Peroxiredoxin Research Articles

Telomere shortening in hypertensive heart disease depends on NOX2-mediated loss of PRDX1 and oxidative DNA damage

Abstract Background Heart failure (HF) coincides with cardiomyocyte telomere shortening. Arterial hypertension is the most prominent risk factor for HF. Both HF and arterial hypertension are associated with dysregulation of the neurohormonal axis and increased ROS. However, how neurohormonal activation is linked to telomere shortening in the pathogenesis of HF is incompletely understood. Further, recent findings suggest that peroxiredoxin 1 (PRDX1) can act as a telomere specific antioxidant. Methods To induce hypertensive HF, male C57BL/6J mice were subjected to AngII-infusion, uninephrectomy and high-salt (AngII++) for 5 weeks. Cardiac function was assessed by ultrasound. Mouse adult cardiomocytes (CMs) isolated from C57BL/6J mice and mice lacking the NADPH-oxidase (NOX) 2/gp91phox as well as rat ventricular CM-derived cells (H9C2) were stimulated with AngII. Telomere length was quantified by Q-FISH after staining with a C-rich telomere probe (TelC). DNA/RNA-damage was evaluated after staining for Oxo-8-Gua (8-oxo-7,8-dihydroguanine) and Oxo-8-G (8-oxo-7,8-dihydroguanosine). Superoxide (O2-) was quantified by 2-hydroxyethidium (2-HE) using HPLC analysis. Colocalization of Oxo-8-Gua and TelC was quantified by automated image analysis. (Sub)cellular and tissue expression of gp91phox/Nox2 and PRDX1 was evaluated by ICC/IHC. All image quantification was conducted semiautomatically. Results In mice subjected to hypertensive HF, CM telomere shortening correlated significantly with both left ventricular (LV) dilatation and impairment of LV systolic function, paralleled by a significant loss of myocardial PRDX1 and significantly increased myocardial DNA/RNA-damage. Similarly, CMs stimulated with AngII exhibited significant telomere shortening, significant loss of PRDX1 and significantly increased DNA/RNA-damage, together with significantly increased CM superoxide production and significantly increased expression of the superoxide generating enzyme gp91phox/NOX2. Correspondingly, deficiency of gp91phox/NOX2 significantly prevented AngII-induced CM telomere shortening, DNA/RNA-damage and PRDX1-depletion. A similar effect could be observed upon stimulation of CMs with the specific histone deacetylase (HDAC) 6-inhibitor tubastatin, which prevents deacetylation of PRDX1. Conclusion We could provide first evidence that in heart failure, ROS originating from increased NOX2-activity leads to depletion of the telomere-targeted antioxidant and repair-protein PRDX1, which results in damage to the telomeric DNA. Both depletion of NOX2 as well as harnessing the intrinsic antioxidant defense by stabilizing PRDX1 via HDAC6-inhibition prevented CM telomere shortening and could be further evaluated as potential therapeutic targets to address heart failure. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): German Federal Ministry for Education and Research

MiR-375 suppresses the growth and metastasis of esophageal squamous cell carcinoma by targeting PRDX1.

Esophageal cancer (EC) is one of the most lethal cancers. Esophageal squamous cell carcinoma (ESCC) is the most common histological subtype in Asian people. Diverse microRNAs, such as miR-375, have been confirmed to be involved in the process of tumorigenesis and metastasis. However, the underlying mechanism through which miR-375 acts in ESCC patients remains unknown. We used The Cancer Genome Atlas (TCGA) database to analyze the association between miR-375 and the survival rate in patients with esophageal squamous cell carcinoma. Real Time quantitative PCR (RT-qPCR) analysis was performed to evaluate the level of miR-375 in EC tissues and cells. A luciferase reporter assay was used to confirm the target gene of miR-375. A colony formation assay as well as flow cytometric and transwell invasion experiments were employed to examine the effects of miR-375 and peroxiredoxin 1 (PRDX1) on ESCC cells. A tumor xenograft mouse model was then used to investigate the role of miR-375 on tumor growth in vivo. Moreover, we performed rescue experiments to evaluate the effect of PRDX1 on ESCC progression. miR-375 expression was significantly downregulated in both ESCC clinical tissues and serum, and the reduction of miR-375 was remarkably linked to a poor prognosis in ESCC. Further investigation illustrated that aberrant expression of miR-375 dampened the growth and infiltration of ESCC cells both in vitro and in vivo. Bioinformatics and luciferase reporter analysis verified that the transcript of PRDX1 is a direct target of miR-375 and its expression in ESCC cells was found to be inversely modulated by miR-375. Moreover, the tumor formation experiment in nude mice confirmed that miR-375 can effectively dampen tumor growth in xenograft tumor mice models. Notably, over-expression of PRDX1 effectively counteracted the tumor-suppressing capabilities of miR-375. We demonstrated the antitumor effect of miR-375 on ESCC by targeting PRDX1 both in vitro and in vivo.

