Faculty Bibliography

Objectives: Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disorder that is characterized by selective degeneration of both upper and lower motor neurons, resulting in paralysis of skeletal muscle and respiratory failure, with death occurring within 2-5 years of diagnosis. 90% of cases are sporadic, and of the 10% that are familial more than 20 genes (>150 mutations) have been found to be associated with ALS, most notably copper/zinc superoxide dismutase (SOD1). SOD1 mutant proteins are believed to cause toxicities in degenerating neurons. Studies suggest that the neuronal and non-neuronal cell contributions to the onset and progression of ALS are complex. It was proposed that there are two phases of neuroinflammation in the spinal cord - the first being an early neuroprotective phase followed by a second late neurotoxic phase. One of the challenges in the study of neuroinflammation is that it is difficult to serially track the disease process, as there are no bona fide biomarkers for onset and progression in ALS. For this reason, we use PET with [¹¹C]PBR28 to track microglial neuroinflammation in the brain and spinal cord. We and others have shown that the receptor for advanced glycation end products (RAGE) is highly expressed in human ALS spinal cord, particularly in microglia, and to an increased degree compared to that of age-matched control subjects. Our previous studies with myeloid/microglia deletion of Ager and treatment with sRAGE (soluble RAGE) suggested that RAGE impairs survival and motor function in Sod1G93Amice. The ultimate goal is to test the hypothesis that RAGE inhibition in either initiation or progression phases of the ALS will prolong survival and maintain motor function in adult Sod1G93Amice.
Method(s): MicroPET/CT (Inveon, Siemens) with [¹¹C]PBR28 was used to track and compare microglial neuroinflammation in the brain and spinal cord of WT vs. ALS mice (110-120 day old), also after treatment with RAGE inhibitors (subject identity was blind to study investigator and data analyst). Using IRW (Inveon Research Workplace, Siemens), several ROIs in the thoracic and lumbar spinal cord (T13, L1, L2, L3) were drawn on the fused PET/CT images to obtain the regional SUVs. An automated atlasbased methodology using Firevoxel (https://urldefense.proofpoint.com/v2/url?u=https-3A__wp.nyu.edu_Firevoxel&d=DwIBAg&c=j5oPpO0eBH1iio48DtsedeElZfc04rx3ExJHeIIZuCs&r=CY_mkeBghQnUPnp2mckgsNSbUXISJaiBQUhM-Uz9W58&m=_uGsTvUTTD_GxqvwK245ZUiiSbzVraIboytFijFDOwU&s=RlC-AQtmqr84rzBwvDmgK_FCVdvbCfsFvuN-dVODTpM&e= ) that we previously developed was used for brain mapping and segmentation to derive regional timeactivity curves (TAC) and SUVs for 20 brain regions.
Result(s): Dynamic regional SUV [¹¹C]PBR28 binding data were obtained and averaged SUVs derived from the last 5 frames (with steady and less variable intensity levels) were compared. Results derived from both spinal cord and brain regions displayed a similar trend with two obvious clusters. Reduced binding was observed for ALS group as compared to WT. RAGE inhibitor-treated ALS mi ce showed increased binding (brain SUV avg. 0.402+/-0.0382 over 20 ROIs) as compared to vehicle-treated (0.157+/-0.0339), suggesting that RAGE inhibition may contribute to the restoration of homeostasis in ALS animals (i.e., their bindings after treatment were closer to those in WT (0.485+/-0.171)). Notably, hypothalamus, brain stem, and olfactory bulb consistently exhibited higher binding, suggesting their role in this regulation.
Conclusion(s): Inconsistent outcomes have been reported in the literature when comparing TSPO ligand binding for imaging neuroinflammation. Our data are consistent with findings from several recent studies; i.e., reduced PBR28 binding was associated with disease state (e.g., in patients with PTSD or alcoholism). A notion that the reduced binding might reflect competition from endogenous TSPO ligands such as cholesterol can't be excluded. The strategies described here will test the hypothesis that pharmacological antagonism of RAGE signal transduction in either initiation or progression phases of the ALS will prolong survival and maintain motor function in adult Sod1G93Amice

