Examining the Endogenous Antioxidant Response Through Immunofluorescent Analysis of Nrf2 in Tissue

Abstract

As organisms designed to depend upon oxygen to sustain life, humans are necessarily and continually exposed to damaging oxidizing agents. As a vital protective measure, oxygen-dependent organisms have developed a highly evolutionarily conserved mechanism for preventing oxidative stress. NF-E2 (nuclear factor (erythroid-derived 2))-related faetor-2 (Nrf2) is the primary regulator of this endogenous antioxidant response. Many diseases that plague human society, ranging from various cancers to neurodegenerative diseases, have oxidative stress as a component of their etiology, and thus, much disease research has focused on Nrf2, both as a potential point of biological failure and as a promising therapeutic target. As a transcription factor, Nrf2 is active only when it is nuclear, and is regulated largely by its subcellular distribution. Thus, Nrf2 protein levels and subcellular localization are both key pieces of information when studying the endogenous antioxidant response. Immunofluorescent analysis (IFA) of Nrf2 in human tissue is a particularly powerful tool in the study of Nrf2 in disease, because it allows examination of both of these regulatory mechanisms that modulate Nrf2 activity.

The drug monosodium luminol (GVT) preserves crypt-villus epithelial organization and allows survival of intestinal T cells in mice infected with the ts1 retrovirus

“…In mice infected with ts1 and also treated with GVT (ts1-GVT), however, crypt-villus cytoarchitecture is intact at 30 dpi, intestinal epithelial cell gradients are in place, no T cell apoptosis is occurring, and the gPr80 env protein is not accumulated by any cell type in the tissue. In the intact small intestines of GVT-protected mice, most epithelial cells are negative for gp70, and thus do not appear to be infected, but they do contain large amounts of the transcription factor Nrf2, which orchestrates protective antioxidant responses [41,42]. These effects of ts1-GVT treatment are remarkably similar to those occurring in the thymi of ts1-GVT mice, whose epithelial cells also become loaded with Nrf2 [25].…”

“…Like many other cell types, intestinal epithelial cells maintain ambient intracellular levels of NFE-2-related factor 2 (Nrf2), which is a transcriptional factor that coordinately upregulates many genes that participate in antioxidant defenses [41][42][43][44][45]. Under steady-state conditions, Nrf2 is held in an inactive state as part of a complex with another protein called Keap-1.…”

Seabuckthorn Paste Protects Lipopolysaccharide‐Induced Acute Lung Injury in Mice through Attenuation of Oxidative Stress

“…Nuclear factor erythroid 2-related factor 2 (Nrf2) nuclear translocation in lung cells was determined by immunofluorescence technique described by Lindl and Sciutto [ 11 ]. The sections of the lung tissue prepared as mentioned above were deparaffinized and rehydrated through submersion in graded alcohols.…”

Cystamine protects from 3-nitropropionic acid lesioning via induction of nf-e2 related factor 2 mediated transcription

“…In human postmortem brain, Nrf2 activation was detected with immunostaining in Alzheimer’s disease, Lewy body variant of Alzheimer’s disease, and Parkinson’s disease (Ramsey, et al, 2007). Recently, the immunostaining protocol for Nrf2 in human brain has been standardized, however the investigators note the difficulty with which this low abundance protein is detected (Lindl and Jordan-Sciutto, 2008). Furthermore, the same group has been thus far unsuccessful using their protocol to specifically detect Nrf2 in murine tissue (personal communication).…”