Activation Of Receptor For Advanced Glycation Endproducts Research Articles

MiR-23a-5p alleviates chronic obstructive pulmonary disease through targeted regulation of RAGE-ROS pathway

BackgroundChronic obstructive pulmonary disease (COPD) is a common respiratory disease and represents the third leading cause of death worldwide. This study aimed to investigate miRNA regulation of Receptor for Advanced Glycation End-products (RAGE), a causal receptor in the pathogenesis of cigarette smoke (CS)-related COPD, to guide development of therapeutic strategies.MethodsRAGE expression was quantified in lung tissue of COPD patients and healthy controls, and in mice with CS-induced COPD. RNA-sequencing of peripheral blood from COPD patients with binding site prediction was used to screen differentially expressed miRNAs that may interact with RAGE. Investigation of miR-23a-5p as a potential regulator of COPD progression was conducted with miR-23a-5p agomir in COPD mice in vivo using histology and SCIREQ functional assays, while miR-23a-5p mimics or RAGE inhibitor were applied in 16-HBE human bronchial epithelial cells in vitro. RNA-sequencing, ELISA, and standard molecular techniques were used to characterize downstream signaling pathways in COPD mice and 16-HBE cells treated with cigarette smoke extract (CSE).ResultsRAGE expression is significantly increased in lung tissue of COPD patients, COPD model mice, and CSE-treated 16-HBE cells, while inhibiting RAGE expression significantly reduces COPD severity in mice. RNA-seq analysis of peripheral blood from COPD patients identified miR-23a-5p as the most significant candidate miRNA interaction partner of RAGE, and miR-23a-5p is significantly downregulated in mice and cells treated with CS or CSE, respectively. Injection of miR-23a-5p agomir leads to significantly reduced airway inflammation and alleviation of symptoms in COPD mice, while overexpressing miR-23a-5p leads to improved lung function. RNA-seq with validation confirmed that reactive oxygen species (ROS) signaling is increased under CSE-induced aberrant upregulation of RAGE, and suppressed in CSE-stimulated cells treated with miR-23a-5p mimics or overexpression. ERK phosphorylation and subsequent cytokine production was also increased under RAGE activation, but inhibited by increasing miR-23a-5p levels, implying that the miR-23a-5p/RAGE/ROS axis mediates COPD pathogenesis via ERK activation.ConclusionsThis study identifies a miR-23a-5p/RAGE/ROS signaling axis required for pathogenesis of COPD. MiR-23a-5p functions as a negative regulator of RAGE and downstream activation of ROS signaling, and can inhibit COPD progression in vitro and in vivo, suggesting therapeutic targets to improve COPD treatment.Graphical

Modulating the RAGE-Induced Inflammatory Response: Peptoids as RAGE Antagonists.

While the primary pathology of Alzheimer's disease (AD) is defined by brain deposition of amyloid-β (Aβ) plaques and tau neurofibrillary tangles, chronic inflammation has emerged as an important factor in AD etiology. Upregulated cell surface expression of the receptor for advanced glycation end-products (RAGE), a key receptor of innate immune response, is reported in AD. In parallel, RAGE ligands, including Aβ aggregates, HMGB1, and S100B, are elevated in AD brain. Activation of RAGE by these ligands triggers release of inflammatory cytokines and upregulates cell surface RAGE. Despite such observation, there are currently no therapeutics that target RAGE for treatment of AD-associated neuroinflammation. Peptoids, a novel class of potential AD therapeutics, display low toxicity, facile blood-brain barrier permeability, and resistance to proteolytic degradation. In the current study, peptoids were designed to mimic Aβ, a ligand that binds the V-domain of RAGE, and curtail RAGE inflammatory activation. We reveal the nanomolar binding capability of peptoids JPT1 and JPT1a to RAGE and demonstrate their ability to attenuate lipopolysaccharide-induced pro-inflammatory cytokine production as well as upregulation of RAGE cell surface expression. These results support RAGE antagonist peptoid-based mimics as a prospective therapeutic strategy to counter neuroinflammation in AD and other neurodegenerative diseases.

RAGE signaling regulates the progression of diabetic complications.

