Faculty Bibliography

INTRODUCTION AND AIMS/OBJECTIVE:Oxytocin (OT) is a neuropeptide hormone secreted by the posterior pituitary gland. Deficits in OT action have been observed in patients with behavioral and mood disorders, some of which correlate with an increased risk of cardiovascular disease (CVD). Recent research has revealed a wider systemic role that OT plays in inflammatory modulation and development of atherosclerotic plaques. This study investigated the role that OT plays in cholesterol transport and foam cell formation in LPS-stimulated THP-1 human macrophages. METHODS:THP-1 differentiated macrophages were treated with media, LPS (100 ng/ml), LPS + OT (10 pM), or LPS + OT (100 pM). Changes in gene expression and protein levels of cholesterol transporters were analyzed by real time quantitative PCR (RT-qPCR) and Western blot, while ox-LDL uptake and cholesterol efflux capacity were evaluated with fluorometric assays. RESULTS:RT-qPCR analysis revealed a significant increase in ABCG1 gene expression upon OT + LPS treatment, compared to LPS alone (p = 0.0081), with Western blotting supporting the increase in expression of the ABCG1 protein. Analysis of ox-LDL uptake showed a significantly lower fluorescent value in LPS + OT (100pM) -treated cells when compared to LPS alone (p < 0.0001). While not statistically significant (p = 0.06), cholesterol efflux capacity increased with LPS + OT treatment. CONCLUSION/CONCLUSIONS:We demonstrate here that OT can attenuate LPS-mediated lipid accumulation in THP-1 macrophages. These findings support the hypothesis that OT could be used to reduce pro-inflammatory and potentially atherogenic changes observed in patients with heightened CVD risk. This study suggests further exploration of OT effects on monocyte and macrophage cholesterol handling in vivo.

Animal studies have shown that the gut microbiome can influence memory, social behavior, and anxiety-like behavior. Several human studies show similar results where variation in the gut microbiome is associated with dementia, depression, and personality traits, though most of these studies are limited by small sample size and other biases. Here, we analyzed fecal samples from 313 participants in the Wisconsin Longitudinal Study, a randomly selected population-based cohort of older adults, with measured psycho-cognitive dimensions (cognition, mood, and personality) and key confounders. 16s V4 sequencing showed that Megamonas is associated with all measured psycho-cognitive traits, Fusobacterium is associated with cognitive and personality traits, Pseudoramibacter_Eubacterium is associated with mood and personality traits, Butyvibrio is associated with cognitive traits, and Cloacibacillus is associated with mood traits. These findings are robust to sensitivity analyses and provide novel evidence of shared relationships between the gut microbiome and multiple psycho-cognitive traits in older adults, confirming some of the animal literature, while also providing new insights. While we addressed some of the weaknesses in prior studies, further studies are necessary to elucidate temporal and causal relationships between the gut microbiome and multiple psycho-cognitive traits in well-phenotyped, randomly-selected population-based samples.

PURPOSE/OBJECTIVE:Cardiovascular disease (CVD) is the leading cause of death, globally, and its prevalence is only expected to rise due to the increasing incidence of co-morbidities such as obesity and diabetes. Medical treatment of CVD is directed primarily at slowing or reversing the underlying atherosclerotic process by managing circulating lipids with an emphasis on control of low-density lipoprotein (LDL) cholesterol. However, over the past several decades, there has been increasing recognition that chronic inflammation and immune system activation are important contributors to atherosclerosis. This shift in focus has led to the elucidation of the complex interplay between cholesterol and cellular secretion of cytokines involved in CVD pathogenesis. Of the vast array of cytokine promoting atherosclerosis, interferon (IFN)-γ is highly implicated and, therefore, of great interest. METHODS:Literature review was performed to further understand the effect of IFN-γ on the development of atherosclerotic CVD. RESULTS:IFN-γ, the sole member of the type II IFN family, is produced by T cells and macrophages, and has been found to induce production of other cytokines and to have multiple effects on all stages of atherogenesis. IFN-γ activates a variety of signaling pathways, most commonly the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway, to induce oxidative stress, promote foam cell accumulation, stimulate smooth muscle cell proliferation and migration into the arterial intima, enhance platelet-derived growth factor expression, and destabilize plaque. These are just a few of the contributions of IFN-γ to the initiation and progression of atherosclerotic CVD. CONCLUSION/CONCLUSIONS:Given the pivotal role of IFN-γ in the advancement of CVD, activation of its signaling pathways is being explored as a driver of atherosclerosis. Manipulation of this key cytokine may lead to novel therapeutic avenues for CVD prevention and treatment. A number of therapies are being explored with IFN-γ as the potential target.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive disease with high mortality that commonly occurs in middle-aged and older adults. IPF, characterized by a decline in lung function, often manifests as exertional dyspnea and cough. Symptoms result from a fibrotic process driven by alveolar epithelial cells that leads to increased migration, proliferation, and differentiation of lung fibroblasts. Ultimately, the differentiation of fibroblasts into myofibroblasts, which synthesize excessive amounts of extracellular matrix proteins, destroys the lung architecture. However, the factors that induce the fibrotic process are unclear. Diagnosis can be a difficult process; the gold standard for diagnosis is the multidisciplinary conference. Practical biomarkers are needed to improve diagnostic and prognostic accuracy. High-resolution computed tomography typically shows interstitial pneumonia with basal and peripheral honeycombing. Gas exchange and diffusion capacity are impaired. Treatments are limited, although the anti-fibrotic drugs pirfenidone and nintedanib can slow the progression of the disease. Lung transplantation is often contraindicated because of age and comorbidities, but it improves survival when successful. The incidence and prevalence of IPF has been increasing and there is an urgent need for improved therapies. This review covers the detailed cellular and molecular mechanisms underlying IPF progression as well as current treatments and cutting-edge research into new therapeutic targets.

