Effects of cerivastatin on adrenergic pathways, hypertrophic growth and TGFbeta expression in adult ventricular cardiomyocytes

The effects of statin treatment in the setting of heart failure have already been shown. Nevertheless, there is little knowledge about its influence on adrenergic pathways in cardiomyocytes. Therefore, this study investigated the impact of cerivastatin on adrenoceptor-mediated signalling pathways in isolated adult ventricular cardiomyocytes. It focused on two endpoints: hypertrophic growth and TGFbeta expression. Cultured cardiomyocytes were used to study rac activation (analysed by its translocation into the membrane fraction), ROS formation (H(2)DCF fluorescence) and hypertrophic growth ((14)C-phenylalanine incorporation). Alpha- and beta-adrenoceptor stimulation showed significant differences regarding rac activation, ROS formation, and p38 MAP kinase activation. Both alpha- and beta-adrenoceptor stimulation induced TGFbeta expression. Upon activation of alpha-adrenergic signalling - although ROS formation was not influenced by cerivastatin - TGFbeta expression decreased. Following beta stimulation, TGFbeta expression as well as rac and p38 MAP kinase activation were reduced after pre-treatment with cerivastatin. Statin treatment did not show any influence on hypertrophic growth. In summary, this study clearly demonstrates the ability of adrenoceptor stimulation to increase TGFbeta expression. One component of the beneficial effects of statin therapy on heart failure might therefore be due to a dominant reduction and inhibition of TGFbeta, which is involved in many pathophysiological processes in cardiomyocytes.     

Environmentally safe removal/disposal of Coomassie Brilliant Blue from gel destain and used gel stain

Gel destaining following Coomassie Brilliant Blue (CBB) staining involves the use of toxic reagents. Here we demonstrate the efficacy of various paper adsorbents in adsorbing CBB. Kimwipes adsorbed the best, followed by Teri towels, multifold towels, and Whatman numbers 1 and 3 filter papers. Three Kimwipes completely adsorbed the dye released from a CBB-stained mini-gel. Nonradioactive destain solution can, therefore, be recycled for destaining CBB-stained gels. Stain removal with Kimwipes helps in reducing destain use and in reducing organic liquid waste, and it is 7.5-fold cheaper compared with an available method for CBB disposal. Following this, we determined the suitability of this procedure to remove the dye from a used CBB staining solution awaiting proper disposal by our Institutional Safety Office. The dye from a 0.05% CBB staining solution could be removed in 5 to 10 min using 75 Kimwipes. The CBB-adsorbed Kimwipes did not release the stain when squeezed dry even after incubation in various salts over 1 week and in water for 5 weeks. The CBB removed allows its easy disposal as solid waste and will not leach out from solid landfills. Thus, stain removal with Kimwipes helps in disposing CBB in an environmentally friendly manner and allows recycling of destaining solution.     

Problems with Multiple Use of Transfer Buffer in Protein Electrophoretic Transfer

Two-dimensional gel electrophoresis (2DE) and SDS-PAGE are the two most useful methods in protein separation. Proteins separated by 2DE or SDS-PAGE are usually transferred to membranes using a variety of methods, such as electrophoretic transfer, heat-mediated transfer, or nonelectrophoretic transfer, for specific protein detection and/or analysis. In a recent study, Pettegrew et al.¹ claim to reuse transfer buffer containing methanol for at least five times for transferring proteins from SDS-PAGE to polyvinylidene difluoride. They add 150–200 ml fresh transfer solution each time for extended use as a result of loss of transfer buffer. Finally, they test efficiency of each protein transfer by chemiluminescence detection. Here, we comment on this report, as we believe this method is not accurate and useful for protein analysis, and it can cause background binding as well as inaccurate protein analysis.     

Punicic acid: A potential compound of pomegranate seed oil in Type 2 diabetes mellitus management

Diabetes is one of the most prevalent diseases in the worldwide. Type 2 diabetes mellitus (T2DM), the most common form of the disease, has become a serious threat to public health and is a growing burden on global economies. Due to the unexpected adverse effects of antidiabetic medicines, the use of nutraceuticals as a complementary therapy has drawn extensive attention by investigators. In this issue, a novel nutraceutical, Punicic acid (PA)—the main ingredient of pomegranate seed oil (PSO) that has potential therapeutic effects in T2DM—has been investigated. PA is a peroxisome proliferator-activated receptor gamma agonist, and unlike synthetic ligands, such as thiazolidinediones, it has no side effects. PA exerts antidiabetic effects via various mechanisms, such as reducing inflammatory cytokines, modulating glucose homeostasis, and antioxidant properties. In this review, we discussed the potential therapeutic effects of PSO and PA and represented the related mechanisms involved in the management of T2DM.     

