Ameliorating ER-stress attenuates Aeromonas hydrophila-induced mitochondrial dysfunctioning and caspase mediated HKM apoptosis in Clarias batrachus.

Endoplasmic reticulum (ER)-stress and unfolding protein response (UPR) has not been implied in Aeromonas hydrophila-pathogenicity. We report increased expression of the ER-stress markers: CHOP, BiP and phospho-eIF2α in A. hydrophila-infected headkidney macrophages (HKM) in Clarias batrachus. Pre-treatment with ER-stress inhibitor, 4-PBA alleviated ER-stress and HKM apoptosis suggesting ER-UPR critical for the process. The ER-Ca(2+) released via inositol-triphosphate and ryanodine receptors induced calpain-2 mediated superoxide ion generation and consequent NF-κB activation. Inhibiting NF-κB activation attenuated NO production suggesting the pro-apoptotic role of NF-κB on HKM pathology. Calpain-2 activated caspase-12 to intensify the apoptotic cascade through mitochondrial-membrane potential (ψm) dissipation and caspase-9 activation. Altered mitochondrial ultra-structure consequent to ER-Ca(2+) uptake via uniporters reduced ψm and released cytochrome C. Nitric oxide induced the cGMP/PKG-dependent activation of caspase-8 and truncated-Bid formation. Both the caspases converge onto caspase-3 to execute HKM apoptosis. These findings offer a possible molecular explanation for A. hydrophila pathogenicity.

Identification of autoantibodies against transthyretin for the screening and diagnosis of rheumatoid arthritis.

Rheumatoid arthritis (RA) is a chronic, autoimmune, systemic and inflammatory rheumatic disease that leads to inflammation of the joints and surrounding tissues. Identification of novel protein(s) associated with severity of RA is a prerequisite for better understanding of pathogenesis of this disease that may also have potential to serve as novel biomarkers in the diagnosis of RA. Present study was undertaken to compare the amount of autoantigens and autoantibodies in the plasma of RA patients in comparison to healthy controls. Plasma samples were collected from the patients suffering from RA, Osteoarthritis (OA), Systemic lupus erythematosus (SLE) and healthy volunteers. The screening of plasma proteins were carried out using 2-dimensional gel electrophoresis followed by identification of differentially expressed protein by MALDI-TOF MS/MS. Among several differentially expressed proteins, transthyretin (TTR) has been identified as one of the protein that showed significantly up regulated expression in the plasma of RA patients. The results were further validated by Western blot analysis and ELISA. In comparison to OA synovium, an exclusive significantly high expression of TTR in RA has been validated through IHC, Western blotting and IEM studies. Most importantly, the increase in expression of TTR with the progression of severity of RA condition has been observed. The autoantibodies against TTR present in the RA plasma were identified using immunoprecipitation-Western methods. The significant production of autoantibodies was validated by ELISA and Western blot analysis using recombinant pure protein of TTR. Hence, these novel observations on increase in TTR expression with the increase in severity of RA conditions and significant production of autoantibodies against TTR clearly suggest that a systematic studies on the role TTR in the pathogenesis of RA is immediately required and TTR may be used as a serum diagnostic marker together with other biochemical parameters and clinical symptoms for RA screening and diagnosis.

Magnetite (Fe3O4) nanocrystals affect the expression of genes involved in the TGF-beta signalling pathway.

An understanding of interaction of nanomaterials with living systems is fundamental to address nanosafety issues, which, in turn will dictate the future prospects of nanomedicine. Herein, we examine the molecular effects of uptake of Magnetite (Fe(3)O(4)) Nanocrystals (MNC) using a transcriptomics approach. The uptake of MNC was studied by electron microscopy. This was followed by transcriptional profiling using whole genome microarrays, functional analysis of microarray data, real time PCR and biochemical assay for CASP9. Transcriptional profiling revealed 69 genes to be differentially expressed upon MNC treatment. Many of these genes are associated with TGF-beta signaling and include ID1, ID2, ID3, CASP9, SMAD6 and SMAD7, which are important negative regulators of signaling pathways involved in development and tumorigenesis. Moreover, upon treatment with MNC, expression of CASP9 was also found to decrease in a dose dependent manner. This approach could help us to identify specific effects of MNC upon cells and give us simultaneous clues about their biocompatibility and therapeutic potential. The MNC can specifically interfere with TGF-beta signaling by inhibiting the expression of ID and SMAD genes. As TGF-beta signaling invokes different responses in undifferentiated cells and adult tissues in a cell-type specific manner, our findings have far reaching implications in cellular development, differentiation and cancer.

