PEGylated human plasma fibronectin is proteolytically stable,supports cell adhesion,cell migration,focal adhesion assembly,and fibronectin fibrillogenesis

Delayed wound healing in many chronic wounds has been linked to the degradation of fibronectin (FN) by abnormally high protease levels. We sought to develop a proteolytically stable and functionally active form of FN. For this purpose, we conjugated 3.35 kDa polyethylene glycol diacrylate (PEGDA) to human plasma fibronectin (HPFN). Conjugation of PEGDA to HPFN or HPFN PEGylation was characterized by an increase of approximately 16 kDa in the average molecular weight of PEGylated HPFN compared to native HPFN in SDS‐PAGE gels. PEGylated HPFN was more resistant to α chymotrypsin or neutrophil elastase digestion than native HPFN: after 30 min incubation with α chymotrypsin, 56 and 90% of native and PEGylated HPFN respectively remained intact. PEGylated HPFN and native HPFN supported NIH 3T3 mouse fibroblast adhesion and spreading, migration and focal adhesion formation in a similar manner. Fluorescence microscopy showed that both native and PEGylated HPFN in the culture media were assembled into extracellular matrix (ECM) fibrils. Interestingly, when coated on surfaces, native but not PEGylated HPFN was assembled into the ECM of fibroblasts. The proteolytically stable PEGylated HPFN developed herein could be used to replenish FN levels in the chronic wound bed and promote tissue repair. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29: 493–504, 2013     

Proteome profile of a pandemic Vibrio parahaemolyticus SC192 strain in the planktonic and biofilm condition

Vibrio parahaemolyticus is one of the leading causative agents of foodborne diseases in humans. In this study, the proteome profiles of the pandemic strain V. parahaemolyticus SC192 belonging to the O3:K6 serovar during the planktonic and biofilm stages were analyzed by two-dimensional liquid chromatography coupled to tandem mass spectrometry. This non-gel-based multidimensional protein identification technology approach identified 45.5% of the proteome in the reference genome V. parahaemolyticus RIMD 2210633. This is the largest proteome coverage obtained so far in V. parahaemolyticus and provides evidence for expression of 27% of the hypothetical proteins. Comparison of the planktonic and biofilm proteomes based on their cluster of orthologous groups, gene ontologies and KEGG pathways provides basic information on biofilm specific functions and pathways. To the authors’ knowledge, this is the first study to generate a global proteome profile of the pandemic strain of V. parahaemolyticus and the method reported here could be used to rapidly obtain a snapshot of the proteome of any microorganism at a given condition.     

3-Nitrotoluene dioxygenase from Diaphorobacter sp. strains: cloning,sequencing and evolutionary studies

The first step in the degradation of 3-nitrotoluene by Diaphorobacter sp. strain DS2 is the dihydroxylation of the benzene ring with the concomitant removal of nitro group. This is catalyzed by a dioxygenase enzyme system. We report here the cloning and sequencing of the complete dioxygenase gene with its putative regulatory sequence from the genomic DNA of Diaphorobacter sp. strains DS1, DS2 and DS3. Analysis of the 5 kb DNA stretch that was cloned, revealed five complete open reading frames (ORFs) encoding for a reductase, a ferredoxin and two dioxygenase subunits with predicted molecular weights (MW) of 35, 12, 50 and 23 kDa respectively. A regulatory protein was also divergently transcribed from the reductase subunit and has a predicated MW of 34 kDa. Presence of parts of two functional ORFs in between the reductase and the ferredoxin subunits reveals an evolutionary route from a naphthalene dioxygenase like system of Ralstonia sp. strain U2. Further a 100 % identity of its ferredoxin subunit reveals its evolution via dinitrotoluene dioxygenase like system present in Burkholderia cepacia strain R34. A modeled structure of oxygenase_3NT from strain DS2 was generated using nitrobenzene dioxygenase as a template. The modeled structure only showed minor changes at its active site. Comparison of growth patterns of strains DS1, DS2 and DS3 revealed that Diaphorobacter sp. strain DS1 has been evolved to degrade 4-nitrotoluene better by an oxidative route amongst all three strains.     

