Oxidative stress management in the filamentous, heterocystous, diazotrophic cyanobacterium, Anabaena PCC7120

Reactive oxygen species (ROS) are inevitably generated as by-products of respiratory/photosynthetic electron transport in oxygenic photoautotrophs. Unless effectively scavenged, these ROS can damage all cellular components. The filamentous, heterocystous, nitrogen-fixing strains of the cyanobacterium, Anabaena, serve as naturally abundant contributors of nitrogen biofertilizers in tropical rice paddy fields. Anabaena strains are known to tolerate several abiotic stresses, such as heat, UV, gamma radiation, desiccation, etc., that are known to generate ROS. ROS are detoxified by specific antioxidant enzymes like superoxide dismutases (SOD), catalases and peroxiredoxins. The genome of Anabaena PCC7120 encodes two SODs, two catalases and seven peroxiredoxins, indicating the presence of an elaborate antioxidant enzymatic machinery to defend its cellular components from ROS. This article summarizes recent findings and depicts important perspectives in oxidative stress management in Anabaena PCC7120.     

Schizophyllum commune: A New Organism in Eye Infection

Purpose

We report a case of mycotic keratitis caused by a rare fungus Schizophyllum commune.

Methods

Clinical examination, slit-lamp examination, and microbiological evaluation of the corneal ulcer were done, and its treatment outcome was studied. The fungal etiology was established by conventional microbiological techniques, polymerase chain reaction and speciation by DNA sequencing.

Results

Corneal scraping showed the presence of fungal filaments. The fungus was identified as S. commune based on DNA sequence analysis of the internal transcribed spacer region. The organism was susceptible to amphotericin B and voriconazole and demonstrated resistance to anidulafungin, itraconazole, and fluconazole. Therapeutic keratoplasty was performed but there was recurrence of the infection in the graft, which was controlled with topical voriconazole and intracameral amphotericin B. At the end of 3 months, the affected eye had developed phthisis bulbi.

Conclusion

The best of our knowledge, this is the first reported case of keratitis caused by the rare fungus S. commune. Management of these cases is difficult, and surgical procedures may be needed.     

Association of IL-6, TNF-α and IL-10 gene polymorphisms with type 2 diabetes mellitus

Type 2 diabetes mellitus (T2DM) is a metabolic pro-inflammatory disorder characterized by chronic hyperglycemia and increased levels of circulating cytokines suggesting a causal role of inflammation in its etiology. Polymorphism of cytokine genes including interleukin-6 (IL-6), tumour necrosis factor-α (TNF-α) and interleukin-10 (IL-10) were studied in T2DM patients as well as in normal healthy controls. Genomic DNA was isolated from both T2DM patients and controls followed by quantification and genotyping by polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) using suitable primers. The genotypic, allelic and carriage rate frequency distribution in patients and controls were analyzed by SPSS (version 15.0). Odd ratios with 95 % confidence interval was determined to describe the strength of association by logistic regression model. Double and triple combinations of genotypes were analyzed by χ² test. Gene–gene interaction and linkage disequilibrium tests were performed using SHEsis software. Individually, IL-6, TNF-α and IL-10 did not show any association. In double combination, IL-6 ?597 GA and TNF-α ?308 GG genotypes increased the risk up to 21 times and in triple combination IL-6 ?597 AA, TNF-α ?308 GG and IL-10 ?592 CA increased the risk of T2DM up to 314 times. In gene–gene interaction allele ‘A’ of all studied polymorphisms increased the risk of T2DM up to 1.41 times. Our results suggest that individuals having a haplotype combination of AA, GG and CA for IL-6, TNF-α and IL-10 gene polymorphisms will have higher susceptibility and be at greater risk of developing T2DM.     