Diabetes, the ninth leading cause of death globally, is expected to affect 642 million people by 2040. With the advancement of an aging society, the number of patients with diabetes having multiple underlying diseases, such as hypertension, obesity, and chronic inflammation, is increasing. Thus, the concept of diabetic kidney disease (DKD) has been accepted worldwide, and comprehensive treatment of patients with diabetes is required. Receptor for advanced glycation endproducts (RAGE), a multiligand receptor, belonging to the immunoglobulin superfamily is extensively expressed throughout the body. Various types of ligands, including advanced glycation endproducts (AGEs), high mobility group box 1, S100/calgranulins, and nucleic acids, bind to RAGE, and then induces signal transduction to amplify the inflammatory response and promote migration, invasion, and proliferation of cells. Furthermore, the expression level of RAGE is upregulated in patients with diabetes, hypertension, obesity, and chronic inflammation, suggesting that activation of RAGE is a common denominator in the context of DKD. Considering that ligand-and RAGE-targeting compounds have been developed, RAGE and its ligands can be potent therapeutic targets for inhibiting the progression of DKD and its complications. Here, we aimed to review recent literature on various signaling pathways mediated by RAGE in the pathogenesis of diabetic complications. Our findings highlight the possibility of using RAGE-or ligand-targeted therapy for treating DKD and its complications.

Dieckol, Derived from the Edible Brown Algae Ecklonia cava, Attenuates Methylglyoxal-Associated Diabetic Nephropathy by Suppressing AGE–RAGE Interaction

The formation of advanced glycation end products (AGE) is linked to the pathogenesis of diabetic nephropathy. The aim of this work was to assess the therapeutic potential and underlying mechanism of action of dieckol (DK), isolated from Ecklonia cava, on renal damage induced by methylglyoxal (MGO) in mouse glomerular mesangial cells. The antiglycation properties of DK were evaluated using ELISA. We conducted molecular docking, immunofluorescence analysis, and Western blotting to confirm the mechanism by which DK prevents AGE-related diabetic nephropathy. DK treatment exhibited antiglycation properties through the inhibition of AGE production, inhibition of cross-linking between AGE and collagen, and breaking of its cross-linking. DK pretreatment exhibited protective effects on renal cells by suppressing MGO-induced intracellular reactive oxygen species (ROS) formation, intracellular MGO and AGE accumulation, activation of the apoptosis cascade and apoptosis-related protein expression, activation of receptor for AGE (RAGE) protein expression, and suppression of the glyoxalase system. Furthermore, DK exhibited a stronger binding affinity for RAGE than AGE, which was confirmed as exerting a competitive inhibitory effect on the AGE-RAGE interaction. These results demonstrated that DK is a potential natural AGE inhibitor that can be utilized to prevent and treat AGE-induced diabetic nephropathy.

Peptoids as potential therapeutics to reduce S100B-induced RAGE-associatedneuroinflammation in Alzheimer's disease.

Receptor for advanced glycation end-products (RAGE)-associated chronic neuroinflammation has been linked with Alzheimer's disease (AD) pathology. RAGE ligand S100B, which is elevated in AD brain, can induce release of inflammatory cytokines and upregulate RAGE. Consequently, targeting RAGE to reduce chronic inflammation represents a novel therapeutic strategy for AD. Peptoids are ideal potential AD therapeutics as they resist proteolytic degradation by repositioning side-chains from the α-carbon to the amide nitrogen and exhibit facile blood-brain barrier permeability. Peptoids designed to mimic RAGE ligands demonstrated strong binding affinity to RAGE in our previous studies. We hypothesize that these peptoids can attenuate S100B-induced activation of RAGE, thus reducing pro-inflammatory cytokine release associated with chronic neuroinflammation. Differentiated THP-1 macrophages were exposed to chronic low-levels of S100B to confirm upregulation of cytokine release. Macrophages were then treated with 0-50-μM peptoids (JPT1 or JPT1a) in the presence of S100B. To observe peptoid modulation of S100B-induced inflammation, cytokine analysis via immunoassay was performed using cell culture supernatants. Additionally, RAGE was incubated in the presence of 0-500-nM peptoid to achieve equilibrium. Unbound RAGE was quantified via modified ELISA to determine the binding affinity. S100B-treated THP-1 macrophages elicited anticipated inflammatory response evidenced by dose-dependent upregulation of cytokines. When THP-1 macrophages were incubated with peptoids and S100B, peptoids attenuated these S100B-induced inflammatory responses. Moreover, the peptoids demonstrated nanomolar binding affinity for RAGE. Peptoids modulate inflammatory responses induced by RAGE ligand S100B, including inflammatory cytokine release. These results implicate peptoids as a potential therapeutic agent for not only AD but also other inflammation-related illnesses. Because these peptoids have additionally exhibited the ability to modulate Aβ aggregation and transform the morphology of the aggregates formed, they may function as dual-target therapeutics for AD.

