Therapeutic strategies targeting the endothelial glycocalyx

Abstract

PURPOSE OF REVIEW

This review will highlight recent studies that have examined the endothelial glycocalyx in a variety of health conditions, as well as potential glycocalyx-targeted therapies.

RECENT FINDINGS

A degraded glycocalyx is present in individuals that consume high sodium diet or have kidney disease, diabetes, preeclampsia, COVID-19, or sepsis. Specifically, these conditions are accompanied by elevated glycocalyx components in the blood, such as syndecan-1, syndecans-4, heparin sulfate and enhanced heparinase activity. Impaired glycocalyx barrier function is accompanied by decreased nitric oxide bioavailability, increased leukocyte adhesion to endothelial cells, and vascular permeability. Glycocalyx degradation appears to play a key role in the progression of cardiovascular complications. However, studies that have used glycocalyx-targeted therapies to treat these conditions are scarce. Various therapeutics can restore the glycocalyx in kidney disease, diabetes, COVID-19, and sepsis. Exposing endothelial cells to glycocalyx components, such as heparin sulfate and hyaluronan protects the glycocalyx.

SUMMARY

We conclude that the glycocalyx is degraded in a variety of health conditions, although it remains to be determined whether glycocalyx degradation plays a causal role in disease progression and severity, and whether glycocalyx-targeted therapies improve patient health outcomes. Future studies are warranted to investigate therapeutic strategies that target the endothelial glycocalyx.

INTRODUCTION

The endothelial glycocalyx is a potential target to prevent or treat cardiovascular diseases (Figure 1). The glycocalyx is a negatively charged gel-like structure that is bound to the luminal endothelium that protects the endothelium from many of the contents in flowing blood (1). Currently, there are several direct and indirect methodologies used to quantify the glycocalyx in vivo and in vitro, many of which are reviewed extensively elsewhere (2–4). We have shown that the endothelial glycocalyx is deteriorated in advanced age (5, 6) and disease (7). Importantly, glycocalyx deterioration in advanced age accompanies arterial dysfunction (i.e., aortic stiffening and endothelial dysfunction), which is reversed after dietary supplementation with glycocalyx precursors (5). In our previous Current Opinion in Pharmacology review, we highlighted recent studies that examined the role of the endothelial glycocalyx in advanced age and cardiovascular disease (8). Thus, in this review, we will take it a step further by highlighting recent studies that have examined the glycocalyx in a variety of health conditions (i.e., high salt sodium diet, kidney disease, diabetes, preeclampsia, COVID-19, and sepsis). The glycocalyx is a relatively new therapeutic target, and glycocalyx-targeted therapies have not been employed in some of these conditions reviewed here. Therefore, one of the goals of this review is to highlight existing therapies that warrant further investigation in other conditions that have a degraded glycocalyx. Please note that not all glycocalyx-targeted therapies are discussed here either, refer to Banerjee et al. for a more extensive review outlining glycocalyx-targeted therapies (9).

Figure 1:

Image of the endothelial glycocalyx in a mouse soleus capillary captured via electron microscopy (scale bar = 500 μm).

