Accumulating evidence shows that oxidative stress is involved in a wide variety of human diseases: rheumatoid arthritis, Alzheimer’s disease, Parkinson’s disease, cancers, etc. Here, we discuss the significance of oxidative conditions in different disease, with the focus on neurodegenerative disease including Parkinson’s disease, which is mainly caused by oxidative stress. Reactive oxygen and nitrogen species (ROS and RNS, respectively), collectively known as RONS, are produced by cellular enzymes such as myeloperoxidase, NADPH-oxidase (nicotinamide adenine dinucleotide phosphate-oxidase) and nitric oxide synthase (NOS). Natural antioxidant systems are categorized into enzymatic and non-enzymatic antioxidant groups. The former includes a number of enzymes such as catalase and glutathione peroxidase, while the latter contains a number of antioxidants acquired from dietary sources including vitamin C, carotenoids, flavonoids and polyphenols. There are also scavengers used for therapeutic purposes, such as 3,4-dihydroxyphenylalanine (L-DOPA) used routinely in the treatment of Parkinson’s disease (not as a free radical scavenger), and 3-methyl-1-phenyl-2-pyrazolin-5-one (Edaravone) that acts as a free radical detoxifier frequently used in acute ischemic stroke. The cell surviving properties of L-DOPA and Edaravone against oxidative stress conditions rely on the alteration of a number of stress proteins such as Annexin A1, Peroxiredoxin-6 and PARK7/DJ-1 (Parkinson disease protein 7, also known as Protein deglycase DJ-1). Although they share the targets in reversing the cytotoxic effects of H₂O₂, they seem to have distinct mechanism of function. Exposure to L-DOPA may result in hypoxia condition and further induction of ORP150 (150-kDa oxygen-regulated protein) with its concomitant cytoprotective effects but Edaravone seems to protect cells via direct induction of Peroxiredoxin-2 and inhibition of apoptosis.

Proteomics is a powerful tool to understand the molecular mechanisms causing the production of high penicillin titers by industrial strains of the filamentous fungus Penicillium chrysogenum as the result of strain improvement programs. Penicillin biosynthesis is an excellent model system for many other bioactive microbial metabolites. The recent publication of the P. chrysogenum genome has established the basis to understand the molecular processes underlying penicillin overproduction. We report here the proteome reference map of P. chrysogenum Wisconsin 54-1255 (the genome project reference strain) together with an in-depth study of the changes produced in three different strains of this filamentous fungus during industrial strain improvement. Two-dimensional gel electrophoresis, peptide mass fingerprinting, and tandem mass spectrometry were used for protein identification. Around 1000 spots were visualized by “blue silver” colloidal Coomassie staining in a non-linear pI range from 3 to 10 with high resolution, which allowed the identification of 950 proteins (549 different proteins and isoforms). Comparison among the cytosolic proteomes of the wild-type NRRL 1951, Wisconsin 54-1255 (an improved, moderate penicillin producer), and AS-P-78 (a penicillin high producer) strains indicated that global metabolic reorganizations occurred during the strain improvement program. The main changes observed in the high producer strains were increases of cysteine biosynthesis (a penicillin precursor), enzymes of the pentose phosphate pathway, and stress response proteins together with a reduction in virulence and in the biosynthesis of other secondary metabolites different from penicillin (pigments and isoflavonoids). In the wild-type strain, we identified enzymes to utilize cellulose, sorbitol, and other carbon sources that have been lost in the high penicillin producer strains. Changes in the levels of a few specific proteins correlated well with the improved penicillin biosynthesis in the high producer strains. These results provide useful information to improve the production of many other bioactive secondary metabolites.

