Introduction

Endothelin is a 21-amino acid long peptide that is a vasoconstrictor produced from endothelial cells, vascular smooth muscle cells (VSMC), macrophages, and the renal medulla. They are known to produce endothelin-1 (ET-1) that acts on the receptors ETA and ETB. These receptors are G-protein coupled cell-surface receptors, and ETA is mainly present on smooth muscle cells. ETB receptors appear on endothelial and renal epithelial cells. Both are present in the lungs. ET-1 is known as a potent vasoconstrictor with proliferative, pro-fibrotic, pro-oxidative, pro-inflammatory properties, and maintenance of the tone of VSMC’s. Other ET’s discovered include ET-2 and ET-3. Endothelins are involved in neurovascular uncoupling, affects cognitive functions and blood pressure. Its influence of blood pressure is an essential part of diseases such as postmenopausal hypertension, pre-eclampsia, and pulmonary hypertension in which it plays a major role in its pathogenesis and hence is a vital cornerstone for appropriate management.[1][2][3] In this review, we cover physiology, biosynthesis, and pathophysiology of various diseases that involve endothelin.

Cellular Level

Biosynthesis[4]:

The synthesis of ET gets regulated at the level of transcription. The first protein transcribed would undergo processing by a furin-type proprotein convertase. This process yields a molecule called ‘big ET-1', an inactive intermediate. Endothelin converting enzyme (ECE) acts on big ET-1, converting it into ET-1. ECE conversion consists of two steps: Big ET-1 to ET-1(1-31) by chymase and then to ET-1 by neprilysin.

Cellular Signaling: 

Arterial vasoconstriction is mainly by ETA, while venous vasoconstriction is by ETB. Vasodilation is otherwise mediated via ETB receptor activation. The same vessel undergoes constriction when the ET-1 passes deeper into the smooth muscle layer and activates the ETA receptors.

Regulation: 

Angiotensin-II, transforming growth factor-b, thrombin, bradykinin, hypoxia, and low-density lipoprotein (either oxidized or acetylated) induces the expression of ET-1. An inhibitor of expression is nitric oxide (NO).

Endothelin Converting Enzyme: 

Endothelin converting enzyme-1 (ECE-1) belongs in the family of metalloproteases. The ECE-1 gene is on chromosome 1. The same ECE-1 has four isoforms (ECE-1a, 1b, 1c, 1d), among which ECE-1a and ECE-1c have antagonistic activities of suppressing and increasing the invasiveness of cancer cells.

Endothelin Receptors: 

There are two types of G-protein coupled transmembrane receptors detected - ETA and ETB.[5] Both these receptors produce variant effects corresponding to the binding subtype of endothelin. They also differ in their affinity to each type of endothelin, ie. The binding affinity of ETA: ET-1 is greater than ET-2, which is greater than ET-3, whereas ETB has an equal affinity to all subtypes. ETA and ETB are present variably over different locations. ETA receptors are mainly present on VSMC’s and cause vasoconstriction on ET-1 activation, contrary to ETB that results in a vasodilatory effect. ETB’s on the endothelium also adds to vasodilation in the vessel by releasing vasodilators that act on the VSMC’s. One of the primary functions of the ETB receptor is the ETB-mediated-clearance mechanism in the lungs, which enables clearance of about 80% of the ET-1 in the system.[6]

Development

The first endothelin was identified by culturing aortic endothelial cells and was named as endothelin-1 (ET-1) from the name ‘endothelium-derived-vasoconstrictor.’ Over time ET-2 and ET-3 were discovered from snakes. They are genetically encrypted into the human genome in chromosomes 6, 1, and 20, respectively. Promoters of endothelin production include factors such as thrombin, insulin, epinephrine, angiotensin II, cortisol, and other extrinsic factors such as cyclosporine, vascular shear stress, and hypoxia. The factors that reduce ET include nitric oxide and its inducers, natriuretic peptides, and prostanoids. ET is produced in an immature form called preproendothelin-1 in the cell and undergoes maturation, which involves the enzyme endothelin converting enzyme-1 (ECE-1), which is responsible for cleaving and thus maturing the molecule to 21-amino acids. A newly discovered endothelin produced from mast cells has 31-amino acids (therefore called ET-11-31) in length and has a more constrictive potency.[7] Their pressor responses were unable to be significantly controlled by ET-receptor blockers. ET-2 gets expressed in the ovary and intestinal epithelial cells. ET-3 is present on endothelial cells and intestinal epithelial cells.[8] The half-life of endothelin is less than 5 minutes in human plasma.[9]

