Abstracts from the 20th International Symposium on Signal Transduction at the Blood-Brain Barriers

https://deepblue.lib.umich.edu/bitstream/2027.42/138963/1/12987_2017_Article_71.pd

Macrophage polarization by the microenvironment of atherosclerotic plaques

Macrophages are key cellular mediators of innate immunity: they are positionally and transcriptionally programmed to respond to pathogens and environmental challenges. When activated by inflammatory signals in their microenvironment they develop into functionally and phenotypically distinct polarized subpopulations: classically activated macrophages, M1, characterized by cytotoxix/proinflammatory activity; alternatively activated macrophages, M2, characterized by anti-inflammatory/wound repair activity. M1 pro-inflammatory macrophages drive atherosclerotic plaques progression towards instability, cap fragilization and rupture. Our study provide new informations about the role exherted by IL-23 and its receptor in human carotid atherosclerotic plaque progression. We show the presence of IL-23 immunoreactivity, mRNA and protein in macrophages infiltrating human carotid atherosclerotic plaques. Our immunohistochemical analysis demonstrated a strong IL-23 immunoreactivity within the inflammatory infiltrate at the shoulder of the plaques, and at the level of cells lining the fibrous cap. FISH analysis confirmed the expression of IL-23 detected by immunohistochemistry. Immunofluorescence, followed by FISH analysis, showed that cells positive for IL-23 mRNA bind anti-CD68 mAb, thus indicating that these cells belong to the macrophage components of the inflammatory infiltrate. This result was further confirmed by double labelling experiments. IL-23 immunoreactivity was detected within the fibrous layer and co-localized with cells belonging to the monocyte-macrophage lineage as shown by their strong CD68- and CD14-related reaction. Clusters of double-positive cells were found at the border of the plaque, as well as in the subendothelial space. Immunohistochemistry and immunofluorescence showed a strong immunoreactivity for IL-23R at the level of inflammatory mononuclear cells accumulated within the plaque. In vitro, only M1 pro-inflammatory, but not M2 anti-inflammatory macrophages produced IL-23, upon stimulation with zymosan or bacterial lipopolysaccharide. Our results suggests that a hyperactive and highly pathogenic IL-23-IL-23R system drives chronic inflammation in atherosclerosis, while the presence of IL-23 proximal to the fibrous cap may contribute to the atherosclerotic plaque instability

IS RAGE THE SO FAR UNIDENTIFIED TRAIT D'UNION BETWEEN VASCULAR RISK FACTORS AND ALZHEIMER'S DISEASE?

889 IS RAGE THE SO FAR UNIDENTIFIED TRAIT D'UNION BETWEEN VASCULAR RISK FACTORS AND ALZHEIMER'S DISEASE? Rita Businaro1, L. Capriotti1, M. Corsi1, M. Leopizzi1, V. Nicolia2, A. Fuso2 1Medico-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Latina, 2Surgery 'P. Valdoni', Sapienza University of Rome, Rome, Italy Aim: The receptor for advanced glycation end products (RAGE), is considered to be a key mediator of atherogenesis. Recent findings indicate the important role played by RAGE in the progression of Alzheimer's disease (AD). It has been shown that RAGE signaling contributes to the production of proinflammatory cytokines, leading to the impairment of neuronal functions and to the amyloid accumulation. Moreover, atherosclerosis, stroke and cardiac disease in the aging individual, could result in cerebrovascular dysfunction and trigger AD pathology. It is known that diet depleted in folate and vitamins B6 and B12 promotes an enhanced expression of RAGE and leads to hyperhomocysteinemia, a well-known risk factor for the development of cardiovascular disease as well as for late onset AD. This work aims at evaluating the effects of hyperhomocysteinemia, induced by B vitamin deficiency, on RAGE expression. Methods: TgCNRD8 mice carrying a Indiana/Swedish mutated APP transgene, an animal model of AD, were grown either with control or B vitamin deficient diet. We measured RAGE by immunohistochemistry, western blots, real-time PCR. Results: B vitamin deficiency enhances RAGE expression in particular at the level of frontal and parietal microvasculature. Both neurons and endothelium in the hippocampus were stained, showing an increase in the number of positive cells as well as in the amount of reaction, compared to the animals fed with a standard diet. Conclusions: B vitamin deficiency and hyperomocysteinemia are able to modulate RAGE expression, promoting in this way atherosclerosis progression and the transport of beta Amyloid across the BBB

