Co-treatment with interferon-γ and 1-methyl tryptophan ameliorates cardiac fibrosis through cardiac myofibroblasts apoptosis

Molecular and Cellular Biochemistry - Electron transfer occurs through heme-Fe across the cytochrome c protein. The current models of long range electron transfer pathways in proteins include...     

Determination of 3-keto-4-ene steroids and their hydroxylated metabolites catalyzed by recombinant human cytochrome P450 1B1 enzyme using gas chromatography-mass spectrometry with trimethylsilyl derivatization

A protocol utilizing gas chromatography with selected ion monitoring mass spectrometric detection (GC-SIM-MS) using a simplified trimethylsilyl (TMS) derivatization protocol was developed and validated for the determination of hydroxylated metabolites of 3-keto-4-ene steroids such as testosterone, progesterone and androstenedione. Hydroxylated metabolites catalyzed by human CYP1B1 were extracted with methylene chloride and derivatized with BSTFA-10% TMCS. To get an optimal derivatizing condition, the effect of various incubation times and temperatures was evaluated. When the incubation temperature and time in the presence of the TMS derivatizing agent were increased, the 3-keto group became derivatized with TMS to form a 3-TMS derivative. To minimize the formation of the TMS ether on the 3-keto group, a reaction condition of 56 degrees C for 10 min was used for the routine measurement of the steroids and their hydroxylated metabolite. Performance studies including linearity of calibration curves, extraction efficiency and precision were performed. Linearity of the calibration curves was satisfactory from 0.125 to 5 microM for most compounds except 21-hydroxyprogesterone and 16alpha-hydroxyandrostenedione which deviated from linearity at the lower concentrations. Mean percentage extraction recoveries were greater than 80% for all compounds. Most compounds showed good precisions with C.V.s of within-day precision of less than 5% and C.V.s of between-day precision of less than 10%. The selected ion chromatograms from the recombinant human CYP1B1 incubations with testosterone, progesterone and androstenedione showed evidence of 6beta-, 16alpha-, 2alpha-, and 15alpha-hydroxytestosterone, 6alpha- and 16alpha-hydroxyprogesterone and 6alpha- and 16alpha-hydroxyandrostenedione, respectively. There was no significant interference associated with Escherichia coli membrane extracts in detecting hydroxylated metabolites. This procedure provides a rapid and sensitive method for the evaluation of steroid hydroxylation by CYP isoenzymes.     

The Presence of Thyroid-Stimulation Blocking Antibody Prevents High Bone Turnover in Untreated Premenopausal Patients with Graves’ Disease

Osteoporosis-related fractures are one of the complications of Graves’ disease. This study hypothesized that the different actions of thyroid-stimulating hormone receptor (TSHR) antibodies, both stimulating and blocking activities in Graves’ disease patients might oppositely impact bone turnover. Newly diagnosed premenopausal Graves’ disease patients were enrolled (n = 93) and divided into two groups: patients with TSHR antibodies with thyroid-stimulating activity (stimulating activity group, n = 83) and patients with TSHR antibodies with thyroid-stimulating activity combined with blocking activity (blocking activity group, n = 10). From the stimulating activity group, patients who had matched values for free T4 and TSH binding inhibitor immunoglobulin (TBII) to the blocking activity group were further classified as stimulating activity-matched control (n = 11). Bone turnover markers BS-ALP, Osteocalcin, and C-telopeptide were significantly lower in the blocking activity group than in the stimulating activity or stimulating activity-matched control groups. The TBII level showed positive correlations with BS-ALP and osteocalcin levels in the stimulating activity group, while it had a negative correlation with the osteocalcin level in the blocking activity group. In conclusion, the activation of TSHR antibody-activated TSH signaling contributes to high bone turnover, independent of the actions of thyroid hormone, and thyroid-stimulation blocking antibody has protective effects against bone metabolism in Graves’ disease.     

The Importance of Normalization References in Interpreting Life Cycle Assessment Results

A normalization step is widely exercised in life cycle assessment (LCA) studies in order to better understand the relative significance of impact category results. In the normalization stage, normalization references (NRs) are the characterized results of a reference system, typically a national or regional economy. Normalization is widely practiced in LCA‐based decision support and policy analysis (e.g., LCA cases in municipal solid waste treatment technologies, renewable energy technologies, and environmentally preferable purchasing programs, etc.). The compilation of NRs demands significant effort and time as well as an intimate knowledge of data availability and quality. Consequently only one set of published NRs is available for the United States, and has been adopted by various studies. In this study, the completeness of the previous NRs was evaluated and significant data gaps were identified. One of the reasons for the significant data gaps was that the toxic release inventory (TRI) data significantly underestimate the potential impact of toxic releases for some sectors. Also the previous NRs did not consider the soil emissions and nitrogen (N) and phosphorus (P) runoffs to water and chemical emissions to soils. Filling in these data gaps increased the magnitude of NRs for “human health cancer,” “human health noncancer,” “ecotoxicity,” and “eutrophication” significantly. Such significant changes can alter or even reverse the outcome of an LCA study. We applied the previous and updated NRs to conventional gasoline and corn ethanol LCAs. The results demonstrate that NRs play a decisive role in the interpretation of LCA results that use a normalization step.     

Biophysical effects in off‐resonant gold nanoparticle mediated (GNOME) laser transfection of cell lines,primary‐ and stem cells using fs laser pulses

Gold nanoparticle mediated (GNOME) laser transfection is a powerful technique to deliver small biologically relevant molecules into cells. However, the transfection of larger and especially negatively charged DNA remains challenging. The efficiency for pDNA was 0.57% using parameter that does not influence the endo‐ and exogenous DNA. In order to gain a deeper understanding of the actual molecule uptake process, the uptake efficiency was determined using molecules of different sizes. It was evaluated that uncharged dextran molecules (2000 kDa) were delivered with an efficiency of 68%. The intracellular distribution of injected molecules was visualized and larger molecules were primary found in the cytoplasm. Patch clamp measurements suggested a permeabilization time up to 15 minutes. The uptake efficiency depended on the size and charge of the molecule to deliver as well as the cell size. A minor role for transfection plays the cell type since primary stem cells were successfully transfected.

The perforation efficiency of semi‐adherent and suspension cells is influenced by the cell and molecule size.     

Comparison of the MicroScan,VITEK 2, and Crystal GP with 16S rRNA sequencing and MicroSeq 500 v2.0 analysis for coagulase-negative Staphylococci

Background  

Three phenotypic identification systems (MicroScan, VITEK 2, and Crystal GP) were evaluated for their accuracy to identify coagulase-negative staphylococci (CNS). A total of 120 clinical isolates confirmed to be CNS via 16S rRNA sequencing and analysis with the MicroSeq 500 v2.0 database were assessed.