Current Vascular Pharmacology - Volume 12, Issue 2, 2014

GPCR-mediated receptor transactivation of EGFR, is one of the effector mechanisms by which GPCR ligands, such as Ang II, thrombin and ET -1, catecholamine, SII-angiotensin, PAF, and uPA that are released at the arterial injury sites, can potentiate intimal hyperplasia. The process of EGFR transactivation can be cognate ligand-dependent or independent. In cognate ligand- dependent transactivation, ligand-bound GPCR results in the activation of metalloproteases, which sheds membrane tethered growth factor that binds to EGFR on an extracellular ligand-binding domain causing its transactivation. Whereas, in cognate ligand independent transactivation, ligand bound GPCR activates EGFR intracellularly via second messenger system. The mechanism of EGFR transactivation depends on cell type, GPCR ligand and the type of GPCR. In this review article, such cross-talks are critically discussed. The EGFR transactivation generates mitogenic signals leading to various pathological conditions. The goal of this review article is to identify potential targets for therapeutic intervention in clinical conditions related to intimal hyperplasia in cardiovascular system.

The discovery of endothelin (ET) in 1988 has led to considerable effort to unravel its implication in health and disease and the mechanisms evoked by ET. ET-1 and related signaling aberrancies are believed to be implicated in the pathogenesis of diverse cardiovascular diseases, such as hypertension, atherosclerosis, hypertrophy and diabetes. The endothelin system consists of three potent vasoconstrictive isopeptides, ET-1, ET-2 and ET-3, signaling through two G protein coupled receptors, ETA and ETB, which are linked to multiple signaling pathways. Activated signaling transduction pathways include the modulation of the adenylyl cyclase/cAMP pathway through stimulatory (Gs) and inhibitory (Gi) G proteins, activation of the phosphoinositide pathway through the activation of proteins Gq/11, generation of oxidative stress, growth factor receptor-related mitogenic events, such as the activation of phosphatidylinositol-3 kinase pathway, phosphoinositide pathway and activation of the mitogen-activated protein (MAP) kinase cascade. The levels of ETA and ETB receptors as well as the signaling pathways activated by these receptors are altered in several cardiovascular diseases including hypertension, hypertrophy, atherosclerosis, diabetes, etc. In this review, we provide an overview of the signaling events modulated by ET-1 in vascular smooth muscle cells in both physiological and pathological conditions.

G-protein coupled receptors (GPCRs) are commonly present at the plasma membrane and their signaling modulates excitation-contraction coupling and excitation-secretion coupling of excitable and non-excitable cells of the cardiovascular system. Their effect on excitation-gene expression coupling was attributed, in part, to the nuclear translocation of their signaling and/or to the entry into the nuclear membrane of the internalized GPCRs. However, the recently established paradigm showed that, in addition to plasma membrane G-proteins, GPCRs exist as native nuclear membranes receptors and they modulate nuclear ionic homeostasis and function. These nuclear membrane GPCRs could function independently of plasma membrane GPCRs. Growing evidence also shows that these nuclear membrane GPCRs contribute to protein synthesis and also undergo changes in pathological conditions. The presence of a GPCR at both the plasma and nuclear membranes and/or only at the nuclear membranes represents a new challenge to better understand their contribution to cell physiology and pathology and, consequently, to the development of new therapeutic drugs targeting this category of receptors.