Autologous leuco-platelet preparation promotes neoangiogenesis and wound healing

The rationale of our study was to test the potential of the autologous elements of peripheral white blood cells and platelets to promote neoangiogenesis, thereby overcoming hypoxemia, the major obstacle to the regeneration of the tissue. An highly innovative technique, based on an autologous leuco-platelet preparation, applied by direct infiltration to the ulcer was employed to treat 65 patients with ulcers of different etiologies. Hyaluronic acid was used as a scaffold, supplemented with autologous cells. Patients were followed up until complete healing of the lesions. Morphometric analysis was perfomed on histologic sections to determine the presence of neoangiogenesis. The wound healing was obtained in all cases after 2 until 22 months with this treatment (median time is 11.4 months). More than half of the patients recovered completely within the first 12 months. Morphometric analysis on histologic sections determined the presence of an abundant neovasculature in the close proximity of the infiltrations of leuco-platelet concentrate. Our data show that autologous leucoplatelet preparation layered onto ischemic ulcers may improve neoangiogenesis leading to wound healing. These findings suggest that this cell-based therapy may be a useful tool for the treatment for intractable skin ulcers resulting from diabetes, ischemia and collagen diseases

Modulation of inflammation as a way of delaying alzheimer’s disease progression: The diet’s role

Background: Most of the recent reports suggest that inflammatory mediators play a central role in the etiopathogenesis of Alzheimer’s disease (AD) and that the conditions leading to a chronic low-grade inflammation, such as stress, depression, obesity and metabolic syndrome, increase the odds of developing Mild Cognitive Impairment (MCI) and AD. Microglia cells are the main actors in the AD process: stimuli from the microenvironment may induce microglia cells to switch to a classically activated inflammatory phenotype M1, or, on the contrary to an alternatively activated M2 phenotype characterized by the secretion of different types of cytokines. Many attempts are currently being made in order to delay the progression of AD by reducing inflammatory mechanisms underlying the disease. Several studies support a relationship among neuroinflammation and nutrients, foods or dietary patterns, taking into account the synergistic or antagonistic biochemical interactions among nutrients as well as the different food sources of the same nutrient. Natural antioxidant and anti-inflammatory compounds found in plant foods, such as fruits, particularly berries (such as strawberry, blueberry, blackcurrant, blackberry, blueberry and mulberry) have been shown to exert neuroprotective activity. It is still unclear whether the dietary bioactive compounds enter the Blood Brain Barrier (BBB) playing a direct antiinflammatory or pro-inflammatory effect on microglia and/or other Central Nervous System (CNS) cells. Another hypothesis is that they may trigger a peripheral reaction that induce indirectly a CNS’ response. The subsequent synthesis of cytokines may drive microglia polarization by different ways. So, via an indirect route microglia detects and responds to immune-to-brain signaling. Conclusion: This review summarizes current evidence about the potential mechanisms of the interaction among diet, neuroinflammation and AD

Hunting the risk NPY and ACE Polymorphisms as predictors of cardiovascular diseases: case report and review of the literature

Many research efforts were addressed to identify individuals at high risk for multifactorial diseases, such as cardiovascular alterations and related diseases. Great interest was paid to investigate the genetic liability in multifactorial illnesses. The prognosis of high-risk patients might be greatly ameliorated using genetic predisposition risk factors, such as the polymorphisms of neuropeptide Y (NPY) and Angiotensin converting enzyme (ACE) genes. Epidemiologic results suggest that selected polymorphisms of both NPY and ACE might be helpful to improve the evaluation of patients, offering a powerful prognostic tool and paving the way to novel molecular therapeutic strategies. We present a case report of a male, sudden-death from myocardial infarction, presenting with left ventricular hypertrophy. The patient carried polymorphisms of ACE and NPY genes, respectively ACE genotype ID and NPY genotype T-399C, actually considered as risk factors

Interleukin-18 modulation in autism spectrum disorders

Background: Autism spectrum disorder (ASD) is a neurodevelopmental disease which affects 1 in 88 children. Its etiology remains basically unknown, but it is apparent that neuroinflammation is involved in disease development. Great attention has been focused on pro-inflammatory cytokines, and several studies have reported their dysfunction unbalance in serum as well as in the brain. The present work aimed at evaluating putative dysregulation of interleukin-18 (IL-18), a pro-inflammatory cytokine of the IL-1 family in the sera of patients with ASD of different grades, compared to healthy controls, as well as in postmortem brain samples obtained from patients with tuberous sclerosis as well as acute inflammatory diseases. Moreover, quantitative analysis of IL-18 was performed in the sera and brain obtained from Reeler mice, an experimental model of autism. METHODS: Serum IL-18 levels were measured by ELISA. IL-18 was localized by immunohistochemical analysis in brain sections obtained from tuberous sclerosis and encephalitis patients, as well as from gender- and age-matched controls, and in the brain sections of both Reeler and wild-type mice. IL-18 was also quantified by Western blots in homogenates of Reeler and wild-type mice brains. IL-18 binding protein (IL-18BP) was evaluated in Reeler and wild-type mice plasma as well as in their brains (sections and homogenates). RESULTS: IL-18 content decreased in the sera of patients with autism compared to healthy subjects and in Reeler sera compared to wild-type controls. IL-18 was detected within glial cells and neurons in the brain of subjects affected by tuberous sclerosis and encephalitis whereas in healthy subjects, only a weak IL-18 positivity was detected at the level of glial cells. Western blot identified higher amounts of IL-18 in Reeler brain homogenates compared to wild-type littermates. IL-18BP was expressed in higher amounts in Reeler brain compared to the brain of wild-type mice, whereas no significant difference was detected comparing IL-18BP plasma levels. CONCLUSIONS: IL-18 is dysregulated in ASD patients. Further studies seemed necessary to clarify the molecular details behind IL-18 increase in the brain and IL-18 decrease in the sera of patients. An increase in the size of the patient cohort seems necessary to ascertain whether decreased IL-18 content in the sera can become a predictive biomarker of ASD and whether its measure, in combination with other markers (e.g., increased levels of brain-derived neurotrophic factor (BDNF)), may be included in a diagnostic panel