Potential of Raloxifene in reversing osteoarthritis-like alterations in rat chondrocytes: An in vitro model study

The aim of this study was to investigate the effects of Raloxifene (Ral) on degeneration-related changes in osteoarthritis (OA)-like chondrocytes using two- and three-dimensional models. Five-azacytidine (Aza-C) was used to induce OA-like alterations in rat articular chondrocytes and the model was verified at molecular and macrolevels. Chondrocytes were treated with Ral (1, 5 and 10 μM) for 10 days. Caspase-3 activity, gene expressions of aggrecan, collagen II, alkaline phosphatase (ALP), collagen X, matrix metalloproteinases (MMP-13, MMP-3 and MMP-2), and MMP-13, MMP-3 and MMP-2 protein expressions were studied in two-dimensional model. Matrix deposition and mechanical properties of agarose-chondrocyte discs were evaluated in three-dimensional model. One μM Ral reduced expression of OA-related genes, decreased apoptosis, and MMP-13 and MMP-3 protein expressions. It also increased aggrecan and collagen II gene expressions relative to untreated OA-like chondrocytes. In three-dimensional model, 1 μM Ral treatment resulted in increased collagen deposition and improved mechanical properties, although a significant increase for sGAG was not observed. In summation, 1 μM Ral improved matrix-related activities, whereas dose increment reversed these effects except ALP gene expression and sGAG deposition. These results provide evidence that low-dose Ral has the potential to cease or reduce the matrix degeneration in OA.     

Synthesis of metallic nanoparticles using plant extracts

Biomolecules present in plant extracts can be used to reduce metal ions to nanoparticles in a single-step green synthesis process. This biogenic reduction of metal ion to base metal is quite rapid, readily conducted at room temperature and pressure, and easily scaled up. Synthesis mediated by plant extracts is environmentally benign. The reducing agents involved include the various water soluble plant metabolites (e.g. alkaloids, phenolic compounds, terpenoids) and co-enzymes. Silver (Ag) and gold (Au) nanoparticles have been the particular focus of plant-based syntheses. Extracts of a diverse range of plant species have been successfully used in making nanoparticles. In addition to plant extracts, live plants can be used for the synthesis. Here we review the methods of making nanoparticles using plant extracts. Methods of particle characterization are reviewed and potential applications of the particles in medicine are discussed.     

New Wrinkles on Polynucleotide Duplexes

Abstract

Most fibrous polynucleotides of general sequence exhibit secondary structures that are described adequately by regular helices with a repeated motif of only one nucleotide. Such helices exploit the fact that A:T, T:A, G:C, and C:G pairs are essentially isomorphous and have dyadically-related glycosylic bonds. Polynucleotides with regularly repeated base-sequences sometimes assume secondary structures with larger repeated motifs which reflect these base-sequences. The dinucleotide units of the Z-like forms of poly d(As⁴T):poly d(As⁴T), poly d(AC):poly d(GT) and poly d(GC):poly d(GC) are dramatic instances of this phenomenon. The wrinkled B and D forms of poly d(GC):poly d(GC) and poly d(AT):poly d(AT) are just as significant but more subtle examples. It is possible also to trap more exotic secondary structures in which the molecular asymmetric unit is even larger. There is, for example, a tetragonal form of poly d(AT):poly d(AT) which has unit cell dimensions a = b = 1.71nm, c= 7.40nm, γ = 90°. The C dimension corresponds to the pitch of a molecular helix which accommodates 24 successive nucleotide pairs arranged as a 4₃ helix of hexanucleotide duplexes. The great variety of nucleotide conformations which occur in these large asymmetric units has prompted us to describe them as pleiomeric, a term used in botany to describe whorls having more than the usual number of structures. Pleiomeric DNAs need not contain nucleotide conformations that are very different from one another. On the other hand, DNAs carrying nucleotides of very different conformation must be pleiomeric. This is because 4 nucleotides of different conformation are needed to join patches of secondary structure which are as different as A or B or Z. Differences in nucleotide structures may occur also between chains rather than within chains. In poly d(A):poly d(T), the purine nucleotides all contain Ci'-endo furanose rings and the pyrimidine nucleotides C2 '-endo rings. Analogous heteronomous structures may exist in DNA-RNA hybrids although these duplexes are also found to have symmetrical A-type conformations.     

