Large-scale identification of proteins in human salivary proteome by liquid chromatography/mass spectrometry and two-dimensional gel electrophoresis-mass spectrometry

Human saliva contains a large number of proteins and peptides (salivary proteome) that help maintain homeostasis in the oral cavity. Global analysis of human salivary proteome is important for understanding oral health and disease pathogenesis. In this study, large-scale identification of salivary proteins was demonstrated by using shotgun proteomics and two-dimensinal gel electrophoresis-mass spectrometry (2-DE-MS). For the shotgun approach, whole saliva proteins were prefractionated according to molecular weight. The smallest fraction, presumably containing salivary peptides, was directly separated by capillary liquid chromatography (LC). However, the large protein fractions were digested into peptides for subsequent LC separation. Separated peptides were analyzed by on-line electrospray tandem mass spectrometry (MS/MS) using a quadrupole-time of flight mass spectrometer, and the obtained spectra were automatically processed to search human protein sequence database for protein identification. Additionally, 2-DE was used to map out the proteins in whole saliva. Protein spots 105 in number were excised and in-gel digested; and the resulting peptide fragments were measured by matrix-assisted laser desorption/ionization-mass spectrometry and sequenced by LC-MS/MS for protein identification. In total, we cataloged 309 proteins from human whole saliva by using these two proteomic approaches.     

Genomic evidence of contemporary hybridization between Schistosoma species

Hybridization between different species of parasites is increasingly being recognised as a major public and veterinary health concern at the interface of infectious diseases biology, evolution, epidemiology and ultimately control. Recent research has revealed that viable hybrids and introgressed lineages between Schistosoma spp. are prevalent across Africa and beyond, including those with zoonotic potential. However, it remains unclear whether these hybrid lineages represent recent hybridization events, suggesting hybridization is ongoing, and/or whether they represent introgressed lineages derived from ancient hybridization events. In human schistosomiasis, investigation is hampered by the inaccessibility of adult-stage worms due to their intravascular location, an issue which can be circumvented by post-mortem of livestock at abattoirs for Schistosoma spp. of known zoonotic potential. To characterise the composition of naturally-occurring schistosome hybrids, we performed whole-genome sequencing of 21 natural livestock infective schistosome isolates. To facilitate this, we also assembled a de novo chromosomal-scale draft assembly of Schistosoma curassoni. Genomic analyses identified isolates of S. bovis, S. curassoni and hybrids between the two species, all of which were early generation hybrids with multiple generations found within the same host. These results show that hybridization is an ongoing process within natural populations with the potential to further challenge elimination efforts against schistosomiasis.     

Effect of doxorubicin on heart mitochondrial enzymes in rats: a protective role for alpha-tocopherol.

Effect of doxorubicin on heart mitochondrial enzymes was studied in rats with or without the administration of alpha-tocopherol. Rats were treated with doxorubicin 2.5 mg/kg, ip body wt once a week for 8 weeks. Alpha-tocopherol was co-administered orally for 2 months (400 mg/kg body wt daily). TCA cycle enzyme, NADH-dehydrogenase, cytochrome-C-oxidase and Na+,K(+)-ATPase activities were found to be decreased in doxorubicin treatment. A significant decrease in protease activity was observed with a concomitant increase in mitochondrial protein level. Mitochondrial lipid peroxide level was found to be increased with a decrease in thiol content. Alpha-tocopherol co-administration was found to maintain the mitochondrial enzyme activities as well as the thiol content. The results are discussed with reference to the antioxidant nature of alpha-tocopherol.     

Anti-biofilm activity of LC-MS based Solanum nigrum essential oils against multi drug resistant biofilm forming P. mirabilis

Urinary tract infections are second most important diseases worldwide due to the increased amount of antibiotic resistant microbes. Among the Gram negative bacteria, P. mirabilis is the dominant biofilm producer in urinary tract infections next to E. coli. Biofilm is a process that produced self-matrix of more virulence pathogens on colloidal surfaces. Based on the above fact, this study was concentrated to inhibit the P. mirabilis biofilm formation by various in-vitro experiments. In the current study, the anti-biofilm effect of essential oils was recovered from the medicinal plant of Solanum nigrum, and confirmed the available essential oils by liquid chromatography-mass spectroscopy analysis. The excellent anti-microbial activity and minimum biofilm inhibition concentration of the essential oils against P. mirabilis was indicated at 200 µg/mL. The absence of viability and altered exopolysaccharide structure of treated cells were showed by biofilm metabolic assay and phenol–sulphuric acid method. The fluorescence differentiation of P. mirabilis treated cells was showed with more damages by confocal laser scanning electron microscope. Further, more morphological changes of essential oils treated cells were differentiated from normal cells by scanning electron microscope. Altogether, the results were reported that the S. nigrum essential oils have anti-biofilm ability.     

