Inhibition of the thioredoxin system is a basis for the antileukemic potential of 13-hydroxy-15-oxo-zoapatlin

The mammalian thioredoxin (Trx) system, composed of Trx, Trx reductase (TrxR), and NADPH, is the most important thiol system involved in the redox control of signaling and regulatory proteins in apoptosis and cell proliferation. Here we addressed the inhibition of the Trx system by 13-hydroxy-15-oxo-zoapatlin (OZ), a nor-kaurane diterpene previously shown to possess proapoptotic potential and to cause cell cycle arrest in leukemia cells. OZ was found, by both biochemical and mass spectrometry-based approaches, to target Trx1 and TrxR in a cell-free system. In particular, the formation of reversible OZ adducts to Trx1 Cys35, Cys62, and Cys73 was demonstrated. We next showed that OZ efficiently inhibited Trx and TrxR catalytic activity in Molt4 cells. The occurrence of oxidative modifications of Trx molecules was assessed by “redoxWestern blot” analyses. OZ-mediated Trx oxidation resulted in apoptosis signaling kinase-1 release and activation of downstream JNK and p38 pathways. By means of specific inhibitors of these two stress-activated protein kinases, we demonstrated that the JNK pathway plays a major role in determining the apoptotic fate of OZexposed cells, whereas p38 activation seems to be involved mainly in OZ-induced G2/M block

Rimonabant-induced apoptosis in leukemia cell lines: Activation of caspase-dependent and -independent pathways

Rimonabant (SR141716), a cannabinoid CB1 receptor antagonist known for anti-obesity activity, has more recently been shown to inhibit tumor cell growth. Here we demonstrated the antitumor potential of SR141716 in leukemia-derived cell lines and its low toxicity in normal cells (PBMC). SR141716 (1-20 mu M range of doses) reduced Jurkat and U937 cell number by activating death signals as well as affecting cell cycle progression. The most prominent response in U937 to SR141716 was a G(0)/G(1) block, while in Jurkat cells there was activation of cell death processes. SR141716-treated cells exhibited the morphological and biochemical features of apoptosis and to some extent necrosis. Apoptotic mode of cell death was confirmed in both cell lines by analysis of cell morphology, phosphatidylserine exposure and DNA fragmentation. Moreover, the drug was found to induce an early and robust mitochondrial membrane depolarization. In Jurkat cells the apoptotic process was typically caspase-dependent, while in U937 caspase-independent pathways were also activated. The contribution of PARP activation to SR141716-induced apoptosis in U937 was suggested by protein PARylation, AIF release and apoptosis reversal by PARP inhibitors. Moreover. SR141716 negatively modulated, especially in U937, the PI3K/AKT pathways. In conclusion, our data indicate that SR141716 elicits alternative response and/or cell death pathways depending on the cell type affected. (C) 2010 Elsevier Inc. All rights reserved

COD and TPH analysis in slops experimental treatment plants: analytical problems.

COD AND TPH ANALYSIS IN SLOPS EXPERIMENTAL TREATMENT PLANTS: ANALYTICAL PROBLEMS. In the last years, the persistence and accumulation of xenobiotic compounds in the environment created many disposal problems of oily wastewater generated by ships, mainly in engine-rooms (bilge waters) and by cleaning of tanker (slops). The high salinity levels (up to 25.000 mgL-1) and the pollutants concentration, both limit the chances of discharge into the sewer systems and address the disposal of these wastewaters to the sea. For these reasons it is necessary to treat such wastewater efficiently before discharging [1]. As a part of a broader project concerning slops treatments, this work addresses issues related to the analytical methods of the COD and TPH parameters, chosen under the provisions of Italian Legislative Decree 152/06, that implements the European directives on environmental protection. In the COD analysis the greatest difficulty was the high salinity levels corresponding to high chlorides levels. Chlorides cause a positive interference in the measurement, and this interference during the analysis of COD was investigated at various concentrations of mercury in order to try to minimize the use of this reagent that causes many problems of safety and disposal, and at different times reaction [2][3][4]. The major problems in TPH analysis concerned correct setup of the gas chromatographic separation, high variability in sample composition and the high capacity of the sample to form emulsions during the liquid-liquid extraction procedure. This paper reports the results of the analysis of COD and TPH and the problems related to the development of a suitable analytical method for the analysis of these specimens. This study is part of STITAM European project, created with the aim to develop innovative technologies for the treatment of liquid wastes of navigation, in order to better safeguard marine environment. [1] G. Mancini, S. Cappello, M.M. Yakimov, A. Polizzi, M. Torregrossa, Chemical Engineering Transactions 27 (2012) 37-42. [2] I. Vyrides, D.C. Stuckey, Bioresource Technology 100 (2009) 979–982 [3] Alexandra M.E. Viana da Silva, Ricardo J.N. Bettencourt da Silva , M. Filomena G.F.C. Camoes, Analytica Chimica Acta 699 (2011) 161– 169 [4] D.D.C. Freire, G.L. Sant’anna jr, Environmental Technology 19 (1998) 1243- 124

