Feed-Forward and Feed-Back Circuits of the NRF2/AP-1 Composite Pathway

Being the central regulator of oxidative status of the cell, NRF2 must be regulated so that its activity can be rapidly and strongly induced when needed and quickly suppressed when not. Moreover, for the cell, NRF2 means much more than just antioxidant defense. Numerous general functions rely on NRF2 and related factors. All this implies that the NRF2 pathway has peculiar and powerful mechanisms of control of its activity. To a great extent, these mechanisms are based on feed-forward and feed-back circuits. These circuits, more than a dozen, are in the focus of this chapter

Oxidative Status Pathways: Systemic Biomarkers

Systemic biomarkers (i.e. biomarkers of functioning of cellular pathways) offer a broad spectrum of diagnostic capabilities. There are several approaches to using systemic biomarkers that derive from exact needs of a researcher or a clinical specialist. First, analyzing a multifunctional and multi-systemic pathway in circulating cells (e.g. leukocytes) allows to gather generalized information on functioning of the organism. Second, there are numerous pathways that, even still in circulating cells, allow to assess risks of developing or stage of development of numerous diseases, including the leaders of non-infection diseases mortality—cardiovascular diseases. Third, biopsy specimens can readily be used to assess the exact signaling type of a disease (especially cancer) thus helping in selecting the best treatment option. Due to unique properties of the human oxidative status pathways that are discussed in the present chapter, diagnostics specialists are now acquiring an all-in-one toolbox for profiling and detecting almost any non-infectious and a broad range of infectious diseases. In addition to properties of the human oxidative status pathways opening these possibilities, this chapter considers exact systemic biomarkers deriving from this approach, reveals some examples of usage of the resulting diagnostic technology and provides instances of successful clinical application of the systemic biomarker approach

Alpha-Synuclein Cell-to-Cell Transfer and Seeding in Grafted Dopaminergic Neurons In Vivo

Several people with Parkinson’s disease have been treated with intrastriatal grafts of fetal dopaminergic neurons. Following autopsy, 10–22 years after surgery, some of the grafted neurons contained Lewy bodies similar to those observed in the host brain. Numerous studies have attempted to explain these findings in cell and animal models. In cell culture, α-synuclein has been found to transfer from one cell to another, via mechanisms that include exosomal transport and endocytosis, and in certain cases seed aggregation in the recipient cell. In animal models, transfer of α-synuclein from host brain cells to grafted neurons has been shown, but the reported frequency of the event has been relatively low and little is known about the underlying mechanisms as well as the fate of the transferred α-synuclein. We now demonstrate frequent transfer of α-synuclein from a rat brain engineered to overexpress human α-synuclein to grafted dopaminergic neurons. Further, we show that this model can be used to explore mechanisms underlying cell-to-cell transfer of α-synuclein. Thus, we present evidence both for the involvement of endocytosis in α-synuclein uptake in vivo, and for seeding of aggregation of endogenous α-synuclein in the recipient neuron by the transferred α-synuclein. Finally, we show that, at least in a subset of the studied cells, the transmitted α-synuclein is sensitive to proteinase K. Our new model system could be used to test compounds that inhibit cell-to-cell transfer of α-synuclein and therefore might retard progression of Parkinson neuropathology

Nanocomposites for X-Ray Photodynamic Therapy

Photodynamic therapy (PDT) has long been known as an effective method for treating surface cancer tissues. Although this technique is widely used in modern medicine, some novel approaches for deep lying tumors have to be developed. Recently, deeper penetration of X-rays into tissues has been implemented, which is now known as X-ray photodynamic therapy (XPDT). The two methods differ in the photon energy used, thus requiring the use of different types of scintillating nanoparticles. These nanoparticles are known to convert the incident energy into the activation energy of a photosensitizer, which leads to the generation of reactive oxygen species. Since not all photosensitizers are found to be suitable for the currently used scintillating nanoparticles, it is necessary to find the most effective biocompatible combination of these two agents. The most successful combinations of nanoparticles for XPDT are presented. Nanomaterials such as metal–organic frameworks having properties of photosensitizers and scintillation nanoparticles are reported to have been used as XPDT agents. The role of metal–organic frameworks for applying XPDT as well as the mechanism underlying the generation of reactive oxygen species are discussed

Approaches to probiotics properties testing using Caco-2 cells

Adequate biological models are a prerequisite to screening and development of probiotic drugs. In the present study, Caco-2 cell line is reviewed as a model for analyzing properties of probiotics. This cell culture possesses all the characteristics necessary for evaluating the effects of probiotic drugs on a wide range of both intracellular processes and intercellular interactions. Informativeness of Caco-2 cells is in the focus of the present review

