Human Urinary mRNA as a Biomarker of Cardiovascular Disease

Abstract

Background mRNA in urine supernatant (US-mRNA) might encode information about renal and cardiorenal pathophysiology, including hypertension. H, whether the US-mRNA transcriptome reflects that of renal tissues and whether changes in renal physiology are detectable using US-mRNA is unknown. Methods We compared transcriptomes of human urinary extracellular vesicles and human renal cortex. To avoid similarities attributable to ubiquitously expressed genes, we separately analyzed ubiquitously expressed and highly kidney-enriched genes. To determine whether US-mRNA reflects changes in renal gene expression, we assayed cell-depleted urine for transcription factor activity of mineralocorticoid receptors (MR) using probe-based quantitative polymerase chain reaction. The urine was collected from prehypertensive individuals (n=18) after 4 days on low-sodium diet to stimulate MR activity and again after suppression of MR activity via sodium infusion. Results In comparing this US-mRNA and human kidney cortex, expression of 55 highly kidney-enriched genes correlated strongly (r_s=0.82) while 8457 ubiquitously expressed genes correlated moderately (r_s=0.63). Standard renin-angiotensin-aldosterone system phenotyping confirmed the expected response to sodium loading. Cycle threshold values for MR-regulated targets ( SCNN1A, SCNN1G, TSC22D3) changed after sodium loading, and MR-regulated targets ( SCNN1A, SCNN1G, SGK1, and TSC22D3) correlated significantly with serum aldosterone and inversely with urinary sodium excretion. Conclusions RNA-sequencing of urinary extracellular vesicles shows concordance with human kidney. Perturbation in human endocrine signaling (MR activation) was accompanied by changes in mRNA in urine supernatant. Our findings could be useful for individualizing pharmacological therapy in patients with disorders of mineralocorticoid signaling, such as resistant hypertension. More generally, these insights could be used to noninvasively identify putative biomarkers of disordered renal and cardiorenal physiology.

Keywords: RNA, messenger; blood pressure; kidney; mineralocorticoids; urine.

Figure 1.

Correlation of gene expression between transcriptomes of human urinary extracellular vesicles and a human kidney cortex, or a human kidney cortex and a second human kidney cortex sample. A and B, Correlation for highly kidney-enriched genes; (C and D), correlation for ubiquitously expressed (housekeeping) genes. E, Correlation coefficients of gene expression between transcriptomes of human urinary extracellular vesicles and of a series of kidney (top) and brain and bladder samples (lower 2 rows, respectively, as a control). Correlations are calculated as Spearman ρ.

Figure 2.

Scatterplots of cycle threshold (C_t) values according to log-transformed serum aldosterone, with Pearson correlation coefficients. The C_t values’ sign has been reversed (from positive to negative) so that a larger C_t value corresponds with greater gene expression for more intuitive interpretation.

Figure 3.

Scatterplots of log-transformed urinary sodium-creatinine ratio according to cycle threshold (C_t) value, with Pearson correlation coefficients. The C_t values’ sign has been reversed (from positive to negative) so that a larger C_t value corresponds with greater gene expression for more intuitive interpretation.