Decrease of CD68 Synovial Macrophages in Celastrol Treated Arthritic Rats

Background

Rheumatoid arthritis (RA) is a chronic immune-mediated inflammatory disease characterized by cellular infiltration into the joints, hyperproliferation of synovial cells and bone damage. Available treatments for RA only induce remission in around 30% of the patients, have important adverse effects and its use is limited by their high cost. Therefore, compounds that can control arthritis, with an acceptable safety profile and low production costs are still an unmet need. We have shown, in vitro, that celastrol inhibits both IL-1β and TNF, which play an important role in RA, and, in vivo, that celastrol has significant anti-inflammatory properties. Our main goal in this work was to test the effect of celastrol in the number of sublining CD68 macrophages (a biomarker of therapeutic response for novel RA treatments) and on the overall synovial tissue cellularity and joint structure in the adjuvant-induced rat model of arthritis (AIA).

Methods

Celastrol was administered to AIA rats both in the early (4 days after disease induction) and late (11 days after disease induction) phases of arthritis development. The inflammatory score, ankle perimeter and body weight were evaluated during treatment period. Rats were sacrificed after 22 days of disease progression and blood, internal organs and paw samples were collected for toxicological blood parameters and serum proinflammatory cytokine quantification, as well as histopathological and immunohistochemical evaluation, respectively.

Results

Here we report that celastrol significantly decreases the number of sublining CD68 macrophages and the overall synovial inflammatory cellularity, and halted joint destruction without side effects.

Conclusions

Our results validate celastrol as a promising compound for the treatment of arthritis.     

A chalcid wasp acts chiefly as a hyperparasitoid by mostly using small uncommon hosts

Although ovipositing insects may predominantly use resources that lead to high offspring quality, exceptions to this rule have considerably aided understanding of oviposition decisions. We report the frequency of host species use by a solitary facultative hyperparasitoid, Brachymeria subrugosa Blanchard (Hymenoptera: Chalcididae). In our samples, the wasp attacks the large pupae of the moth Gonioterma indecora Zeller (Lepidoptera: Elachistidae), as well as the considerably smaller, and rarer, pupae of two of its other parasitoids. Consistent with conditional sex allocation models, the wasp produced mainly female offspring on the largest (moth) host, an unbiased sex ratio on the middle‐sized (parasitoid) host, and only males on the smallest (parasitoid) host. Adult offspring size was correlated with the size of the host attacked. These features strongly suggest that the two smaller, primary parasitoid, hosts produce lower‐quality offspring. Despite being more common, the proportion of hosts from which parasitoids emerged was lowest (14%) on the largest host species, and highest on the rarer middle‐sized (34%) and smallest (30%) hosts. This suggests that costs or constraints on attacking high‐quality primary hosts may be a selective force favouring the evolution of hyperparasitism.     

Worldwide Occurrence of Integrative Conjugative Element Encoding Multidrug Resistance Determinants in Epidemic Vibrio cholerae O1

In the last decades, there has been an increase of cholera epidemics caused by multidrug resistant strains. Particularly, the integrative and conjugative element (ICE) seems to play a major role in the emergence of multidrug resistant Vibrio cholerae. This study fully characterized, by whole genome sequencing, new ICEs carried by multidrug resistant V. cholerae O1 strains from Nigeria (2010) (ICEVchNig1) and Nepal (1994) (ICEVchNep1). The gene content and gene order of these two ICEs are the same, and identical to ICEVchInd5, ICEVchBan5 and ICEVchHai1 previously identified in multidrug resistant V. cholerae O1. This ICE is characterized by dfrA1, sul2, strAB and floR antimicrobial resistance genes, and by unique gene content in HS4 and HS5 ICE regions. Screening for ICEs, in publicly available V. cholerae genomes, revealed the occurrence and widespread distribution of this ICE among V. cholerae O1. Metagenomic analysis found segments of this ICE in marine environments far from the direct influence of the cholera epidemic. Therefore, this study revealed the epidemiology of a spatio-temporal prevalent ICE in V. cholerae O1. Its occurrence and dispersion in V. cholerae O1 strains from different continents throughout more than two decades can be indicative of its role in the fitness of the current pandemic lineage.     

Features of the hmg 1 subfamily of genes encoding HMG-CoA reductase in potato

3-Hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) catalyzes a key step in isoprenoid metabolism leading to a range of compounds that are important for the growth, development and health of the plant. We have isolated 7 classes of genomic clones encoding HMGR from a potato genomic library. Comparison of nucleic acid sequences reveals a high degree of identity between all seven classes of clones and the potato hmg 1 gene described by Choi et al. (Plant Cell 4: 1333, 1992), indicating that all are members of the same subfamily in potato. A representative member (hmg 1.2) of the most abundant class of genomic clones was selected for further characterization. Transgenic tobacco and potato containing the -glucuronidase (GUS) reporter gene under the control of the hmg 1.2 promoter expressed GUS activity constitutively at a low level in many plant tissues. High levels of GUS activity were observed only in the pollen. GUS assays of isolated pollen, correlations of GUS activity with the HMGR activity of anthers, hmg 1.2 promoter deletion studies, and segregation analysis of the expression of hmg 1.2::GUS among the R₂ pollen of R₁ progeny plants demonstrated that the hmg 1.2 promoter controls pollen expression.     