Functional role and ribosomal position of the unique N-terminal region of DHX29, a factor required for initiation on structured mammalian mRNAs

Translation initiation on structured mammalian mRNAs requires DHX29, a DExH protein that comprises a unique 534-aa-long N-terminal region (NTR) and a common catalytic DExH core. DHX29 binds to 40S subunits and possesses 40S-stimulated NTPase activity essential for its function. In the cryo-EM structure of DHX29-bound 43S preinitiation complexes, the main DHX29 density resides around the tip of helix 16 of 18S rRNA, from which it extends through a linker to the subunit interface forming an intersubunit domain next to the eIF1A binding site. Although a DExH core model can be fitted to the main density, the correlation between the remaining density and the NTR is unknown. Here, we present a model of 40S-bound DHX29, supported by directed hydroxyl radical cleavage data, showing that the intersubunit domain comprises a dsRNA-binding domain (dsRBD, aa 377–448) whereas linker corresponds to the long α-helix (aa 460–512) that follows the dsRBD. We also demonstrate that the N-terminal α-helix and the following UBA-like domain form a four-helix bundle (aa 90–166) that constitutes a previously unassigned section of the main density and resides between DHX29’s C-terminal α-helix and the linker. In vitro reconstitution experiments revealed the critical and specific roles of these NTR elements for DHX29’s function.     

Applicability of airlift draft-tube fluidized bioreactors for binary protein mixture bioseparation

An airlift draft-tube fluidized bioreactor has been designed and tested for applications in protein bioseparation. Operating parameters and geometrical dimensions of the bioreactor were optimized to ensure fluid circulation in a defined cyclic pattern between the riser and the downcomer. The overall directionality of liquid flow generates homogeneous field of low shear and achieves good mixing efficiency. Bioseparation of proteins was achieved from solutions containing both BSA and BHb at different initial concentrations and at pH 7. Similar adsorption capacities of both proteins were observed in single protein adsorption experiments at pH 7. Compressibility of BHb allowed for high adsorption capacity, in addition to the hydrophobic interaction forces. Apparently the homogeneous and lower shear generated by the airlift bioreactor reduces the compressibility of adsorbed BHb. This allowed for higher BSA adsorption from solutions containing BSA and BHb mixtures. Conventional batch adsorption experiments showed more adsorption of BHb, which reduces bioseparation efficiency.     

In silico analysis of antibody triggering biofilm associated protein in Acinetobacter baumannii

Acinetobacter baumannii surface protein, commonly known as biofilm associated protein (Bap), is involved in biofilm formation. A high propensity among the clinical isolates to form biofilm and a significant association of biofilms with multiple drug resistance has been demonstrated. Production of antibodies can be used for inhibition of biofilm and control of the diseases caused by A. baumannii. Large molecular mass of Bap justifies an approach to identifying A. baumannii effective antigens. It has a core domain of seven repeat modules A-G. With the large number of available biofilm gene sequences, bioinformatic tools are needed to identify the genes encoding the antigens. Proteins containing these tandem repeats of Bap domains have high propensities to attach to each other to form biofilm. We hypothesized that conserved and functional domains of tandem repeat could be identified with a search and alignment of the repeats for evaluation of antigenic determinants. Here we demonstrate the results of bioinformatics screening and gene scan of the gene sequence database of homolog sequences to identify conserved domains. Higher scoring hits were found in repeat modules mostly D, B, C and A, respectively. Upon the analysis four regions of highly structural and functional conserved regions from Bap sequence of A. baumannii were selected. 3D structure, antigenicity and solubility predictions revealed that these regions were appropriate candidates for antibody production.     

Assessment of hemolytic activity of bee venom against some physicochemical factors

Bee venom (BV) is a biotoxin with biologically active peptides which have cell lysis and hemolytic activity properties. These properties can be affected under different storage conditions or during the production process. In present study, we investigated effects of a number of physicochemical factors, including temperature, pH, UV radiation, ultrasound waves and storage time on hemolytic activity of BV. Maximum absorption and melting temperature of BV solution were obtained as 280 nm and ~70 °C, respectively. Cell hemolysis 50 (CH₅₀) -concentration of BV that can lyse 50% of red blood cells- was determined as 0.94 μg/ml at ambient temperature. CH₅₀ was shown not to be importantly varied at temperature up to 60 °C, pH value 2 to 13 and under UV/ ultrasound radiation. Storage at ?20, 6 and 25 °C for 6 months made about 2.5, 35 and 1000 times increase in CH₅₀. From the results, it may be concluded that BV is a relatively resistant hemolytic agent and can be used in a variety of laboratory research and product manufacturing methods.