Purpose of Study Autism spectrum disorders (ASDs) are neurodevelopmental disorders with lifelong consequences and poorly understood pathophysiology. Dysregulated cholesterol metabolism is implicated in ASD etiology. Cholesterol is essential for neuroactive steroid production, myelin sheath formation, and normal brain development. Early postnatal or in utero exposure to the antiepileptic drug valproic acid (VPA), a branched short-chain fatty acid, causes autism-like neural and behavioral deficits in humans and rodents. This study examines the link between VPA and cholesterol deficit in cultured human neurons and microglia. Methods Used SHSY-5Y human neuroblastoma cells and HMC3 human microglial cells were exposed to VPA at 0, 250, 1000 and 5000 muM for 24h, N=3 per condition. Expression of critical genes that regulate cholesterol transport were quantified by RT-PCR using specific primers for each. These include the efflux proteins ABCA1, ABCG1, 27-hydroxylase (27-OHase) and 24-hydroxylase (24-OHase), and the influx scavenger receptor CD36 - all vital for brain cholesterol balance. Expression of these target genes was normalized to concurrently measured GAPDH mRNA levels. Summary of Results In SH-SY5Y neurons, VPA exposure caused a concentration-dependent increase in ABCA1 (P <0.001), ABCG1, 27-OHase (P <0.001) (figure 1), and CD36 (P=0.015). In HMC3, VPA exposure caused a concentration- dependent increase in ABCG1 (80-fold at highest dose, P<=0.001) and 24-OHase (P < 0.001) with a reduction in ABCA-1 (P=0.002) and an increase in CD36 (P<0.001). Conclusions This study shows that VPA has a dramatic hypocholesterolemic effect on two key cell types that compose the developing brain. The net impact of the changes observed in these cholesterol-related genes would be outflow and metabolism. Further, enhanced 27-OHase activity produces an oxysterol metabolite with neurotoxic effects that include downregulating synaptic proteins and decreasing neurite number and length. Together, our results suggest that VPA impairs brain cholesterol homeostasis. A better understanding of the involvement of cholesterol in the mechanisms by which VPA leads to ASDs may translate into novel preventative therapies for this serious disorder

The idea that trillions of bacteria inhabit our gut is somewhat unnerving, yet these bacteria may have a greater influence on our behavior than previously thought. Accumulating data strongly suggest that these gut commensal organisms have a strong inter-relationship with our brain and behavior, including cognitive function, mood, and personality. In this chapter, we discuss the role of the gut microbiome in the development of human personality, mood and mood disorders, and cognition, with a particular emphasis on the current consensus and controversies in the literature surrounding the behavioral effects of bioactive metabolites, microbial ratio shifts, and neurotransmitter synthesis facilitated by the microbiome.

