Hyperphosphatemia, Phosphoprotein Phosphatases, and Microparticle Release in Vascular Endothelial Cells

Significance Statement

Cardiovascular disease is the leading cause of death in patients with chronic kidney disease. More than half of the patients with kidney diseases develop vascular complications at some stage of their disease progression. Hyperphosphatemia, high serum inorganic phosphate (Pi) levels, is a common biochemical abnormality in kidney patients and can exert damaging effects on endothelial cells lining the blood vessels. Microparticles, small 100-1000nm in diameter cell membrane fragments, are released from cells under stress. While hyperphosphatemia can exert its damaging effect on cells of the vasculature by inducing soft-tissue calcification, we have proposed that Pi can exert functional effects on the cells of the blood vessels (especially endothelial cells), resulting in generation of MPs by a direct inhibition of phosphoprotein phosphatases resulting in cytoskeletal protein dysregulation and subsequent MP formation. The mechanism of this Pi-induced cell stress and microparticle formation is elusive. In this study we have shown a novel mechanism by which Pi induces cell-stress, resulting in the generation of pro-coagulant forms of endothelial MPs which may contribute to acute occlusive events. The major finding of our study is that inorganic phosphate concentrations similar to those observed in hyperphosphatemia in kidney patients trigger an acute release of pro-coagulant microparticles from human endothelial cells. This is of direct interest to nephrologists because it provides a molecular basis for the observed link between hyperphosphatemia and cardiovascular disease in kidney patients. It is also of interest to a wider audience because the proposed mechanism provides a widely applicable explanation for pathological effects of phosphate excess in mammalian cells. Our study provides a novel pathologic link between hyperphosphatemia, generation of MPs, and thrombotic risk.

About the author

Nima Abbasian, Ph.D.

BSc in Biology (Microbiology) Azad University, Iran (2006)

MSc in Infection & Immunity, University of Leicester, UK (2010)

Ph.D. in Infection, Immunity, & Inflammation, University of Leicester, UK (2015)

KRUK Postdoctoral Research Fellow, University of Leicester, UK (2015-Present)

Research interests

My research focuses on cellular and molecular mechanisms of inorganic phosphate (Pi) induced stress on blood cells and endothelial cell dysfunction in kidney disease. I am particularly interested in: a) The regulation of intracellular concentration of inorganic phosphate (Pi) by plasma membrane solute transporter proteins in human endothelial cells in response to higher extracellular Pi concentration (known as hyperphosphatemia) and subsequent signal transductions induced by Pi in the cells, and b) The mechanism of high Pi-induced stress in cells and subsequent release of cell membrane-derived microparticle (MPs). I have focused particularly on the mechanism of inorganic phosphate signalling to cytoskeletal proteins and subsequent clinically important pro-coagulant endothelial MP release that acutely starts budding of the cell membrane during uremic cardiovascular disease.

Journal Reference

J Am Soc Nephrol. 2015;26(9):2152-62.

Abbasian N¹, Burton JO², Herbert KE³, Tregunna BE¹, Brown JR², Ghaderi-Najafabadi M³, Brunskill NJ², Goodall AH³, Bevington A⁴.

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Departments of Infection, Immunity and Inflammation and.
Departments of Infection, Immunity and Inflammation and John Walls Renal Unit, University Hospitals of Leicester, Leicester, United Kingdom; and.
Cardiovascular Sciences, University of Leicester, Leicester, United Kingdom; Leicester National Institute for Health Research Cardiovascular Biomedical Research Unit Cardiovascular Sciences, University of Leicester, United Kingdom.
Departments of Infection, Immunity and Inflammation and [email protected].

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Abstract

Hyperphosphatemia in patients with advanced CKD is thought to be an important contributor to cardiovascular risk, in part because of endothelial cell (EC) dysfunction induced by inorganic phosphate (Pi). Such patients also have an elevated circulating concentration of procoagulant endothelial microparticles (MPs), leading to a prothrombotic state, which may contribute to acute occlusive events. We hypothesized that hyperphosphatemialeads to MP formation from ECs through an elevation of intracellular Pi concentration, which directly inhibits phosphoprotein phosphatases, triggering a global increase in phosphorylation and cytoskeletal changes. In cultured human ECs (EAhy926), incubation with elevated extracellular Pi (2.5 mM) led to a rise in intracellular Pi concentration within 90 minutes. This was mediated by PiT1/slc20a1 Pi transporters and led to global accumulation of tyrosine- and serine/threonine-phosphorylated proteins, a marked increase in cellular Tropomyosin-3, plasma membrane blebbing, and release of 0.1- to 1-μm-diameter MPs. The effect of Pi was independent of oxidative stress or apoptosis. Similarly, global inhibition of phosphoprotein phosphatases with orthovanadate or fluoride yielded a global protein phosphorylation response and rapid release of MPs. The Pi-induced MPs expressed VE-cadherin and superficial phosphatidylserine, and in a thrombin generation assay, they displayed significantly more procoagulant activity than particles derived from cells incubated in medium with a physiologic level of Pi (1 mM). These data show a mechanism of Pi-induced cellular stress and signaling, which may be widely applicable in mammalian cells, and in ECs, it provides a novel pathologic link between hyperphosphatemia, generation of MPs, and thrombotic risk.

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