Cyr61 induces IL-6 production by fibroblast-like synoviocytes promoting Th17 differentiation in rheumatoid arthritis

Cysteine-rich protein 61 (Cyr61)/CCN1 is a product of an immediate early gene and functions in mediating cell adhesion and inducing cell migration. We previously showed that increased production of Cyr61 by fibroblast-like synoviocytes (FLS) in rheumatoid arthritis (RA) promotes FLS proliferation and participates in RA pathogenesis with the IL-17-dependent pathway. However, whether Cyr61 in turn regulates Th17 cell differentiation and further enhances inflammation of RA remained unknown. In the current study, we explored the potential role of Cyr61 as a proinflammatory factor in RA pathogenesis. We found that Cyr61 treatment dramatically induced IL-6 production in FLS isolated from RA patients. Moreover, IL-6 production was attenuated by Cyr61 knockdown in FLS. Mechanistically, we showed that Cyr61 activated IL-6 production via the αvβ5/Akt/NF-κB signaling pathway. Further, using a coculture system consisting of purified CD4(+) T cells and RA FLS, we found that RA FLS stimulated Th17 differentiation, and the pro-Th17 differentiation effect of RA FLS can be attenuated or stimulated by Cyr61 RNA interference or addition of exogenous Cyr61, respectively. Finally, using the collagen-induced arthritis animal model, we showed that treatment with the anti-Cyr61 mAb led to reduction of IL-6 levels, decrease of Th17 response, and attenuation of inflammation and disease progression in vivo. Taken together, our results reveal a novel role of Cyr61 in promoting Th17 development in RA via upregulation of IL-6 production by FLS, thus adding a new layer into the functional interplay between FLS and Th17 in RA pathogenesis. Our study also suggests that targeting of Cyr61 may represent a novel strategy in RA treatment.     

Complex Evolutionary Relationships Among Four Classes of Modular RNA-Binding Splicing Regulators in Eukaryotes: The hnRNP, SR, ELAV-Like and CELF Proteins

Alternative RNA splicing in multicellular organisms is regulated by a large group of proteins of mainly unknown origin. To predict the functions of these proteins, classification of their domains at the sequence and structural level is necessary. We have focused on four groups of splicing regulators, the heterogeneous nuclear ribonucleoprotein (hnRNP), serine?Carginine (SR), embryonic lethal, abnormal vision (ELAV)-like, and CUG-BP and ETR-like factor (CELF) proteins, that show increasing diversity among metazoa. Sequence and phylogenetic analyses were used to obtain a broader understanding of their evolutionary relationships. Surprisingly, when we characterised sequence similarities across full-length sequences and conserved domains of ten metazoan species, we found some hnRNPs were more closely related to SR, ELAV-like and CELF proteins than to other hnRNPs. Phylogenetic analyses and the distribution of the RRM domains suggest that these proteins diversified before the last common ancestor of the metazoans studied here through domain acquisition and duplication to create genes of mixed evolutionary origin. We propose that these proteins were derived independently rather than through the expansion of a single protein family. Our results highlight inconsistencies in the current classification system for these regulators, which does not adequately reflect their evolutionary relationships, and suggests that a domain-based classification scheme may have more utility.     

Cynandione A mitigates ischemic injuries in rats with cerebral ischemia

Cynandione A, an acetophenone from the roots of Cynanchum auriculatum and other species in the genus attenuates neurotoxicity of a variety of neurotoxic agents such as l-glutamate in vitro. In this study, we sought to further characterize the neuroprotective effects of cynandione A and other acetophenones from the roots of C. auriculatum in pheochromocytoma tumor cell line PC12 and investigate whether cynandione A protected against ischemic injuries in rats with experimentally induced cerebral ischemia. Viability assays using the 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophen-yl)-2H-tetrazolium monosodium salt method and lactate dehydrogenase (LDH) release assays showed that cynandione A dose-dependently attenuated glutamate-induced cytotoxicity. Comparative proteomic analysis by two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization-time of flight MS/MS of PC12 cells treated with cynandione A showed 10 μM cynandione A caused broad changes in protein expression in PC12 cells including down-regulation of high mobility group box 1 (HMGB1) and dihydropyrimidinase-like 2 (DPYSL2). Immunoblotting studies showed that 10 μM cynandione A aborted glutamate-induced increase in DPYSL2 and HMGB1 levels in PC12 cells and 30 mg/kg cynandione A also attenuated the rise in HMGB1 levels and mitigated DPYSL2 cleavage in brain tissues of rats with cerebral ischemia. Furthermore, rats with cerebral ischemia treated with 30 mg/kg cynandione A exhibited markedly improved neurological deficit scores at 24 and 72 h compared with control and a 7.2% reduction in cerebral infarction size at 72 h (p < 0.05 vs. control). Our findings demonstrated that cynandione A mitigated ischemic injuries and should be further explored as a neuroprotective agent for ischemic stroke.     