The impact of successful cataract surgery on quality of life, household income and social status in South India

Background

To explore the hypothesis that sight restoring cataract surgery provided to impoverished rural communities will improve not only visual acuity and vision-related quality of life (VRQoL) but also poverty and social status.

Methods

Participants were recruited at outreach camps in Tamil Nadu, South India, and underwent free routine manual small incision cataract surgery (SICS) with intra-ocular lens (IOL) implantation, and were followed up one year later. Poverty was measured as monthly household income, being engaged in income generating activities and number of working household members. Social status was measured as rates of re-marriage amongst widowed participants. VRQoL was measured using the IND-VFQ-33. Associations were explored using logistic regression (SPSS 19).

Results

Of the 294 participants, mean age ± standard deviation (SD) 60±8 years, 54% men, only 11% remained vision impaired at follow up (67% at baseline; p<0.001). At one year, more participants were engaged in income generating activities (44.7% to 77.7%; p<0.001) and the proportion of households with a monthly income <1000 Rps. decreased from 50.5% to 20.5% (p<0.05). Overall VRQoL improved (p<0.001). Participants who had successful cataract surgery were less likely to remain in the lower categories of monthly household income (OR 0.05–0.22; p<0.02) and more likely to be engaged in income earning activities one year after surgery (OR 3.28; p = 0.006). Participants widowed at baseline who had successful cataract surgery were less likely to remain widowed at one year (OR 0.02; p = 0.008).

Conclusion

These findings indicate the broad positive impact of sight restoring cataract surgery on the recipients’ as well as their families’ lives. Providing free high quality cataract surgery to marginalized rural communities will not only alleviate avoidable blindness but also - to some extent - poverty in the long run.     

Presynaptic learning and memory with a persistent firing neuron and a habituating synapse: a model of short term persistent habituation

Our paper explores the interaction of persistent firing axonal and presynaptic processes in the generation of short term memory for habituation. We first propose a model of a sensory neuron whose axon is able to switch between passive conduction and persistent firing states, thereby triggering short term retention to the stimulus. Then we propose a model of a habituating synapse and explore all nine of the behavioral characteristics of short term habituation in a two neuron circuit. We couple the persistent firing neuron to the habituation synapse and investigate the behavior of short term retention of habituating response. Simulations show that, depending on the amount of synaptic resources, persistent firing either results in continued habituation or maintains the response, both leading to longer recovery times. The effectiveness of the model as an element in a bio-inspired memory system is discussed.     

Glycosaminoglycans alter the conformation of interferon-gamma

Interferon-gamma (IFN-gamma) is a potent immunomodulatory cytokine whose physiological roles are modulated by the extracellular matrix. Here, circular dichroism and fluorescence spectroscopic techniques are used to demonstrate that low molecular weight heparin and chondroitin sulfate cause significant changes in secondary and tertiary structure of IFN-gamma. The results suggest that heparin and chondroitin sulfate modulate IFN-gamma activity by causing structural changes in the IFN-gamma dimer.     

Computational and Structural Investigation of Deleterious Functional SNPs in Breast Cancer BRCA2 Gene

In this work, we have analyzed the genetic variation that can alter the expression and the function in BRCA2 gene using computational methods. Out of the total 534 SNPs, 101 were found to be non synonymous (nsSNPs). Among the 7 SNPs in the untranslated region, 3 SNPs were found in 5′ and 4 SNPs were found in 3′ un-translated regions (UTR). Of the nsSNPs 20.7% were found to be damaging by both SIFT and PolyPhen server among the 101 nsSNPs investigated. UTR resource tool suggested that 2 SNPs in the 5′ UTR region and 4 SNPs in the 3′ UTR regions might change the protein expression levels. The mutation from asparagine to isoleucine at the position 3124 of the native protein of BRCA2 gene was most deleterious by both SIFT and PolyPhen servers. A structural analysis of this mutated protein and the native protein was made which had an RMSD value of 0.301 nm. Based on this work, we proposed that this most deleterious nsSNP with an SNPid rs28897759 is an important candidate for the cause of breast cancer by BRCA2 gene.     

