Human epidermal growth factor receptor bispecific ligand trap RB200: abrogation of collagen-induced arthritis in combination with tumour necrosis factor blockade

Introduction  

Rheumatoid arthritis (RA) is a chronic disease associated with inflammation and destruction of bone and cartilage. Although inhibition of TNFα is widely used to treat RA, a significant number of patients do not respond to TNFα blockade, and therefore there is a compelling need to continue to identify alternative therapeutic strategies for treating chronic inflammatory diseases such as RA. The anti-epidermal growth factor (anti-EGF) receptor antibody trastuzumab has revolutionised the treatment of patients with EGF receptor-positive breast cancer. Expression of EGF ligands and receptors (known as HER) has also been documented in RA. The highly unique compound RB200 is a bispecific ligand trap that is composed of full-length extracellular domains of HER1 and HER3 EGF receptors. Because of its pan-HER specificity, RB200 inhibits responses mediated by HER1, HER2 and HER3 in vitro and in vivo. The objective of this study was to assess the effect of RB200 combined with TNF blockade in a murine collagen-induced arthritis (CIA) model of RA.     

The promoter of filamentation (POF1) protein from <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> is an ATPase involved in the protein quality control process

Background

The gene YCL047C, which has been renamed promoter of filamentation gene (POF1), has recently been described as a cell component involved in yeast filamentous growth. The objective of this work is to understand the molecular and biological function of this gene.

Results

Here, we report that the protein encoded by the POF1 gene, Pof1p, is an ATPase that may be part of the Saccharomyces cerevisiae protein quality control pathway. According to the results, Δpof1 cells showed increased sensitivity to hydrogen peroxide, tert-butyl hydroperoxide, heat shock and protein unfolding agents, such as dithiothreitol and tunicamycin. Besides, the overexpression of POF1 suppressed the sensitivity of Δpct1, a strain that lacks a gene that encodes a phosphocholine cytidylyltransferase, to heat shock. In vitro analysis showed, however, that the purified Pof1p enzyme had no cytidylyltransferase activity but does have ATPase activity, with catalytic efficiency comparable to other ATPases involved in endoplasmic reticulum-associated degradation of proteins (ERAD). Supporting these findings, co-immunoprecipitation experiments showed a physical interaction between Pof1p and Ubc7p (an ubiquitin conjugating enzyme) in vivo.

Conclusions

Taken together, the results strongly suggest that the biological function of Pof1p is related to the regulation of protein degradation.

     

Visfatin is regulated by rosiglitazone in type 2 diabetes mellitus and influenced by NFκB and JNK in human abdominal subcutaneous adipocytes

Visfatin has been proposed as an insulin-mimicking adipocytokine, predominantly secreted from adipose tissue and correlated with obesity. However, recent studies suggest visfatin may act as a proinflammatory cytokine. Our studies sought to determine the significance of this adipocytokine and its potential role in the pathogenesis of T2DM. Firstly, we examined the effects of diabetic status on circulating visfatin levels, and several other adipocytokines, demonstrating that diabetic status increased visfatin*, TNF-α*** and IL-6*** compared with non-diabetic subjects (*p<0.05, **p<0.01, ***p<0.001, respectively). We then assessed the effects of an insulin sensitizer, rosiglitazone (RSG), in treatment naïve T2DM subjects, on circulating visfatin levels. Our findings showed that visfatin was reduced post-RSG treatment [vs. pre-treatment (*p<0.05)] accompanied by a reduction in HOMA-IR**, thus implicating a role for insulin in visfatin regulation. Further studies addressed the intracellular mechanisms by which visfatin may be regulated, and may exert pro-inflammatory effects, in human abdominal subcutaneous (Abd Sc) adipocytes. Following insulin (Ins) and RSG treatment, our in vitro findings highlighted that insulin (100 nM), alone, upregulated visfatin protein expression whereas, in combination with RSG (10 nM), it reduced visfatin*, IKKβ** and p-JNK1/2*. Furthermore, inhibition of JNK protein exacted a significant reduction in visfatin expression (**p<0.01), whilst NF-κB blockade increased visfatin (*p<0.05), thus identifying JNK as the more influential factor in visfatin regulation. Additional in vitro analysis on adipokines regulating visfatin showed that only Abd Sc adipocytes treated with recombinant human (rh)IL-6 increased visfatin protein (*p<0.05), whilst rh visfatin treatment, itself, had no influence on TNF-α, IL-6 or resistin secretion from Sc adipocytes. These data highlight visfatin''s regulation by insulin and RSG, potentially acting through NF-κB and JNK mechanisms, with only rh IL-6 modestly affecting visfatin regulation. Taken together, these findings suggest that visfatin may represent a pro-inflammatory cytokine that is influenced by insulin/insulin sensitivity via the NF-κB and JNK pathways.     

Genetic diversity and mycelial compatibility groups of the plant-pathogenic fungus Sclerotinia sclerotiorum in Brazil

The genetic variability of 40 Sclerotinia sclerotiorum isolates from various fields widely distributed throughout Brazil and different host crops was analyzed using RAPD markers and mycelial compatibility groupings (MCGs). The isolates were characterized using 16 random primers of the OPERON series, which produced 121 DNA fragments. UPGMA cluster analysis using Jaccard's genetic distance and MCGs allowed separation of the isolates into three clusters, with similarity indices of 68.2, 61.8, and 61.8%, and five MCGs. The haplotypes obtained with RAPD markers provided very characteristic groupings of S. sclerotiorum isolates according to MCG, but did not show any relationship with geographic origin or host type. Furthermore, analysis of molecular variance demonstrated that 99.1% of the observed variation was a result of genetic differences between individuals; the host culture did not have a significant effect. This is the first report of high level variability of S. sclerotiorum in Brazil based on the study of isolates of wide geographical origin, supported by RAPD markers and MCGs. These results endorse the prevalence of sexual reproduction in tropical and subtropical regions in contrast to clonal reproduction in temperate regions.     