RAGE pathway activation and function in chronic kidney disease and COVID-19.

The multi-ligand receptor for advanced glycation end-products (RAGE) and its ligands are contributing factors in autoimmunity, cancers, and infectious disease. RAGE activation is increased in chronic kidney disease (CKD) and coronavirus disease 2019 (COVID-19). CKD may increase the risk of COVID-19 severity and may also develop in the form of long COVID. RAGE is expressed in essentially all kidney cell types. Increased production of RAGE isoforms and RAGE ligands during CKD and COVID-19 promotes RAGE activity. The downstream effects include cellular dysfunction, tissue injury, fibrosis, and inflammation, which in turn contribute to a decline in kidney function, hypertension, thrombotic disorders, and cognitive impairment. In this review, we discuss the forms and mechanisms of RAGE and RAGE ligands in the kidney and COVID-19. Because various small molecules antagonize RAGE activity in animal models, targeting RAGE, its co-receptors, or its ligands may offer novel therapeutic approaches to slowing or halting progressive kidney disease, for which current therapies are often inadequate.

MLKL Regulates Rapid Cell Death-independent HMGB1 Release in RSV Infected Airway Epithelial Cells.

Respiratory syncytial virus (RSV)-induced bronchiolitis is a significant contributor to infant morbidity and mortality. Previously, we identified that necroptosis, a pro-inflammatory form of cell death mediated by receptor-interacting serine/threonine-protein kinase 1 (RIPK1) and RIPK3, and mixed lineage kinase domain like protein (MLKL), occurs in RSV-infected human airway epithelial cells (hAECs), mediating the release of the alarmin high mobility group box 1 (HMGB1). Here, we show that RSV infection of hAECs induces the biphasic release of HMGB1 at 6 (“early”) and 24 (“late”) hours post infection (hpi). The early phase of HMGB1 release at 6 hpi is cell death-independent, however, this release is nonetheless attenuated by inhibition of MLKL (primarily associated with necroptosis). The early release of HMGB1 promotes the late phase of HMGB1 release via the activation of RAGE (receptor for advanced glycation endproducts) and occurs with cell death. Treatment of hAECS with exogenous HMGB1 combined with a pan-caspase inhibitor induces hAEC necroptosis, and is attenuated by the RAGE antagonist, FPS-ZM1. Together, these findings demonstrate that RSV infection of hAECs leads to the early release of HMGB1, followed by a paracrine feed-forward amplification loop that further increases HMGB1 levels and promotes cell death. As the inhibition of MLKL or targeting of HMGB1/RAGE pathway attenuates the release of pro-inflammatory HMGB1 and decreases viral load, this suggests that the pharmacological targeting of these pathways may be of benefit for the treatment of severe RSV bronchiolitis.

The Role of AGE-RAGE Signalling as a Modulator of Gut Permeability in Diabetes

There is increasing evidence for the role of intestinal permeability as a contributing factor in the pathogenesis of diabetes; however, the molecular mechanisms are poorly understood. Advanced glycation endproducts, of both exogenous and endogenous origin, have been shown to play a role in diabetes pathophysiology, in part by their ligation to the receptor for advanced glycation endproducts (RAGE), leading to a proinflammatory signalling cascade. RAGE signalling has been demonstrated to play a role in the development of intestinal inflammation and permeability in Crohn’s disease and ulcerative colitis. In this review, we explore the role of AGE-RAGE signalling and intestinal permeability and explore whether activation of RAGE on the intestinal epithelium may be a downstream event contributing to the pathogenesis of diabetes complications.

Non-enzymatic glycoxidation linked with nutrition enhances the tumorigenic capacity of prostate cancer epithelia through AGE mediated activation of RAGE in cancer associated fibroblasts

The molecular implications of food consumption on cancer etiology are poorly defined. The rate of nutrition associated non-enzymatic glycoxidation, a reaction that occurs between reactive carbonyl groups on linear sugars and nucleophilic amino, lysyl and arginyl groups on fats and proteins, is rapidly increased by food cooking and manufacturing processes. In this study, we assign nutrition-associated glycoxidation with significant oncogenic potential, promoting prostate tumor growth, progression, and metastasis in vivo. Advanced glycation end products (AGEs) are the final irreversible product of non-enzymatic glycoxidation. Exogenous treatment of prostate tumor cells with a single AGE peptide replicated glycoxidation induced tumor growth in vivo. Mechanistically, receptor for AGE (RAGE) deficiency in the stroma inhibited AGE mediated tumor growth. Functionally, AGE treatment induced RAGE dimerization in activated fibroblasts which sustained and increased the migratory potential of tumor epithelial cells. These data identify a novel nutrition associated pathway that can promote a tissue microenvironment conducive for aggressive tumor growth. Targeted and/or interventional strategies aimed at reducing AGE bioavailability as a consequence of nutrition may be viewed as novel chemoprevention initiatives.