Endogenous kinins are important vasoactive peptides whose effects are mediated by two G-Protein-coupled receptors (R), named B2R (constitutive) and B1R (inducible). They are involved in vascular homeostasis, ischemic pre- and post- conditioning, but also in cardiovascular diseases. They contribute to the therapeutic effects of angiotensin-1 converting enzyme inhibitors (ACEI) and angiotensin AT1 receptor blockers. Benefits derive primarily from vasodilatory, antiproliferative, antihypertrophic, antifibrotic, antithrombic and antioxidant properties, which are associated with the release of endothelial factors such as nitric oxide, prostacyclin and tissue plasminogen activator. Uncontrolled production of kinins or the inhibition of their metabolism may lead to unwanted pro-inflammatory side effects. Thus, B2R antagonism is salutary in angioedema, septic shock, stroke, and Chagas vasculopathy. B1R is virtually absent in healthy tissues, yet this receptor is induced by the cytokine pathway and the oxidative stress via the transcriptional nuclear factor NF-κB. The B1R may play a compensatory role for the lack of B2R, and its up-regulation during tissue damage may be a useful mechanism of host defense. Activation of both receptors may be beneficial, notably in neovascularisation, angiogenesis, heart ischemia and diabetic nephropathy. At the same time, B1R is a potent activator of inducible nitric oxide and NADPH oxidase, which are associated with vascular inflammation, increased permeability, insulin resistance, endothelial dysfunction and diabetic complications. The dual beneficial and deleterious effects of kinin receptors and, particularly B1R, raise an unsettled issue on the therapeutic value of B1R/B2R agonists versus antagonists in cardiovascular diseases. Hence, the Janus-face of kinin receptors needs to be seriously addressed in the upcoming clinical trials for each pathological setting.

Vasoactive peptides such as angiotensin II and endothelin-1 as well as growth factors regulate vascular homeostasis through signaling pathways that are triggered in both normal and disease states. These vasoactive peptides and growth factors also increase the cellular levels of calcium which, through calcium binding effector systems initiating the downstream signaling and physiological responses in target cells. A multifunctional calcium-calmodulin-dependent protein kinase II (CaMKII) has emerged as an important transducer of vasoactive peptide-induced responses in vascular smooth muscle cells (VSMC). The catalytic activity of CaMKII can be stimulated by autophosphorylation and oxidation leading to the activation of signaling events that mediate growth, proliferation, migration, and gene transcription in VSMC. Pharmacological and gene deletion approaches have demonstrated a requirement of CaMKII in mediating the mitogen- activated protein kinase and phosphatidyl-inositol 3-kinase/protein kinase B signalling, as well as the proliferative, migratory and transcriptional responses of vasoactive peptides. In addition, a potential involvement of hyperactive CaMKII in animal models of vascular disease has also been reported. Therefore, this review aims to highlight the role of CaMKII in mediating signaling and physiological responses in VSMC and discuss its potential role in vascular pathophysiology.

G protein signaling is an extremely complex event that is involved in almost every cellular process. As such, G protein-coupled receptors are the most commonly found type of transmembrane receptors used by cells to initiate intracellular signaling events. However, the widely accepted model of cyclical GDP-GTP exchange in response to ligand binding to 7TMRs, followed by dissociation of the G protein subunits and activation of intracellular signaling cascades, has repeatedly been challenged in recent years. Some of the exceptions that have been brought forth include signaling by a non-dissociated, rearranged heterotrimer and the existence of “reverse-mode”, active G proteins that interact with active receptors. Here, we focus on Gαi/o, one of the common Gα classes, and outline a major exception to the classical model, that of G protein coupling to RTKs. We then describe a novel concept in Gαi/o signaling, namely that the pathways induced by agonist binding circumvent the typical signaling pathways responsive to decreases in the second messenger cAMP, via adenylyl cyclase inhibition.

The integrity of the vitamin D endocrine system is essential for human health. Nutritional vitamin D deficiency in otherwise healthy individuals, associates with a higher risk of mortality for all causes, despite normal serum calcitriol. These deadly causes extend beyond the recognized adverse impact of vitamin D deficiency on calcium and phosphate homeostasis predisposing to secondary hyperparathyroidism, bone loss and vascular calcification. Vitamin D deficiency also associates with an early onset of disorders of aging, including hypertension, proteinuria, insulin resistance, immune abnormalities that enhance the propensity for viral and bacterial infections, autoimmune disorders, cancer, and multiple organ damage due to excessive systemic inflammation causing atherosclerosis, vascular stiffness, renal lesions, and impaired DNA-damage responses. The frequency and severity of all of these disorders markedly increase in chronic kidney disease (CKD) because the kidney is essential to maintain serum levels of calcitriol, the most potent endogenous endocrine activator of the vitamin D receptor (VDR), and also of 25-hydroxyvitamin D, for local rather than systemic VDR activation. The goal of this review is to update the current understanding of the pathophysiology behind the classical and non-classical actions of VDR activation that help prevent the onset and/or attenuate the progression of renal and cardiovascular damage in CKD. This knowledge is essential to identify non-invasive, sensitive and accurate biomarkers of the severity of these disorders, a first step to generate evidence-based recommendations for a safe correction of vitamin D and/or calcitriol deficiency in the course of CKD that effectively improves outcomes.

