Utilization of Jatropha deoiled seed cake for production of cellulases under solid-state fermentation

Toxic waste generated by Jatropha seed cake after utilization of biodiesel on one hand has stimulated the need to develop new technologies to treat the waste and on the other, forced us to reevaluate the efficient utilization of its nutritive potential for production of various high-value compounds and its conversion to non-toxic forms which could be used as animal feed stock. In this study, Jatropha seed cake was used for production of cellulases by new isolate of Thermoascus aurantiacus under solid-state fermentation. The interaction of nitrogen source concentration, moisture ratio, initial pH of the medium and inoculum size was investigated and modelled using response surface methodology (RSM) using Box-Behnken Design (BBD). Under optimized conditions endo-β-1,4-glucanase, β-glucosidase and filter paper activities were found to be 124.44, 28.86, 4.87?U/g of substrate, respectively. Characterization of endo-β-1,4-glucanase, β-glucosidase was done after partial purification by ammonium sulfate fractionation followed by desalting. The endo-β-1,4-glucanase and β-glucosidase showed maximum activity at 70?°C and pH 4. Saccharification studies performed with different lignocellulosic substrates showed that sugar cane bagasse was most susceptible to enzymatic hydrolysis. The study suggests that Jatropha seed cake can be used as a viable nutrient source for cellulase production without any pretreatment under solid-state fermentation by T. aurantiacus.     

Overexpression of SIPK in tobacco enhances ozone-induced ethylene formation and blocks ozone-induced SA accumulation

Ozone induces rapid activation of SIPK, a mitogen-activated protein kinase (MAPK) in tobacco. Through transgenic manipulation it has previously been shown that overexpression of SIPK leads to enhanced ozone-induced lesion formation with concomitant accumulation of ROS. In spite of this hypersensitive phenotype, the effect of this altered SIPK expression on the levels of various hormones that regulate ozone-induced cell death has remained unexplored. The response of both salicylate and ethylene, the major phytohormones that modulate ozone-induced cell death, have now been analysed in SIPK-OX tobacco plants. Ozone treatment strongly induced ethylene formation in the sensitive SIPK-OX plants at ozone concentrations that failed to elicit stress ethylene release in WT plants. By contrast, SIPK-overexpressing plants displayed no ozone-induced SA accumulation, whereas WT plants accumulated SA upon ozone exposure. Epistatic analysis of SIPK-OX function suggests that the ozone-induced cell death observed in SIPK-OX plants is either independent, or upstream, of SA accumulation.     

A Homochiral Diphenylalanine Analog Based Mechanoresponsive Hydrogel: An Insight Towards Its Wound Healing Efficacy

Deciphering the most promising strategy for the evolution of potential wound-healing therapeutics is one of the greatest challenging affairs to date. The development of peptide-based smart scaffolds with innate antimicrobial, anti-inflammatory, and antioxidant properties is an appealing way out. Aligned to the goal a set of Hydrogelators I–IV were developed utilizing the concept of chiral orchestration in diphenylalanine fragment, such that the most potent construct with all the bench marks namely mechanoresponsiveness, biocompatibility, consistent antimicrobial and antioxidant properties, could be fished out from the design. Interestingly, our in vitro Antifungal and Lipid peroxidation analysis identified the homochiral isomer Boc-δ-Ava-L-Phe-L-Phe-OH (Hydrogelator I ), as an ideal candidate for the wound healing experiment, so we proceeded for the in vivo histopathological and antioxidant measurements in Wister rats. Indeed the wound images obtained from the different sets of animals on the 14^th day of treatment demonstrated that with increased recovery time, hydrogelator I displayed a significant reduction in the lesion diameter compared to the marketed drug, and negative control. Even the histopathological measurements using H & E staining demonstrated diminished tissue destruction, neutrophil infiltration necrosis, and lymphatic proliferation in the hydrogelators, in comparison to others, backed by in vivo lipid peroxidation data. Overall our investigation certifies hydrogelator I as an effective therapeutic for managing the wound healing complication.     