Telomere Shortening in Hypertensive Heart Disease Depends on Oxidative DNA Damage and Predicts Impaired Recovery of Cardiac Function in Heart Failure.

Heart failure (HF) coincides with cardiomyocyte telomere shortening. Arterial hypertension is the most prominent risk factor for HF. Both HF and arterial hypertension are associated with dysregulation of the neurohormonal axis. How neurohormonal activation is linked to telomere shortening in the pathogenesis of HF is incompletely understood. Cardiomyocyte telomere length was assessed in a mouse model of hypertensive HF induced by excess neurohormonal activation (AngII [angiotensin II] infusion, high salt diet, and uninephrectomy), in AngII-stimulated cardiomyocytes and in endomyocardial biopsies from patients with HF. Superoxide production, expression of NOX2 (NADPH oxidase 2) and PRDX1 (peroxiredoxin 1) and HDAC6 (histone deacetylase 6) activity were assessed. Telomere shortening occurred in vitro and in vivo, correlating with both left ventricular (LV) dilatation and LV systolic function impairment. Telomere shortening coincided with increased superoxide production, increased NOX2 expression, increased HDAC6 activity, loss of the telomere-specific antioxidant PRDX1, and increased oxidative DNA-damage. NOX2 knockout prevented PRDX1 depletion, DNA-damage and telomere shortening confirming this enzyme as a critical source of reactive oxygen species. Cotreatment with the NOX inhibitor apocynin ameliorated hypertensive HF and telomere shortening. Similarly, treatment with the HDAC6 inhibitor tubastatin A, which increases PRDX1 bioavailability, prevented telomere shortening in adult cardiomyocytes. To explore the clinical relevance of our findings, we examined endomyocardial biopsies from an all-comer population of patients with HF with reduced ejection fraction. Here, cardiomyocyte telomere length predicted the recovery of cardiac function. Cardiomyocyte telomere shortening and oxidative damage in heart failure with reduced ejection fraction induced by excess neurohormonal activation depends on NOX2-derived superoxide and may help to stratify HF therapy.

Identification of differentially expressed ferroptosis-related genes in abdominal aortic aneurysm: Bioinformatics analysis.

PurposeFerroptosis plays a crucial role in the development and progression of abdominal aortic aneurysm (AAA). The aim of this study was to identify differentially expressed genes associated with ferroptosis in AAA through bioinformatics analysis combined with experimental validation.Materials and methodsFirstly, the mRNA expression profile datasets GSE57691 and GSE47472 from Gene Expression Omnibus database were screened, and principal component analysis was carried out. Next, the R software (version 4.0.0) was used to analyze potentially differentially expressed genes associated with AAA and ferroptosis. Subsequently, protein–protein interaction analysis, gene ontology enrichment analysis, and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis were performed on the selected candidate genes. Finally, quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression levels of the first five selected abnormal ferroptosis-related genes in clinical samples obtained from patients with AAA and healthy controls.ResultsBased on the information contained in the two datasets, a total of 20 differentially expressed ferroptosis-related genes (three upregulated genes and 17 downregulated genes) were selected. Protein–protein interaction analysis demonstrated interaction between these genes, while gene ontology enrichment analysis of ferroptosis genes with differential expression indicated that some enrichment items were associated with oxidative stress. The qRT-PCR results showed that the expression levels of interleukin-6 (IL-6), peroxiredoxin 1 (PRDX1), and stearoyl-CoA desaturase (SCD) were consistent with the bioinformatics prediction results obtained from the mRNA chip.ConclusionBioinformatics analysis identified 20 potential ferroptosis-related differentially expressed genes in AAA. Further verification by qRT-PCR showed that IL-6, PRXD1, and SCD might affect the process of AAA by regulating ferroptosis. Our results might assist in further understanding the pathogenesis of AAA and guiding treatment.