The Rare-Earth Elements Doping of BaGdF₅ Nanophosphors for X-ray Photodynamic Therapy

It is known that the initiation of photodynamic therapy (PDT) in deep-seated tumors requires the use of X-rays to activate the reactive oxygen species generation in deep tissues. The aim of this paper is to synthesize X-ray nanophosphors and analyze their structural and luminescence characteristics to push the PDT process deep into the body. The article deals with BaGdF5:Eu3+, BaGdF5:Sm3+, and BaGdF5:Tb3+ nanophosphors synthesized using microwave synthesis. It is found that the nanoparticles are biocompatible and have sizes 5–17 nm. However, according to the analysis of X-ray excited optical luminescence, BaGdF5:Sm3+ nanophosphors will not be effective for treating deep-seated tumors. Thus, BaGdF5:Eu3+ and BaGdF5:Tb3+ nanoparticles meet the requirements for the subsequent production of nanocomposites based on them that can be used in X-ray photodynamic therapy

AAV3-Mediated Transfer and Expression of the Pyruvate Dehydrogenase E1 Alpha Subunit Gene Causes Metabolic Remodeling and Apoptosis of Human Liver Cancer Cells

Most cancers rely disproportionately on glycolysis for energy even in the presence of adequate oxygen supply, a condition known as aerobic glycolysis , or the Warburg effect. Pharmacological reversal of the Warburg effect has been shown to cause selective apoptosis of tumor cells, presumably by stimulating mitochondrial respiratory chain activity and production of reactive oxygen species that, in turn, induce a caspase-mediated series of reactions leading to cell death. We reasoned that a similar effect on tumor cells might result from up-regulation of the E1α subunit gene (pda1) of the pyruvate dehydrogenase complex (PDC) that catalyzes the rate-limiting step in aerobic glucose oxidation and thus plays a major role in the control of oxidative phosphorylation. To test this postulate, we employed a self-complementary adeno-associated virus (scAAV)-based delivery and expression system for targeting pda1 to the mitochondria of primary cultures of human hepatoblastoma (HB) and hepatocellular carcinoma (HCC) cells. Serotypes 1-10 scAAV vectors that included enhanced green fluorescent (egfp) reporter gene driven by either cytomegalovirus (CMV) or chicken beta-actin (CBA) promoters were analyzed for transduction ability of HB (Huh-6) and HCC (Huh-7 and HepG2) cell lines and primary cultures of normal human hepatocytes. Serotype 3 scAAV-egfp (scAAV3-egfp) vector was the most efficient and transduced up to 90% of cells. We limited the transgene expression primarily to liver cancer cells by generating scAAV3 vectors that contained the human alpha-fetoprotein promoter (AFP)-driven reporter gene (scAAV3.AFP-egfp) and the potentially therapeutic gene scAAV3.AFP-pda1. Infection of Huh-6 cells by the scAAV3.AFP-pda1 vector increased protein expression of E1α, PDC catalytic activity, and late-stage apoptotic cell death. Apoptosis was also associated with increased protein expression of Bcl-X/S, an early marker of apoptosis, and release of cytochrome c into the cytosol of infected HB cells. These data indicate that molecular targeting of mitochondrial oxidative metabolism in liver cancer cells by AAV3-mediated delivery of pda1 holds promise as a novel and effective therapeutic approach for human hepatic tumors

Individual expression features of GPX2, NQO1 and SQSTM1 transcript variants induced by hydrogen peroxide treatment in HeLa cells

Abstract Pathway activity assessment-based approaches are becoming highly influential in various fields of biology and medicine. However, these approaches mostly rely on analysis of mRNA expression, and total mRNA from a given locus is measured in the majority of cases. Notably, a significant portion of protein-coding genes produces more than one transcript. This biological fact is responsible for significant noise when changes in total mRNA transcription of a single gene are analyzed. The NFE2L2/AP-1 pathway is an attractive target for biomedical applications. To date, there is a lack of data regarding the agreement in expression of even classical target genes of this pathway. In the present paper we analyzed whether transcript variants of GPX2, NQO1 and SQSTM1 were characterized by individual features of expression when HeLa cells were exposed to pro-oxidative stimulation with hydrogen peroxide. We found that all the transcripts (10 in total) appeared to be significantly individually regulated under the conditions tested. We conclude that individual transcripts, rather than total mRNA, are best markers of pathway activation. We also discuss here some biological roles of individual transcript regulation