LL-37 and HMGB1 induce alveolar damage and reduce lung tissue regeneration via RAGE.

The receptor for advanced glycation end-products (RAGE) has been implicated in the pathophysiology of chronic obstructive pulmonary disease (COPD). However, it is still unknown whether RAGE directly contributes to alveolar epithelial damage and abnormal repair responses. We hypothesize that RAGE activation not only induces lung tissue damage but also hampers alveolar epithelial repair responses. The effects of the RAGE ligands LL-37 and HMGB1 were examined on airway inflammation and alveolar tissue damage in wild-type and RAGE-deficient mice and on lung damage and repair responses using murine precision cut lung slices (PCLS) and organoids. In addition, their effects were studied on the repair response of human alveolar epithelial A549 cells, using siRNA knockdown of RAGE and treatment with the RAGE inhibitor FPS-ZM1. We observed that intranasal installation of LL-37 and HMGB1 induces RAGE-dependent inflammation and severe alveolar tissue damage in mice within 6 h, with stronger effects in a mouse strain susceptible for emphysema compared with a nonsusceptible strain. In PCLS, RAGE inhibition reduced the recovery from elastase-induced alveolar tissue damage. In organoids, RAGE ligands reduced the organoid-forming efficiency and epithelial differentiation into pneumocyte-organoids. Finally, in A549 cells, we confirmed the role of RAGE in impaired repair responses upon exposure to LL-37. Together, our data indicate that activation of RAGE by its ligands LL-37 and HMGB1 induces acute lung tissue damage and that this impedes alveolar epithelial repair, illustrating the therapeutic potential of RAGE inhibitors for lung tissue repair in emphysema.

Caspase-Dependent HMGB1 Release from Macrophages Participates in Peripheral Neuropathy Caused by Bortezomib, a Proteasome-Inhibiting Chemotherapeutic Agent, in Mice.

Given the role of macrophage-derived high mobility group box 1 (HMGB1) in chemotherapy-induced peripheral neuropathy (CIPN) caused by paclitaxel, we analyzed the role of HMGB1 and macrophages in the CIPN caused by bortezomib, a proteasome-inhibiting chemotherapeutic agent used for the treatment of multiple myeloma. Repeated administration of bortezomib caused CIPN accompanied by early-stage macrophage accumulation in the dorsal root ganglion. This CIPN was prevented by an anti-HMGB1-neutralizing antibody, thrombomodulin alfa capable of accelerating thrombin-dependent degradation of HMGB1, antagonists of the receptor for advanced glycation end-products (RAGE) and C-X-C motif chemokine receptor 4 (CXCR4), known as HMGB1-targeted membrane receptors, or macrophage depletion with liposomal clodronate, as reported in a CIPN model caused by paclitaxel. In macrophage-like RAW264.7 cells, bortezomib as well as MG132, a well-known proteasome inhibitor, caused HMGB1 release, an effect inhibited by caspase inhibitors but not inhibitors of NF-κB and p38 MAP kinase, known to mediate paclitaxel-induced HMGB1 release from macrophages. Bortezomib increased cleaved products of caspase-8 and caused nuclear fragmentation or condensation in macrophages. Repeated treatment with the caspase inhibitor prevented CIPN caused by bortezomib in mice. Our findings suggest that bortezomib causes caspase-dependent release of HMGB1 from macrophages, leading to the development of CIPN via activation of RAGE and CXCR4.

Advanced Glycation End Products Impair Cardiac Atrial Appendage Stem Cells Properties.