Regulatory effect of peroxiredoxin 1 (PRDX1) on doxorubicin-induced apoptosis in triple negative breast cancer cells

Patients with triple negative breast cancer (TNBC) lack the estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2; thus, conventional hormone and targeted therapies have minimal effect on them. Therefore, clinical treatment of TNBC is still based on chemotherapy and supplemented by other methods. Doxorubicin (DOX), a common drug used in TNBC chemotherapy, has high affinity for cardiolipin, and the nematosomes are rich in cardiolipin; therefore, DOX has high mitochondria-targeting ability. DOX accumulates and plunders the electrons of nicotinamide adenine dinucleotide phosphate (NADPH) and cytochrome C in mitochondria to produce semiquinone DOX. Under the action of oxygen molecules, semiquinone DOX is reduced to DOX and reactive oxygen species (ROS) are generated. The accumulation of ROS can cause mitochondrial dysfunction and lead to mitochondrial dependent apoptosis. Bioinformatic analysis of samples from TNBC patients revealed that peroxiredoxin 1 (PRDX1) was highly expressed in TNBC tissues, and the poor prognosis of patients with high PRDX1 expression was considerably increased. Previous studies determined that DOX can upregulate the expression of the PRDX1 protein in the human TNBC cell line (MDA-MB-231). Thus, we speculate that PRDX1 plays an important role in the process of DOX-induced TNBC cell apoptosis. In this study, we aimed to explore the role of PRDX1 in the process of DOX-induced TNBC cell apoptosis. We found that PRDX1 deletion increased the sensitivity of MDA-MB-231 cells to DOX, which was mainly due to mitochondrial oxidative stress caused by intracellular ROS accumulation, leading to mitochondria-dependent apoptosis. Deletion of PRDX1 promotes the PI3K/Akt signaling pathway to mediate the expression of GSK3β. Gsk3β is an upstream signal of mitochondria-dependent apoptosis, and is also an important target of ROS. PRDX1 participates in adriamycin-induced apoptosis of TNBC cells by regulating the expression level of GSK3β. Our findings present new insights to treat breast cancer and TNBC, outlines the clinical use of DOX, and provides a basic theory to develop PRDX1 gene function.

Effects of Aloe vera extract on growth, viability, ultrastructure and expression of mRNA for antioxidant enzymes in bovine secondary follicles cultured in vitro

This study aimed to investigate the effects of Aloe vera extract on follicular growth, viability, ultrastructure, and mRNA levels for superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase 1 (GPX1) and peroxiredoxin 6 (PRDX6) in bovine secondary follicles cultured in vitro. To this end, secondary follicles were mechanically isolated from the ovarian cortex and cultured at 38.5 °C, with 5% CO2 in air, for 18 days in TCM-199+ alone or supplemented with 2.5%, 5.0%, 10.0% and 20.0% Aloe vera extract. Follicular growth, morphology and antrum formation were evaluated every 6 days, while ultrastructure was evaluated at the end of culture. Analysis of viability was performed by calcein-AM and ethidium homodimer-1, while mRNA levels for SOD, CAT, GPX1 and PRDX6 were evaluated by real-time PCR at the end of culture. The results show that follicles cultured with 2.5% Aloe vera had increased the rate of antrum formation, while 2.5% and 5.0% Aloe vera improved follicular viability rate. Follicles cultured with 2.5% and 10.0% Aloe vera increased the levels of mRNA for SOD and GPX1 respectively, but the levels of CAT were reduced in follicles cultured with 2.5%, 5.0%, 10.0% and 20.0%. Additionally, follicles cultured with 2.5% of Aloe vera had their ultrastructure well preserved, while those cultured with 5.0%, 10.0% and 20.0% exhibited increased oocyte vacuolization and damaged organelles. In conclusion, 2.5% Aloe vera increases antrum formation, viability and expression of mRNA for SOD in cultured secondary follicles, but higher concentrations of Aloe vera have negative effects on follicular ultrastructure.

