Proteomics Profiling of Heterozygous and Homozygous Patients with ABCA1 Gene Mutation: A Tangier Disease Molecular Map

Case Report

A Proteomics. 2015;2(1): 1007.

Proteomics Profiling of Heterozygous and Homozygous Patients with ABCA1 Gene Mutation: A Tangier Disease Molecular

Ucciferri N1, Rocchiccioli S1*, Puntoni M1,2 Bigazzi F2, Cecchettini A1,3, Citti L1 and Sampietro T2

1Institute of Clinical Physiology-CNR Pisa, Italy

2Fondazione Toscana Gabriele Monasterio Pisa, Italy

3Department of Clinical and Experimental Medicine, University of Pisa, Italy

*Corresponding author: Rocchiccioli S, Institute of Clinical Physiology-CNR, Via Moruzzi 1, 56124 Pisa, Italy

Received: November 18, 2014; Accepted: February 04, 2015; Published: February 10, 2015

Abstract

Tangier Disease (TD) is a rare inherited disorder with approximately 100 worldwide identified cases. Alpha-lipoprotein deficiency is the main characteristic of this disease, associated with a virtual absence of High Density Lipoproteins (HDL) in blood. Additional symptoms are mild hypertriglyceridemia, neuropathy and enlarged, orange-colored tonsils. Genetically TD is caused by mutations in the ABCA1 gene, which prevent the release of cholesterol and phospholipids from cells, leading to the accumulation of lipids within cells and body tissues.

In this work a TD patient and his parental heterozygous were examined from a proteomics point of view. Plasma as well as proteome and secretome of circulating monocytes were analyzed.

Plasma proteins underlined in TD the imbalance of lipid trafficking and metabolism, associated with the stimulation of pro-inflammatory pathways. Proteome and secretome of monocytes highlighted an extensive down regulation of mitochondrial enzymes and vesicular trafficking agents along with a substantial cytoskeletal rearrangement, suggesting a reduced activation state of monocytes from TD homozygous patient.

This work is the first proteomics profiling of heterozygous and homozygous TD phenotypes and it suggests a TD case as a model to understand general mechanisms of lipid transport and metabolism and their linkage to inflammatory processes.

Keywords: Tangier disease; Protemics profiling; Monocyte proteomics; Rare disease

Case Presentation

Tangier Disease (TD) is an autosomal recessive genetic disorder, described for the first time in 1961 [1] and characterized by impaired HDL-mediated cholesterol efflux and abnormal intracellular lipid trafficking and turnover. TD patients accumulate cholesterol in body tissues; show a reduced level of HDL, disturbances of nerve functions, premature atherosclerosis and a high incidence of Coronary Artery Disease (CAD). TD is a rare genetic disorder, with less than 100 cases reported in the literature, and patients present both alleles of ABCA1 gene mutated; this gene encodes a member of the ATPBinding Cassette (ABC) transporter family [2, 3]. Carriers of a single ABCA1 mutation (heterozygotes) display an intermediate phenotype with a 50% reduction in the ABCA1-mediated cell cholesterol efflux [4]. Despite the commitment of a unique documented gene, TD is characterized by high variability in phenotypic manifestations, either qualitatively or quantitatively, in terms of clinical severity and organ involvement. In brief there is not a direct correlation between the gene mutation and the numerous and various clinical features described in TD patients.

ABCA1 is the major responsible transporter for clearing cholesterol from macrophages and, since cholesterol accumulation in arterial macrophages is atherogenic, this pathway has a clear involvement in the progression and/or regression of cardiovascular diseases [5]. Due to its central role in the modulation of cholesterol homeostasis, ABCA1 is an attractive target for drug development, but the molecular actors of the ABCA1 lipid pathway are not completely revealed. Studies are needed to understand how lipids are translocated across the plasma membrane and to identify associated proteins that modulate this pathway. Proteomics studies would help to disclose intracellular and secreted factors involved in dysfunction of lipid trafficking and in its clinical manifestation suggesting new targets for therapeutic interventions that might modulate ABCA1 activity assisting the traffic of cholesterol from cells and tissues.

In this paper, proteomics analyses of monocytes and plasma of a TD patient compared with his heterozygous father are reported. The clinical characterization, proband’s kindred and plasma lipoprotein profiles of these two cases have been already published in Sampietro et al. [6] and in Puntoni et al. [7] (a brief description is also presented in the Supplementary Information).

In brief, molecular factors and activation pathways directly responsible of the pathological phenotype are highlighted and the TD case has been suggested as model to obtain insights in possible mechanisms responsible for lipid dysfunctions and eventually, as a consequence, for atherosclerosis initiation.

Proteomics Profiling

TD homozygous proteomics profile was compared with his parental heterozygous (for the ABCA1 mutation). This choice was suggested by the fact that genetic background is akin and both patients are subjected to the same clinical treatments for cardiovascular disease, even if the heterozygous is mildly affected. In this way it was possible to approximate differences in the proteomics profiles due mostly to the different expression of the pathology so as to extrapolate factors involved in dyslipidemia as a main cause of early onset of atherosclerosis.

Using a shot-gun proteomics strategy (for details see Supplementary Information), we were able to identify 197 proteins in depleted plasma samples of both patients (Table S1).

46 plasma proteins (Table 1) were found differentially expressed between TD patient and his heterozygous father. In particular 22 proteins resulted down-regulated and 24 up-regulated in homozygous with respect to heterozygous. Among the downregulated in the homozygous, 13 different apolipoproteins were particularly interesting. In addition, also Paraoxonase 1 has been found down-regulated in homozygous patient plasma. This enzyme has been suggested to be involved in the protection against oxidative modification of lipoproteins and consequently against pivotal events leading to atheroma formation [8,9].These results, although preliminary, confirm that plasma proteomics can evidence the imbalance of lipid trafficking and metabolism in TD, suggesting a distinctive characteristic fingerprint, provided with a panel of correlated elements. Among the differentially expressed proteins some are linked to inflammation, such as Orosomucoid 1 and 2 and Kininogen 1. Kininogen 1 has been found over expressed in plasma of homozygous patient and it seems to be a key mediator of inflammation [10]. On the other hand, Orosomucoid 1 and 2 resulted down-expressed in homozygous plasma and, besides being involved in pro-inflammatory responses [11,12], they also exert a possible role as lipid carrier in blood circulation [13,14]. Moreover and of note, 4 differentially expressed proteins are related to different growth factor pathways: Vasorin may act as inhibitor of TGFbeta signaling [15], Insulin like Growth Factor Binding Protein 4 (IGFBP4) and Insulin-like Growth Factor-Binding Protein Complex Acid Labile Subunits (IGFALS) are key elements of the IGF pathway and Hepatocyte Growth Factor Activator (HGFA) that seems to play role in inflammatory processes [16]. In conclusion, plasma proteome of TD patient proves a general dysregulation of lipid trafficking and metabolism, associated with the stimulation of growth factor and inflammatory pathways.