Background: During myocardial infarction (MI), billions of cardiomyocytes are lost. The optimal therapy should effectively replace damaged cardiomyocytes, possibly with stem cells able to engraft and differentiate into adult functional cardiomyocytes. As such, cardiac atrial appendage stem cells (CASCs) are suitable candidates. However, the presence of elevated levels of advanced glycation end products (AGEs) in cardiac regions where CASCs are transplanted may affect their regenerative potential. In this study, we examine whether and how AGEs alter CASCs properties in vitro. Methods and Results: CASCs in culture were exposed to ranging AGEs concentrations (50 µg/mL to 400 µg/mL). CASCs survival, proliferation, and migration capacity were significantly decreased after 72 h of AGEs exposure. Apoptosis significantly increased with rising AGEs concentration. The harmful effects of these AGEs were partially blunted by pre-incubation with a receptor for AGEs (RAGE) inhibitor (25 µM FPS-ZM1), indicating the involvement of RAGE in the observed negative effects. Conclusion: AGEs have a time- and concentration-dependent negative effect on CASCs survival, proliferation, migration, and apoptosis in vitro, partially mediated through RAGE activation. Whether anti-AGEs therapies are an effective treatment in the setting of stem cell therapy after MI warrants further examination.

Nutritional AGEing and RAGEing as a Regulator of the Tumor Microenvironment

Abstract Objectives Advanced glycation end products (AGEs) are reactive metabolites formed endogenously by glyoxidative, oxidative and lipoxidative stresses. Foods associated with modern dietary habits are particularly AGE laden but despite increasing epidemiological evidence for oncogenic potential, cause and effect relationships are lacking. The objective was to provide detailed mechanistic insight and in vivo confirmation that AGEs found in the diet are oncogenic drivers of tumorigenesis. Methods We used the heat driven formation of glyoxidative, oxidative and lipoxidative stresses in experimental mouse chow to reproduce the wide spectrum of the AGEs found in vivo. Syngeneic xenograft and spontaneous prostate and breast cancer mouse models were then fed the AGE specific diets and the effects of chronic AGE consumption on tumor growth assessed. To gain mechanistic insight, human and mouse two compartment co-culture models using primary fibroblasts and matched tumor epithelial cells were then used to assess the effects of AGEs on extracellular crosstalk in the TME. Results A high impact finding from our research is that consumption of AGEs found in our diet promotes prostate tumor growth, aggression and metastasis by functioning as ligand to the transmembrane receptor for AGE (RAGE). Dietary-AGEs promoted neoplastic growth by functioning as ligand to RAGE expressed in the prostate tumor stroma not tumor epithelium. Dietary-AGE activation of stromal RAGE caused a regulatory program of ‘activated fibroblasts’ defined by the increased expression of cancer associated fibroblast markers, NFkB, MYC and pro-tumorigenic paracrine secretion. Fibroblast activation was accompanied by decreased expression of androgen receptor (AR) and the increased expression of neuroendocrine differentiation markers in tumor epithelial cells. AGE exposed primary fibroblasts isolated from patient tissue conferred tumor promoting abilities when cultured with patient matched tumor epithelial cells. Conclusions For the first time these data demonstrate a direct cause and effect relationship between dietary-AGEs and neoplastic growth. This may lay the foundation for strategic self-management strategies aimed at reducing AGE exposure in the diet to reduce cancer incidence and mortality. Funding Sources NIH/NCI; ACS.

Receptor for Advanced Glycation End Products Acts as a Fuel to Colorectal Cancer Development.

Receptor for advanced glycation end-products (RAGE) is a multiligand binding and single-pass transmembrane protein taken in diverse chronic inflammatory conditions. RAGE behaves as a pattern recognition receptor, which binds and is engaged in the cellular response to a variety of damage-associated molecular pattern molecules, as well as HMGB1, S100 proteins, and AGEs (advanced glycation end-products). The RAGE activation turns out to a formation of numerous intracellular signaling mechanisms, resulting in the progression and prolongation of colorectal carcinoma (CRC). The RAGE expression correlates well with the survival of colon cancer cells. RAGE is involved in the tumorigenesis, which increases and develops well in the stressed tumor microenvironment. In this review, we summarized downstream signaling cascade activated by the multiligand activation of RAGE, as well as RAGE ligands and their sources, clinical studies, and tumor markers related to RAGE particularly in the inflammatory tumor microenvironment in CRC. Furthermore, the role of RAGE signaling pathway in CRC patients with diabetic mellitus is investigated. RAGE has been reported to drive assorted signaling pathways, including activator protein 1, nuclear factor-κB, signal transducer and activator of transcription 3, SMAD family member 4 (Smad4), mitogen-activated protein kinases, mammalian target of rapamycin, phosphoinositide 3-kinases, reticular activating system, Wnt/β-catenin pathway, and Glycogen synthase kinase 3β, and even microRNAs.