HIGH-SALT DIET

In nearly all industrialized societies, excess dietary sodium consumption is a major contributor to cardiovascular disease. Because the glycocalyx is negatively charged, it is believed to buffer sodium in the blood, which has been supported by in vitro studies (10). While the glycocalyx can buffer sodium, there is a limit, and as sodium concentration exceeds the buffering capacity, the glycocalyx rapidly deteriorates and likely initiates arterial dysfunction. It is not clear how much sodium is required to degrade the glycocalyx. However, a recent study indicates that high sodium consumption was accompanied by poor red blood cell sodium sensitivity, which implies a deteriorated glycocalyx (11). Recently, our laboratory demonstrated that 12-weeks of high-sodium diet in mice deteriorates the endothelial glycocalyx (12) and induces arterial dysfunction, indicated by elevated systolic blood pressure, aortic stiffening, and endothelial dysfunction (13). In support of this, high-sodium diet-induced glycocalyx deterioration has also been shown to be accompanied by the loss of key glycocalyx components such as syndecan-1, sydecan-4, and heparin sulfate (14). High-sodium diet also results in the alterations in immunoglobulins, apolipoproteins, fatty acid-binding proteins, integrins, and macrophages that promote endothelial dysfunction (15). Although a high-sodium diet is consumed by the vast majority of industrialized societies, there are no current studies that have investigated glycocalyx-targeted therapies to restore the glycocalyx in people that consume a high-sodium diet. However, given that most of the population consumes excess dietary sodium, it is likely that therapeutics for other conditions reviewed here could be effective in reversing high-sodium diet-induced glycocalyx deterioration and subsequent cardiovascular disease risk.

KIDNEY DISEASE

In the renal circulation, the glycocalyx lines the glomerular endothelium and plays a critical role in its integrity and function. When the glomerular endothelial glycocalyx is damaged, the endothelium and podocytes are exposed to the circulating factors within the blood, leading to albuminuria (16). Indeed, patients with podocyte disorders have an increase in syndecan-1, heparin sulfate in the blood, as well as enhanced heparinase activity, indicating glycocalyx degradation (17). Glycocalyx degradation in these conditions is accompanied by von Willebrand factor and thrombomodulin release into the blood that may also contribute to podocyte damage (17). Similarly, acute kidney damage with transient proteinuria could be caused by locally activated heparinases in kidney that degrade the glycocalyx leading to a diminished glomerular filtration barrier and vascular leakage (18). In patients with chronic kidney disease (CKD), peritubular capillaries undergo rarefaction and increased permeability that is at least partly due to a reduced glycocalyx thickness (19) and increased glycocalyx stiffness (20). Indeed, exposing endothelial cells to serum from patients with CKD degrades the glycocalyx, inducing a dysfunctional, inflammatory endothelial cells with lower nitric oxide synthase expression and increased monocyte adhesion (20). Extracellularly, glycocalyx alterations in CKD are partially dependent on the activation of mineralocorticoid receptors and its downstream target epithelial sodium channel on the surface of endothelial cells. In diabetic nephropathy, glycocalyx degradation is associated with mineralocorticoid receptor activation (21) and cholesterol deposition in glomerular endothelial cells (22), which may lead to increased glomerular albumin permeability. When exposing endothelial cells to glycocalyx precursors, such as fucoidan and hyaluronan, the glycocalyx is not degraded by from serum of patients with CKD. The preventative effect of glycocalyx precursors have been shown to be dependent on intracellular ERK/MAPK signaling pathway, indicating the preservation of glycocalyx may be dependent on the exocytosis of components in preformed vesicles rather than direct incorporation from the circulation (23). However, it is unclear if the ERK/MAPK pathway is directly involved in CKD-related glycocalyx deterioration. Nevertheless, glomerular endothelial glycocalyx integrity appears to be predictive of the glomerular function, as kidney diseases are often accompanied by glycocalyx degradation.