IL-23 involvement in human carotid atherosclerosis progression

Atherosclerosis is now considered an autoimmune-inflammatory disease associated with lipoprotein metabolism alterations. Atheroma begins with the accumulations of oxidized low-density lipoprotein in the subendothelial matrix and the recruitment of monocytes which then differentiate to different functional phenotypes depending on the surrounding microenvironment. Plaque macrophages represent the majority of leukocytes in the atherosclerotic lesions, and their secretory activity, including proinflammatory cytokines and matrix-degrading proteases, is related to the progression as well as to the fragilization and rupture of the plaque (1). The balance between pro- and anti-inflammatory molecules is a major determinant of disease progression (2). In this connection an important role is played by cytokines secreted by 2 major populations of macrophages: M1 or classically activated and M2 or alternatively activated (3). Advanced plaques contain a majority of pro-inflammatory cells and processes leading to plaque rupture are probably dependent on mediators released by these cells in the close proximity of the fibrous cap. The identification of molecules involved in such a mechanism should provide new target for an effective prevention of main tromboembolic complications. Recently, among the many inflammatory cells and cytokines involved in atherosclerosis, an emerging role has been assigned to Th17 and IL-17 cytokine family. IL-23, a heterodimeric cytokine, member of IL-12 family of cytokines, represents the most important cytokine for maintaining and expanding Th17 cells and contributes to local inflammation. Interestingly, very recent meta-analysis correlated autoimmune diseases with excessive cardiovascular events (4), and previous studies have shown the presence of autoantibodies directed to autoantigens of the plaque in patients with carotid atherosclerosis (5,6). Aim of our study was to localize IL-23- and IL-23 receptor-positive cells within human carotid atherosclerotic plaques and to determine the levels of this cytokine in plaque supernatants and in sera from patients with carotid atherosclerosis. We investigated the presence of IL-23 and IL-23 receptor in human carotid atherosclerotic plaques by immunohistochemistry (30 samples), fluorescent in situ hybridization (FISH) (10 samples) and western blot (11 samples). 53% of the supernatant of cultered carotid plaques (19 samples) as well as 60% of carotid atherosclerosis patient sera (25 samples) have shown to contain discrete amounts of IL-23, as measured by enzyme-linked immunosorbent assay (ELISA). In particular our results demonstrated the presence of CD 68 and IL-23-double positive cells at the border and within the fibrous cap of complicated plaques. Moreover IL-23 receptor was detected at the surface of T lymphocytes distributed within the inflammatory infiltrate of the plaque. Of note, only human M1 pro-inflammatory macrophages obtained in vitro from monocytes induced to differentiate in the presence of granulocyte-macrophage colony stimulating factor were able to release IL-23, in response to toll-like receptor 2 and 4 agonists. In addition 53% of the supernatant of cultured carotid plaques (19 samples) as well as 60% of patient sera (25 samples) contained detectable amounts of IL-23, as measured by ELISA. All these findings suggest a relevant role for IL-23 in the inflammatory processes of atherosclerosis, identifying a new potential therapeutic target for immune modulation of atherosclerosis. 1) Businaro R. J Neuroimmune Pharmacol. 2013;8:15-27. 2) Businaro R. et al. Ann N Y Acad Sci. 2012;1262:134-41. 3) Taghavie-Moghadam et al. Ann N Y Acad Sci. 2014;1319:19-37. 4) Amaya-Amaya et al.Biomed Res Int. 2014;2014:367359. 5) Riganò R. et al. Ann N Y Acad Sci. 2007;1107:1-10 6) Businaro R. et al Atherosclerosis. 2009;207:74-8