PRDX6: A protein bridging S-palmitoylation and diabetic neuropathy

Diabetic neuropathy is regarded as one of the most debilitating outcomes of diabetes. It can affect both the peripheral and central nervous systems, leading to pain, decreased motility, cognitive decline, and dementia. S-palmitoylation is a reversible posttranslational lipid modification, and its dysregulation has been implicated in metabolic syndrome, cancers, neurological disorders, and infections. However, the role of S-palmitoylation in diabetic neuropathy remains unclear. Here we demonstrate a potential association between activating protein palmitoylation and diabetic neuropathy. We compared the proteomic data of lumbar dorsal root ganglia (DRG) of diabetes mice and palmitoylome profiling data of the HUVEC cell line. The mapping results identified peroxiredoxin-6 (PRDX6) as a novel target in diabetic neuropathy, whose biological mechanism was associated with S-palmitoylation. Bioinformatic prediction revealed that PRDX6 had two palmitoylation sites, Cys47 and Cys91. Immunofluorescence results indicated PRDX6 translocating between the cytoplasm and cell membrane. Protein function analysis proposed that increased palmitoylation could competitively inhibit the formation of disulfide-bond between Cys47 and Cys91 and change the spatial topology of PRDX6 protein. Cl–HCO3- anion exchanger 3 (AE3) was one of the AE family members, which was proved to express in DRG. AE3 activity evoked Cl- influx in neurons which was generally associated with increased excitability and susceptibility to pain. We demonstrated that the S-palmitoylation status of Cys47 could affect the interaction between PRDX6 and the C-terminal domain of AE3, thereby regulating the activity of AE3 anion exchanger enzyme in the nervous system. The results highlight a central role for PRDX6 palmitoylation in protection against diabetic neuropathy.

1643TiP First-in-human dose-escalation and expansion study (MITOPE) to evaluate mitochondrial PRX3 inhibition by RSO-021 in patients with mesothelioma and other advanced solid tumors

Tumor cells generate elevated levels of reactive oxygen species (ROS) and therefore exhibit increased expression and activity of critical ROS scavenging pathways, including the mitochondrial peroxide scavenging enzyme peroxiredoxin 3 (PRX3). RSO–021 is a novel PRX3 inhibitor that inactivates PRX3 through direct covalent adduction of active site cysteine residues, in turn, inducing oxidative stress to levels incompatible with tumor cell survival. Preclinical studies of RSO-021 show that it selectively inhibits mitochondrial PRX3 in transformed cells and is well-tolerated in animal studies, warranting its clinical phase 1/2 evaluation.

Redox regulation of RAD51 Cys319 and homologous recombination by peroxiredoxin 1

RAD51 is a critical recombinase that functions in concert with auxiliary mediator proteins to direct the homologous recombination (HR) DNA repair pathway. We show that Cys319 RAD51 possesses nucleophilic characteristics and is important for irradiation-induced RAD51 foci formation and resistance to inhibitors of poly (ADP-ribose) polymerase (PARP). We have previously identified that cysteine (Cys) oxidation of proteins can be important for activity and modulated via binding to peroxiredoxin 1 (PRDX1). PRDX1 reduces peroxides and coordinates the signaling actions of protein binding partners. Loss of PRDX1 inhibits irradiation-induced RAD51 foci formation and represses HR DNA repair. PRDX1-deficient human breast cancer cells and mouse embryonic fibroblasts display disrupted RAD51 foci formation and decreased HR, resulting in increased DNA damage and sensitization of cells to irradiation. Following irradiation cells deficient in PRDX1 had increased incorporation of the sulfenylation probe DAz-2 in RAD51 Cys319, a functionally-significant, thiol that PRDX1 is critical for maintaining in a reduced state. Molecular dynamics (MD) simulations of dT-DNA bound to a non-oxidized RAD51 protein showed tight binding throughout the simulation, while dT-DNA dissociated from an oxidized Cys319 RAD51 filament. These novel data establish RAD51 Cys319 as a functionally-significant site for the redox regulation of HR and cellular responses to IR.

Exploring urine:serum fractional excretion ratios as potential biomarkers for lupus nephritis.

ObjectivesThe goal of this exploratory study is to determine if urine:serum fractional excretion ratios can outperform the corresponding urinary biomarker proteins in identifying active renal disease in systemic lupus erythematosus (SLE).MethodsThirty-six adult SLE patients and twelve healthy controls were examined for serum and urine levels of 8 protein markers, namely ALCAM, calpastatin, hemopexin, peroxiredoxin 6 (PRDX6), platelet factor 4 (PF4), properdin, TFPI and VCAM-1, by ELISA. Fractional excretion of analyzed biomarkers was calculated after normalizing both the urine and serum biomarker levels against creatinine. A further validation cohort of fifty SLE patients was included to validate the initial findings.ResultsThe FE ratios of all 8 proteins interrogated outperformed conventional disease activity markers such as anti-dsDNA, C3 and C4 in identifying renal disease activity. All but VCAM-1FE were superior to the corresponding urine biomarkers levels in differentiating LN activity, exhibiting positive correlation with renal SLEDAI. ALCAMFE, PF4FE and properdinFE ratios exhibited the highest accuracy (AUC>0.9) in distinguishing active LN from inactive SLE. Four of the FE ratios exhibited perfect sensitivity (calpastatin, PRDX6, PF4 and properdin), while ALCAMFE, PF4FE and properdinFE exhibited the highest specificity values for active LN. In addition, several of these novel biomarkers were associated with higher renal pathology activity indices. In the validation cohort ALCAMFE, PF4FE and properdinFE once again exhibited higher accuracy metrics, surpassing corresponding urine and serum biomarkers levels, with ALCAMFE exhibiting 95% accuracy in distinguishing active LN from inactive SLE.ConclusionsWith most of the tested proteins, urine:serum fractional excretion ratios outperformed corresponding urine and serum protein measurements in identifying active renal involvement in SLE. Hence, this novel class of biomarkers in SLE ought to be systemically evaluated in larger independent cohorts for their diagnostic utility in LN assessment.

