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Division of Endocrinology and Metabolism, 1st Department of Internal Medicine, AHEPA University Hospital, Aristotle University of Thessaloniki, AUTH, Greece
Division of Endocrinology and Metabolism, 1st Department of Internal Medicine, AHEPA University Hospital, Aristotle University of Thessaloniki, AUTH, Greece
The overall effect of valproic acid on bone is an increased fracture rate.
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Valproic acid decreases osteoblast proliferation and alters the collagen synthesis.
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Vitamin D catabolic enzymes are induced by valproic acid.
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Valproic acid increases indirectly bone fragility through endocrine side effects.
Abstract
Purpose
Valproic acid (VPA) is a broad-spectrum antiepileptic drug, which is widely used as a first line treatment for idiopathic and symptomatic generalized epilepsy, as well as in non-epileptic psychiatric disorders in adult and pediatric patients. Although valproic acid is considered to be a generally well–tolerated drug, numerous studies have shown an increased bone loss and fracture risk in patients treated with VPA. The purpose of this review is to outline recent findings on VPA molecular mechanisms and their action on bone metabolism.
Methods
Unrestricted electronic search of medical databases, complemented by additional manual searches, was performed by August 2016.
Results/conclusion
The main effects of VPA on bone metabolism involve a decrease in osteoblast proliferation, changes in collagen synthesis as well as an induction of vitamin D catabolism. Apart from these direct actions of VPA in bone, indirect effects affecting other endocrine organs also contribute to VPA-induced bone loss.
Valproic acid (2-propylpentanoic acid, N-dipropylacetic acid) is a branched short-chain fatty acid, which derived from valeric acid and was synthesized by Burton in 1882 [
]. Valproic acid (VPA) was initially used as molecule carrier. It was in 1963 that Meunier, while he was studying the antiepileptic effects of new molecules against seizures induced by pentelenetetrazole in experimental animals, reported that VPA prevented pentylenetetrazol-induced convulsions in rodents [
] carried out the first human study leading to the acknowledgement of VPA antiepileptic properties, which was approved in 1978 by FDA as a first-line anti-epileptic drug.
Despite its well-known anti-convulsive activity, VPA is also effectively used in non-epileptic conditions, such as migraine and bipolar disorders, while it has also been recently explored for its use as an adjuvant anti-cancer agent. More precisely, VPA is found to suppress tumor growth and tumor angiogenesis because of its action as histone deacetylase (HDAC) inhibitor. Histone acetylation increases gene transcription, while histone deacetylation suppresses the transcription process. VPA induces HDAC inhibition, histone acetylation and hyperacetylation accumulation, reverse HDAC-mediated transcriptional repression and subsequently mediates in various cell functions such as cell differentiation and cell apoptosis [
VPA is considered to be a generally well-tolerated drug with serious side effects, such as hepatotoxicity, pancreatitis and teratogenicity rarely being reported. Among its long-term side effects, osteoporosis and increased fracture risk have been extensively studied in humans presenting inconsistent results, while the underlying mechanisms remain largely unknown. In this review we outline recent studies concerning the effect of VPA on bone cells and the molecular mechanisms implicated.
2. Overview of bone metabolism
The human skeleton is composed of cortical and trabecular bone, which undergoes continuous renewal throughout lifetime. Bone remodeling occurs through highly tuned and concerted actions of the bone cells, which resorb damaged, old bone (osteoclasts) and form and lay down new bone matrix (osteoblasts) under the tight regulation of the osteocytes, which act as the bone mechanostat and orchestrate bone renewal according to what is needed in every bone unit (Fig. 1).
Osteoclasts are large multi-nucleated cells deriving from the mononuclear hematopoietic cell lineage. Differentiation of monocytes into active resorbing osteoclasts is regulated positively by the receptor activator of nuclear factor kappa B ligand (RANKL) and the macrophage-stimulating colony factor (M-CSF), and negatively by osteoprotegerin (OPG) [
]. Mature osteoclasts present a ruffled border, which is a series of deep folds in the area of the plasma membrane in contact with the bone matrix, secreting lysosomal enzymes, such as tartrate-resistant acid phosphatase (TRAP) and cathepsin K [
]. During bone resorption signals secreted by osteoclasts, osteocytes or the resorbed bone matrix itself attract osteoblasts, promoting the coupling of bone resorption with bone formation.
Osteoblasts are descendants of the mesenchymal stem cell (MSC) lineage, along with adipocytes, chondrocytes, myoblasts and fibroblasts. Osteoblast differentiation from multipotent MSCs is mainly dependent on the transcription factors runt-related transcription factor 2 or osterix [
]. Bone formation takes place in three stages: 1) production of osteoid matrix, 2) maturation of osteoid matrix, and 3) mineralization of the matrix. Osteoblasts secrete various autocrine and paracrine factors, such as transforming growth factor-beta, bone morphogenetic proteins, RANKL, M-CSF and OPG [
]. After fulfilling their bone formation role, osteoblasts assume one of three fates: 1) undergo apoptosis, 2) remain on the bone surface as flat bone lining cells, or 3) become entombed in the newly-formed bone matrix as terminally-differentiated osteocytes. While the exact role of bone lining cells is not well understood, it has been suggested that they may be involved in the initiation of bone remodeling [
]. Osteocytes are buried in the bone matrix in lacunae, while their long slender cell processes connect to other osteocytes and the bone surface through narrow channels called canaliculi. The resulting interconnected network is known as the lacunar-canalicular system. In contrast to short living osteoclasts (2–3 weeks) and osteoblasts (1 month), osteocytes are the longest living cell population in bone and are responsible for sensing mechanical forces imposed on bone and translating them to biochemical cues, which ultimately regulate bone turnover.
