Proteomic Changes in Chick Brain Proteome Post Treatment with Lathyrus Sativus Neurotoxin, ß-N-Oxalyl-L-α,ß-Diaminopropionic Acid (L-ODAP): A Better Insight to Transient Neurolathyrism.

Neurolathyrism is a neurodegenerative disorder characterized by spastic paraplegia resulting from the excessive consumption of Lathyrus sativus (Grass pea). ß-N-Oxalyl-L-α,ß-diaminopropionic acid (L-ODAP) is the primary neurotoxic component in this pea. The present study attempted to evaluate the proteome-wide alterations in chick brain 2 hr and 4 hr post L-ODAP treatment. Proteomic analysis of chick brain homogenates revealed several proteins involved in cytoskeletal structure, signaling, cellular metabolism, free radical scavenging, oxidative stress and neurodegenerative disorders were initially up-regulated at 2 hr and later recovered to normal levels by 4 hr. Since L-ODAP mediated neurotoxicity is mainly by excitotoxicity and oxidative stress related dysfunctions, this study further evaluated the role of L-ODAP in apoptosis in vitro using human neuroblastoma cell line, IMR-32. The in vitro studies carried out at 200 µM L-ODAP for 4 hr indicate minimal intracellular ROS generation and alteration of mitochondrial membrane potential though not leading to apoptotic cell death. L-ODAP at low concentrations can be explored as a stimulator of various reactive oxygen species (ROS) mediated cell signaling pathways not detrimental to cells. Insights from our study may provide a platform to explore the beneficial side of L-ODAP at lower concentrations. This study is of significance especially in view of the Government of India lifting the ban on cultivation of low toxin Lathyrus varieties and consumption of this lentil.

Continuous evaluation of changes in the serum proteome from early to late stages of sepsis caused by Klebsiella pneumoniae.

Serum protein profiles of patients with bacterial sepsis from the day of diagnosis until recovery/mortality were compared from early to late stages in response to severe sepsis using two dimensional electrophoresis. The proteins exhibiting changes during the course of sepsis (20­28 day mortality) were selected and identified by matrix­assisted laser desorption ionization­time of flight­tandem mass spectrometry. Among the proteins identified, haptoglobin (Hp), transthyretin (TTR), orosomucoid 1/α1 acid glycoprotein (ORM1), α1 antitrypsin (A1AT), serum amyloid A (SAA) and S100A9 exhibited differential expression patterns between survivors (S; n=6) and non­survivors (NS; n=6), particularly during the early stages of sepsis. Expression factors (EFs), taken as the ratio between the NS and S during early stages, showed ratios of Hp, 0.39 (P≤0.012); TTR, 3.96 (P≤0.03); ORM1, 0.69 (P≤0.79); A1AT, 0.92 (P≤0.87) and SAA, 0.69 (P≤0.01). S100A9, an acute phase protein, exhibited an EF ratio of 1.68 (P≤0.004) during the end stages of sepsis. A delayed rise in levels was observed in Hp, A1AT, ORM1, S100A9 and SAA, whereas TTR levels increased during the early stages of sepsis in NS. Analysis of inflammatory responses in the early stages of sepsis revealed increased mRNA expression in leukocytes of interleukin (IL)­6 (EF, 2.50), IL­10 (EF, 1.70) and prepronociceptin (EF, 1.6), which is a precursor for nociceptin in NS compared with S, and higher Toll­like receptor­4 (EF, 0.30) levels in S compared with NS. Therefore, a weaker acute phase response in the early stages of sepsis in NS, combined with an inefficient inflammatory response, may contribute to sepsis mortality.

Role of cytoplasmic deadenylation and mRNA decay factors in yeast apoptosis.

Strains of Saccharomyces cerevisiae lacking factors involved in 5' to 3' mRNA decay pathway (DCP1, DCP2, DHH1, PAT1, LSM1 and LSM4) exhibit caspase-dependent apoptosis and accelerated chronological aging. In the present study, yeast strains lacking mRNA decapping activation factors (DCP2 and LSM1), cytoplasmic exosome function (SKI2) or cytoplasmic deadenylases (double deletion of CCR4 and PAN2) showed typical markers of eukaryotic apoptosis such as increased cellular reactive oxygen species levels, externalization of phosphatidyl serine, chromatin fragmentation, enhanced caspase gene (YCA1) expression and protein activity in mid-log phase cultures. The transcript levels of negative regulators of mRNA decapping (eIF4E and Pab1) were considerably elevated in strains defective in cytoplasmic deadenylation and reduced in strains lacking cytoplasmic 3' to 5' exosome function or decapping activators. Among the yeast strains studied, lsm1Δ and ccr4Δpan2Δ mutants displayed strongest apoptotic phenotype followed by mutants lacking DCP2 or SKI2. Among yeast strains exhibiting deadenylation defects, slight apoptotic phenotype was observed in ccr4Δ mutants and cell death markers imperceptible in pan2Δ mutants.

Efficient pH dependent drug delivery to target cancer cells by gold nanoparticles capped with carboxymethyl chitosan.

Doxorubicin (DOX) was immobilized on gold nanoparticles (AuNPs) capped with carboxymethyl chitosan (CMC) for effective delivery to cancer cells. The carboxylic group of carboxymethyl chitosan interacts with the amino group of the doxorubicin (DOX) forming stable, non-covalent interactions on the surface of AuNPs. The carboxylic group ionizes at acidic pH, thereby releasing the drug effectively at acidic pH suitable to target cancer cells. The DOX loaded gold nanoparticles were effectively absorbed by cervical cancer cells compared to free DOX and their uptake was further increased at acidic conditions induced by nigericin, an ionophore that causes intracellular acidification. These results suggest that DOX loaded AuNPs with pH-triggered drug releasing properties is a novel nanotheraputic approach to overcome drug resistance in cancer.

Cellular uptake, cytotoxicity, apoptosis and DNA-binding investigations of Ru(II) complexes.

Three new compounds, [Ru(Hdpa)2PyIP](ClO4)2·2H2O (1) [Ru(Hdpa)2FyIP](ClO4)2·2H2O (2) and [Ru(Hdpa)2IIP](ClO4)2·2H2O (3) have been synthesized and characterized by spectroscopic techniques such as elemental analysis, UV/Vis, FT-IR, (1)H NMR, (13)C NMR and mass spectra. The CT-DNA binding properties of 1-3 have been investigated by absorption, emission spectroscopy and viscosity measurements. Experimental results suggested that they can interact with DNA through intercalative mode with different binding strengths. These were found to promote the cleavage of plasmid DNA. Cell viability results indicated that all compounds showed significant dose dependent cytotoxicity in selected cell lines and 1 shown higher cytotoxicity than cisplatin on HeLa cells. Cellular uptake studies were studied by flow cytometry and confocal microscopy.