DIABETES

The glycocalyx likely plays a major role in the pathogenesis of cardiovascular complications in diabetes, such as diabetic retinopathy, cardiomyopathy, and nephropathy. In the absence of overt cardiovascular disease, glycocalyx barrier function is impaired following an oral glucose tolerance test in individuals with impaired glucose tolerance or first-degree relatives of patients with diabetes (24). Moreover, impaired glycocalyx barrier function is accompanied by a low nitrite/nitrate plasma levels, implying an acute endothelial dysfunction. Thus, individuals at risk for diabetes have an abnormal response to a glucose challenge that results in a deteriorated glycocalyx and markers of endothelial dysfunction. Interestingly, there have been conflicting observations on alterations to endothelial glycocalyx with diet-induced obesity (12, 25). A less thick, but stiffer glycocalyx has been observed in mice fed a Western diet, which was accompanied by a downregulated inwardly rectifying K⁺ channels on endothelial cells. These findings indicated an impairment in sensitivity of endothelial cells to shear stress that leads to endothelial dysfunction (25). In contrast, our laboratory observed an increased glycocalyx thickness and barrier function in mice fed a Western diet (12) that was also accompanied by endothelial dysfunction (26). An increase in glycocalyx thickness was also observed in the cerebrovasculature after high-fat diet consumption by others (27). Currently, it is unclear how the glycocalyx is affected by diabetes and likely multifactorial dependent on site of measurement and length/severity of diabetes. It appears that an early adaptation in a prediabetic state may result in a thickening of the glycocalyx, whereas late and/or more severe disease may result in a deteriorated glycocalyx.

In patients with diabetes, retinopathy is a major health concern, and the ocular glycocalyx may play a pivotal role in preventing retinopathy. Indeed, the ocular glycocalyx is diminished in both type-1 and type-2 diabetes, which may be due to chronic elevations in blood glucose levels (28). In vitro findings indicate that hyperglycemia results in elevated shedding of glycocalyx components, such as heparin sulfate, chondroitin sulfate, and hyaluronan when retinal endothelial cells are exposed to high-glucose media (29). Another under-recognized complication of diabetes is diabetic cardiomyopathy. To investigate the undefined role of endothelial glycocalyx in coronary circulation and hence diastolic function in diabetes, Qiu et al. utilized both type-1 and type-2 diabetes mice models and found that diastolic dysfunction was preceded by a loss of endothelial glycocalyx in the coronary microvascular endothelial cells. Loss of glycocalyx was accompanied by perivascular edema, indicating an increased microvascular permeability (30). Importantly, administration of angiopoietin-1 was able to restore glycocalyx, reduce perivascular edema, and improve diastolic function (30). Restoration of the glycocalyx has also been shown with empagliflozin, a sodium-glucose cotransporter inhibitor used in the treatment of type 2 diabetes and has been shown to ameliorate endothelial dysfunction and arterial stiffness in a mouse model of vascular aging (31). In regard to the glycocalyx, empagliflozin attenuates inflammation and endoplasmic reticulum stress of aortic endothelial cells caused by heparin sulfate degradation in endothelial glycocalyx by reducing oxidative stress and unfolded protein responses (32). In summary, a compromised endothelial glycocalyx is a precursor to impaired glucose handling ability and end-organ complications in patients with diabetes, thus, early detection and treatments may be of clinical importance.

PREECLAMPSIA

Preeclampsia is a condition that can occur in pregnant women after 20 weeks of gestation (early-onset: week 20–34; late-onset: week 34+) and results in elevated blood pressure. Although the cause of preeclampsia is unclear, other symptoms often include elevated aortic stiffness, endothelial dysfunction, oxidative stress, inflammation, and glycocalyx degradation (33). In recent years, it has been suggested that endothelial glycocalyx degradation may contribute to endothelial dysfunction in preeclampsia, as glycocalyx properties are lower in preeclampsia compared to non-preeclamptic pregnant women (34, 35). This is supported by findings from multiple investigators that patients with early-onset preeclampsia have elevated blood concentrations of hyaluronan and syndecan-1 compared to normotensive pregnant women (33, 34, 36). Hyaluronan and syndecan-1 are components of the glycocalyx, thus, elevated concentrations in the blood indicated a degraded glycocalyx. It should be noted that glycocalyx properties are lower in healthy pregnant women compared to nonpregnant women (35). These findings suggest that pregnancy results in glycocalyx degradation may occur naturally but is further degraded in preeclampsia. The reduction in the glycocalyx layer could play a crucial role in the pathophysiology of preeclampsia and is a potential target for therapies. Due to the complex pathogenesis of preeclampsia and safety concerns concerning drug usage during pregnancy, there are currently no effective pharmacological treatments. Nonetheless, researchers are exploring the potential advantages of dietary intervention in lowering the risk of preeclampsia, where the daily requirement of dietary intake in a healthy pregnant woman is higher than that of a healthy nonpregnant woman. It is suggested by these findings that glycocalyx degradation may have a significant role in the pathophysiology of preeclampsia, and further study is required to investigate the mechanism in greater detail.