Molecular cloning, prokaryotic expression and the anti-inflammatory activity of porcine PRDX5

Peroxiredoxin 5 (PRDX5) is the sole member of the atypical 2-Cys subfamily of mammalian PRDXs, a family of thiol-dependent peroxidases. In addition to its antioxidant effect, PRDX5 has been implicated in modulating the inflammatory response. In this study, the full-length cDNA encoding porcine PRDX5 (pPRDX5) was cloned. Subsequently, using porcine alveolar macrophages (PAMs), the target cells of PRRSV infection in vivo, we found that the recombinant pPRDX5 protein inhibited inflammatory responses induced by tumor necrosis factor alpha (TNF-α) or porcine reproductive and respiratory syndrome virus (PRRSV), a virus causing severe interstitial pneumonia in pigs. By contrast, knockdown of endogenous pPRDX5 with specific siRNA enhanced inflammatory responses induced by TNF-α or PRRSV. We also demonstrated that the involvement of pPRDX5 in inflammation regulation depended on its peroxidase activity. Taken together, these results showed that pPRDX5 is an anti-inflammatory molecule, which may play an important immune-regulation role in the pathogenicity of PRRSV.

Effects of cereblon on stress-activated redox proteins and core behavior

The mechanisms underlying the vulnerability and resilience of an individual to stress are only partly understood. Response to stress is determined by behavioral and biochemical changes in the brain. Chronic ultra-mild stress (CUMS) induces an anhedonic-like state in mice that resembles symptoms of human depression. This study reports the role of cereblon (CRBN) in regulating the metabolic and antioxidant status of neuronal tissues in the mouse model of CUMS. Intriguingly, Crbn−/− (KO) mice showed resilient responsiveness, both at the behavioral and proteomic levels. Several core behaviors were also differentially altered by CUMS in KO mice. Liquid chromatography with tandem mass spectrometry (LC-MS/MS)-based proteome analysis of whole brain lysate (WBL) showed an enriched chaperonic, metabolic, and antioxidant status in the brains of KO subjects, including several members of DNAJ chaperones, creatine kinase, quinone oxidoreductase, superoxide dismutase (SOD1), glutathione S-transferase Mu (GSTM), peroxiredoxin-6 (PRDX6), and thioredoxin. Pathological phosphorylation as characterized by aggregation of tau and α-synuclein (α-syn) was significantly reduced in the neuronal tissues of KO mouse model of CUMS as compared to wild type (WT) mice. Furthermore, significantly increased SOD1 activity and reduced lipid peroxidation were observed in Crbn-KO systems. Integrated signaling pathways were also identified in CRBN-specific sub-networks constructed from protein-protein interaction analysis by STRING. The present study highlights the roles of CRBN in regulating the stress response (SR) and reshaping metabolic status in the brains of mice exposed to CUMS. A better understanding of the molecular mechanisms of depression and neurodegeneration can improve the development of novel treatments.

ANXA2, SP17, SERPINA5, PRDX2 genes, and sperm DNA fragmentation differentially represented in male partners of infertile couples with normal and abnormal sperm parameters.