Conformational and thermal stability of mature dimeric human myeloperoxidase and a recombinant monomeric form from CHO cells

Myeloperoxidase (MPO) is a lysosomal heme enzyme present in the azurophilic granules of human neutrophils and monocytes. It is a critical element of the human innate immune system by exerting antimicrobial effects. It is a disulfide bridged dimer with each monomer containing a light and a heavy polypeptide and its biosynthesis and intracellular transport includes several posttranslational processing steps. By contrast, MPO recombinantly produced in Chinese hamster ovary cell lines is monomeric, partially unprocessed and contains a N-terminal propeptide (proMPO). It mirrors a second form of MPO constitutively secreted from normal bone marrow myeloid precursors. In order to clarify the impact of posttranslational modifications on the structural integrity and enzymology of these two forms of human myeloperoxidase, we have undertaken an investigation on the conformational and thermal stability of leukocyte MPO and recombinant proMPO by using complementary biophysical techniques including UV-Vis, circular dichroism and fluorescence spectroscopy as well as differential scanning calorimetry. Mature leucocyte MPO exhibits a peculiar high chemical and thermal stability under oxidizing conditions but is significantly destabilized by addition of dithiothreitol. Unfolding of secondary and tertiary structure occurs concomitantly with denaturation of the heme cavity, reflecting the role of three MPO-typical heme to protein linkages and of six intra-chain disulfides for structural integrity by bridging N- and C-terminal regions of the protein. Recombinant monomeric proMPO follows a similar unfolding pattern but has a lower conformational and thermal stability. Spectroscopic and thermodynamic data of unfolding are discussed with respect to the known three-dimensional structure of leukocyte MPO as well as to known physiological roles.     

Loss of allostery and coenzyme B12 delivery by a pathogenic mutation in adenosyltransferase

ATP-dependent cob(I)alamin adenosyltransferase (ATR) is a bifunctional protein: an enzyme that catalyzes the adenosylation of cob(I)alamin and an escort that delivers the product, adenosylcobalamin (AdoCbl or coenzyme B(12)), to methylmalonyl-CoA mutase (MCM), resulting in holoenzyme formation. Failure to assemble holo-MCM leads to methylmalonic aciduria. We have previously demonstrated that only 2 equiv of AdoCbl bind per homotrimer of ATR and that binding of ATP to the vacant active site triggers ejection of 1 equiv of AdoCbl from an adjacent site. In this study, we have mimicked in the Methylobacterium extorquens ATR, a C-terminal truncation mutation, D180X, described in a patient with methylmalonic aciduria, and characterized the associated biochemical penalties. We demonstrate that while k(cat) and K(M)(Cob(I)) for D180X ATR are only modestly decreased (by 3- and 2-fold, respectively), affinity for the product, AdoCbl, is significantly diminished (400-fold), and the negative cooperativity associated with its binding is lost. We also demonstrate that the D180X mutation corrupts ATP-dependent cofactor ejection, which leads to transfer of AdoCbl from wild-type ATR to MCM. These results suggest that the pathogenicity of the corresponding human truncation mutant results from its inability to sequester AdoCbl for direct transfer to MCM. Instead, cofactor release into solution is predicted to reduce the capacity for holo-MCM formation, leading to disease.     

The complexities of obesity and diabetes with the development and progression of pancreatic cancer

Pancreatic cancer (PC) is one of the most lethal malignant diseases with the worst prognosis. It is ranked as the fourth leading cause of cancer-related deaths in the United States. Many risk factors have been associated with PC. Interestingly, large numbers of epidemiological studies suggest that obesity and diabetes, especially type-2 diabetes, are positively associated with increased risk of PC. Similarly, these chronic diseases (obesity, diabetes, and cancer) are also a major public health concern. In the U.S. population, 50 percent are overweight, 30 percent are medically obese, and 10 percent have diabetes mellitus (DM). Therefore, obesity and DM have been considered as potential risk factors for cancers; however, the focus of this article is restricted to PC. Although the mechanisms responsible for the development of these chronic diseases leading to the development of PC are not fully understood, the biological importance of the activation of insulin, insulin like growth factor-1 (IGF-1) and its receptor (IGF-1R) signaling pathways in insulin resistance mechanism and subsequent induction of compensatory hyperinsulinemia has been proposed. Therefore, targeting insulin/IGF-1 signaling with anti-diabetic drugs for lowering blood insulin levels and reversal of insulin resistance could be useful strategy for the prevention and/or treatment of PC. A large number of studies have demonstrated that the administration of anti-diabetic drugs such as metformin and thiazolidinediones (TZD) class of PPAR-γ agonists decreases the risk of cancers, suggesting that these agents might be useful anti-tumor agents for the treatment of PC. In this review article, we will discuss the potential roles of metformin and TZD anti-diabetic drugs as anti-tumor agents in the context of PC and will further discuss the complexities and the possible roles of microRNAs (miRNAs) in the pathogenesis of obesity, diabetes, and PC.