Postharvest Application of Aloe vera Gel-Based Edible Coating to Improve the Quality and Storage Stability of Fresh-cut Papaya

Ready-to-eat products are damaged by various factors, including exposure to O2 and CO2, extreme temperatures, and rapid decay, due to trauma during processing. +e use of natural antimicrobial agents and antioxidants might extend the shelf-life of the fruits. +e aim of this work is to investigate the effects of four different antibrowning and gelling agents added into the Aloe vera gelbased edible coatings and applied to fresh-cut papaya. EC1 treatment consists of Aloe vera gel (30% v/v), EC2 contains CaCl2 (5% v/v), EC3 contains K carrageenan (0.5% v/v), and EC4 contains sodium alginate (1.5% v/v) and K carrageenan (0.5% v/v). +e fruits treated with EC2 showed the best results while maintaining high values in terms of firmness (that differ from the control of 42.5%), soluble solid content (that differ from the control of 14.6%), and titratable acidity (that differ from the control of 49%). Hence, the addition of CaCl2 also reduces the ripening rate and loss of color without altering the product’s sensory qualities. EC3 and EC4 treatments have provided an oxygen barrier and reduced respiratory rate, increasing the firmness retention and keeping a high C∗ value thanks to K carrageenan and sodium alginate

Photoplethysmography (PPG)-determined heart rate variability (HRV) and extracellular water (ECW) in the evaluation of chronic stress and inflammation

Background Heart rate variability (HRV) is the physiological variation in the time interval between consecutive heartbeats. Extracellular water (ECW) is the aqueous compartment surrounding cells and has been used as an inflammation index. Medically unexplained symptoms (MUS) refer to persistent psychosomatic bodily complaints, and their presence may be employed as a clinical index of chronic stress and inflammation, useful in distinguishing suffering patients from healthy subjects. Aim To evaluate the clinical performance of SDNN (standard deviation of intracardiac beat time interval of normal sinus beats) and RMSSD (square root of the mean squared differences of successive NN intervals) as indices of HRV, and their correlation with ECW and MUS. Patients and methods A large multicenter retrospective study was conducted in 37 Italian medical practices in Caucasian men and women aged 20 to 70 years. SDNN and RMSSD were measured with the PPG Stress Flow (R) device (BioTekna, Italy), while ECW was determined with the BIA-ACC (R) device (BioTekna, Italy). All subjects filled in a MUS (R) questionnaire with 19 “nonspecific” symptom questions. The study sample was stratified by decade of age into five groups. Results Data from 9246 subjects comprising 3127 males and 6119 females, with a median age of 47 years, were analyzed. HRV index SDNN and RMSSD distributions in the entire sample and in each of the five age groups were significantly greater in subjects with a limited number of MUS (0-5) than in subjects with six or more symptoms, while both distributions correlated negatively with ECW. Conclusion SDNN and RMSSD and ECW were predictors of MUS and were successfully used to objectively evaluate chronic stress and inflammation

BAG3 protects Bovine Papillomavirus type 1-transformed equine fibroblasts against pro-death signals