This study aims to evaluate the expression of genes associated with the fertilisation potential and embryo development, sperm DNA fragmentation (SDF), and acrosome reaction in male partners of infertile couples with different sperm parameters compared to fertile men. First, male partners of infertile couples with abnormal (N= 25) and normal sperm parameters (N= 25), and fertile men (N= 10) were included in experimental groups I, II, and controls respectively. The mRNA levels of the Annexin A2 (ANXA2), Sperm protein 17 (SP17), Plasma serine protease inhibitor (SERPINA5), and Peroxiredoxin-2 (PRDX2) genes and SDF were evaluated. To evaluate the maturity of the sperm and oxidative stress, the acrosome reaction, the lipid peroxidation, and total antioxidant were measured. As result, SP17 showed a significantly lower expression in both experimental groups. SERPINA5 was significantly down-regulated in experimental group I that was aligned with the low rate of acrosome reaction. Significant overexpression of PRDX2 was found between experimental group II and controls. Significant higher rates of SDF were seen in both experimental groups compared to the controls. Finally, our data suggest that differentially gene expression of SP17 is a potential diagnostic biomarker in infertile men either with normal or abnormal sperm parameters. SDF is one of the causes of male infertility, independent of the sperm parameters.

Identification of novel biomarkers and therapeutic target candidates for stasis-heat symptom pattern of acute intracerebral hemorrhage by quantitative plasma proteomics.

To explore the novel biomarkers and therapeutic target candidates related to the stasis-heat syndrome of acute intracerebral hemorrhage (AICH). Applying an isobaric tagging for relative and absolute quantitation-(iTRAQ-) based quantitative proteomic approach, plasma samples from AICH patients with stasis-heat, and AICH patients with non-stasis-heat and healthy control subjects were collected and analyzed to distinguish differentially expressed proteins (DEPs) correlated to AICH with stasis-heat in this block design. The standard Western blot was applied to verify DEPs. Additionally, DEPs were analyzed via bioinformatic platforms and further approved via Ingenuity Pathway Analysis (IPA). A total of 26 DEPs were found among AICH with the stasis-heat, AICH with non-stasis-heat, and healthy control group. The seven DEPs compared with the non-stasis-heat group are closely related to the pathogenesis of stasis heat. These proteins showed three different protein expression patterns. The alpha-1-b glycoprotein (A1BG) and copper-protein (CP) were up-regulated in the stasis-heat group, but down-regulated in the non-stasis-heat group. Compared with the non-stasis-heat group, the expression abundance of actinin, alpha 1 (ACTN1), carbonic anhydrase I (CA1), peroxiredoxin 2 (PRDX2), and vinculin (VCL) is higher in the stasis-heat group, while the CD44 is the opposite. These differences reflect that stasis-heat syndrome has more severe inflammatory immune response, coagulation disorders and damage. Bioinformatics analysis revealed that a wide variety of cellular and metabolic processes and some signaling pathways were involved in the pathophysiology of AICH with stasis-heat. AICH with stasis-heat syndrome showed more severe inflammatory reactions, tissue damage, and coagulation disorders than non-stasis heat syndrome. There are differences in the protein expression patterns between the stasis-heat syndrome and non-stasis-heat syndrome. These differences reflect that stasis-heat syndrome has more severe damage. CD44, CP, ACTN1, CA1, VCL, PRDX2, and A1BG could be the potential biomarkers and therapeutic target candidates of the stasis-heat subtype. This study provides a reasonable explaination for Liangxue Tongyu decoction through anti-inflammatory and brain protection treatment.

Tumor suppressor gene ARMC5 controls adrenal redox state through NRF1 turnover.

is a tumor suppressor gene frequently mutated in primary bilateral macronodular adrenal hyperplasia (PBMAH), an adrenal cause of Cushing's syndrome. The function of ARMC5 is poorly understood, aside from the fact that it regulates cell viability and adrenal steroidogenesis by mechanisms still unknown. Tumor suppressor genes play an important role in modifying intracellular redox response, which in turn regulates diverse cell signaling pathways. In this study, we demonstrated that inactivation in adrenocortical cells increased the expression of actors scavenging reactive oxygen species, such as superoxide dismutases (SOD) and peroxiredoxins (PRDX) by increasing the transcriptional regulator NRF1. Moreover, ARMC5 is involved in the NRF1 ubiquitination and in its half-life. Finally, inactivation alters adrenocortical steroidogenesis through the activation of p38 pathway and decreases cell sensitivity to ferroptosis participation to increase cell viability. Altogether, this study uncovers a function of ARMC5 as a regulator of redox homeostasis in adrenocortical cells, controlling steroidogenesis and cell survival.

