Up-Regulation of Mcl-1 and Bak by Coronavirus Infection of Human, Avian and Animal Cells Modulates Apoptosis and Viral Replication

Virus-induced apoptosis and viral mechanisms that regulate this cell death program are key issues in understanding virus-host interactions and viral pathogenesis. Like many other human and animal viruses, coronavirus infection of mammalian cells induces apoptosis. In this study, the global gene expression profiles are first determined in IBV-infected Vero cells at 24 hours post-infection by Affymetrix array, using avian coronavirus infectious bronchitis virus (IBV) as a model system. It reveals an up-regulation at the transcriptional level of both pro-apoptotic Bak and pro-survival myeloid cell leukemia-1 (Mcl-1). These results were further confirmed both in vivo and in vitro, in IBV-infected embryonated chicken eggs, chicken fibroblast cells and mammalian cells at transcriptional and translational levels, respectively. Interestingly, the onset of apoptosis occurred earlier in IBV-infected mammalian cells silenced with short interfering RNA targeting Mcl-1 (siMcl-1), and was delayed in cells silenced with siBak. IBV progeny production and release were increased in infected Mcl-1 knockdown cells compared to similarly infected control cells, while the contrary was observed in infected Bak knockdown cells. Furthermore, IBV infection-induced up-regulation of GADD153 regulated the expression of Mcl-1. Inhibition of the mitogen-activated protein/extracellular signal-regulated kinase (MEK/ERK) and phosphoinositide 3-kinase (PI3K/Akt) signaling pathways by chemical inhibitors and knockdown of GADD153 by siRNA demonstrated the involvement of ER-stress response in regulation of IBV-induced Mcl-1 expression. These results illustrate the sophisticated regulatory strategies evolved by a coronavirus to modulate both virus-induced apoptosis and viral replication during its replication cycle

A Recombinant Avian Infectious Bronchitis Virus Expressing a Heterologous Spike Gene Belonging to the 4/91 Serotype

We have shown previously that replacement of the spike (S) gene of the apathogenic IBV strain Beau-R with that from the pathogenic strain of the same serotype, M41, resulted in an apathogenic virus, BeauR-M41(S), that conferred protection against challenge with M41 [1]. We have constructed a recombinant IBV, BeauR-4/91(S), with the genetic backbone of Beau-R but expressing the spike protein of the pathogenic IBV strain 4/91(UK), which belongs to a different serogroup as Beaudette or M41. Similar to our previous findings with BeauR-M41(S), clinical signs observations showed that the S gene of the pathogenic 4/91 virus did not confer pathogenicity to the rIBV BeauR-4/91(S). Furthermore, protection studies showed there was homologous protection; BeauR-4/91(S) conferred protection against challenge with wild type 4/91 virus as shown by the absence of clinical signs, IBV RNA assessed by qRT-PCR and the fact that no virus was isolated from tracheas removed from birds primarily infected with BeauR-4/91(S) and challenged with IBV 4/91(UK). A degree of heterologous protection against M41 challenge was observed, albeit at a lower level

Legionella pneumophila Secretes a Mitochondrial Carrier Protein during Infection

The Mitochondrial Carrier Family (MCF) is a signature group of integral membrane proteins that transport metabolites across the mitochondrial inner membrane in eukaryotes. MCF proteins are characterized by six transmembrane segments that assemble to form a highly-selective channel for metabolite transport. We discovered a novel MCF member, termed Legionella nucleotide carrier Protein (LncP), encoded in the genome of Legionella pneumophila, the causative agent of Legionnaire's disease. LncP was secreted via the bacterial Dot/Icm type IV secretion system into macrophages and assembled in the mitochondrial inner membrane. In a yeast cellular system, LncP induced a dominant-negative phenotype that was rescued by deleting an endogenous ATP carrier. Substrate transport studies on purified LncP reconstituted in liposomes revealed that it catalyzes unidirectional transport and exchange of ATP transport across membranes, thereby supporting a role for LncP as an ATP transporter. A hidden Markov model revealed further MCF proteins in the intracellular pathogens, Legionella longbeachae and Neorickettsia sennetsu, thereby challenging the notion that MCF proteins exist exclusively in eukaryotic organisms

Distal locking stem for revision femoral loosening and peri-prosthetic fractures

Revision total hip arthroplasty in the setting of a large proximal femoral deficiency or a peri-prosthetic fracture remains a challenging problem. We describe the development, surgical technique and the use of cementless revision stems with distal inter-locking screws to provide immediate stability of the femoral implant. Results were assessed in a large multicentre French study conducted with the french hip and knee surgery society (SFHG). We retrospectively reviewed 725 revisions using interlocking stems from 14 French orthopaedic departments. Seven different stems were used in this series. In-patient records were retrieved, and in addition to demographic data the indication for revision, the preoperative and postoperative PMA and Harris hip scores were documented. The bone deficiency was classified on the basis of the French National Orthopaedic Meeting (SOFCOT) classification. Intraoperative complications and problems if any were retrieved from operative notes. Clinical status and radiographs at the final follow-up were evaluated, paying special attention to the metaphyseal filling index. Average follow-up was 4.5 years. As for the clinical results, the mean Harris hip score at last follow-up was 81. Therefore, it increased by an average of 31 points. Bone reconstruction was assessed on the cortico-medullary index in the metaphyseal area and at mid-shaft increasing from 36 to 45 and 54 to 63, respectively. Radiologically, 637 implants were stable, and 40 demonstrated subsidence. Forty-eight implants have been revised. We found a significant relation between the metaphyseal filling index, the stability of the stem and the quality of bone reconstruction. Results were analysed with respect to three groups of stems: group 1 was a straight, partially HA-coated implant; group 2 was a curved, fully HA-coated implant; and group 3 was a curved, partially-coated implant. Group 1 showed a significantly higher rate of failure when compared with the others types of implants. Group 2 had better functional results than group 3, which in turn reported better results than group 1. With regard to implant fixation, group 2 had significantly better results. Both groups 2 and 3 induced less thigh pain than group 1. The distal interlocking stem has shown promising results for femoral revisions. The advantages are initial axial and rotational stability and consistent bony in-growth owing to hydroxyapatite coating. Distal locked stems are mainly indicated to treat complex femoral revision with severe bone loss and peri-prosthetic fractures