Association of classical semen parameters, sperm DNA fragmentation index, lipid peroxidation and antioxidant enzymatic activity of semen in ram-lambs

The objective was to determine relationships among classical semen characteristics, sperm chromatin structure assay (SCSA), lipid peroxidation and antioxidant enzymatic activity in ram-lamb semen. Fifty-seven ram-lambs were electroejaculated, and routine semen evaluation was conducted (as part of a breeding soundness evaluation). The percentage of sperm DNA fragmentation index (%DFI) and the percentage of sperm with abnormally high DNA stainability (HDS; immature spermatozoa) were determined by SCSA using the metachromatic properties of acridine orange. Semen was centrifuged at 800 x g for 15 min to separate spermatozoa and seminal plasma and the aliquots were stored at -70 degrees C until analyzed. Lipid peroxidation, superoxide dismutase (SOD), and glutathione peroxidase (GPx) levels in seminal plasma and spermatozoa were measured by spectrophotometric assays. The classical semen parameters were negatively related to lipid peroxidation and GPx activity in spermatozoa; motility and morphology were negatively related to %DFI (P < 0.05). Based on Kruskal-Wallis pair-wise comparison of median values among breeding soundness outcome groups, %DFI was lower in the satisfactory group compared to other groups (P < 0.05) and the lipid peroxidation and GPx activity in seminal plasma and spermatozoa were lower in satisfactory and questionable groups (P < 0.05). However, the SOD was lower in the unsatisfactory group (P < 0.05). In summary, classical semen parameters were negatively related to % DFI, lipid peroxidation and GPx activity in ram-lamb spermatozoa and seminal plasma. There were indications that SOD and GPx have crucial protective roles against the toxic effect of reactive oxygen species (ROS) in ram-lamb semen.     

Transient Accumulation of Metabolic Intermediates of p-Cresol in the Culture Medium by a Pseudomonas sp. Strain A Isolated from a Sewage Treatment Plant

Summary Activated sludge from a sewage treatment plant in Kanpur, India, was screened for bacterial strains metabolizing p-cresol exclusively under aerobic conditions. One such isolate was identified to be belonging to the genus Pseudomonas based on morphological and physiological criteria as well as 16S ribosomal RNA gene sequence analysis. Two intermediates were identified from the culture medium during the growth phase of Pseudomonas sp. strain A that indicated that the strain degraded p-cresol via the protocatechuate (PCA) pathway. p-Cresol was rapidly converted into p-hydroxybenzaldehyde (PHB) during early growth phase, which was later utilized after p-cresol depletion. p-Hydroxybenzoate (PHBA) accumulation was observed during the later stages of exponential growth phase. Kinetic constants for the degradation of p-cresol were determined using Haldane’s model. High μ_max and inhibitory constant (K_I) values along with the observed accumulation of significant amounts of PHB in culture filtrates seem to indicate that the isolated Pseudomonas sp. strain A may be of potential use in biotransformations.     

Translocation of metals in pea plants grown on various amendment of electroplating industrial sludge

A pot-culture experiment was conducted to observe the effects of acidic sludge addition to the soils on bioavailability and uptake of heavy metals in different parts of pea plant as well as its influence on the growth of that plant. It is observed from our result the abundances of total and bio-available heavy metals in sludge vary as follows: Fe>Mn>Cr>Ni>Cu>Pb>Zn>Cd and Fe>Ni>Mn>Cr>Cu>Zn>Pb>Cd. Sludge applications increased both the total metals, DTPA-extractable metals and total N in the soils. On the other hand lime application has decreased the bioavailability of heavy metals with no change in total N in sludge amended soils. Organic carbon showed positive correlation with all metals except Zn, Cr and Pb. CEC also showed a strong positive correlation (R(2)>0.7) with the low translocation efficiency of pea plants. The value of translocation factor from shoot to seed was found to be smaller than root to shoot of pea plants. Our study thus shows that pea plants were found to be well adapted to the soil amended with 10% sludge with 0.5% lime treatment, minimizing most of the all metal uptake in the shoot of that plant. So, on the basis of the present study, possible treatment may be recommended for the secure disposal of acidic electroplating sludge.