Effect of some essential oils on in vitro methane emission

The objectives of this study were to characterise four essential oils (EO) chemically and to evaluate their effect on ruminal fermentation and methane emission in vitro. The investigated EO were isolated from Achillea santolina, Artemisia judaica, Schinus terebinthifolius and Mentha microphylla, and supplemented at four levels (0, 25, 50 and 75 microl) to 75 ml of buffered rumen fluid plus 0.5 g of substrate. The main components of the EO were piperitone (49.1%) and camphor (34.5%) in A. judaica, 16-dimethyl 15-cyclooactdaiene (60.5%) in A. santolina, piperitone oxide (46.7%) and cis-piperitone oxide (28%) in M. microphylla, and gamma-muurolene (45.3%) and alpha-thujene (16.0%) in S. terebinthifolius. The EO from A. santolina (at 25 and 50 j1), and all levels of A. judaica increased the gas production significantly, but S. terebinthifolius (at 50 and 75 microl), A. santolina (at 75 microl) and all levels of M. microphylla decreased the gas production significantly in comparison with the control. The highest levels of A. santolina and A. judaica, and all doses from M. microphylla EO inhibited the methane production along with a significant reduction in true degradation of dry matter and organic matter, protozoa count and NH3-N concentration. It is concluded that the evaluated EO have the potential to affect ruminal fermentation efficiency and the EO from M. microphylla could be a promising methane mitigating agent.     

Two new species of Acacia (Leguminosae-Mimosoideae) from Yemen

The new species Acacia harala and A. mahrana , both from southern Yemen, are described and illustrated. A. harala belongs to A. subgen. Acacia and is known from the Abyan and Shabwah Regions, whereas A. mahrana belongs to A. subgen. Aculeiferum and is known only from the Mahrah Region.     

Features of visual function in the naked mole-rat <Emphasis Type="Italic">Heterocephalus glaber</Emphasis>

The eyes and visual capacity of the naked mole-rat, Heterocephalus glaber, a subterranean rodent, were evaluated using anatomical, biochemical, and functional assays, and compared to other rodents of similar body size (mouse and gerbil). The eye is small compared to mouse, yet possesses cornea, lens, and retina with typical mammalian organization. The optic nerve cross-sectional area and fiber density are ~10% and ~50% that of gerbil, respectively. Levels per unit retinal area of 11-cis and all-trans retinal, derivatives of vitamin A associated with the visual cycle, are comparable to mouse. The corneal electroretinogram (ERG) exhibits early and late negative components that scale with flash strength; raising the body temperature of this poikilothermic animal from 30°C (normal for H. glaber ) to 37°C (normal for mouse) revealed an ERG response with typically mammalian features, but greatly attenuated and with slower kinetics. Leaving the nest chamber was a behavior correlated with light onset displayed preferentially by breeding females. Optical models of five mole-rat eyes suggest reasonable, but variable, image formation at the retina, possibly related to age. Results are consistent with amorphous light detection, possibly useful for circadian entrainment or escape behavior in the event of tunnel breeches.     

Angiotensin AT2 receptor deficiency after myocardial infarction

Hearts of normotensive angiotensin II type 2 receptor (AT₂)-deficient mice do not develop fibrosis after angiotensin II-induced chronic hypertension. Thus, the goal of our study was to clarify whether AT₂ knockouts (KOs) are also characterized by altered left ventricular (LV) function and modified remodeling of the extracellular matrix (ECM) after induction of myocardial infarction (MI). MI was induced in 5-mo-old female AT₂-deficient mice and controls by occlusion of the left coronary artery. Time-matched sham-operated animals served as controls. After 48 h, the first sets of mice were hemodynamically characterized using a pressure-tip catheter (n=8/group). We also obtained pressure volume loops using a microconductance catheter in additional sets of animals 3 wk after induction of MI (n=7/group). Finally, the collagen index was illustrated by Sirius red staining and quantified by digital analysis. Whereas the LV function of sham-operated animals did not differ between both genotypes, the collagen index was 44% lower in KO animals. Forty-eight hours and 3 wk post-MI, systolic and diastolic LV function were impaired in both AT₂-deficient and wild-type (WT) animals to the same extent by approx 45%. No differences were found between the two genotypes with respect to LV hypertrophy and the fibrosis index in the infarcted and noninfarcted areas 3 wk post-MI. While AT₂-KO mice had less cardiac collagen content under basal conditions, the receptor deficiency had no significant influence on LV function at the two investigated time points after induction of MI or on the remodeling of ECM at the latter time point. Thus, hypetension-induced fibrosis is probably triggered by other control mechanisms than fibrosis induced by MI.     

Synthesis of some new 1,3/ or 1,4-bis(glucopyranosyl-1,2,4-triazol-5-ylthio)propanes/ or butanes as potential antimicrobial agents

Glucosidation of the appropriate 1,3 or 1,4-bis(4-amino or arylideneamino-2,4-dihydro-3-thioxo-3H-1,2,4-triazol-5-ylthio)propanes or butanes with 2,3,4,6-tetra-O-acetyl-alpha-D-glucopyranosyl bromide followed by chromatographic separation gave the corresponding N-, S-, and N,S-bis(glucosides). Chemical transformation leading to new functionalities has been achieved. Antimicrobial screening of 10 selected compounds resulted in their activity against Aspergillus fumigatus, Penicillium italicum, Syncephalastrum racemosum, Candida albicans, Staphylococcus aureus, Pseudomonas aeruginosa, Bacillus subtilis, and Escherichia coli.