Abstract C029: Consumption of dietary AGEs during puberty and increased breast cancer risk: A link between lifestyle and cancer disparity

Abstract Introduction. The focus of this study is on early lifestyle factors and their effect on mammary development during puberty and how they relate to increased breast cancer risk and disparities. At this time we do not understand what biological changes occur during pubertal mammary development which leads to a greater risk of developing cancer in later life. Identifying the molecular mechanisms that cause aberrant pubertal mammary development may lead to defined strategies to reduce breast cancer burden in later life. As our bodies use the sugars that we consume for energy they generate waste chemicals known as advanced glycation end products or AGEs for short. Significantly, low income, obesity and a sedentary lifestyle are established factors driving health disparity that also contribute to increased AGE accumulation levels in our bodies. In particular, AGE content in the Western Diet has consistently increased over the last 50 years due to increased consumption of sugar-laden and cheap processed/manufactured foods which are high in reactive AGE metabolites and can promote obesity. Methods. We use a dietary mouse model to assess impact of AGE on normal mammary development. Wild type FVB/n and RAGE null (RAGE-/-) mice are fed the respective diets from weaning until 7 (pubertal) or 12 (adult) weeks of age. Mammary glands are extracted for whole mounting and paraffin embedded for histology. Fibroblasts were isolated from mammary glands and cultured ex vivo. Transwell migration assays were performed with isolated fibroblasts and HC11 mouse mammary epithelial cells. qPCR was performed on the isolated fibroblasts to assess their activation status. Results. Early life exposures during mammary development influence the breast microenvironment to increase breast cancer risk. We show that due to an innate ability to influence the cellular matrix, dietary AGEs disrupt developmental programs during puberty and promote breast tumor growth. Through receptor for AGE (RAGE) dependent and independent mechanisms, chronic AGE consumption delayed ductal extension, increased ductal branching and caused aberrant terminal end bud (TEB) morphology. Dietary AGE activation of RAGE mediated a program of activated stroma leading to hyperplastic growth and the formation of pre-neoplastic lesions which persisted into adulthood. Importantly, AGE mediated effects remained even after diet intervention after puberty. In dietary-AGE breast tumor models, AGE mediated changes in tissue architecture and cell function were recapitulated and resulted in 3-fold increase in neoplastic growth. Through the perpetual activation of a reactive stroma, AGEs derived from diet represent a common early life exposure which can influence tumor behavior. Conclusions. A greater mechanistic understanding of the link between AGE intake during puberty and increased breast cancer risk may define novel potential strategies for lifestyle and pharmacological intervention aimed at reducing breast cancer risk and cancer disparities. Citation Format: Callan C Frye, Bradley A Krisanits, Reid Schuster, Jaime Randise, Lourdes M Nogueira, Kristi Helke, Amanda C LaRue, David P Turner, Victoria J Findlay. Consumption of dietary AGEs during puberty and increased breast cancer risk: A link between lifestyle and cancer disparity [abstract]. In: Proceedings of the Twelfth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2019 Sep 20-23; San Francisco, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl_2):Abstract nr C029.

Consumption of Dietary Advanced Glycation End Products Promotes Prostate Tumor Growth by Creating a Tumor Enhancing Stromal Microenvironment

Abstract Objectives The literature regarding the role of advanced glycation end products (AGEs) on tumor biology has shown only moderate promise reflected by increases in cell growth, migration and invasion in vitro which is not supported by increased tumor growth in vivo14-16– A caveat to these studies is that they are centered upon a single AGE peptide and a subsequent assessment of their molecular effects on tumor epithelial cells. The objective is to show that by feeding mice a high AGE diet we can recapitulate a microenvironment comprising of a wide spectrum of AGEs which can influence neoplastic growth. Methods We recapitulated a dietary-AGE induced microenvironment in syngeneic xenograft and spontaneous breast and prostate mouse cancer models and the effects on tumor growth assessed. The mechanistic consequences of dietary-AGEs on the tumor microenvironment were further defined using mouse and human primary and immortalized two-compartment co-culture ex vivo culture models. Results Dietary-AGE consumption in breast and prostate tumor models significantly accelerated tumor growth by functioning as ligand to the transmembrane receptor for AGE (RAGE). Our studies demonstrate that AGEs promote neoplastic growth by functioning as ligand to RAGE expressed in the tumor stroma not the tumor epithelial cells. Dietary-AGE activation of RAGE in both breast and prostate tumors caused a regulatory program of ‘activated fibroblasts’ defined by increased expression of cancer associated fibroblast markers, NFkB and MYC upregulation, and pro-tumorigenic paracrine secretion. Complementary to this, our published studies show that high intake of dietary AGE after BCa diagnosis increases risk of mortality in postmenopausal women. Conclusions These data demonstrate, for the first time, the oncogenic potential of dietary-AGEs in promoting neoplastic growth. This lays the foundation for strategic changes aimed at reducing cancer incidence and mortality as pharmacological, educational and/or interventional strategies aimed at reducing the dietary-AGE accumulation pool may one day be viewed as universal cancer preventative and/or therapeutic initiatives especially when combined with existing therapies. Funding Sources David P. Turner was supported by grants from the NIH/NCI, R21 CA194469 and U54 CA21096..

