Assessment of eye doses to staff involved in interventional cardiology procedures in Kuwait

In this study, which is the first of its kind in the gulf region, eye doses of interventional cardiologists and nurses were measured using active dosimeters for left and right eyes, in 60 percutaneous coronary interventions in three main hospitals in Kuwait. The dose given in terms of H_p(0.07) per procedure when ceiling suspended screens were used by main operators ranged from 18.5 to 30.3 µSv for the left eye and from 12.6 to 23.6 µSv for the right eye. Taking into account typical staff workload, the results show that the dose limit of 20 mSv/year to the eyes can be exceeded for interventional cardiologists in some situations, which demonstrates the need of using additional effective radiation protection tools, e.g. protective eye spectacles, in addition to the regular and proper use of ceiling suspended screens. With indications of increase in workload, the need for availability of a dedicated active dosimeter for the regular monitoring of eye doses is emphasized.

     

Essential requirement of cytochrome c release for caspase activation by procaspase-activating compound defined by cellular models

Mitochondrial cytochrome c (cyt. c) release and caspase activation are often impaired in tumors with Bcl-2 overexpression or Bax and Bak-defective status. Direct triggering of cell death downstream of Bax and Bak is an attractive strategy to kill such cancers. Small molecule compounds capable of direct caspase activation appear to be the best mode for killing such tumors. However, there is no precise model to screen such compounds. The currently employed cell-free systems possess the inherent drawback of lacking cellular contents and organelles that operate in integrating cell death signaling. We have developed highly refined cell-based approaches to validate direct caspase activation in cancer cells. Using this approach, we show that PAC-1 (first procaspase-activating compound), the first direct activator of procaspases identified in a cell-free system, in fact requires mitochondrial cyt. c release for triggering caspase activation similar to other antitumor agents. It can induce significant caspase activation and cell death in the absence of Bax and Bak, and in cells overexpressing Bcl-2 and Bcl-xL. This study for the first time defines precise criteria for the validation of direct caspase-activating compounds using specialized cellular models that is expected to accelerate the discovery of potential direct caspase activators.     

Autocrine transforming growth factor alpha regulates cell adhesion by multiple signaling via specific phosphorylation sites of p70S6 kinase in colon cancer cells

Recently, we showed that autocrine transforming growth factor alpha (TGFalpha) controls the epidermal growth factor receptor (EGFR)-mediated basal expression of integrin alpha2, cell adhesion and motility in highly progressed HCT116 colon cancer cells. We also reported that the expression of basal integrin alpha2 and its biological effects are critically controlled by the constitutive activation of the ERK/MAPK pathway (Sawhney, R. S., Sharma, B., Humphrey, L. E., and Brattain, M. G. (2003) J. Biol. Chem. 278, 19861-19869). In the present report, we further examine the downstream signaling mechanisms underlying EGFR/ERK signaling and integrin alpha2 function in HCT116 cells. Selective MEK inhibitors attenuated TGFalpha-mediated basal activation of p70S6K (S6K) specifically at Thr-389, indicating that this S6K site is downstream of ERK/MAPK signaling. Cells were treated with the selective protein kinase C (PKC) inhibitor bisindolylmaleimide to determine the role of PKC in S6K activation. The Thr-421 and Ser-424 phosphorylation sites of S6K were specifically inhibited by bisindolylmaleimide, which also blocked integrin alpha2 expression, cell adhesion, and motility. These data establish a novel cell motility function of S6K via PKC activation in a cancer cell. In addition, we examined whether mammalian target of rapamycin signaling controls S6K activation. Rapamycin inhibited constitutive S6K phosphorylation specifically at Thr-389, Thr-421, and Ser-424 sites. The assignment of these phosphorylation sites on S6K to biological functions was unequivocally confirmed by transfection of cells with specific single phosphorylation site dominant negative mutants. These experiments show for the first time that autocrine TGFalpha regulates cell adhesion function by multiple signaling pathways via specific phosphorylation sites of S6K in cancer cells.     