Expression of antioxidant enzymes in lesions of multiple sclerosis and its models

Oxidative stress promotes tissue injury in the central nervous system in neurological disorders such as multiple sclerosis (MS). To protect against this, antioxidant enzymes including superoxide dismutase-1 (SOD1), heme oxygenase-1 (HO-1), peroxiredoxin-5 (PRDX5) and glutathione peroxidase-4 (GPX4) may be upregulated. However, whether antioxidant enzyme elevation in mouse models of neurodegeneration corresponds to their expression in human diseases such as MS requires investigation. Here, we analyzed and compared the expression of SOD1, HO-1, PRDX5 and GPX4 in the murine spinal cord of three models of MS: focal lesions induced by (1) oxidized phosphatidylcholine or (2) lysophosphatidylcholine (lysolecithin), and (3) diffuse lesions of experimental autoimmune encephalomyelitis. Notably, CD68+ microglia/macrophages were the predominant cellular populations that expressed the highest levels of the detected antioxidant enzymes. Overall, the expression patterns of antioxidant enzymes across the models were similar. The increase of these antioxidant enzymes was corroborated in MS brain tissue using spatial RNA sequencing. Collectively, these results show that antioxidant capacity is relatively conserved between mouse models and MS lesions, and suggest a need to investigate whether the antioxidant elevation in microglia/macrophages is a protective response during oxidative injury, neurodegeneration, and MS.

Structure-based analysis and rational design of human peroxiredoxin-1's C-terminus-derived peptides to target sulfiredoxin-1 in pancreatic cancer

Human peroxiredoxin (PRX) family of antioxidant enzymes reduces hydrogen peroxide and alkyl hydroperoxide involved in the redox signaling, among which the widely documented PRX1 is a versatile molecule regulating cell growth, differentiation and apoptosis, and has been implicated in the tumorigensis of pancreatic cancer. In this study, we systematically examined the complex crystal structure of PRX1 with its cognate interacting partner sulfiredoxin-1 (SRX1) at molecular level, and found that the PRX1–SRX1 association is a typical peptide-mediated protein–protein interaction, where a 18-mer C-terminal tail (CTT) segment of PRX1 was identified to be primarily responsible for the interaction, which contributes ~80% and ~ 55% to the total binding potency of SRX1 to PRX1 monomer and homodimer, respectively. We also demonstrated that the SRX1 exhibits a strong global selectivity for PRX1 CTT tail over other PRX family proteins. Next, the intermolecular interaction between PRX1 CTT tail and SRX1 was investigated at structural, energetic and dynamic levels, from which a 9-mer core region of PRX1 CTT tail was defined as the SRX1-binding hotspot. Biophysical assays substantiated that the CTT and CTTc peptides (out of PRX1 protein context) can bind in an independent manner and possess a close affinity to SRX1. Based on the CTTc sketch a computational combinatorial library consisting of 216 designed peptide derivatives was rationally generated, from which the top-5 hits were found to have comparable affinity with CTT peptide and improved affinity relative to CTTc peptide. They can be used as structurally reduced lead molecular entities to further develop new peptidic agents for therapeutic purpose to disrupt the native PRX1–SRX1 interaction by competing with PRX1 CTT tail for the peptide-binding pocket of SRX1.

Structural New Data for Mitochondrial Peroxiredoxin From Trypanosoma cruzi Show High Similarity With Human Peroxiredoxin 3: Repositioning Thiostrepton as Antichagasic Drug.

Trypanosoma cruzi, the causal agent of Chagas disease, has peroxiredoxins (PRXs) expressed in all stages of the parasite and whose function is to detoxify oxidizing agents, such as reactive oxygen species (ROS). These proteins are central for the survival and replication of the parasite and have been proposed as virulence factors. Because of their importance, they have also been considered as possible therapeutic targets, although there is no specific drug against them. One of them, the mitochondrial PRX (TcMPX), is important in the detoxification of ROS in this organelle and has a role in the infectivity of T. cruzi. However, their structural characteristics are unknown, and possible inhibitors have not been proposed. The aim was to describe in detail some structural characteristics of TcMPX and compare it with several PRXs to find possible similarities and repositioning the antibiotic Thiostrepton as a potential inhibitor molecule. It was found that, in addition to the characteristic active site of a 2-cys PRX, this protein has a possible transmembrane motif and motifs involved in resistance to hyper oxidation. The homology model suggests a high structural similarity with human PRX3. This similarity was corroborated by cross-recognition using an anti-human PRX antibody. In addition, molecular docking showed that Thiostrepton, a potent inhibitor of human PRX3, could bind to TcMPX and affect its function. Our results show that Thiostrepton reduces the proliferation of T. cruzi epimastigotes, cell-derived trypomastigotes, and blood trypomastigotes with low cytotoxicity on Vero cells. We also demonstrated a synergic effect of Thriostepton and Beznidazol. The convenience of seeking treatment alternatives against T. cruzi by repositioning compounds as Thiostrepton is discussed.