Complete Genome Sequence of a Vero Cell-Adapted Isolate of Porcine Epidemic Diarrhea Virus in Eastern China

In early 2012, a widespread porcine epidemic diarrhea virus (PEDV) occurred in eastern China. A cell-adapted isolate, SD-M, was at the four-passage level of virulent field strain SD, which was isolated from a 2-day-old dead suckling piglet that had suffered from severe diarrhea in Shandong Province, China. We report here the complete genome sequence of SD-M. This sequence will promote a better understanding of the molecular pathogenesis of PEDV.     

The N-Terminal Region of IFITM3 Modulates Its Antiviral Activity by Regulating IFITM3 Cellular Localization

Interferon-inducible transmembrane (IFITM) protein family members IFITM1, -2, and -3 restrict the infection of multiple enveloped viruses. Significant enrichment of a minor IFITM3 allele was recently reported for patients who were hospitalized for seasonal and 2009 H1N1 pandemic flu. This IFITM3 allele lacks the region corresponding to the first amino-terminal 21 amino acids and is unable to inhibit influenza A virus. In this study, we found that deleting this 21-amino-acid region relocates IFITM3 from the endosomal compartments to the cell periphery. This finding likely underlies the lost inhibition of influenza A virus that completes its entry exclusively within endosomes at low pH. Yet, wild-type IFITM3 and the mutant with the 21-amino-acid deletion inhibit HIV-1 replication equally well. Given the pH-independent nature of HIV-1 entry, our results suggest that IFITM3 can inhibit viruses that enter cells via different routes and that its N-terminal region is specifically required for controlling pH-dependent viruses.     

A 205-Nucleotide Deletion in the 3′ Untranslated Region of Avian Leukosis Virus Subgroup J,Currently Emergent in China,Contributes to Its Pathogenicity

In the past 5 years, an atypical clinical outbreak of avian leukosis virus subgroup J (ALV-J), which contains a unique 205-nucleotide deletion in its 3′ untranslated region (3′UTR), has become epidemic in chickens in China. To determine the role of the 205-nucleotide deletion in the pathogenicity of ALV-J, a pair of viruses were constructed and rescued. The first virus was an ALV-J Chinese isolate (designated HLJ09SH01) containing the 205-nucleotide deletion in its 3′UTR. The second virus was a chimeric clone in which the 3′UTR contains a 205-nucleotide sequence corresponding to a region of the ALV-J prototype virus. The replication and pathogenicity of the rescued viruses (rHLJ09SH01 and rHLJ09SH01A205) were investigated. Compared to rHLJ09SH01A205, rHLJ09SH01 showed a moderate growth advantage in vitro and in vivo, in addition to exhibiting a higher oncogenicity rate and lethality rate in layers and broilers. Increased vascular endothelial growth factor A (VEGF-A) and vascular endothelial growth receptor subtype 2 (VEGFR-2) expression was induced by rHLJ09SH01 more so than by rHLJ09SH01A205 during early embryonic vascular development, but this increased expression disappeared when the expression levels were normalized to the viral levels. This finding suggests that the expression of VEGF-A and VEGFR-2 is associated with viral replication and may also represent a novel molecular mechanism underlying the oncogenic potential of ALV-J. Overall, our findings not only indicate that the unique 205-nucleotide deletion in the ALV-J genome occurred naturally in China and contributes to increased pathogenicity but also point to the possible mechanism of ALV-J-induced oncogenicity.     

Complete Genome Sequence of Staphylococcus aureus Bacteriophage GH15

GH15 is a polyvalent phage that shows activity against a wide range of Staphylococcus aureus strains. In this work, the complete genome sequence of GH15 was determined. With a genome size of 139,806 bp (double-stranded DNA), GH15 is the largest staphylococcal phage sequenced to date. The complete genome encodes 214 open reading frames (ORFs) and 4 tRNAs. The closest relatives are the class III staphylococcal myobacteriophages, including K, A5W, ISP, Sb-1, and G1. Interestingly, although corresponding gene sequences demonstrate very high similarity, all the introns and inteins present in the phages listed above are absent in GH15. As such, GH15 can be considered phylogenetically unique among the staphylococcal myobacteriophages, indicating the diversity of this family.