Receptor for advanced glycation end products modulates oxidative stress and mitochondrial function in the soleus muscle of mice fed a high-fat diet.

Accumulation of advanced glycation end products (AGEs) and activation of the receptor for AGEs (RAGE) are implicated in the progression of pathologies associated with aging, chronic inflammation, diabetes, and cellular stress. RAGE activation is also implicated in cardiovascular complications of type 2 diabetes, such as nephropathy, retinopathy, accelerated vascular diseases, and cardiomyopathy. Studies investigating the effects of AGE/RAGE axis activation on skeletal muscle oxidative stress and metabolism are more limited. We tested whether a high-fat diet (HFD) would alter circulating AGE concentration, skeletal muscle AGE accumulation, and oxidative stress in wild-type and RAGE-deficient mice. The physiological significance of AGE/RAGE axis activation in HFD-fed mice was evaluated in terms of exercise tolerance and mitochondrial respiratory chain complex activity. HFD elicited adiposity, abnormal fat distribution, and oral glucose intolerance. HFD also induced accumulation of Nε-carboxymethyl-l-lysine, increased protein carbonyl levels, and impaired respiratory chain complex activity in soleus muscle. Ablation of RAGE had no effects on weight gain and oral glucose tolerance in HFD-fed mice. Peak aerobic capacity and mitochondrial cytochrome-c oxidase activity were restored in HFD-fed RAGE-/- mice. We concluded that RAGE signaling plays an important role in skeletal muscle homeostasis of mice under metabolic stress. Novelty HFD in mice induces accumulation of AGEs, oxidative stress, and mitochondrial dysfunction in the soleus muscle. RAGE, the multi-ligand receptor for AGEs, modulates oxidative stress and mitochondrial electron transport chain function in the soleus muscle of HFD-fed mice.

Inhibitors of Advanced Glycation End Product (AGE) Formation and Accumulation.

A range of chemically different compounds are known to inhibit the formation and accumulation of advanced glycation end products (AGEs) or disrupt associated signalling pathways. There is evidence that some of these agents can provide end-organ protection in chronic diseases including diabetes. Whilst this group of therapeutics are structurally and functionally different and have a range of mechanisms of action, they ultimately reduce the deleterious actions and the tissue burden of advanced glycation end products. To date it remains unclear if this is due to the reduction in tissue AGE levels per se or the modulation of downstream signal pathways. Some of these agents either stimulate antioxidant defence or reduce the formation of reactive oxygen species (ROS), modify lipid profiles and inhibit inflammation. A number of existing treatments for glucose lowering, hypertension and hyperlipidaemia are also known to reduce AGE formation as a by-product of their action. Targeted AGE formation inhibitors or AGE cross-link breakers have been developed and have shown beneficial effects in animal models of diabetic complications as well as other chronic conditions. However, only a few of these agents have progressed to clinical development. The failure of clinical translation highlights the importance of further investigation of the advanced glycation pathway, the diverse actions of agents which interfere with AGE formation, cross-linking or AGE receptor activation and their effect on the development and progression of chronic diseases including diabetic complications. Advanced glycation end products (AGEs) are (1) proteins or lipids that become glycated as a result of exposure to sugars or (2) non-proteinaceous oxidised lipids. They are implicated in ageing and the development, or worsening, of many degenerative diseases, such as diabetes, atherosclerosis, chronic kidney and Alzheimer's disease. Several antihypertensive and antidiabetic agents and statins also indirectly lower AGEs. Direct AGE inhibitors currently investigated include pyridoxamine and epalrestat, the inhibition of the formation of reactive dicarbonyls such as methylglyoxal as an important precursor of AGEs via increased activation of the detoxifying enzyme Glo-1 and inhibitors of NOX-derived ROS to reduce the AGE/RAGE signalling.

