Fabry disease peripheral blood immune cells release inflammatory cytokines: Role of globotriaosylceramide
Highlights
► PBMC from Fabry patients present a higher proinflammatory cytokine profile. ► Dendritic cells and monocytes from Fabry patients secrete proinflammatory cytokines. ► Gb3 is the responsible of this proinflammatory profile, likely mediated by TLR4.
Introduction
Fabry disease is an X-linked lysosomal disorder (LD) due to deficiency of the enzyme α-galactosidase A (αGal, EC 3.2.1.22, codified by the gene GLA) [1], [2]. Absent or reduced enzyme activity results in impaired catabolism of neutral glycosphingolipids, particularly globotriaosylceramide (Gb3), which in turn leads to intracellular deposition of such lipids [3]. Although tissue and organ dysfunctions can be partly attributed to direct buildup of Gb3, the exact molecular mechanisms that link this accumulation to the eventual cell and tissue damage have not yet been clearly established. In a number of different LDs, other concurrent pathological mechanisms are elicited, which together contribute to the phenotypic expression of each disease [4].
Growing evidence shows immune system irregularities associated to lysosomal disorders [5]. In several different LDs, immune cells from affected patients display a constitutive proinflammatory pattern of cytokine expression [6], [7], [8]. For Fabry disease in particular, a number of immune alterations have been described. Leucocytes and endothelial cells have been shown to exhibit an activated profile. Deposition of globotriaosylceramide occurs in numerous cell types including vascular endothelial and smooth muscle cells. Fabry disease is associated to vascular injury and a prothrombotic state, that could be related to the finding of higher surface expression of integrins involved in cell adhesion [9]. The evidence for endothelium activation was confirmed lately by Shen et al. [10]. These authors showed that accumulated Gb3 induces oxidative stress and up-regulates adhesion molecules. The mechanisms of endothelial compromise may be associated to the decreased nitric oxide bioavailability detected in a Gla null mouse model, which could account for aberrant vascular phenotypes [11]. A pro-oxidative and proinflammatory state that correlates with urinary Gb3 levels has been recently described [12]. Our group has previously reported alterations in the number of leukocyte subpopulations, as well as in their surface marker levels [13]. Later on, we have described a higher apoptotic rate in peripheral blood mononuclear cells (PBMC), which could be directly attributed to the effects of elevated Gb3 levels [14].
In this work, we investigate the existence of an underlying proinflammatory state in the PBMC of Fabry patients, in particular in the subsets related to the innate immune system, and the possible effect of excessive Gb3 levels on the initiation and maintenance of this state.
Section snippets
Patient's and normal control's samples
Blood samples from 29 Fabry patients, 15 males and 14 females (mean age: 30.2 ± 18.4 years old; range: 3–74 years old) were taken for this study. Diagnosis of Fabry disease was established by clinical examination reduced enzymatic activity and genetic test. Clinical manifestations of the patients included angiokeratoma, acroparesthesia, cornea verticillata, abdominal pain, proteinuria and hypohidrosis; however none of the patients suffered from renal insufficiency, heart failure or cerebrovascular
Expression and production of proinflammatory cytokines in Fabry PBMC
To assess the existence of a proinflammatory state in PBMC from Fabry patients, we first analyzed the expression and production of proinflammatory cytokines in PBMC from patients and normal controls.
Unstimulated, freshly isolated PBMC from Fabry patients showed a basal increased expression of the proinflammatory cytokines IL-1β and TNF-α compared to normal controls (p = 0.0098 and p = 0.0496, respectively), whereas the expression levels of IL-1α, IL-6, and IFN-γ were unaltered (Fig. 1).
When the
Discussion
The idea of a chronic stimulation of the immune system in LDs has been introduced a long time ago [17], through findings of increase cytokine levels in sera from affected patients. However, the mechanism that explained these observations was far from being elucidated. In the last few years, much work has been done in an attempt to clarify this key aspect. Production of proinflammatory cytokines such as IL-1β and TNF-α seems to be a common phenomenon among a wide range of LDs [5]. Particularly,
Conclusions
In summary, PBMC from Fabry patients display a proinflammatory cytokine expression and production profile. In particular, dendritic cells and monocytes, two innate immune subsets, could be the main source of this basal proinflammatory state. Additionally, Gb3 can induce the same cytokine profile on normal monocyte-derived macrophages and dendritic cells, indicating that it could be responsible for the generation of this proinflammatory profile. Finally, Gb3 seems to depend on TLR4 to achieve
Role of the funding source
The funding source had no involvement neither in the study design; in the collection, analysis, interpretation of data; in the writing of the report; nor in the decision to submit the paper for publication.
The following are the supplementary data related to this article.
Acknowledgments
We wish to thank the Banco de Sangre, Instituto de Hemoterapia de la Provincia de Buenos Aires, La Plata for kindly providing the buffy coats used in this study.
This work was supported by Shire HGT (USA) [research grant 2007/2008 to PR]; and Agencia Nacional de Promoción Científica y Tecnológica (Argentina) [PICT 2007 No. 01070 to PR].
PR acts as a consultant and has received research grants from Shire HGT.
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