In human cancer cells, BAG3 protein is known to sustain cell survival. Here, for the first time, we demonstrated the expression of BAG3 protein in equine sarcoids in vivo as well as in an in vitro model of sarcoid-derived equine fibroblasts. Evidence of a possible involvement of BAG3 in equine sarcoid carcinogenesis was obtained by immunohistochemistry analysis of tumour samples. We found that the most of tumour samples stained positive for BAG3, even though to a different grade, while normal dermal fibroblasts from a healthy horse displayed very weak staining pattern for BAG3 expression. By siRNA technology, we demonstrated the role of BAG3 in counteracting basal as well as chemical-triggered pro-death signals. BAG3 down-modulation in EqSO4b, a sarcoid-derived fully transformed cell line harbouring bovine papilloma virus (BPV)-1 genome, promotes cell death and cell cycle arrest in G0/G1. In addition, we found that BAG3 silencing sensitized cells to phenethylisothiocyanate (PEITC), a promising cancer chemopreventive/chemotherapeitic agent present in edible cruciferous vegetables. Notably, such a pro-survival role of BAG3 was less relevant in E.Derm cells, taken as normal counterpart, thus suggesting a mutual cooperation between BAG3 and viral oncoproteins to sustain cell survival

Powerful tumor cell growth-inhibiting activity of a synthetic derivative of atractyligenin: Involvement of PI3K/Akt pathway and thioredoxin system

Background: The semi-synthetic ent-kaurane 15-ketoatractyligenin methyl ester (SC2017) has been previously reported to possess high antiproliferative activity against several solid tumor-derived cell lines. Our study was aimed at investigating SC2017 tumor growth-inhibiting activity and the underlying mechanisms in Jurkat cells (T-cell leukemia) and xenograft tumor models. Methods: Cell viability was evaluated byMTT assay. Cell cycle progression, reactive oxygen species (ROS) elevation and apoptotic hallmarks were monitored by flow cytometry. Inhibition of thioredoxin reductase (TrxR) by biochemical assays. Levels and/or activation status of signaling proteins were assessed by western blotting. Xenograft tumors were generated with HCT 116 colon carcinoma cells. Results: SC2017 displayed cell growth-inhibiting activity against Jurkat cells (halfmaximal inhibitory concentration values (IC50) b 2 μM), but low cell-killing potential in human peripheral blood mononuclear cells (PBMC). The primary response of Jurkat cells to SC2017was an arrest in G2 phase followed by caspase-dependent apoptosis. Inhibition of PI3K/Akt pathway and TrxR activity by SC2017 was demonstrated by biochemical and pharmacological approaches. At least, SC2017 was found to inhibit xenograft tumor growth. Conclusions: Our results demonstrate that SC2017 inhibits tumor cell growth in in vitro and in vivo models, but displays moderate toxicity against PBMC. We also demonstrate that SC2017 promotes caspase-dependent apoptosis in Jurkat cells by affecting Akt activation status and TrxR functionality. General significance: Our observations suggest the semi-synthetic ent-kaurane SC2017 as a promising chemotherapeutic compound. SC2017 has, indeed, shown to possess tumor growth inhibiting activity and be able to counteract PI3K/Akt and Trx system survival signaling

Structural characterization of tetranortriterpenes from Pseudrocedrela kotschyi and Trichilia emetica and study of their activity towards the chaperone Hsp90

Investigation of roots extracts Pseudrocedrela kotschyi and Trichilia emetica led to identification of 5 limonoid derivatives, Kotschyins D–H, and 11 known compounds. Their structures were elucidated by extensive 1D and 2D NMR experiments in conjunction with mass spectrometry. A surface plasmon resonance (SPR) approach was adopted to screen their Hsp90 binding capability and kotschyin D showed a significant affinity for the chaperone. Therefore, the characterization of the biological activity of kotschyin D by means of a panel of chemical and biological approaches, including limited proteolysis, molecular docking and biochemical and cellular assays, was performed. Our result indicated this compound as a type of client selective Hsp90 inhibitor, directly binding to the middle domain of the protein and possibly preventing its interaction with the activator of Hsp90 ATPase 1 (Aha1)