhythm-dependent. Here, we showed for the first time that Smad3 and Snail show circadian expression in human gingival fibroblasts (HGF-1) and human mesenchymal stem cells (MSC) after serum shock. They also showed circadian expression in the mouse liver. We confirmed that BMAL1/2, DEC1/2, VEGF, and PER1/2/3 also show circadian expression in both HGF-1 and MSC. The mRNA peaks and phases in circadian expression of these genes differed between HGF-1 and MSC. In a luciferase assay, Smad3 promoter activity was upregulated by CLOCK/BMAL1. These findings suggest that Smad3 and Snail have circadian rhythm in vitro and vivo, and that circadian expression of Smad3 depends on CLOCK/BMAL1.     

Expression Profiling of the <Emphasis Type="Italic">CSDP</Emphasis> Transcription Factor Gene Family Points to Roles in Organ Development and Abiotic Stress Response in Tomato (<Emphasis Type="Italic">Solanum lycopersicum</Emphasis> L.)

The cold shock domain proteins (CSDPs) are small group of nucleic acid-binding proteins that act as RNA chaperones in growth regulation, development, and stress adaptation in plants. The functions of CSDPs have been studied in Arabidopsis (Arabidopsis thaliana), rice (Oryza sativa), wheat (Triticum aestivum), and Chinese cabbage (Brassica rapa). To gain insight into the function of CSDPs in tomato (Solanum lycopersicum), we performed a genome-wide analysis of CSDPs through in silico characterization and expression profiling in different organs and in response to different abiotic stress and phytohormone treatments. We identified five non-redundant SlCSDP genes. The evolutionary analysis and phylogenetic classification indicated that tomato CSDPs are more closely related to potato than those of others. The five SlCSDP genes are distributed on four of the 12 tomato chromosomes and no segmental or tandem duplication events are detected among them. Expression analysis showed broad expression patterns with strong expression in fruit development and ripening. Expression of individual SlCSDP genes was significantly altered by stress and phytohormone treatments. SlCSDP2, SlCSDP3, and SlCSDP4 were highly induced by all four abiotic stresses and by phytohormone treatment in tomato. These findings provide a foundation for future research towards functional biological roles of CSDP gene in particular to develop tomato cultivars with large size, early ripening, and abiotic stress tolerance.     

Ferulic acid (FA) abrogates γ-radiation induced oxidative stress and DNA damage by up-regulating nuclear translocation of Nrf2 and activation of NHEJ pathway

The present study was aimed to evaluate the radioprotective effect of ferulic acid (FA), a naturally occurring plant flavonoid in terms of DNA damage and damage related alterations of repair pathways by gamma radiation. FA was administered at a dose of 50?mg/kg body weight for five consecutive days prior to exposing the swiss albino mice to a single dose of 10?Gy gamma radiation. Ionising radiation induces oxidative damage manifested by decreased expression of Cu, Zn-SOD (SOD stands for super oxide dismutase), Mn-SOD and catalase. Gamma radiation promulgated reactive oxygen species (ROS) mediated DNA damage and modified repair pathways. ROS enhanced nuclear translocation of p53, activated ATM (ataxia telangiectasia-mutated protein), increased expression of GADD45a (growth arrest and DNA-damage-inducible protein) gene and inactivated Non homologous end joining (NHEJ) repair pathway. The comet formation in irradiated mice peripheral blood mononuclear cells (PBMC) reiterated the DNA damage in IR exposed groups. FA pretreatment significantly prevented the comet formation and regulated the nuclear translocation of p53, inhibited ATM activation and expression of GADD45a gene. FA promoted the nuclear translocation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and activated NHEJ repair pathway to overcome ROS mediated oxidative stress and DNA damage. Therefore, the current study stated that FA can challenge the oxidative stress by (i) inducing nuclear translocation of Nrf2, (ii) scavenging ROS, and (iii) activating NHEJ DNA repair process.     

Trypanosome Alternative Oxidase Possesses both an N-Terminal and Internal Mitochondrial Targeting Signal

Recognition of mitochondrial targeting signals (MTS) by receptor translocases of outer and inner membranes of mitochondria is one of the prerequisites for import of nucleus-encoded proteins into this organelle. The MTS for a majority of trypanosomatid mitochondrial proteins have not been well defined. Here we analyzed the targeting signal for trypanosome alternative oxidase (TAO), which functions as the sole terminal oxidase in the infective form of Trypanosoma brucei. Deleting the first 10 of 24 amino acids predicted to be the classical N-terminal MTS of TAO did not affect its import into mitochondria in vitro. Furthermore, ectopically expressed TAO was targeted to mitochondria in both forms of the parasite even after deletion of first 40 amino acid residues. However, deletion of more than 20 amino acid residues from the N terminus reduced the efficiency of import. These data suggest that besides an N-terminal MTS, TAO possesses an internal mitochondrial targeting signal. In addition, both the N-terminal MTS and the mature TAO protein were able to target a cytosolic protein, dihydrofolate reductase (DHFR), to a T. brucei mitochondrion. Further analysis identified a cryptic internal MTS of TAO, located within amino acid residues 115 to 146, which was fully capable of targeting DHFR to mitochondria. The internal signal was more efficient than the N-terminal MTS for import of this heterologous protein. Together, these results show that TAO possesses a cleavable N-terminal MTS as well as an internal MTS and that these signals act together for efficient import of TAO into mitochondria.     

Cytotoxic and Pathogenic Properties of Klebsiella oxytoca Isolated from Laboratory Animals

Klebsiella oxytoca is an opportunistic pathogen implicated in various clinical diseases in animals and humans. Studies suggest that in humans K. oxytoca exerts its pathogenicity in part through a cytotoxin. However, cytotoxin production in animal isolates of K. oxytoca and its pathogenic properties have not been characterized. Furthermore, neither the identity of the toxin nor a complete repertoire of genes involved in K. oxytoca pathogenesis have been fully elucidated. Here, we showed that several animal isolates of K. oxytoca, including the clinical isolates, produced secreted products in bacterial culture supernatant that display cytotoxicity on HEp-2 and HeLa cells, indicating the ability to produce cytotoxin. Cytotoxin production appears to be regulated by the environment, and soy based product was found to have a strong toxin induction property. The toxin was identified, by liquid chromatography-mass spectrometry and NMR spectroscopy, as low molecular weight heat labile benzodiazepine, tilivalline, previously shown to cause cytotoxicity in several cell lines, including mouse L1210 leukemic cells. Genome sequencing and analyses of a cytotoxin positive K. oxytoca strain isolated from an abscess of a mouse, identified genes previously shown to promote pathogenesis in other enteric bacterial pathogens including ecotin, several genes encoding for type IV and type VI secretion systems, and proteins that show sequence similarity to known bacterial toxins including cholera toxin. To our knowledge, these results demonstrate for the first time, that animal isolates of K. oxytoca, produces a cytotoxin, and that cytotoxin production is under strict environmental regulation. We also confirmed tilivalline as the cytotoxin present in animal K. oxytoca strains. These findings, along with the discovery of a repertoire of genes with virulence potential, provide important insights into the pathogenesis of K. oxytoca. As a novel diagnostic tool, tilivalline may serve as a biomarker for K oxytoca-induced cytotoxicity in humans and animals through detection in various samples from food to diseased samples using LC-MS/MS. Induction of K. oxytoca cytotoxin by consumption of soy may be in part involved in the pathogenesis of gastrointestinal disease.