Combination of RAGE Inhibitors and Gemcitabine Impedes Tumor Growth by Reducing Autophagy and Facilitating Apoptosis in Pancreatic Cancer

Pancreatic cancer (PC) is a lethal disease with the current 5-year survival of only about 8%. Despite decades of research, gemcitabine remains the standard of therapy for PC. However, after an initial good response, eventual development of chemo-resistance is a major challenge faced by gemcitabine therapy. One principal cause of chemo-resistance is the release of high mobility group box 1 (HMGB1) protein from dying tumor cells, upon treatment with gemcitabine. When released, HMGB1 interacts with the receptor for advanced glycation end-products (RAGE), stimulating distinct signaling pathways leading to increased autophagy and tumor cell proliferation along with a reduction in apoptosis. RAGE activation by HMGB1 thereby supports tumor cell survival, making the tumor less sensitive towards gemcitabine. We hypothesized that inhibiting RAGE-HMGB1 interaction, in combination with gemcitabine therapy could reduce autophagy and tumor cell proliferation as well as facilitate apoptosis. This would ultimately reduce chemo-resistance and make pancreatic tumor more sensitive towards gemcitabine therapy. In our study, we intended to determine the efficacy of RAGE inhibitors (anti-RAGE antibody IgG2A11 and small molecule RAGE inhibitor FPS-ZM1) in combination with gemcitabine in PC. We employed an orthotopic mouse model of PC to test our hypothesis. In this model, the murine #5508 KPC pancreatic cancer cell line was implanted in the pancreas of C57BL/6 mice. The implanted mice received different treatments (saline, RAGE inhibitor, gemcitabine or gemcitabine with RAGE inhibitor). At the end of the study, tumors obtained from different treatment groups were assessed for their weight. Additionally, these tumors were analyzed for the expression of markers of autophagy, cell proliferation and apoptosis by western blot. We observed smaller tumors in mice treated with the combination of anti-RAGE antibody IgG2A11 & gemcitabine as well as lower levels of phosphorylated ERK in the combination treated group. This data was presented at “EB 2018” conference. We now show that the smaller tumors in the combination group are also associated with an increase in p62 levels, supporting a reduction in autophagy. In addition, we observed higher levels of cleaved PARP in this combination group suggesting higher levels of apoptosis in tumor cells. In our second study where we used the small molecule RAGE inhibitor FPS-ZM1 in combination with gemcitabine, we also observed significantly smaller tumors in the combination group as compared to the gemcitabine alone treatment group. In conclusion, our data strongly suggests that inhibiting RAGE in combination with gemcitabine could be a valid therapeutic approach for the treatment of pancreatic adenocarcinoma. Support or Funding Information This work was in part supported by the College of Health Professions at NDSU and by the NIH Grant Number P20GM109024 from the National Institute of General Medicine (NIGMS). This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Abstract # 3095 Anxiety, pro-inflammatory Cytokines, and RAGE-associated S100A8/A9 levels in breast cancer patients

Introduction The link between anxiety and breast cancer outcomes has recently been a topic of interest in behavioral medicine, and threat-associated distress states may be associated with increased inflammation, which is relevant for breast cancer recurrence. The purpose of this study was to examine the relationship between post-surgical anxiety levels, pro-inflammatory cytokines IL-1- β , IL-6, and TNF- α , and ligands associated with the receptor for advanced glycation end-products (RAGE). Activation of RAGE can trigger pathways that promote inflammation. To date, no studies have linked anxiety with RAGE ligands in breast cancer patients. Method Stage 0-III breast cancer patients (ages 32–69) were recruited 2–10 weeks post-surgery and interviewed with the Hamilton Rating Scale for Anxiety (HRSA) and provided blood samples before initiating adjuvant chemotherapy or radiation. Results Using linear regression, we found that after controlling for treatment condition, age, stage, body mass index (BMI), time since surgery, and type of surgery, anxiety severity related to significantly greater levels of S100A8/A9 (p = .035) and IL-1- β (p = .022), and marginally with IL-6 (p =.077) and TNF- α (p = .051). Conclusion Anxiety may relate to markers of inflammation in breast cancer patients at an early point in their treatment. Because inflammatory processes may increase odds of metastasis, addressing anxiety early in the post-surgical period may have future health effects in breast cancer patients.