Mechanism

In normal states, the usual function of ET is the maintenance of the stability of the vasculature. The most important interaction with ET is nitric oxide (NO). NO gets released after the utilization of L-arginine by the endothelial nitric oxide synthase (eNOS) to, therefore, increase cyclic GMP.[10] NO production can get defective in situations of endothelial dysfunction due to high oxidative stress or inflammation. Mental stress, anger, or cold can also result in abnormal NO metabolism and later global vasoconstriction. Any oxidative stress can jeopardize the functioning of the NO synthase enzyme.[11] In cases when NO is inadequate or defective, other mechanisms involving prostacyclin, natriuretic peptide, and cytochrome-dependent factors come into play to maintain vasodilation. The diseased state tilts the balance towards unbalanced vasoconstriction where there is an upregulation of ET and down-regulation of NO, consequentially, followed by COX-1 and COX-2 mechanisms assist in vasoconstriction.

Dysfunctional Endothelium

There is a plethora of research supporting the loss of NO bioavailability or activity to be a sentinel event in endothelial dysfunction leading to a cardiovascular compromise.[12] Several factors can result in reduced NO activity: reduced expression of eNOS, reduced substrates or cofactors of eNOS, inhibitors against NOS, or increased destruction of NOS. If there occurs destruction of NO pathway, then the unbalanced increase in ET-1 would result in a pro-inflammatory and pro-coagulant environment in the affected vessels. This event is enabled by the increased expression of Von Willibrand Factor (vWF). The dysfunction of endothelium also causes a reduction in pro-fibrinolytic activity, the cumulative effect being to promote coagulation.

Pathophysiology

Pathophysiologic Basis of Diseases

 Endothelin functions in an autocrine and/or paracrine fashion. It virtually affects almost all organs of the body. The most important diseases to know, which have established associations with endothelin as a part of their pathophysiology are pulmonary hypertension, heart failure, post-menopausal hypertension, pre-eclampsia, ovarian cancer, and chronic kidney diseases.[12]

Pulmonary hypertension (PH)

ET-1 is known to be a responsible contributor to the development of pulmonary hypertension (PH) by increasing the vascular tone and promoting vascular remodeling via its mitogenic activity.[13] Although pulmonary artery hypertension (PAH) is a predominantly female disease, there exist sex-related differences in its pathophysiology and prognosis.[12] This fact was established by identifying responses to ET receptor-blockers, which demonstrated a significantly better response in women compared to that of men. Men, on the other hand, demonstrated a better response to phosphodiesterase inhibitors, thus eliciting a sexual dimorphism in pathophysiology. 

The etiology of the PH also bifurcates in the mechanisms involved — thromboembolism induced PH results in an upregulation of ETB.[14] Identification of these pathways paved the development of treatment strategies with ET receptor blockers such as bosentan (non-selective) and sitaxentan (selective) which is now a part of the guidelines of treatment for both NYHA Class 3 and 4 patients whose symptoms do not subside or are non-responsive to immediate vasodilators.

Heart failure (HF)

 The relationship between heart failure and endothelin had been discovered earlier and has begun to show importance in assessing the prognosis of a patient with HF - especially ET-1(1-31).[15] The ET levels in plasma are directly proportional to the degree of failure. Surprisingly, the effect of ET-1 differs based on the state of failure. In a failing heart, ET-1 increases contractility, whereas of that in a normal heart, it reduces contractility. HF also increases the surface expression of ETA receptors and reduces that of ETB.[16]

 Although there exists a linear relation with endothelin and HF, studies on receptor blockade have failed to show any improvement in patients.

Post-menopausal hypertension (PMH)

 Menopause results in multiple changes in the body of a female that begins on average at the age of 52  and can last for up to 2-10 years. Along with these changes also occurs an increase in the prevalence of hypertension in this population. Research has noted that there is an elevation of ET-1 levels after menopause.[17] With a reducing estrogen level and rising ET-1 level, there occurs arterial stiffening, thus contributing to cardiovascular comorbidity. Estradiol is a suppressor for the release of ET-1.[18] ETB, which is responsible for vasodilatory action, also elicits a reduced response after menopause.