COVID-19

COVID-19, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is a systemic disease associated with vascular inflammation and endothelial injury, both directly by the virus and indirectly due its inflammatory response (37, 38). In COVID-19, elevated systemic inflammation, specifically TNF-a, CRP, and IL-6, have been found to correlate with disease severity (37, 38). Recent studies have shown a link between COVID-19 and endothelial glycocalyx damage, as patients with COVID-19 have impaired glycocalyx barrier function (39). The level of glycocalyx damage may be dependent on COVID-19 severity, as in vitro findings indicate that blood derived from patients with mild symptoms reduced the glycocalyx by approximately 49%, whereas sera from patients with severe symptoms reduced the height of the glycocalyx by >60% (40). COVID-19 severity is also related to the magnitude of elevation in blood concentrations of syndecan-1, heparinase, heparin sulfate, and hyaluronan (37, 40–45). The loss of syndecan-1 from the endothelial glycocalyx leads to increased adhesion of leukocytes to endothelial cells, enhanced vascular permeability, and intravascular coagulation (39, 40, 46), which is a major concern as glycocalyx damage could increase the severity of inflammatory response to COVID-19, as well as the length of recovery. Indeed, endothelial damage has been found to persist even months after infection (i.e., long COVID), with studies indicating residual glycocalyx damage and endothelial dysfunction four months after infection (46, 47). This is in line with others that observed an acute elevation in aortic stiffness and blood pressure (48, 49) that did not return to normal values until 4–6 months after COVID-19 infection (50). Although the timeline is unclear, in some individuals with long COVID, the complete restoration of glycocalyx properties was not achieved until ~18 months post-infection (51).

Given the central role of the glycocalyx in maintaining vascular homeostasis, it represents an attractive therapeutic target for patients with COVID-19. Corticosteroid administration has been shown to have glycocalyx protective potency, stabilizing endothelial barrier function and reducing inflammation associated with COVID-19-induced endotheliitis (52, 53). There have also been several successful strategies aimed at restoring the glycocalyx to accelerate COVID-19 recovery. The mineralocorticoid receptor antagonist spironolactone has been shown to improve glycocalyx thickness in mild COVID-19 cases (40). Another study tested whether glycocalyx degradation could be restored, cultured human primary pulmonary microvascular endothelial cells in the presence of both COVID patient sera and the heparin sulfate mimetic fucoidan. Supplementation of fucoidan restored the glycocalyx and decreased heparinase, an enzyme that degrades heparin sulfate. These results highlighting the potential of glycocalyx preservation or restoration in preventing progression to severe COVID-19 (54). In support of heparin as a target for glycocalyx restoration in COVID-19, the heparinase inhibitor administration has been shown to prevent the decline in glycocalyx thickness in response to COVID-19 serum (39). In conclusion, endothelial cell dysfunction and loss of the glycocalyx appear to drive worse outcomes in severe COVID-19 patients. Further validation of glycocalyx preservation or restoration could encourage future clinical trials to evaluate the efficacy of this approach in treating more severe forms of COVID-19. A mechanistic understanding of the direct and indirect effects of SARS-CoV-2 on the vasculature is critical for clarifying the pathobiology of COVID-19.