Caspase-2 triggers Bax-Bak-dependent and -independent cell death in colon cancer cells treated with resveratrol

Polyphenol phytoalexin (resveratrol), found in grapes and red wine is a strong chemopreventive agent with promising safety records with human consumption and unique forms of cell death induction in a variety of tumor cells. However, the mechanism of resveratrol-induced apoptosis upstream of mitochondria is still not defined. The results from this study suggest that caspase-2 activation occurs upstream of mitochondria in resveratrol-treated cells. The upstream activation of caspase-2 is not dependent on its antioxidant property or NF-kappaB inhibition. The activated caspase-2 triggers mitochondrial apoptotic events by inducing conformational changes in Bax/Bak with subsequent release of cytochrome c, apoptosis-inducing factor, and endonuclease G. Caspase-8 activation seems to be independent of these events and does not appear to be mediated by classical death receptor processing or downstream caspases. Both caspase-2 and caspase-8 contribute toward the mitochondrial translocation of Bid, since neither caspase-8 inhibition nor caspase-2 inhibition could prevent translocation of Bid DsRed into mitochondria. Caspase-2 inhibitors or antisense silencing of caspase-2 prevented cell death induced by resveratrol and partially prevented processing of downstream caspases, including caspase-9, caspase-3, and caspase-8. Studies using mouse embryonic fibroblasts deficient for both Bax and Bak indicate the contribution of both Bax and Bak in mediating cell death induced by resveratrol and the existence of Bax/Bak-independent cell death possibly through caspase-8- or caspase-2-mediated mitochondria-independent downstream caspase processing.     

Reverse micelles‐mediated transport of lipase in liquid emulsion membrane for downstream processing

This work deals with the downstream processing of lipase (EC 3.1.1.3, from Aspergillus niger) using liquid emulsion membrane (LEM) containing reverse micelles for the first time. The membrane phase consisted of surfactants [cetyltrimethylammonium bromide (CTAB) and Span 80] and cosolvents (isooctane and paraffin light oil). The various process parameters for the extraction of lipase from aqueous feed were optimized to maximize activity recovery and purification fold. The mechanism of lipase transport through LEM consisted of three steps namely solubilization of lipase in reverse micelles, transportation of reverse micelles loaded with lipase through the liquid membrane, and release of the lipase into internal aqueous phase. The results showed that the optimum conditions for activity recovery (78.6%) and purification (3.14‐fold) were feed phase ionic strength 0.10 M NaCl and pH 9.0, surfactants concentration (Span 80 0.18 M and CTAB 0.1 M), volume ratio of organic phase to internal aqueous phase 0.9, ratio of membrane emulsion to feed volume 1.0, internal aqueous phase concentration 1.0 M KCl and pH 7.0, stirring speed 450 rpm, and contact time 15 min. This work indicated the feasibility of LEM for the downstream processing of lipase. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012     

An evaluation of small-molecule p53 activators as chemoprotectants ameliorating adverse effects of anticancer drugs in normal cells

Pharmacological activation of wild-type p53 has been found to protect normal cells in culture from cytotoxicity and nuclear aberrations caused by conventional cancer therapeutics. Hence, small-molecule p53 activators could have clinical benefits as chemoprotectants for cancer patients bearing p53-mutant tumors. We have evaluated 16 p53-based cyclotherapy regimes combining p53 activators tenovin-6, leptomycin B, nutlin-3 and low dose actinomycin D, with clinically utilized chemotherapeutic agents (S- and M-phase poisons), vinblastine, vinorelbine, cytosine arabinoside and gemcitabine. All the p53 activators induce reversible cell-cycle arrest in primary human fibroblasts and protect them from both S- and M-phase poisons. Furthermore, studies with p53-mutant cancer cell lines show that nutlin-3 and low dose actinomycin D do not affect the sensitivity of these cells to any of the chemotherapeutics tested. Thus, these two small molecules could be suitable choices for cyclotherapy regimes involving S- or M-phase poisons. In contrast, pre-incubation of p53-mutant cells with tenovin-6 or leptomycin B reduces the efficacy of vinca alkaloids, suggesting that these p53 activators could be effective as chemoprotectants if combined with S- but not M-phase poisons. Discrepancies were observed between the levels of protection detected immediately after treatment and following recovery in fresh medium. This highlights the need to assess both short- and long-term effects when evaluating compounds as potential chemoprotectants for cancer therapy.     