Phytochemical screening and anti-oxidant activity of Sargassum wightii enhances the anti-bacterial activity against Pseudomonas aeruginosa

In this study, the phytochemical, phenolic, flavonoid and bioactive compounds were successfully screened from crude extract of Sargassum wightii by LC-MS analysis after NIST interpretation. Bacterial growth inhibition study result was shown with 24 mm zone inhibition at 200 µg/mL concentration against P. aeruginosa. The increased phenolic content was much closed to gallic acid and the range was observed at 250 μg/mL concentration. In addition, flavonoid contents of the algae extract was indicated more significant with rutin at 200 μg/mL. In result, both the phenolic and flavonoid contents of the extract were more correlated with gallic acid and rutin. Further, the total anti-oxidant and DPPH radical scavenging activities were shown increased activity at 200 μg/mL concentrations. Furthermore, the excellent anti-bacterial alteration result was observed at 200 μg/mL concentration by minimum inhibition concentration. Therefore, the result was revealed that the marine algae Sargassum wightii has excellent phytochemical and anti-oxidant activities, and it has improved anti-bacterial activity against P. aeruginosa.     

Performance of established native seedlings in relation to invasive Lantana camara, rainfall and species’ habitat preferences in a seasonally dry tropical forest

Native species’ response to the presence of invasive species is context specific. This response cannot be studied in isolation from the prevailing environmental stresses in invaded habitats such as seasonal drought. We investigated the combined effects of an invasive shrub Lantana camara L. (lantana), seasonal rainfall and species’ microsite preferences on the growth and survival of 1,105 naturally established seedlings of native trees and shrubs in a seasonally dry tropical forest. Individuals were followed from April 2008 to February 2010, and growth and survival measured in relation to lantana density, seasonality of rainfall and species characteristics in a 50-ha permanent forest plot located in Mudumalai, southern India. We used a mixed effects modelling approach to examine seedling growth and generalized linear models to examine seedling survival. The overall relative height growth rate of established seedlings was found to be very low irrespective of the presence or absence of dense lantana. 22-month growth rate of dry forest species was lower under dense lantana while moist forest species were not affected by the presence of lantana thickets. 4-month growth rates of all species increased with increasing inter-census rainfall. Community results may be influenced by responses of the most abundant species, Catunaregam spinosa, whose growth rates were always lower under dense lantana. Overall seedling survival was high, increased with increasing rainfall and was higher for species with dry forest preference than for species with moist forest preference. The high survival rates of naturally established seedlings combined with their basal sprouting ability in this forest could enable the persistence of woody species in the face of invasive species.     

A2B adenosine receptor blockade inhibits growth of prostate cancer cells

The role of the A_2B adenosine receptor (AR) in prostate cell death and growth was studied. The A_2B AR gene expression quantified by real-time quantitative RT-PCR and Western blot analysis was the highest among four AR subtypes (A₁, A_2A, A_2B, and A₃) in all three commonly used prostate cancer cell lines, PC-3, DU145, and LNCaP. We explored the function of the A_2B AR using PC-3 cells as a model. The A_2B AR was visualized in PC-3 cells by laser confocal microscopy. The nonselective A_2B AR agonist NECA and the selective A_2B AR agonist BAY60-6583, but not the A_2A AR agonist CGS21680, concentration-dependently induced adenosine 3′,5′-cyclic monophosphate (cyclic AMP) accumulation. NECA diminished lactate dehydrogenase (LDH) release, TNF-α-induced increase of caspase-3 activity, and cycloheximide (CHX)-induced morphological changes typical of apoptosis in PC-3 cells, which were blocked by a selective A_2B AR antagonist PSB603. NECA-induced proliferation of PC-3 cells was diminished by siRNA specific for the A_2B AR. The selective A_2B AR antagonist PSB603 was shown to inhibit cell growth in all three cell lines. Thus, A_2B AR blockade inhibits growth of prostate cancer cells, suggesting selective A_2B AR antagonists as potential novel therapeutics.     

NMR assignment of a PDZ domain in complex with a HPV51 E6 derived N-terminally pyroglutamic acid modified peptide

The resonance assignment of an amino-terminal pyroglutamic acid containing peptide derived from the E6 protein of human papillomavirus (HPV) type 51 in complex with PDZ domain 2 of hDlg/SAP-97 is reported. The assignments include ¹H, ¹³C and ¹⁵N resonances for the protein and peptide in the complex and all of the peptide’s pyroglutamic acid nuclei.     

Kidney Transplantation Is Associated with Catastrophic Out of Pocket Expenditure in India

Kidney transplantation (KT) is only viable renal replacement option for most patients in India. Most patients do not have health insurance and meet treatment expenditure from their own resources. We prospectively evaluated the expenses associated with KT and its impact on the socioeconomic status of families in a public hospital. All direct and indirect expenses incurred by the patients from the time of diagnosis of chronic kidney disease to KT were recorded. Direct expenses included physician fees, cost of drugs and disposables, dialysis, and expenses on investigations and hospitalization. Indirect expenses included travel, food, stay, and loss of income suffered by the family. Educational dropout and financial loss were also recorded. There were 43 males and 7 females between the ages of 12 and 57 years. Direct expenses ranged from US$ 2,151–23,792 and accounted for two-thirds of the total expenses. Pre-referral hospitalization, dialysis and medication accounted for majority of direct expense. Indirect expenses ranged from US$ 226–15,283. Travel expenses and loss of income accounted for most of indirect expense. About 54%, 8%, and 10% of families suffered from severe, moderate, and some financial crisis respectively. A total of 38 families had job losses, and 1 patient and 12 caregivers dropped out of studies. To conclude, KT is associated with catastrophic out-of-pocket expenditure and pushes a majority of the patients who come for treatment to public hospitals into severe financial crisis. Educational dropout and loss of jobs are other major concerns. Systematic efforts are required to address these issues.