Possible role of amyloid-beta, adenine nucleotide translocase and cyclophilin-D interaction in mitochondrial dysfunction of Alzheimer's disease.

Alzheimer's disease (AD) is a common neurodegenerative disease characterized by both extra- as well as intracellular deposition of amyloid beta peptides (Abeta). The accumulation of Abeta in mitochondria is associated with mitochondrial dysfunction and oxidative stress in AD. Recent evidences suggest the involvement of Abeta interaction with mitochondrial proteins such as cyclophilin-D (CypD) in oxidative stress, mitochondrial permeability transition (MPT) and Alzheimer's associated neurodegeneration. The present study is an effort to elucidate the molecular interaction of Abeta with other proteins involved in MPT like adenine nucleotide translocase (ANT). Based on our prediction for sub-cellular localization using WolfPSORT and other experimental evidences, we suggest that Abeta molecules localize in mitochondrial inner membrane in close vicinity with ANT. Our simulation study for protein-protein interaction clearly suggests that the ANT-Abeta interaction is stronger than CypD-Abeta interaction. Further the lipophilic nature and evidences regarding the localization of Abeta in the mitochondrial inner-membrane also support the possibility of strong interaction between ANT and Abeta. Interaction between ANT and Abeta may affect normal physiological function of ANT i.e. transport of ATP and ADP. Since both the CypD-Abeta as well as ANT-Abeta interaction are energetically favorable and both CypD and ANT are associated with the regulation of MPT, the functional impact of both these interactions warrants more in-depth investigations for elucidating the mechanisms involved in Abeta-induced oxidative stress.

Tubular aggregate myopathy: a rare form of myopathy.

Tubular aggregate myopathy (TAM) is a rare form of myopathy with an autosomal dominant or recessive pattern. Four rare cases of TAM are described. All patients presented with muscle aches and pains, sometimes cramps. Muscle biopsies were snap frozen and processed for routine, special, enzyme, and immunohistochemistry. Tissue was also processed for electron microscopy. Muscle biopsy revealed similar changes characterized by subsarcolemmal accumulation of granular material that stained red with modified Gomori trichrome stain, intense blue with nicotinamide adenine dinucleotide-tetrazolium reductase, but was non-reactive to succinyl dehydrogenase and cytochrome oxidase stains. Ultrastructural examination showed aggregates of hexagonal tubules in the subsarcolemmal region, which are pathognomonic of this entity. This report highlights the importance of histochemistry and electron microscopy for accurate diagnosis; otherwise TAM can be misdiagnosed on clinical grounds as a metabolic or mitochondrial myopathy.

High salinity reduces the content of a highly abundant 23-kDa protein of the mangrove Bruguiera parviflora.

A significant decrease in the amount of a protein, whose migration in two-dimensional gel electrophoresis corresponds to an apparent molecular mass of 23 kDa and pI = 6.5, was observed in leaves of NaCl-treated Bruguiera parviflora (Roxb.) Wt. & Arn. ex Griff. seedlings. This particular salt-sensitive protein, designated as SSP-23, almost disappeared after 45 days of treatment in 400 mM NaCl as compared to untreated seedlings (0 mM NaCl) where the presence of the protein was significant. A polyclonal antibody raised against the 23-kDa protein was used to determine the subcellular localization of this protein in leaves by cross-reaction with proteins from isolated chloroplasts, mitochondria, peroxisomes and cytosol fractions on Western blots. SSP-23 was confirmed to be localized in the cytosol by immunoblotting. The disappearance of SSP-23 as a result of high NaCl treatment suggests that this protein is salt-sensitive and has a possible role in salt adaptation.