Effect of viscosity and dielectric constant variation on fractional fluorescence quenching analysis of coumarin dye in binary solvent mixtures

Photo physical properties of fluorescent organic compounds give an immense improved knowledge on characteristics of excited state that is beneficial to devise innovate molecules and understand their performance in particular applications. Coumarin derivatives have been extensively investigated in this regard. This article narrates steady state fluorescence quenching measurements of a coumarin derivative namely 3‐hydroxy‐3‐[2‐oxo‐2‐(3‐oxo‐3H‐benzo[f]chromen‐2‐yl)‐ethyl]‐1,3‐dihydro‐indol‐2‐one (3HBCD) in a binary mixture of acetonitrile and 1,4‐dioxane. Aniline is used as quencher. Fluorescence intensity is large in acetonitrile and decreases as the percentage of 1,4‐dioxane in the solvent mixture increases. With modest quencher concentration a deviation towards the x axis is noticed in the Stern–Volmer (S–V) plots. This downward curvature is interpreted as due to the presence of 3HBCD in different conformers in the lowest energy level. Ground state intramolecular hydrogen bonding formation is observed due to the conformational changes in the solute. Figured estimations of various quenching parameters recommend that, while dynamic quenching prompts linearity in S–V plot at lower quencher concentration, increasing quenching efficiency with increasing medium viscosity suggests that reaction is not entirely controlled by material diffusion. Stern–Volmer constant increases with decreasing medium dielectric constant.     

Extracellular Histones Induce Chemokine Production in Whole Blood Ex Vivo and Leukocyte Recruitment In Vivo

The innate immune system relies to a great deal on the interaction of pattern recognition receptors with pathogen- or damage-associated molecular pattern molecules. Extracellular histones belong to the latter group and their release has been described to contribute to the induction of systemic inflammatory reactions. However, little is known about their functions in the early immune response to an invading pathogen. Here we show that extracellular histones specifically target monocytes in human blood and this evokes the mobilization of the chemotactic chemokines CXCL9 and CXCL10 from these cells. The chemokine induction involves the toll-like receptor 4/myeloid differentiation factor 2 complex on monocytes, and is under the control of interferon-γ. Consequently, subcutaneous challenge with extracellular histones results in elevated levels of CXCL10 in a murine air pouch model and an influx of leukocytes to the site of injection in a TLR4 dependent manner. When analyzing tissue biopsies from patients with necrotizing fasciitis caused by Streptococcus pyogenes, extracellular histone H4 and CXCL10 are immunostained in necrotic, but not healthy tissue. Collectively, these results show for the first time that extracellular histones have an important function as chemoattractants as their local release triggers the recruitment of immune cells to the site of infection.     

Physiology,biochemistry and possible applications of microbial caffeine degradation

Caffeine, a purine alkaloid is a constituent of widely consumed beverages. The scientific evidence which has proved the harm of this alkaloid has paved the way for innumerable research in the area of caffeine degradation. In addition to this, the fact that the by-products of the coffee and tea industry pollute the environment has called for the need of decaffeinating coffee and tea industry’s by-products. Though physical and chemical methods for decaffeination are available, the lack of specificity for removal of caffeine in these techniques and their non-eco-friendly nature has opened the area of microbial and enzymatic degradation of caffeine. Another important application of microbial caffeine degradation apart from its advantages like specificity, eco-friendliness and cost-effectiveness is the fact that this process will enable the production of industrially and medically useful components of the caffeine degradation pathway like theobromine and theophylline. This is a comprehensive review which mainly focuses on caffeine degradation, large-scale degradation of the same and its applications in the industrial world.