Toxicity of cadmium and zinc on two microalgae, <Emphasis Type="Italic">Scenedesmus obliquus</Emphasis> and <Emphasis Type="Italic">Desmodesmus pleiomorphus</Emphasis>, from Northern Portugal

Aquatic environments often contain toxic heavy metals that may enter the food web via uptake by microalgae and eventually cause severe poisoning problems at higher trophic levels. The effects of Cd and Zn cations upon growth of two native green microalgal species, Scenedesmus obliquus and Desmodesmus pleiomorphus (previously isolated from a polluted site in Northern Portugal), were accordingly evaluated. Growth inhibition of the microalgal cells was determined following exposure for 96 h to several initial concentrations of aqueous solutions of either of those two metals. At the higher end of Cd and Zn experimental concentration ranges, a significant reduction in cell density was observed in the cultures; EC₅₀ values, calculated after fitting a Weibull model to the experimental data, were 0.058 and 1.92 mg L⁻¹ for Cd and 16.99 and 4.87 mg L⁻¹ for Zn in the case of S. obliquus and D. pleiomorphus, respectively. One observed that S. obliquus can tolerate higher Zn concentrations than D. pleiomorphus, but the reverse holds regarding exposure to Cd.     

Dasatinib Reduces Lung Inflammation and Fibrosis in Acute Experimental Silicosis

Silicosis is an occupational lung disease with no effective treatment. We hypothesized that dasatinib, a tyrosine kinase inhibitor, might exhibit therapeutic efficacy in silica-induced pulmonary fibrosis. Silicosis was induced in C57BL/6 mice by a single intratracheal administration of silica particles, whereas the control group received saline. After 14 days, when the disease was already established, animals were randomly assigned to receive DMSO or dasatinib (1 mg/kg) by oral gavage, twice daily, for 14 days. On day 28, lung morphofunction, inflammation, and remodeling were investigated. RAW 264.7 cells (a macrophage cell line) were incubated with silica particles, followed by treatment or not with dasatinib, and evaluated for macrophage polarization. On day 28, dasatinib improved lung mechanics, increased M2 macrophage counts in lung parenchyma and granuloma, and was associated with reduction of fraction area of granuloma, fraction area of collapsed alveoli, protein levels of tumor necrosis factor-α, interleukin-1β, transforming growth factor-β, and reduced neutrophils, M1 macrophages, and collagen fiber content in lung tissue and granuloma in silicotic animals. Additionally, dasatinib reduced expression of iNOS and increased expression of arginase and metalloproteinase-9 in silicotic macrophages. Dasatinib was effective at inducing macrophage polarization toward the M2 phenotype and reducing lung inflammation and fibrosis, thus improving lung mechanics in a murine model of acute silicosis.     

RXLR-mediated entry of Phytophthora sojae effector Avr1b into soybean cells does not require pathogen-encoded machinery

Effector proteins secreted by oomycete and fungal pathogens have been inferred to enter host cells, where they interact with host resistance gene products. Using the effector protein Avr1b of Phytophthora sojae, an oomycete pathogen of soybean (Glycine max), we show that a pair of sequence motifs, RXLR and dEER, plus surrounding sequences, are both necessary and sufficient to deliver the protein into plant cells. Particle bombardment experiments demonstrate that these motifs function in the absence of the pathogen, indicating that no additional pathogen-encoded machinery is required for effector protein entry into host cells. Furthermore, fusion of the Avr1b RXLR-dEER domain to green fluorescent protein (GFP) allows GFP to enter soybean root cells autonomously. The conclusion that RXLR and dEER serve to transduce oomycete effectors into host cells indicates that the >370 RXLR-dEER-containing proteins encoded in the genome sequence of P. sojae are candidate effectors. We further show that the RXLR and dEER motifs can be replaced by the closely related erythrocyte targeting signals found in effector proteins of Plasmodium, the protozoan that causes malaria in humans. Mutational analysis of the RXLR motif shows that the required residues are very similar in the motifs of Plasmodium and Phytophthora. Thus, the machinery of the hosts (soybean and human) targeted by the effectors may be very ancient.     

Nematicidal Bacteria Associated to Pinewood Nematode Produce Extracellular Proteases

Bacteria associated with the nematode Bursaphelenchus xylophilus, a pathogen of trees and the causal agent of pine wilt disease (PWD) may play a role in the disease. In order to evaluate their role (positive or negative to the tree), strains isolated from the track of nematodes from infected Pinus pinaster trees were screened, in vitro, for their nematicidal potential. The bacterial products, from strains more active in killing nematodes, were screened in order to identify and characterize the nematicidal agent. Forty-seven strains were tested and, of these, 21 strains showed capacity to produce extracellular products with nematicidal activity. All Burkholderia strains were non-toxic. In contrast, all Serratia strains except one exhibited high toxicity. Nematodes incubated with Serratia strains showed, by SEM observation, deposits of bacteria on the nematode cuticle. The most nematicidal strain, Serratia sp. A88copa13, produced proteases in the supernatant. The use of selective inhibitors revealed that a serine protease with 70 kDa was majorly responsible for the toxicity of the supernatant. This extracellular serine protease is different phylogenetically, in size and biochemically from previously described proteases. Nematicidal assays revealed differences in nematicidal activity of the proteases to different species of Bursaphelenchus, suggesting its usefulness in a primary screen of the nematodes. This study offers the basis for further investigation of PWD and brings new insights on the role bacteria play in the defense of pine trees against B. xylophilus. Understanding all the factors involved is important in order to develop strategies to control B. xylophilus dispersion.