We addressed the involvement of the receptor for advanced glycation end products (RAGE) in the impairment of the cellular cholesterol efflux elicited by glycated albumin. Albumin was isolated from type 1 (DM1) and type 2 (DM2) diabetes mellitus (HbA1c > 9%) and non-DM subjects (C). Moreover, albumin was glycated in vitro (AGE-albumin). Macrophages from Ager null and wild-type (WT) mice, or THP-1 transfected with siRNA-AGER, were treated with C, DM1, DM2, non-glycated or AGE-albumin. The cholesterol efflux was reduced in WT cells exposed to DM1 or DM2 albumin as compared to C, and the intracellular lipid content was increased. These events were not observed in Ager null cells, in which the cholesterol efflux and lipid staining were, respectively, higher and lower when compared to WT cells. In WT, Ager, Nox4 and Nfkb1, mRNA increased and Scd1 and Abcg1 diminished after treatment with DM1 and DM2 albumin. In Ager null cells treated with DM-albumin, Nox4, Scd1 and Nfkb1 were reduced and Jak2 and Abcg1 increased. In AGER-silenced THP-1, NOX4 and SCD1 mRNA were reduced and JAK2 and ABCG1 were increased even after treatment with AGE or DM-albumin. RAGE mediates the deleterious effects of AGE-albumin in macrophage cholesterol efflux.

BACKGROUND: Chronic kidney disease (CKD) is associated with a high cardiovascular mortality due to increased rates of vascular lesions and thrombotic events, as well as serum accumulation of uremic toxins. A subgroup of these toxins (advanced glycation end products [AGEs] and S100 proteins) can interact with the receptor for AGEs (RAGE). In this study, we analyzed the impact of CKD on platelet function and arterial thrombosis, and the potential role of RAGE in this process. METHODS:mice). RESULTS: < 0.0001). CONCLUSION/CONCLUSIONS: Our results show that CKD induces platelet hyperactivation, accelerates thrombus formation in a murine model of arterial thrombosis, and that RAGE deletion has a protective role. We propose that RAGE ligands binding to RAGE is involved in CKD-induced arterial thrombosis.

Despite advances in lipid-lowering therapies, people with diabetes continue to experience more limited cardiovascular benefits. In diabetes, hyperglycemia sustains inflammation and preempts vascular repair. We tested the hypothesis that the receptor for advanced glycation end-products (RAGE) contributes to these maladaptive processes. We report that transplantation of aortic arches from diabetic, Western diet-fed Ldlr-/- mice into diabetic Ager-/- (Ager, the gene encoding RAGE) versus WT diabetic recipient mice accelerated regression of atherosclerosis. RNA-sequencing experiments traced RAGE-dependent mechanisms principally to the recipient macrophages and linked RAGE to interferon signaling. Specifically, deletion of Ager in the regressing diabetic plaques downregulated interferon regulatory factor 7 (Irf7) in macrophages. Immunohistochemistry studies colocalized IRF7 and macrophages in both murine and human atherosclerotic plaques. In bone marrow-derived macrophages (BMDMs), RAGE ligands upregulated expression of Irf7, and in BMDMs immersed in a cholesterol-rich environment, knockdown of Irf7 triggered a switch from pro- to antiinflammatory gene expression and regulated a host of genes linked to cholesterol efflux and homeostasis. Collectively, this work adds a new dimension to the immunometabolic sphere of perturbations that impair regression of established diabetic atherosclerosis and suggests that targeting RAGE and IRF7 may facilitate vascular repair in diabetes.

Adipose tissue (AT) plays a central role in both metabolic health and pathophysiology. Its expansion in obesity results in increased mortality and morbidity, with contributions to cardiovascular disease, diabetes mellitus, fatty liver disease, and cancer. Obesity prevalence is at an all-time high and is projected to be 50% in the United States by 2030. AT is home to a large variety of immune cells, which are critical to maintain normal tissue functions. For example, γδ T cells are fundamental for AT innervation and thermogenesis, and macrophages are required for recycling of lipids released by adipocytes. The expansion of visceral white AT promotes dysregulation of its immune cell composition and likely promotes low-grade chronic inflammation, which has been proposed to be the underlying cause for the complications of obesity. Interestingly, weight loss after obesity alters the AT immune compartment, which may account for the decreased risk of developing these complications. Recent technological advancements that allow molecular investigation on a single-cell level have led to the discovery of previously unappreciated heterogeneity in many organs and tissues. In this review, we will explore the heterogeneity of immune cells within the visceral white AT and their contributions to homeostasis and pathology.