Vitamin D is essential for maintaining a healthy mineral and bone metabolism. It stimulates calcium and phosphate absorption by the intestine, regulates bone metabolism, and negatively controls PTH secretion through the endocrine action of its active metabolite calcitriol. Vitamin D also possesses a variety of effects unrelated with mineral and bone metabolism, including the regulation of arterial blood pressure and the prevention of cardiovascular complications, modulation of immunological responses, regulation of insulin production and prevention against diabetes, protection against certain cancers, renoprotection, and other beneficial actions. This article provides an update review of the sources and pharmacological characteristics of natural vitamin D compounds, their most clinical uses, and their most important clinical results.

In patients with chronic kidney disease (CKD) and end-stage renal disease (ESRD), 25OH-vitamin D (calcidiol) deficiency or insufficiency is a common finding with high prevalence. Numerous epidemiological studies have found an independent association of low levels of calcidiol with increased morbidity and mortality. Within different patient cohorts, application of cholecalciferol or ergocalciferol (native vitamin D) as well as calcifediol can help replenish vitamin D levels in patients with and without renal disease. However, it is unclear if such an approach is effective in modifying relevant clinical end-points. Currently available data are insufficient to clearly define situations in which calcifediol therapy might be superior to ergocalciferol or cholecalciferol therapy in terms of increasing calcidiol levels in CKD / ESRD. Similar to ergocalciferol or cholecalciferol application, also calcifediol therapy needs to undergo testing in randomized, controlled trials (RCT) in severe CKD or ESRD with reasonable end-points before recommendations about therapy can be established.

The synthesis of 1α,25-dihydroxyvitamin D3 (Calcitriol) takes place mostly in the kidneys through the action of 1α-hydroxylase (CYP27B1) which converts 25(OH)D into 1,25(OH)2D3. Renal production of calcitriol is stimulated by PTH, low calcium and low phosphate and it is reduced by high phosphate and FGF23. Binding of 1α,25-dihydroxyvitamin D3 to its receptor (VDR) causes gut absorption of calcium and phosphate, decrease in PTH synthesis, stimulation of FGF23. At the bone level calcitriol suppresses pre-osteoblasts and activates mature osteoblasts. VDR is present in a large variety of cells that do not have any direct role in the regulation of mineral metabolism. Calcitriol regulates immune and inflammatory response, cell turnover, cell differentiation, Renin production, reduces proteinuria and others. In patients with Chronic Kidney Disease (CKD) there is a decrease in calcitriol that is apparent at early stages of renal disease; this is probably due to the elevation of FGF23 which is present since very early stage of CKD. In CKD stage, 3-4 moderate doses of calcitriol are effective to control secondary hyperparathyroidism and observational studies suggest that calcitriol therapy increases survival and slows the progression of renal disease as long as phosphate and calcium levels are controlled. Calcitriol (0.5 µg calcitriol twice per week) has been effective in decreasing proteinuria in patients with IgA nephropathy. In dialysis patients, the administration of calcitriol reduces serum PTH levels but it is also known that high doses of calcitriol are associated with hypercalcemia and worse control of hyperphosphatemia. In kidney transplant patients, the administration of calcitriol, 0.5 µg/48h prevents bone mass loss during the first few months after transplantation.

Alfacalcidol is a widely used vitamin D compound, especially in clinical nephrology because it does not require enzymatic activation by the kidneys. For that reason it has been used for decades to treat abnormalities in bone and mineral balance that arise in chronic kidney disease. In this review an overview is provided of available experimental and clinical data that form the basis of its widespread use. Supported by studies on cell lines and animal models, clinical studies have firmly established beneficial effects of alfacalcidol on chronic-kidney disease (CKD-) related bone disease and secondary hyperparathyroidism. Its effects on muscle structure and function are not unambiguous. Neither experimental nor clinical data suggest beneficial effects of alfacalcidol on indicators of cardiovascular disease, including cardiovascular calcification, left ventricular mass function or hypertension. Suggestions of improved mortality in dialysis patients treated with alfacalcidol are based on observational cohort analyses. Integrating all available data leads the conclusion that alfacalcidol is a justifiable compound to prescribe to CKD patients with established bone disease or hyperparathyroidism. Monitoring of calcium and phosphorus levels after its initiation is required and dose adaptations should be made accordingly.