High Throughput Kinomic Profiling of Human Clear Cell Renal Cell Carcinoma Identifies Kinase Activity Dependent Molecular Subtypes

Despite the widespread use of kinase-targeted agents in clear cell renal cell carcinoma (CC-RCC), comprehensive kinase activity evaluation (kinomic profiling) of these tumors is lacking. Thus, kinomic profiling of CC-RCC may assist in devising a classification system associated with clinical outcomes, and help identify potential therapeutic targets. Fresh frozen CC-RCC tumor lysates from 41 clinically annotated patients who had localized disease at diagnosis were kinomically profiled using the PamStation®12 high-content phospho-peptide substrate microarray system (PamGene International). Twelve of these patients also had matched normal kidneys available that were also profiled. Unsupervised hierarchical clustering and supervised comparisons based on tumor vs. normal kidney and clinical outcome (tumor recurrence) were performed and coupled with advanced network modeling and upstream kinase prediction methods. Unsupervised clustering analysis of localized CC-RCC tumors identified 3 major kinomic groups associated with inflammation (A), translation initiation (B), and immune response and cell adhesions (C) processes. Potential driver kinases implicated include PFTAIRE (PFTK1), PKG1, and SRC, which were identified in groups A, B, and C, respectively. Of the 9 patients who had tumor recurrence, only one was found in Group B. Supervised analysis showed decreased kinase activity of CDK1 and RSK1-4 substrates in those which progressed compared to others. Twelve tumors with matching normal renal tissue implicated increased PIM’s and MAPKAPK’s in tumors compared to adjacent normal renal tissue. As such, comprehensive kinase profiling of CC-RCC tumors could provide a functional classification strategy for patients with localized disease and identify potential therapeutic targets.     

In-depth proteomic analyses of ovarian cancer cell line exosomes reveals differential enrichment of functional categories compared to the NCI 60 proteome

Molecular communication between cancer cells and its stromal microenvironment is a key factor for cancer progression. Alongside classic secretory pathways, it has recently been proposed that small membranous vesicles are alternative mediators of intercellular communication. Exosomes carry an effector-rich proteome with the ability to modulate various functional properties of the recipient cell. In this study, exosomes isolated from four epithelial ovarian cancer cell lines (OVCAR3, OVCAR433, OVCAR5 and SKOV3) were characterized using mass spectrometry-based proteomics. Using an optimized workflow consisting of efficient exosome solubilization and the latest generation of proteomic instrumentation, we demonstrate improved detection depth. Systematic comparison of our cancer cell line exosome proteome against public data (Exocarta) and the recently published NCI 60 proteome revealed enrichment of functional categories related to signaling biology and biomarker discovery.     

Photosensitized mefloquine induces ROS-mediated DNA damage and apoptosis in keratinocytes under ambient UVB and sunlight exposure

The present study illustrates the photosensitizing behavior of mefloquine (MQ) in human skin keratinocytes under ambient doses of UVB and sunlight exposure. Photochemically, MQ generated reactive oxygen species superoxide radical, hydroxyl radical, and singlet oxygen through type I and type II photodynamic reactions, respectively, which caused photooxidative damage to DNA and formed localized DNA lesions cyclobutane pyrimidine dimers. Photosensitized MQ reduced the viability of keratinocytes to 25 %. Significant level of intracellular reactive oxygen species (ROS) generation was estimated through fluorescence probe DCF-H2. Increased apoptotic cells were evident through AO/EB staining and phosphatidyl serine translocation in cell membrane. Single-stranded DNA damage was marked through single-cell gel electrophoresis. Mitochondrial membrane depolarization and lysosomal destabilization were evident. Upregulation of Bax and p21 and downregulation of Bcl-2 genes and corresponding protein levels supported apoptotic cell death of keratinocyte cells. Cyclobutane pyrimidine dimers (CPDs) were confirmed through immunofluorescence. In addition, hallmarks of apoptosis and G2/M phase cell cycle arrest were confirmed through flow cytometry analysis. Our findings suggest that MQ may damage DNA and produce DNA lesions which may induce differential biological responses in the skin on brief exposure to UVB and sunlight. Figure

Mefloquine is photosensitized by UVB and sunlight exposure at an appropriate dose and generates ROS involving both type I and type II photosensitization mechanisms. These ROS primarily damage DNA, cell membrane, and membrane-bound organelles. MQ differentially affects various biological processes which end into apoptosis of the cell