The escalating problem of obesity and its multiple metabolic and cardiovascular complications threatens the health and longevity of humans throughout the world. The cause of obesity and one of its chief complications, insulin resistance, involves the participation of multiple distinct organs and cell types. From the brain to the periphery, cell-intrinsic and intercellular networks converge to stimulate and propagate increases in body mass and adiposity, as well as disturbances of insulin sensitivity. This review focuses on the roles of the cadre of innate immune cells, both those that are resident in metabolic organs and those that are recruited into these organs in response to cues elicited by stressors such as overnutrition and reduced physical activity. Beyond the typical cast of innate immune characters invoked in the mechanisms of metabolic perturbation in these settings, such as neutrophils and monocytes/macrophages, these actors are joined by bone marrow-derived cells, such as eosinophils and mast cells and the intriguing innate lymphoid cells, which are present in the circulation and in metabolic organ depots. Upon high-fat feeding or reduced physical activity, phenotypic modulation of the cast of plastic innate immune cells ensues, leading to the production of mediators that affect inflammation, lipid handling, and metabolic signaling. Furthermore, their consequent interactions with adaptive immune cells, including myriad T-cell and B-cell subsets, compound these complexities. Notably, many of these innate immune cell-elicited signals in overnutrition may be modulated by weight loss, such as that induced by bariatric surgery. Recently, exciting insights into the biology and pathobiology of these cell type-specific niches are being uncovered by state-of-the-art techniques such as single-cell RNA-sequencing. This review considers the evolution of this field of research on innate immunity in obesity and metabolic perturbation, as well as future directions.

Pulmonary disease after World Trade Center particulate matter(WTC-PM) exposure is associated with dyslipidemia and the receptor for advanced glycation end products (RAGE); however, the mechanisms are not well understood. We utilized a murine model and a multiOMIC assessment to understand the role of RAGE in the pulmonary long-term effects of a single high intensity exposure to WTC-PM. After 1-month(1-M), WTC-PM exposed wild-type(WT) mice had airway hyperreactivity(AHR) while RAGE-deficient(Ager-/-) were protected. PM-exposed WT mice also had histologic evidence of airspace disease while Ager-/- remained unchanged. Inflammatory mediators such as G-CSF, IP-10, and KC were differentially expressed after WTC-PM exposure. WTC-PM induced α-SMA, DIAPH1, RAGE and significant lung collagen deposition in WT compared to Ager-/-. Compared to WT with PM exposure, relative expression of phosphorylated to total CREB and JNK were significantly increased in the lung of PM-exposed Ager-/-, whereas Akt was decreased. Random forests of the refined lung metabolomic profile classified subjects with 92% accuracy; principal components analysis captured 86.7% of the variance in 3 components and demonstrated prominent sub-pathway involvement including known mediators of lung disease such as vitamin B6 metabolites, sphingolipids, fatty acids, and phosphatidylcholines. Treatment with a partial RAGE antagonist, pioglitazone, yielded similar fold-change expression of metabolites(N6-carboxymethyllysine, 1-methylnicotinamide, (N(1)+N(8))-acetylspermidine and Succinylcarnitine(C4-DC)) between WT and Ager-/- exposed to WTC-PM. RAGE can mediate WTC-PM-induced AHR, and warrants further investigation.

Obesity and diabetes are leading causes of cardiovascular morbidity and mortality. Although extensive strides have been made in the treatments for non-diabetic atherosclerosis and its complications, for patients with diabetes, these therapies provide less benefit for protection from cardiovascular disease (CVD). These considerations spur the concept that diabetes-specific, disease-modifying therapies are essential to identify, especially as the epidemics of obesity and diabetes continue to expand. Hence, as hyperglycemia is a defining feature of diabetes, it is logical to probe the impact of the specific consequences of hyperglycemia on the vessel wall, immune cell perturbation, and endothelial dysfunction-all harbingers to the development of CVD. In this context, high levels of blood glucose stimulate the formation of the irreversible advanced glycation end products, the products of non-enzymatic glycation and oxidation of proteins and lipids. AGEs accumulate in diabetic circulation and tissues and the interaction of AGEs with their chief cellular receptor, receptor for AGE or RAGE, contributes to vascular and immune cell perturbation. The cytoplasmic domain of RAGE lacks endogenous kinase activity; the discovery that this intracellular domain of RAGE binds to the formin, DIAPH1, and that DIAPH1 is essential for RAGE ligand-mediated signal transduction, identifies the specific cellular means by which RAGE functions and highlights a new target for therapeutic interruption of RAGE signaling. In human subjects, prominent signals for RAGE activity include the presence and levels of two forms of soluble RAGE, sRAGE, and endogenous secretory (es) RAGE. Further, genetic studies have revealed single nucleotide polymorphisms (SNPs) of the AGER gene (AGER is the gene encoding RAGE) and DIAPH1, which display associations with CVD. This Review presents current knowledge regarding the roles for RAGE and DIAPH1 in the causes and consequences of diabetes, from obesity to CVD. Studies both from human subjects and animal models are presented to highlight the breadth of evidence linking RAGE and DIAPH1 to the cardiovascular consequences of these metabolic disorders.