Compared to such native vitamin D agents as cholecalciferol (D3), egocalciferol (D2), and calcifediol (25- hydroxy vitamin D3, which need 1-alpha hydroxylase to be activated, 1-alpha-ergocalciferol, also known as doxercalciferol, is a synthetically manufactured vitamin D analog used for treatment of secondary hyperparathyroidism (SHPT) in chronic kidney disease (CKD). Doxercalciferol exhibits more structural similarities to plant-based vitamin D2, ergocalciferol, than with animal-based vitamin D3, cholecalciferol. Because doxercalciferol does not have a 25-hydroxy group, it requires 25-hydroxylation by the liver to be activated, a process independent of kidneys. Doxercalciferol shares these features with its D3 equivalent, 1-alpha-hydroxy-cholecalciferol (alphacalcidol), both of which are activated hepatically and independent of renal or extra-renal 1-alpha hydroxylase. In experimental and clinical studies of CKD and SHPT, doxercalciferol effectively reduces parathyroid hormone levels and restores abnormal bone pathology. The efficacy of doxercalciferol is similar to other vitamin D analogs including calcitriol, alphacalcidol, paricalcitol (19-nor-1,25-dihydroxyvitamin D2 ) and maxacalcitol (1,25-dihydroxy-22-oxa-vitamin D3). The frequency and magnitude of hypercalcemia or hyperphosphatemia observed with doxercalciferol treatment may be less than calcitriol or alphacalcidol therapy but not less than such vitamin D-mimetics as paricalcitol or maxacalcitol. Doxercalciferol can be used for the treatment of SHPT across all ranges of CKD, particularly if hypercalcemia is of concern. There are limited data as to whether doxercalciferol confers greater efficacy or better outcome and survival than other vitamin D analogs and D-mimetics. Whereas further studies are warranted, doxercalciferol can safely be used for correction of SHPT in the entire spectrum of CKD.

The activation of vitamin D receptors (VDR) - (including activation by 25-hydroxyvitamin D) - seems to have not only mineral-metabolism beneficial effects but also important extra-skeletal actions. Paricalcitol is a synthetic vitamin D2 agonist of the VDR approved for the prevention and treatment of secondary hyperparathyroidism associated with chronic kidney disease (CKD). As a result of its selectivity, paricalcitol provides a wider therapeutic window for PTH suppression, minimizing deleterious effects of high serum calcium and/or phosphate concentrations. Paricalcitol also shares, and sometimes improves pleiotropic vitamin-D related systemic effects. For instance, paricalcitol has been repeatedly shown to decrease calcium and phosphate deposition in vessels and to decrease the expression of osteogenic factors preventing the active transformation of smooth muscle vascular cells into osteoblast-like cells in experimental models. In patients, paricalcitol has been associated with improved survival of dialysis patients and it may improve residual albuminuria in diabetic patients. Consequently, paricalcitol may enhance the standard of care in these high-risk patients. Although it seems reasonable to use these potential advantages to guide the individual and integral management of the complex CKD-mineral and bone disorder, it is necessary to recognize that many of these observations have not been proven nor confirmed in prospective clinical trials.

22-oxacalcitriol (OCT) is a vitamin D3 analog and a vitamin D receptor activator (VDRA) that is used as a drug for secondary hyperparathyroidism (SHPT) and has been available clinically in Japan since 2000. The pharmacological characteristics of OCT include rapid clearance from the systemic circulation compared to that of calcitriol, good tissue distribution, and relatively long retention in the nucleus in parathyroid cells. In clinical studies, OCT has been shown to decrease the parathyroid hormone (PTH) level with an effect equivalent to that of calcitriol. Other reports show that OCT produces superior improvement of bone metabolism compared to calcitriol. Treatment with ultrasound-guided direct injection of OCT into the parathyroid has also been attempted. In animal studies, OCT does not influence the blood Ca or P level and is less likely to promote progression of calcification in cardiovascular tissue. The influence of VDRAs including OCT on the progression of cardiovascular lesions and survival in SHPT patients requires further studies.

