Sources of Urinary Proteins and their Analysis by Urinary Proteomics for the Detection of Biomarkers of Disease

Abstract

Renal disorders account for a substantial fraction of the budget for health care in many countries. Proteinuria is a frequent manifestation in afflicted patients, but the origin of the proteins varies based on the nature of the disorder. The emerging field of urinary proteomics has the potential to replace kidney biopsy as the diagnostic procedure of choice for patients with some glomerular forms of renal disease. To fully realize this potential, it is vital to understand the basis for the urinary excretion of protein in physiological and pathological conditions. In this review, we discuss the structure of the nephron, the functional unit of the kidney, and the process by which proteins/peptides enter the urine. We discuss several aspects of proteinuria that impact the proteomic analysis of urine of patients with renal diseases.

Figure 1

Kidney and its structural and functional components. Each human kidney (left top) contains ~1,000,000 functional units or nephrons (middle, top) that span the two regions of the kidney: cortex and medulla. Each nephron is composed of a filtration structure (glomerulus) and a downstream tubule comprised of 11 functional segments. An afferent arteriole delivers blood to the encapsulated glomerulus (right, top) where it enters the capillary network, undergoes ultrafiltration and its residual volume exits by the efferent arteriole to return to the systemic circulation. The ultrafiltrate enters the proximal tubule (PT) as the primary urine and is progressively modified as it flows through the remaining segments of the tubule. Glomeruli are located in the cortex, while the tubular portions of the nephrons span the cortex and medulla. This structural feature creates the necessary concentration gradients for extraction of salts, water and various compounds from the urine in the tubular lumens. In the cross-section of the deep cortex (left, bottom), capillaries (C) and tubuli (T) are depicted, as well as cells in the surrounding tissue, the interstitium. A cross-section of a capillary loop in the glomerulus (right, bottom) reveals the structures and resident cell types responsible for formation of the primary urine: endothelial cell (E) line the capillaries and contain openings (fenestrae) that permit water, salts, and small proteins and low-molecular-weight compounds to filter across the glomerular basement membrane (GBM); arrows in the enlarged inset on the right-hand side depict the flow of ultrafiltrate). These substances then pass through slit diaphragms (SD) that interconnect the interdigitating foot processes (FP), or pedicles, of epithelial cells (podocytes, P) that overlie the capillaries. Upon this entry into Bowman’s space (BS), this fluid is termed primary urine. The mesangium is the centrolobular region of the glomerular tuft that helps to maintain patency of the capillary loops. Mesangial cells (MC) regulate glomerular blood pressure, produce cytokines/chemokines and radical oxygen species, and secrete extracellular matrix proteins necessary for the structural integrity of the glomerulus. Larger circulating substances, such as immune complexes, can more easily enter the mesangium than Bowman’s space because the GBM and slit diaphragms are not present in this area. These processes may culminate in glomerular fibrosis with loss of filtration function of the nephron. In addition, release of various chemokines/cytokines may induce interstitial inflammation and scarring, further damaging the integrity of the nephron to compromise its function.

Figure 2

Electron micrographs of glomerular capillary loops. (A) Glomerulus from a patient with nephrotic-range proteinuria. Black arrows indicate areas of effacement of the foot processes of the epithelial podocyte. Effacement is associated with a substantial decrease in the number of slit diaphragms. (B) Normal glomerulus. Black arrows indicate intact foot processes of the epithelial podocyte overlying the glomerular basement membrane. Slit diaphragms are located between the foot processes. Bars indicate 2 micrometers.