Phyllanthus urinaria extract attenuates acetaminophen induced hepatotoxicity: Involvement of cytochrome P450 CYP2E1

Acetaminophen is a commonly used drug for the treatment of patients with common cold and influenza. However, an overdose of acetaminophen may be fatal. In this study we investigated whether mice, administered intraperitoneally with a lethal dose of acetaminophen, when followed by oral administration of Phyllanthus urinaria extract, may be prevented from death. Histopathological analysis of mouse liver sections showed that Phyllanthus urinaria extract may protect the hepatocytes from acetaminophen-induced necrosis. Therapeutic dose of Phyllanthus urinaria extract did not show any toxicological phenomenon on mice. Immunohistochemical staining with the cytochrome P450 CYP2E1 antibody revealed that Phyllanthus urinaria extract reduced the cytochrome P450 CYP2E1 protein level in mice pre-treated with a lethal dose of acetaminophen. Phyllanthus urinaria extract also inhibited the cytochrome P450 CYP2E1 enzymatic activity in vitro. Heavy metals, including arsenic, cadmium, mercury and lead, as well as herbicide residues were not found above their detection limits. High performance liquid chromatography identified corilagin and gallic acid as the major components of the Phyllanthus urinaria extract. We conclude that Phyllanthus urinaria extract is effective in attenuating the acetaminophen induced hepatotoxicity, and inhibition of cytochrome P450 CYP2E1 enzyme may be an important factor for its therapeutic mechanism.     

Sequencing of Cladosporium sphaerospermum, a Dematiaceous fungus isolated from blood culture

Cladosporium sphaerospermum is one of the most widely distributed allergens causing serious problems in patients with respiratory tract disease. We report the 26,644,473-bp draft genome sequence and gene annotation of C. sphaerospermum UM843. Analysis of the genome sequence led to the finding of genes associated with C. sphaerospermum's melanin biosynthesis, allergens, and antifungal drug resistance.     

Draft genome sequence of Daldinia eschscholzii isolated from blood culture

Daldinia eschscholzii is an invasive endophyte that is most commonly found in plant tissues rich in secondary metabolites. We report the draft genome sequence of D. eschscholzii isolated from blood culture. The draft genome is 35,494,957 bp in length, with 42,898,665 reads, 61,449 contigs, and a G+C content of 46.8%. The genome was found to contain a high abundance of genes associated with plant cell wall degradation enzymes, mycotoxin production, and antifungal drug resistance.     

Genome sequence of an unclassified pleosporales species isolated from human nasopharyngeal aspirate

Pleosporales is the largest order in the fungal class Dothideomycetes. We report the 36,814,818-bp draft genome sequence and gene annotation of UM1110, a Pleosporales isolate associated with unclassified genera that is potentially a new fungal species. Analysis of the genome sequence led to the finding of genes associated with fungal adhesive proteins, secreted proteases, allergens, and pseudohyphal development.     

Knockdown of the fat mass and obesity gene disrupts cellular energy balance in a cell-type specific manner

Recent studies suggest that FTO variants strongly correlate with obesity and mainly influence energy intake with little effect on the basal metabolic rate. We suggest that FTO influences eating behavior by modulating intracellular energy levels and downstream signaling mechanisms which control energy intake and metabolism. Since FTO plays a particularly important role in adipocytes and in hypothalamic neurons, SH-SY5Y neuronal cells and 3T3-L1 adipocytes were used to understand how siRNA mediated knockdown of FTO expression alters cellular energy homeostasis. Cellular energy status was evaluated by measuring ATP levels using a luminescence assay and uptake of fluorescent glucose. FTO siRNA in SH-SY5Y cells mediated mRNA knockdown (-82%), increased ATP concentrations by up to 46% (P = 0.013) compared to controls, and decreased phosphorylation of AMPk and Akt in SH-SY5Y by -52% and -46% respectively as seen by immunoblotting. In contrast, FTO siRNA in 3T3-L1 cells decreased ATP concentration by -93% (p<0.0005), and increased AMPk and Akt phosphorylation by 204% and 70%, respectively suggesting that FTO mediates control of energy levels in a cell-type specific manner. Furthermore, glucose uptake was decreased in both SH-SY5Y (-51% p = 0.015) and 3T3-L1 cells (-30%, p = 0.0002). We also show that FTO knockdown decreases NPY mRNA expression in SH-SY5Y cells (-21%) through upregulation of pSTAT3 (118%). These results provide important evidence that FTO-variant linked obesity may be associated with altered metabolic functions through activation of downstream metabolic mediators including AMPk.     