GRASP55 regulates the unconventional secretion and aggregation of mutant huntingtin

Recent studies demonstrated that the Golgi reassembly stacking proteins (GRASPs), especially GRASP55, regulate Golgi-independent unconventional secretion of certain cytosolic and transmembrane cargoes; however, the underlying mechanism remains unknown. Here, we surveyed several neurodegenerative disease–related proteins, including mutant huntingtin (Htt-Q74), superoxide dismutase 1 (SOD1), tau, and TAR DNA–binding protein 43 (TDP-43), for unconventional secretion; our results show that Htt-Q74 is most robustly secreted in a GRASP55-dependent manner. Using Htt-Q74 as a model system, we demonstrate that unconventional secretion of Htt is GRASP55 and autophagy dependent and is enhanced under stress conditions such as starvation and endoplasmic reticulum stress. Mechanistically, we show that GRASP55 facilitates Htt secretion by tethering autophagosomes to lysosomes to promote autophagosome maturation and subsequent lysosome secretion and by stabilizing p23/TMED10, a channel for translocation of cytoplasmic proteins into the lumen of the endoplasmic reticulum–Golgi intermediate compartment. Moreover, we found that GRASP55 levels are upregulated by various stresses to facilitate unconventional secretion, whereas inhibition of Htt-Q74 secretion by GRASP55 KO enhances Htt aggregation and toxicity. Finally, comprehensive secretomic analysis identified novel cytosolic cargoes secreted by the same unconventional pathway, including transgelin (TAGLN), multifunctional protein ADE2 (PAICS), and peroxiredoxin-1 (PRDX1). In conclusion, this study defines the pathway of GRASP55-mediated unconventional protein secretion and provides important insights into the progression of Huntington’s disease.

2D-DIGE-MS Proteomics Approaches for Identification of Gelsolin and Peroxiredoxin 4 with Lymph Node Metastasis in Colorectal Cancer.

Simple SummarySchematic presentation of the histone modification of gelsolin (GSN) and peroxiredoxin-4 (PRDX4) promoters by endogenous EGFR/RhoA/PKC/ERK signaling pathways in DLD-1 cells. Identifying lymph node metastasis (LNM)-associated proteins by two-dimensional difference gel electrophoresis MS/MS in colorectal cells (CRC) indicates the reliability of candidate markers of GSN and PRDX4 and therapeutic targets for CRC.Background/Aims: A combination of fluorescence two-dimensional difference gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization time of flight mass spectrometry approach was used to search for potential markers for prognosis and intervention of colorectal cancer (CRC) at different stages of lymph node metastasis (LMN). This quantitative proteomic survey aimed to investigate the LNM-associated proteins and evaluate the clinicopathological characteristics of these target proteins in CRC from stage I to stage IV. Methods: Sixteen CRC cases were categorized into paired non-LNM and LNM groups, and two-dimensional difference gel electrophoresis and MS proteome analysis were performed. Differential protein expression between non-LNM and LNM CRC was further validated in a tissue microarray, including 40 paraffin-embedded samples by immunohistochemistry staining. Moreover, a Boyden chamber assay, flow cytometry, and shRNA were used to examine the epithelial–mesenchymal transition and mechanism invasiveness of the differentially expressed proteins in DLD-1 cells and in vivo xenograft mouse model. Results: Eighteen differentially expressed proteins were found between non-LNM and LNM CRC tissues. Among them, protein levels of Gelsolin (GSN) and peroxiredoxin 4 (PRDX4) were abundant in node-positive CRC. Downregulation of GSN and PRDX4 markedly suppressed migration and invasiveness and also induced cell cycle G1/S arrest in DLD-1. Mechanistically, the EGFR/RhoA/PKCα/ERK pathways are critical for transcriptional activation of histone modification of H3 lysine 4 trimethylation (H3K4me3) of GSN and PRDX4 promoters, resulting in upregulation of GSN, PRDX4, Twist-1/2, cyclinD1, proliferating cell-nuclear antigen, β-catenin, N-cadherin, and matrix metalloprotein-9. Conclusions: GSN and PRDX4 are novel regulators in CRC lymph node metastasis to potentially provide new insights into the mechanism of CRC progression and serve as a biomarker for CRC diagnosis at the metastatic stage.