In this brief review we point out the specificities of the vitamin D system that are necessary to understand why each change in the molecule can result in significantly different biologic effects. Vitamin D, with a specific receptor in most of the tissues, has innumerable potential therapeutic applications in many clinical fields. However, excessive pharmacologic increments of circulating natural metabolites carry the risk of significant side effects. To avoid this, natural vitamin D molecules have been modified to more selectively stimulate some tissues. Changes have been attempted on particular parts of the molecule in order to affect some specific step of the complex machinery that characterize the vitamin D system. The first modifications were those in the side chain of the molecule, which are expected to affect, either or both, the steps of binding to transfer protein or the interaction with catabolic enzymes. More recently other regions, like A-ring (involved with receptor interaction) or CD bicyclic ring (involved with molecule stability), have been modified to obtain always more selective products. Notably each modification of the molecule also affects its shape thus further and variably modifying its interaction with the VDR, with the transport proteins or the catabolic enzymes. As a consequence, the biologic effects of new molecules become less predictable and require in vitro evaluation, experimental animal studies and a complete and specific clinical validation in specific disease states. With thousands of analogs synthesized in the laboratories, only a minority are approved for clinical employment. Besides secondary hyperparathyroidism and osteoporosis, Vitamin D analogs can be employed in other clinical conditions like cancer and autoimmunity diseases. We briefly report on some new experimental or already approved analogs in their main clinical fields of employment.

Numerous drugs with vitamin D activity are available for clinical use and it may not be easy for the nonspecialist to select the most suitable for the individual patient. In this paper we review the main characteristics of the available drugs and provide evidence about any potential specific clinical indications, with special emphasis on renal patients, in order to facilitate the optimal choice. Natural vitamin D products (i.e. those identical to natural metabolites) are first examined, followed by the most frequently used synthetic molecules (i.e. bioengineered molecules not-existing in nature), which are generally indicated as “ analogs”. Either cholecalciferol, ergocalciferol or calcifediol can be employed in subjects with normal renal function and in CKD stage 3-5 patients to correct vitamin D deficiency and improve, respectively, age- or growth-related bone disease and secondary hyperparathyroidism. Calcifediol can be considered more rapid and effective. In all cases, especially with increasing doses, the risk of hypercalcemia must be taken into account. Calcitriol, which can be regarded as the active hormonal form of vitamin D, has the most potent hypercalcemic effect in both normal and renal failure patients. In renal patients calcitriol is a potent inhibitor of parathyroid activity, but the risk of hypercalcemia, now regarded as harmful, is evident whenever pharmacologic doses are used. Alfacalcidol, requiring 25-hydroxylation to become the active hormonal form of vitamin D3, is prescribed in normal subjects to treat osteoporosis and in renal patients to cure hyperparathyroidism and renal bone disease. Doxercalciferol, transformed into the active hormonal form of vitamin D2 following 25-hydroxylation, is mostly studied in renal patients in whom it cures secondary hyperparathyroidism, possibly with a lower calcemic effect than calcitriol. Paricalcitol, a vitamin D2 analog not requiring activation, has been specifically developed to suppress PTH in renal patients with a limited calcemic effect. As such it is now regarded as a powerful drug useful to treat even severe cases of secondary hyperparathyroidism. Importantly, reno-protective and cardio-protective effects of this analog have been recently evaluated by means of randomized clinical trials in renal patients with partially positive renal effects and negative cardiac results, thus additional studies are needed for confirmation. 22-oxacalcitriol, a vitamin D3 analog with a limited calcemic effect available in Japan, is mostly used in renal patients affected by secondary hyperparathyroidism. The clinical activity of some vitamin D analogs is such that they can be employed in diseases like cancer and autoimmunity. The clinical activity of some vitamin D analogs is such that they can be employed in diseases like cancer and autoimmunity. In summary, available drugs with vitamin D like activity are not all the same either in terms of pharmacological actions, and side-effects. They have specific characteristics that may be useful to know in order to operate the best choice in the individual patient.