Withaferin A Synergizes the Therapeutic Effect of Doxorubicin through ROS-Mediated Autophagy in Ovarian Cancer

Application of doxorubicin (Dox) for the treatment of cancer is restricted due to its severe side effects. We used combination strategy by combining doxorubicin (Dox) with withaferin A (WFA) to minimize the ill effects of Dox. Treatment of various epithelial ovarian cancer cell lines (A2780, A2780/CP70 and CaOV3) with combination of WFA and Dox (WFA/DOX) showed a time- and dose-dependent synergistic effect on inhibition of cell proliferation and induction of cell death, thus reducing the dosage requirement of Dox. Combination treatment resulted in a significant enhancement of ROS production resulting in immense DNA damage, induction of autophagy analyzed by transmission electron microscope and increase in expression of autophagy marker LC3B, and culminated in cell death analyzed by cleaved caspase 3. We validated combination therapy on tumor growth using an in vitro 3Dimension (3D) tumor model and the more classic in vivo xenograft model of ovarian cancer. Both tumor models showed a 70 to 80% reduction in tumor growth compared to control or animals treated with WFA or Dox alone. Immunohistochemical analysis of the tumor tissues from animals treated with WFA/Dox combination showed a significant reduction in cell proliferation and formation of microvessels accompanied by increased in LC3B level, cleaved caspase 3, and DNA damage. Taken together, our data suggest that combining WFA with Dox decreases the dosage requirement of Dox, therefore, minimizing/eliminating the severe side effects associated with high doses of DOX, suggesting the application of this combination strategy for the treatment of ovarian and other cancers with no or minimum side effects.     

Overexpression of hydroperoxide lyase,peroxygenase and epoxide hydrolase in tobacco for the biotechnological production of flavours and polymer precursors

Plants produce short‐chain aldehydes and hydroxy fatty acids, which are important industrial materials, through the lipoxygenase pathway. Based on the information that lipoxygenase activity is up‐regulated in tobacco leaves upon infection with tobacco mosaic virus (TMV), we introduced a melon hydroperoxide lyase (CmHPL) gene, a tomato peroxygenase (SlPXG) gene and a potato epoxide hydrolase (StEH) into tobacco leaves using a TMV‐based viral vector system to afford aldehyde and hydroxy fatty acid production. Ten days after infiltration, tobacco leaves infiltrated with CmHPL displayed high enzyme activities of 9‐LOX and 9‐HPL, which could efficiently transform linoleic acid into C₉ aldehydes. Protein extracts prepared from 1 g of CmHPL‐infiltrated tobacco leaves (fresh weight) in combination with protein extracts prepared from 1 g of control vector‐infiltrated tobacco leaves (as an additional 9‐LOX source) produced 758 ± 75 μg total C₉ aldehydes in 30 min. The yield of C₉ aldehydes from linoleic acid was 60%. Besides, leaves infiltrated with SlPXG and StEH showed considerable enzyme activities of 9‐LOX/PXG and 9‐LOX/EH, respectively, enabling the production of 9,12,13‐trihydroxy‐10(E)‐octadecenoic acid from linoleic acid. Protein extracts prepared from 1 g of SlPXG‐infiltrated tobacco leaves (fresh weight) in combination with protein extracts prepared from 1 g of StEH‐infiltrated tobacco leaves produced 1738 ± 27 μg total 9,12,13‐trihydroxy‐10(E)‐octadecenoic acid isomers in 30 min. The yield of trihydroxyoctadecenoic acids from linoleic acid was 58%. C₉ aldehydes and trihydroxy fatty acids could likely be produced on a larger scale using this expression system with many advantages including easy handling, time‐saving and low production cost.