Untangling the Knot in Diabetic Nephropathy: The Unanticipated Role of Glycocalyx in the Antiproteinuric Effect of Endothelin Receptor Antagonists

Diabetes is a devastating disease that is imposing itself as one of the largest health emergencies of the 21st century, and high blood glucose is the third-highest risk factor for premature mortality (1). In 2015, almost 5 million people aged 20–79 years died from diabetes and its related complications, which accounted for almost 15% of the overall causes of mortality among people in this age-group (1). With no effective prevention and management programs, the incidence of diabetes and its complications is dramatically increasing worldwide, imposing a substantial impact on the economy. Diabetic nephropathy (DN) affects 30–40% of patients with diabetes (2) and induces a progressive deterioration of renal function. Albuminuria independently associates with increased vascular mortality and morbidity in patients with diabetes (3), in patients with hypertension (4), and in the general population (5), and therefore is not only a sign of renal impairment and a key pathogenic element of renal disease progression but also a marker of more generalized vascular damage. Many studies have linked albuminuria with cardiovascular disease (CVD). A linear relationship was observed between albumin excretion and CVD risk (6), but the exact significance and mechanism of such a relationship are not yet understood. Microalbuminuria not only denotes renal capillary damage but also represents an early biological marker of vascular injury (7). Endothelial dysfunction has been implicated as a potential major mechanism for vascular disease, and microalbuminuria has been linked to markers of endothelial dysfunction in patients with and without diabetes (8).

Several experimental studies provided evidence of a pathogenic role for endothelin 1 (ET-1) in DN. Pioneering investigations documented preservation of renal function and limitation of long-term renal histological changes in diabetic rats after administration of selective endothelin A receptor (ET_AR) antagonists (9,10). The effectiveness of ET_AR antagonism has been attributed to the preservation of the renal microvascular architecture together with a protection against podocyte loss (11), a typical feature of overt diabetes. ET-1 directly affects podocytes as revealed by less proteinuria and protection from podocyte dysfunction and glomerulosclerosis in diabetic mice with a podocyte-specific double deletion of ET_AR and ET_BR (12). In this issue of Diabetes, Boels et al. (13) confirm the renoprotective action of atrasentan, a selective ET_AR antagonist, in streptozotocin-induced diabetic apolipoprotein E (apoE) knockout mice. Notably, atrasentan reduced the urinary albumin-to-creatinine ratio without affecting systemic blood pressure, glomerular hypertrophy, or podocyte number. The latter data are not surprising considering that diabetic apoE knockout mice did not faithfully replicate typical features of overt diabetic nephropathy, and, as already observed in experimental early diabetes, renal nitric oxide levels increased in this model (14). The novelty proposed by Boels et al. is the intriguing mechanism through which atrasentan may preserve the glomerular endothelial glycocalyx, thus exerting its antiproteinuric effect. In the diabetic apoE knockout mice, proteinuria was associated with a reduction of glomerular endothelial glycocalyx coverage and glomerular basement membrane (GBM) thickness (13). The endothelial glycocalyx is a mesh-like, hydrated structure comprising glycoproteins and proteoglycans anchored to the endothelium. In different glomerular diseases including DN, low expression of heparan sulfate proteoglycans in the GBM inversely correlated with the degree of proteinuria (15). In line with these findings, Boels et al. argued that the reduction of the endothelial glycocalyx in diabetic apoE knockout mice was mediated by increased glomerular expression of heparanase, the heparan sulfate–degrading enzyme known to be involved in the development of proteinuria in DN (16). By in vitro studies, the authors demonstrated that incubation of human umbilical vein endothelial cells with plasma of patients with diabetes in flow condition decreased glycocalyx thickness and enhanced heparanase production. Although interesting, this finding should be confirmed in glomerular endothelial cells, in which the composition and thickness of the glycocalyx differ from that of the nonfenestrated endothelium (17). Boels at al. also showed that addition of atrasentan to patient plasma normalized both glycocalyx and heparanase expression, supporting a direct link between ET-1 and alteration in endothelial glycocalyx via ET_AR in diabetes. Some concerns arise about the significance of these provocative findings, considering that endothelial cells do not seem to express ET_AR (18). Hypothesizing the mechanism through which ET-1 may promote increased glomerular heparanase expression, the authors suggested the possibility that in diabetic mice proinflammatory M1 macrophages could be involved in heparanase degradation by secreting cathepsin-L, the enzyme responsible for the cleavage/activation of proheparanase. In this context, the shift in the balance from M1 to anti-inflammatory M2 macrophages in diabetic mice treated with atrasentan provides a convincing explanation for the protective effect of the ET_AR antagonist on glyxocalyx. On the other hand, heparanase was found to stimulate macrophage activation in inflammatory bowel disease, thus creating a vicious circle that powered the chronic proinflammation status (19). That such a detrimental mechanism might be critically involved in heparanase-dependent glycocalyx damage induced by ET-1 in DN should be also considered.

Although the study by Boels et al. (13) greatly advances our understanding of how the finely tuned regulation of the GBM may be disrupted during DN, several important questions still remain. In particular, the current study did not investigate the effect of atrasentan on podocyte injury occurring in DN. In this regard, the authors, in a previous collaborative study (20), demonstrated that the combined reduction of endothelial and podocyte glycocalyx is required for the onset of proteinuria in streptozotocin-induced diabetic mice. ET-1 activated podocytes to release heparanase that in turn reduced endothelial glycocalyx thickness causing proteinuria—all phenomena prevented by the targeted disruption of ET_AR/ET_BR in podocytes (20). In the present study by Boels et al., the complete restoration of the endothelial glycocalyx after atrasentan administration was not associated with a full reduction of albumin excretion. This is consistent with the main role of podocytes in regulating heparanase production and endothelial glycocalyx function, suggesting that impairment of endothelial-podocyte cross talk is the main culprit in overt DN (21).

The significance of this study rests on the new mechanism of renoprotection delivered by an ET_AR antagonist acting on heparanase production in DN (Fig. 1). Future studies are needed for the identification of the intracellular mediators through which ET-1 stimulates heparanase production in glomerular endothelial cells and podocytes. This effort would pave the way for exploring further therapeutic strategies to inhibit heparanase besides the heparanase inhibitors that are currently under investigation in clinical trials in cancer research.