CD73 Rather Than CD39 Is Mainly Involved in Controlling Purinergic Signaling in Calcified Aortic Valve Disease.

Igor Kudryavtsev,Vladislav Titov,Olga Irtyuga,Alexey Golovkin,Olga Moiseeva,Maria Serebriakova,Dmitry Isakov,Patimat Murtazalieva,Ekaterina Zhiduleva,Daria Semenova,Lubov Mitrofanova,Anastasya Shishkova,Anna Malashicheva

doi:10.3389/fgene.2019.00604

Abstract

The study aimed to compare composition of peripheral blood T-cell subsets and assess their surface expression of CD39 and CD73 ectonucleotidases in patients with severe and moderate aortic stenosis (AS) as well as to evaluate involvement of T-cell-mediated immune processes in valve calcification. The study was performed with 38 patients suffering from severe calcified aortic stenosis (SAS), 33 patients with MAS, and 30 apparently healthy volunteers (HVs). The relative distribution and percentage of T-cell subsets expressing CD39 and CD73 were evaluated by flow cytometry. T helper (Th) and cytotoxic T-cell subsets (Tcyt) were identified by using CD3, CD4, and CD8 antibodies. Regulatory T cells (Tregs) were characterized by the expression of CD3, CD4, and high IL-2R alpha chain (CD25high) levels. CD45R0 and CD62L were used to assess differentiation stage of Th, Tcyt, and Treg subsets. It was found that MAS and SAS patients differed in terms of relative distribution of Tcyt and absolute number of Treg. Moreover, the absolute number of Tcyt and terminally differentiated CD45RA-positive effector T-cells (TEMRA) subset was significantly higher in SAS vs. MAS patients and HVs. However, the absolute and relative number of naïve Th and the absolute number of Treg were significantly higher in MAS vs. SAS patients; the relative number of naïve Tregs was significantly (p < 0.01) decreased in SAS patients. It was shown that CD73 expression was significantly higher in SAS vs. MAS patients noted in all EM, CM, TEMRA, and naïve Th cell subsets. However, only the latter were significantly increased (p = 0.003) in patients compared with HVs. SAS vs. MAS patients were noted to have significantly higher percentage of CD73+ EM Tcyt (p = 0.006) and CD73+ CM Tcyt (p = 0.002). The expression of CD73 in patients significantly differed in all three Treg populations such as EM (p = 0.049), CM (p = 0.044), and naïve (p < 0.001). No significant differences in CD39 expression level was found in MAS and SAS patients compared with the HV group. Overall, the data obtained demonstrated that purinergic signaling was involved in the pathogenesis of aortic stenosis and calcification potentially acting via various cell types, wherein among enzymes, degrading extracellular ATP CD73 rather than CD39 played a prominent role.

Highlights

IntroductionA significant number of studies have been recently published evidencing about an important role played by the receptors for extracellular ATP and its metabolites as well as enzymes involved in regulating their metabolism in vascular and heart valve calcification (Fish et al, 2013; Towler, 2017; Dou et al, 2018; Golovkin et al, 2018; Kaniewska-Bednarczuk et al, 2018)
We demonstrated that the percentage of peripheral blood Tcyt was insignificantly increased mainly due to TEMRA Tcyt subset (Supplementary Table 1), whereas frequency of naïve Tcyt and naïve Treg subsets was decreased in patients in severe calcified aortic stenosis (SAS) patients vs. healthy volunteers (HVs), respectively (Figure 2)
Cell functions regulated by purinergic signaling seem to be of importance in various pathological conditions including formation of aortic stenosis and its calcification, which may be related to activity of peripheral blood T-cell subsets and aortic valve interstitial cells

Summary

Introduction

A significant number of studies have been recently published evidencing about an important role played by the receptors for extracellular ATP and its metabolites as well as enzymes involved in regulating their metabolism in vascular and heart valve calcification (Fish et al, 2013; Towler, 2017; Dou et al, 2018; Golovkin et al, 2018; Kaniewska-Bednarczuk et al, 2018). CD39 (E-NTPDase1—ectonucleoside triphosphate diphosphohydrolase 1) cleaves ATP to two AMPs and two phosphate molecules; the latter is considered as one of the major cues promoting tissue hydroxyapatite deposition (Fish et al, 2013). Another phosphate molecule is generated by CD73 (Ecto5′NTase—ecto-5′-nucleotidase) converting AMP to adenosine (Fish et al, 2013). Inorganic pyrophosphate (PPi) formed during AMP synthesis mediated by ectonucleotide pyrophosphatase/phosphodiesterase (ENPP) represents a potential inhibitor of hydroxyapatite deposition and subsequent tissue calcification (Fish et al, 2013)

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Discussion

Conclusion