Adenosine is an endogenous nucleoside with a short half-life that regulates many physiological functions involving the heart and cardiovascular system. Among the cardioprotective properties of adenosine are its ability to improve cholesterol homeostasis, impact platelet aggregation and inhibit the inflammatory response. Through modulation of forward and reverse cholesterol transport pathways, adenosine can improve cholesterol balance and thereby protect macrophages from lipid overload and foam cell transformation. The function of adenosine is controlled through four G-protein coupled receptors: A₁, A_2A, A_2B and A₃. Of these four, it is the A_2A receptor that is in a large part responsible for the anti-inflammatory effects of adenosine as well as defense against excess cholesterol accumulation. A_2A receptor agonists are the focus of efforts by the pharmaceutical industry to develop new cardiovascular therapies, and pharmacological actions of the atheroprotective and anti-inflammatory drug methotrexate are mediated via release of adenosine and activation of the A_2A receptor. Also relevant are anti-platelet agents that decrease platelet activation and adhesion and reduce thrombotic occlusion of atherosclerotic arteries by antagonizing adenosine diphosphate-mediated effects on the P2Y12 receptor. The purpose of this review is to discuss the effects of adenosine on cell types found in the arterial wall that are involved in atherosclerosis, to describe use of adenosine and its receptor ligands to limit excess cholesterol accumulation and to explore clinically applied anti-platelet effects. Its impact on electrophysiology and use as a clinical treatment for myocardial preservation during infarct will also be covered. Results of cell culture studies, animal experiments and human clinical trials are presented. Finally, we highlight future directions of research in the application of adenosine as an approach to improving outcomes in persons with cardiovascular disease.

OBJECTIVE:Mycophenolate (MPA) and cyclosporin A (CsA) are two immunosuppressive agents currently used for the treatment of autoimmune diseases. However, reports regarding their effects on inflammation and lipid handling are controversial. Here, we compare the effect of these two drugs on the expression of proteins involved in cholesterol handling and lipid accumulation in a macrophage cell system utilizing M0, M1 and M2 human macrophages and in murine bone marrow-derived macrophages (BMDM). METHODS:. RESULTS:In M0 macrophages, MPA increased expression of ABCA1 and CXCL16 in a concentration-dependent manner. In M1 THP-1 macrophages, MPA caused a significant increase of 27-hydroxylase mRNA and CD36 and SR-A1 receptor mRNAs. Exposure of M2 macrophages to MPA also stimulated expression of 27-hydroxylase, while downregulating all evaluated scavenger receptors. In contrast, CsA had no impact on cholesterol efflux in M0 and M1 macrophages, but significantly augmented expression of ABCA1 and 27-hydroxylase in M2 macrophages. CsA significantly increased expression of the LOX1 receptor in naïve macrophages, downregulated expression of CD36 and SR-A1 in the M1 subpopulation and upregulated expression of all evaluated scavenger receptors. However, CsA enhanced foam cell transformation in M0 and M2 macrophages, while MPA had no effect on foam cell formation unless used at a high concentration in the M2 subtype. CONCLUSIONS:Our results clearly underline the importance of further evaluation of the effects of these drugs when used in atherosclerosis-prone patients with autoimmune or renal disease.

Background andObjectives: Atherosclerotic cardiovascular disease (CVD) remains a major cause of morbidity and mortality in persons with systemic lupus erythematosus (SLE, lupus). Atherosclerosis, which involves interplay between cholesterol metabolism and cellular inflammatory pathways, is primarily treated with statins since statins have lipid-lowering and anti-inflammatory properties. The Lupus Atherosclerosis Prevention Study (LAPS) was designed to investigate the efficacy of statins against CVD in SLE patients. LAPS demonstrated that 2 years of atorvastatin administration did not reduce atherosclerosis progression in lupus patients. In this LAPs substudy, we use cultured macrophages to explore the atherogenic properties of plasma from LAPS subjects to explain the mechanistic rationale for the inability of statins to reduce CVD in lupus. Materials and Methods: THP-1 differentiated macrophages were treated for 18 h with 10% SLE patient plasma obtained pre- and post-atorvastatin therapy or placebo. Gene expression of the following cholesterol transport genes was measured by qRT-PCR. For efflux-ATP binding cassette transporter (ABC)A1 and ABCG1, 27-hydroxylase, peroxisome proliferator-activated receptor (PPAR)γ, and liver X receptor (LXR)α; and for influx-cluster of differentiation 36 (CD36) and scavenger receptor (ScR)A1. Results: Macrophages exposed to plasma from both statin-treated and placebo-treated groups showed a significant decrease in cholesterol efflux proteins ATP binding cassette (ABC) transporters A1 and ABCG1, an increase in 27-hydroxylase, an increase in the LDL receptor and a decrease in intracellular free cholesterol. No change in influx receptors ScRA1 and CD36, nor nuclear proteins LXRα and PPARγ was observed. Conclusions: Statins do not normalize pro-atherogenic changes induced by lupus and these changes continue to worsen over time. This study provides mechanistic insight into LAPS findings by demonstrating that statins are overall ineffective in altering the balance of cholesterol transport gene expression in human macrophages. Furthermore, our study suggests that statins as a CVD treatment may not be useful in attenuating lipid overload in the SLE environment.