Background: KIAA1199is a recently identified novel gene that is up-regulated in human cancer with poor survival. Our proteomic study on signaling polarity in chemotactic cells revealed KIAA1199 as a novel protein target that may be involved in cellular chemotaxis and motility. In the present study, we examined the functional significance of KIAA1199 expression in breast cancer growth, motility and invasiveness.Methods: We validated the previous microarray observation by tissue microarray immunohistochemistry using a TMA slide containing 12 breast tumor tissue cores and 12 corresponding normal tissues. We performed the shRNA-mediated knockdown of KIAA1199 in MDA-MB-231 and HS578T cells to study the role of this protein in cell proliferation, migration and apoptosisin vitro. We studied the effects of KIAA1199 knockdownin vivoin two groups of mice (n = 5). We carried out the SILAC LC-MS/MS based proteomic studies on the involvement of KIAA1199 in breast cancer.Results: KIAA1199 mRNA and protein was significantly overexpressed in breast tumor specimens and cell lines as compared with non-neoplastic breast tissues from large-scale microarray and studies of breast cancer cell lines and tumors. To gain deeper insights into the novel role of KIAA1199 in breast cancer, we modulated KIAA1199 expression using shRNA-mediated knockdown in two breast cancer cell lines (MDA-MB-231 and HS578T), expressing higher levels of KIAA1199. The KIAA1199 knockdown cells showed reduced motility and cell proliferationin vitro. Moreover, when the knockdown cells were injected into the mammary fat pads of female athymic nude mice, there was a significant decrease in tumor incidence and growth. In addition, quantitative proteomic analysis revealed that knockdown of KIAA1199 in breast cancer (MDA-MB-231) cells affected a broad range of cellular functions including apoptosis, metabolism and cell motility.Conclusions: Our findings indicate that KIAA1199 may play an important role in breast tumor growth and invasiveness, and that it may represent a novel target for biomarker development and a novel therapeutic target for breast cancer.

The filamentous fungus Penicillium chrysogenum is well-known by its ability to synthesize β-lactam antibiotics as well as other secondary metabolites. Like other filamentous fungi, this microorganism is an excellent host for secretion of extracellular proteins because of the high capacity of its protein secretion machinery. In this work, we have characterized the extracellular proteome reference map of P. chrysogenum Wisconsin 54–1255 by two-dimensional gel electrophoresis. This method allowed the correct identification of 279 spots by peptide mass fingerprinting and tandem MS. These 279 spots included 328 correctly identified proteins, which corresponded to 131 different proteins and their isoforms. One hundred and two proteins out of 131 were predicted to contain either classical or nonclassical secretion signal peptide sequences, providing evidence of the authentic extracellular location of these proteins. Proteins with higher representation in the extracellular proteome were those involved in plant cell wall degradation (polygalacturonase, pectate lyase, and glucan 1,3-β-glucosidase), utilization of nutrients (extracellular acid phosphatases and 6-hydroxy-d-nicotine oxidase), and stress response (catalase R). This filamentous fungus also secretes enzymes specially relevant for food industry, such as sulfydryl oxidase, dihydroxy-acid dehydratase, or glucoamylase. The identification of several antigens in the extracellular proteome also highlights the importance of this microorganism as one of the main indoor allergens. Comparison of the extracellular proteome among three strains of P. chrysogenum, the wild-type NRRL 1951, the Wis 54–1255 (an improved, moderate penicillin producer), and the AS-P-78 (a penicillin high-producer), provided important insights to consider improved strains of this filamentous fungus as versatile cell-factories of interest, beyond antibiotic production, for other aspects of white biotechnology.

Background and objectives: With regard to their ease of harvest and common developmental origin, dental pulp stem cells (DPSCs) may act as a favorable source of stem cells in generation of nerves. Moreover; cellular migration and differentiation as well as survival, self-renewal, and proliferation of neuroprogenitor species require the presence of the central nervous system (CNS) mitogens including EGF and bFGF. Accordingly, the possibility of the induction of neuronal differentiation of DPSCs by EGF and bFGF was evaluated in the present study. Materials and methods: DPSCs were treated with 20 ng/ml EGF, 20 ng/ml bFGF, and 10 µg/ml heparin. In order to further induce the neuroprogenitor differentiation, DPSC-derived spheres were also incubated in serum-free media for three days. The resulting spheres were then cultured in high-glucose Dulbecco’s Modified Eagle Medium (DMEM) with 10% FBS. The morphology of the cells and the expression of the differentiation markers were correspondingly analyzed by quantitative polymerase chain reaction (qPCR), western blotting, and immunofluorescence (IF). Results: The EGF/bFGF-treated DPSCs showed significant increase in the expression of the neuroprogenitor markers of Nestin and SRY (sex determining region Y)-box 2 (SOX2), 72 h after treatment. The up-regulation of Nestin and SOX2 induced by growth factors was confirmed using western blotting and IF. The cultures also yielded some neuron-like cells with a significant rise in Nestin, microtubule-associated protein 2 (MAP2), and Neurogenin 1 (Ngn1) transcript levels; compared with cells maintained in the control media (p < 0.05). Conclusion: DPSCs seemed to potentially differentiate into neuron-like cells under the herein-mentioned treatment conditions.