Systemic Hypertension (HTN)

 Essential HTN has normal levels of ET-1; however, there is a local rise in the ET-1 levels in the vascular walls. ETA activation demonstrates an increase in blood pressure, and ETB activity ameliorates hypertension. Thus, the use of blockers of ET receptors has shown an overall reduction of blood pressure.[15]

Pre-eclampsia

 Pre-Eclampsia is a pregnancy-specific syndrome that also encompasses endothelial dysfunction along with elevated blood pressure and renal vascular resistance. As mentioned earlier, hypoxia is a triggering factor for ET-1 synthesis, and placental hypoxia is one of the causative factors for elevating the ET-1 levels in the blood leading the evolution of pre-eclampsia. ET-1 also causes increased expression of soluble fms-like-tyrosine kinase-1 (sFlt-1) that is also involved in the pathogenesis. There is a contribution of increased expression of ETA in the development of preeclampsia along with the dysfunction of the ETB receptor.[2]

Ovarian cancer

 The mitogenic effect of ET-1 is of significance for tumor growth in malignancies in ovarian, prostate, colorectal, bladder, and breast.[19] High levels are detected in those of especially ovarian carcinomas in ascitic fluid samples. It acts synergistically with other growth factors such as vascular endothelial growth factor (VEGF) in mostly an autocrine and paracrine fashion. ET-1 also induces the release of cyclooxygenase-2 and prostaglandin-E2 expression. One of the highly studied and associated remedies is the effect of green tea through epigallocatechin-3-gallate.[20] The involvement of ET-1 in tumor progression has revealed treatment strategies that reduce tumor expansion using ET-receptor—inhibitors. Thus, this fact gives promising results in the synergistic effect when given along with paclitaxel. One of the side effects seen in anti-VEGF therapies is although an ET-1 mediated hypertension.[21]

Pathophysiologic activity in the Kidney:

 Renal endothelium produces endothelin, which regulates sodium and water excretion. In a condition of volume overload, the shear stress on the endothelium leads to the expression of ET-1 that acts on the thick ascending limb and collecting duct to reduce water and sodium reabsorption mediated with the action of intermediates like nitric oxide and other pathways.[22] This idea was proven in experiments utilizing ETA blockers that showed natriuresis by its effect on the nephron.[23] Renal ET-1 also stimulates the release of angiotensin-II that, in turn, cyclically increases the expression of ET-1.[24]

Focal Segmental Glomerulosclerosis (FSGS):

 Endothelin induces glomerular injury to the podocytes via activation of ETB, leading to proteinuria and glomerulosclerosis. The exact mechanism remains not clearly understood.[25] FSGS is a clinicopathological entity involving an aberrant glomerular filtration barrier. During the pathogenesis, it also increases ET-1 urinary excretion. The predominant subtype of FSGS involving such a pattern of progression is the age-associated primary FSGS.[26]

Diabetic Nephropathy (DN):

Hyperglycemia is an inducer for ET-1 production, which causes disassembly of the actin cytoskeleton in diabetic nephropathy. ET antagonists have proven to reduce proteinuria in patients having diabetic nephropathy and reversing the disassembly to an extent.[26]

Hypertensive Nephropathy:

 Hypertension leads to increased shear stress and ET-1 production as a cause or effect. Over time, the damage to the kidney ensues and leads to proteinuria. Using ET blockers have shown to be nephroprotective and demonstrated a gradual reduction in glomerulosclerosis and albuminuria along with an increased blood flow with an improved microvessel structure in the cortical kidney.[26]

Clinical Significance

ET-1 is a peptide hormone that is growing significantly due to the detection of involvement in multiple disease processes and its rising trend with age.[27] Other factors found in association with elevated ET-1 were hyperthyroidism, and the tendency to develop atrial fibrillation. Much research takes place on the effects of ET-1 receptor antagonists, which research has gradually revealed to produce a reversal of some diseases such as diabetic and hypertensive nephropathy, to quote a few examples. Drugs such as bosentan and sitaxentan have been life-saving in conditions of pulmonary hypertension, especially among females and other diseases mentioned above. With many of the trials conducted on drugs that act against the peptide, such as for reduction of stroke or subarachnoid hemorrhage with patients on endothelin receptor blockers, it also identifies increasing complications associated with these drugs.[28] One of the significant breakthroughs is its impact on retarding cancer progression. With the pattern of its association with the diseases, ET-1 could likely serve as a marker of cardiovascular stress accounting for endothelial dysfunction. There is yet to understand all of its functions on how it chains itself into many disease processes.