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Several converging lines of evidence from our group support a potential role of RLIP76 (AKA Rlip) in neurodegenerative disorders, including Alzheimer's Disease (AD). However, the role of Rlip in Alzheimer's and other neurodegenerative diseases is not well understood. The purpose of the present study is to determine the role of Rlip in the brains of AD patients and control subjects. To achieve our goals, we used frozen tissues and formalin-fixed paraffin-embedded postmortem brains from AD patients of different Braak stages and age-matched control subjects. Our immunohistology and immunoblotting blotting analysis revealed that expression of Rlip protein gradually and significantly decreased (p = 0.0001) with AD progression, being lowest in Braak stage IV-V. Rlip was colocalized with Amyloid beta (Aβ) and phosphorylated tau (p-Tau) as observed by IHC staining and co-immunoprecipitation studies. Lipid peroxidation (4-HNE generation) and H 2 O 2 production were significantly higher (p = 0.004 and 0.0001 respectively) in AD patients compared to controls, and this was accompanied by lower ATP production in AD (p = 0.0009). Oxidative DNA damage was measured by 8-Hydroxyguanosine (8-OHdG) in tissue lysates by ELISA and COMET assay. AD 8-OHdG levels were significantly higher (p = 0.0001) compared to controls. COMET assay was performed in brain cells, isolated from frozen postmortem samples. The control samples showed minimal DNA in comets representing few DNA strand breaks (<20 %), (score-0-1). However, the AD group showed an average of 50 % to 65 % of DNA in comet tails (score-4-5) indicating numerous DNA strand breaks. The difference between the two groups was significant (p = 0.001), as analyzed by Open Comet by ImageJ. Elevated DNA damage was further examined by western blot analysis for phosphorylated histone variant H2AX (γH2AX). Induction of γH2AX was very significant (p < 0.0001) and confirmed the presence of double-strand breaks in DNA. Overall, our results indicate an important role for Rlip in maintaining neuronal health and homeostasis by suppressing cellular oxidative stress and DNA damage. Based on our findings, we cautiously conclude that Rlip is a promising therapeutic target for Alzheimer's disease., Competing Interests: Declaration of competing interest Authors declare that a pending disclosure is in progress with contents of this study., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Alzheimer's disease (AD) is a neurodegenerative disease that affects a large proportion of the aging population. RalBP1 (Rlip) is a stress-activated protein that plays a crucial role in oxidative stress and mitochondrial dysfunction in aging and neurodegenerative diseases but its precise role in the progression of AD is unclear. The purpose of our study is to understand the role of Rlip in the progression and pathogenesis of AD in mutant APP/amyloid beta (Aβ)-expressed mouse primary hippocampal (HT22) hippocampal neurons. In the current study, we used HT22 neurons that express mAPP, transfected with Rlip-cDNA and/or RNA silenced, and studied cell survival, mitochondrial respiration, mitochondrial function, immunoblotting & immunofluorescence analysis of synaptic and mitophagy protein's and colocalization of Rlip and mutant APP/Aβ proteins and mitochondrial length and number. We also assessed Rlip levels in autopsy brains from AD patients and control subjects. We found cell survival was decreased in mAPP-HT22 cells and RNA-silenced HT22 cells. However, cell survival was increased in Rlip-overexpressed mAPP-HT22 cells. Oxygen consumption rate (OCR) was decreased in mAPP-HT22 cells and RNA-silenced Rlip-HT22 cells. OCR was increased in Rlip-overexpressed in mAPP-HT22 cells. Mitochondrial function was defective in mAPP-HT22 cells and RNA silenced Rlip in HT22 cells, however, it was rescued in Rlip overexpressed mAPP-HT22 cells. Synaptic and mitophagy proteins were decreased in mAPP-HT22 cells, further reducing RNA-silenced Rlip-HT22 cells. However, these were increased in mAPP+Rlip-HT22 cells. Colocalization analysis revealed Rlip is colocalized with mAPP/Aβ. An increased number of mitochondria and decreased mitochondrial length were found in mAPP-HT22 cells. These were rescued in Rlip overexpressed mAPP-HT22 cells. Reduced Rlip levels were found in autopsy brains from AD patients. These observations strongly suggest that Rlip deficiency causes oxidative stress/mitochondrial dysfunction and Rlip overexpression reduced these defects., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing interests., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest with the contents of this article.

Gadolinium-based contrast agents are linked to nephrogenic systemic fibrosis in patients with renal insufficiency. The pathology of nephrogenic systemic fibrosis is characterized by abnormal tissue repair: fibrosis and ectopic ossification. The mechanisms by which gadolinium could induce fibrosis and ossification are not known. We examined in vitro the effect of a gadolinium-based contrast agent on human peripheral blood mononuclear cells for phenotype and function relevant to the pathology of nephrogenic systemic fibrosis using immunofluorescence, flow cytometry, real-time PCR, and osteogenic assays. We also examined tissues from patients with nephrogenic systemic fibrosis, using IHC to identify the presence of cells with phenotype induced by gadolinium. Gadolinium contrast induced differentiation of human peripheral blood mononuclear cells into a unique cellular phenotype--CD163(+) cells expressing proteins involved in fibrosis and bone formation. These cells express fibroblast growth factor (FGF)23, osteoblast transcription factors Runt-related transcription factor 2, and osterix, and show an osteogenic phenotype in in vitro assays. We show in vivo the presence of CD163(+)/procollagen-1(+)/osteocalcin(+) cells in the fibrotic and calcified tissues of nephrogenic systemic fibrosis patients. Gadolinium contrast-induced CD163(+)/ferroportin(+)/FGF23(+) cells with osteogenic potential may play a role in systemic fibrosis and ectopic ossification in nephrogenic systemic fibrosis., (Copyright © 2013 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)