SEPSIS

Sepsis is a life-threatening condition characterized by organ dysfunction due to a dysregulated response to infection. The glycocalyx may play a critical role in sepsis mortality, as glycocalyx degradation occurs in both patients with sepsis and animal models of sepsis (55–59). Patients with sepsis demonstrate glycocalyx degradation, indicated by elevated plasma heparin sulfate and heparinase concentrations in children (57) and elevated plasma hyaluronan and syndecan-1 in adults (58). It is unclear if there are distinct differences in glycocalyx degradation with advanced age in sepsis. Mouse models of sepsis also demonstrate a degraded glycocalyx (59). Indeed, LPS injection in mice, which induces sepsis, leads to elevated plasma heparin sulfate and IL-6 two hours post-injection, indicating an acute inflammatory response that may be linked to glycocalyx degradation (57). Using cecal ligation and puncture method to induce sepsis in mice, plasma syndecan-1 are increased six hours after surgery (56). Treatment with Colivelin, a humanin peptide derivative, lowers plasma syndecan-1, TNF-α, and IL-10 concentrations. Moreover, Colivelin-treated mice had glycocalyx restoration demonstrated by a thicker and complex GAG bundles compared to vehicle-treated septic mice (56). Heparin sulfate proteoglycan expression is also lowered by 90% in mice that receive cecal ligation and puncture compared to control mice. Interestingly, IFN-β+nicotinamide riboside treatment inhibits the reduction of heparin sulfate proteoglycan expression in this model (55). Moreover, Intravenous administration of IFN-β (1,000 units/20 g) 6 h and 18 h after cecal ligation and puncture improved survival rates by 40% compared to control mice (55). Vitamin C and recombinant thrombomodulin have also been investigated to preserve the glycocalyx and improve outcomes in sepsis. In humans, 48 hours of high-dose intravenous vitamin C (50 mg/kg every 6 hours) lowered plasma syndecan-1 concentrations (60). Treatment recombinant thrombomodulin normalized glycocalyx characteristics in septic mice demonstrated by lower plasma syndecan-1 concentrations after treatment (59). Taking steps to protect and restore the glycocalyx may reduce vascular hyperpermeability, inflammation, and organ dysfunction associated with sepsis. The glycocalyx and endothelium are clearly impacted by sepsis, but the degree to which glycocalyx restoration improves outcomes sepsis requires further investigation, which may lead to new therapeutics for sepsis, as well as other conditions that result in glycocalyx degradation.

CONCLUSIONS

In this review, we highlighted recent studies that have examined the glycocalyx in a variety of different health conditions that are accompanied by a deteriorated glycocalyx. Although glycocalyx degradation appears to be a symptom of these health conditions, it remains to be determined if the glycocalyx plays a causal role in disease progression and severity. In addition to reviewing the recent studies that have examined the glycocalyx in these conditions, we also highlighted glycocalyx-targeted therapies used for treatment. It is apparent that glycocalyx-targeted therapies can restore the glycocalyx in some of these conditions. However, futures studies are warranted to determine how glycocalyx restoration impacts patient health outcomes, as well as to determine if maintaining the glycocalyx via regular diet or dietary supplementation prior to the onset of overt disease is a viable preventative strategy.

Figure 2:

In its normal state, the endothelial glycocaly protects the endothelium from the contents of blood. The glycocalyx mechanotransduces shear stress to the endothelium, which promotes nitric oxide-mediated vasodilation. In its degraded state, components of the endothelial glycocalyx can be found in the blood, and the vasoprotective functions of the glycocalyx are inhibited.

KEY POINTS.

• A degraded glycocalyx may play a key role in the progression of cardiovascular complications and is present in individuals that consume high sodium diet or have kidney disease, diabetes, preeclampsia, COVID-19, or sepsis.

• Glycocalyx degradation can be assessed visually by imaging the glycocalyx or by measuring glycocalyx components in the blood.

• Therapeutics that expose endothelial cells to glycocalyx components have been shown to restore the glycocalyx in kidney disease, diabetes, COVID-19, and sepsis.

• It remains to be determined how glycocalyx restoration impacts patient health outcomes.