Functional Role of Uric Acid in Cardiac Stem Cell Homeostasis

Abstract

In New Zealand, cardiovascular disease (CVD) affects 1 in 20 adults and is the leading cause of mortality. Cardiac pathophysiology along with diabetes mellitus is historically associated with glucose metabolism, but recently it has been suggested that elevated serum uric acid (SUA) could be a major driver of these complications. Cardiac stem cells (CSCs) are a new therapy approach for CVD, however, the therapeutic effects are usually mixed. A major contributing factor could be the necrotic environment that the stem cells are being isolated from or transplanted back into. Elevated SUA could contribute significantly to the necrotic environment. Hence, our goal was to characterise the effects of elevated SUA, reflected by changes in extracellular uric acid (EUA) on cardiac progenitor cell (CPC) function in order to improve stem cell therapy, as CPCs are also investigated for use in stem cell therapy and are similar to CSC in terms of physiology and function. We characterised the uric acid (UA) transporter expression profile in human CPCs including known renal transporters: ABCG2, MRP4, GLUT9, URAT1, OAT1, OAT2, OAT4 and NPT4, using qPCR. Human CPCs as well as mRNA from human heart tissue express UA efflux transporters ABCG2 and MRP4 to a greater extent than UA influx transporter GLUT9, irrespective of diabetic status. The expression of GLUT9, ABCG2 and MRP4 were also confirmed at the protein level through western blot analyses of proteins extracted from CPCs. This transporter profile in human CPCs support our new concept of a ‘cellular uric acid homeostasis’ (CUAH), and the fact that a disturbance of CUAH, as in diabetes mellitus, may be detrimental to the function of CPCs and ultimately affect stem cell therapy. From functional assays, it appears that CPCs are sensitive to changes in EUA concentrations. By the end of UA treatment, 300μM UA treatment, which is considered normal SUA, had the most viable cells. The amount of viable cells decreased with 200μM UA and 500μM UA significantly (p<0.01), suggesting that once SUA concentration deviates from optimal levels, ii it begins to affect the functionality of stem cells. Apoptosis assays conducted on CPCs revealed a significant inverse relationship with increased apoptosis in 200μM UA compared to 300μM UA (p<0.05), however, apoptosis at 500μM was not significantly different to 300μM (p=0.1384). Together with the changes in viability, this may suggest that UA may be inducing apoptosis in CPCs if SUA is deviating from normal. Furthermore, the addition of uric acid transporter inhibitors, benzbromarone and probenecid, appeared to cause an amplified effect, with a significant decrease in proliferation compared to control (p<0.0001) for both substances, and a significant increase in apoptosis with the addition of probenecid (p<0.01). In sum, the results support the idea that CUAH is crucial for the functionality of CPCs, and therefore changes in CUAH, or the environment that stem cells and progenitor cells are transplanted into can impair their ability to integrate into the host tissue, and therefore decrease the efficacy of stem cell therapy.