3. The effect of enzyme – inducers and non-enzyme inducers anti-epileptic drugs on bone metabolism
The underlying pathophysiological mechanism of anti-epileptic drugs (AEDs)- effect on bone metabolism is multifactorial, and includes activation of cytochrome p450 enzyme, increased bone turnover and increased urinary loss of calcium and phosphorus [
]. The relationship between AED type and fracture risk, however, remains uncertain.
The conventional enzyme-inducing AEDs(EIAEDs), such as phenytoin, phenobarbital, primidone and carbamazepine, induce the hepatic cytochrome P450 enzyme system and are the AEDs most commonly associated with decreased bone mass and increased fracture risk. The main mechanism is via accelerated catabolism of 1,25 vitamin D to 24,25 vitamin D leading to vitamin D deficiency, decreased intestinal absorption of calcium and secondary hyperparathyroidism [
On the other hand, limited data are available regarding the effect of non-enzyme-inducers (AEDs), such as lamotrigine, oxcarbamazepine, levetiracetam and topiramate, on bone and calcium metabolism [
The impact of VPA, which is an inhibitor of the cytochrome P450 enzyme, on bone metabolism is still controversial. Earlier studies had reported no association between the use of VPA and bone loss while more recent ones demonstrated abnormal biochemical indices of bone and mineral metabolism, and higher fracture rates after long-term treatment in both children and adults [
Clinical trials, investigating the effects of VPA on bone mineral density (BMD) and fracture risk, have reported inconsistent results. In some studies VPA monotherapy in children or young adults did not affect BMD values at the femoral neck and lumbar spine [
In a meta-analysis, which included studies with children on long-term VPA treatment from 0.5 to 8 years, the authors concluded that VPA induces a considerable decrease in BMD [
Furthermore, apart from osteoporosis and fracture risk, VPA has been associated with a significant decrease in growth velocity after one year of treatment, determined by the difference between two consecutive measures of the patient’s height and compared with healthy subjects [
In order to clarify the effects of VPA on bone metabolism and interpret the reduction in BMD values, different bone markers have been conscripted and yet there is also conflict in this aspect (Table 2).
Table 2Studies investigating parameters of calcium homeostasis and bone markers in patients under long-treatment with VPA.
Fish liver oil and propolis as protective natural products against the effect of the anti-epileptic drug valproate on immunological markers of bone formation in rats.
Fish liver oil and propolis as protective natural products against the effect of the anti-epileptic drug valproate on immunological markers of bone formation in rats.
Fish liver oil and propolis as protective natural products against the effect of the anti-epileptic drug valproate on immunological markers of bone formation in rats.
Fish liver oil and propolis as protective natural products against the effect of the anti-epileptic drug valproate on immunological markers of bone formation in rats.
Fish liver oil and propolis as protective natural products against the effect of the anti-epileptic drug valproate on immunological markers of bone formation in rats.
Fish liver oil and propolis as protective natural products against the effect of the anti-epileptic drug valproate on immunological markers of bone formation in rats.
], have shown that VPA treatment in children decrease bone formation markers other than osteocalcin, such as procollagen I carboxyterminal-propeptide (PICP).
On the contrary, three human studies and one animal study demonstrated that VPA treatment increases the serum levels of bone specific ALP (bALP) [
Fish liver oil and propolis as protective natural products against the effect of the anti-epileptic drug valproate on immunological markers of bone formation in rats.
]. Voudris et al. showed that increased bALP values are not necessarily followed by an increase in total ALP and may be a useful marker in diagnosing impairment of bone metabolism in children treated with VPA [
] examined adult patients under long-term treatment with VPA and reported a significant elevation of bone resorption as assessed by higher levels of ionized calcium and the increase in carboxyterminal telopeptide of type I collagen(ICTP) concentration. On the contrary, in other studies treatment with VPA decreased serum levels of tartrate-resistant acid phosphatase (TRACP), which reflect the number of active osteoclasts [
], and ICTP levels. In addition, in the study by Lin et al. decreased TRACP levels in children under treatment with VPA was positively correlated with decreased growth velocity [
Fish liver oil and propolis as protective natural products against the effect of the anti-epileptic drug valproate on immunological markers of bone formation in rats.
], showed that VPA increased bone formation (osteocalcin and bone specific alkaline phosphatase), and bone resorption, such as N-terminal telopeptide, NTx, markers while it also altered the RANKL/OPG ratio in favor of RANKL, thereby promoting osteoclastogenesis.
4.1.3 Calcium homeostasis
Another part of bone metabolism that has been examined in patients under treatment with VPA is calcium (Ca) homeostasis. Most of the studies have reported that VPA treatment did not alter significantly Ca, phosphate (P), 25-(OH)-vitamin D, PTH or ALP values [
], pointing to the development of secondary hyperparathyroidism in these patients, while in two others VPA treatment increased significantly calcium levels [
] categorized young patients under VPA treatment according to their activity and demonstrated that inactive children have significantly lower 25-(OH)-vitamin D concentration accompanied by decreased PTH levels compared to children with higher activity.
5. Molecular mechanisms of VPA effect on bone metabolism
]. Despite the enhancement of osteoblasts differentiation, it was shown that during the maturation process of an osteopr○genitor cell line (hFOB 1.19), treatment with VPA significantly down-regulated synthesis of two bone proteins, collagen type 1 and osteonectin [
], which display critical roles in bone formation and subsequent mineralization by mature osteoblasts. Collagen type 1 is the major protein component of bone matrix and mutations in collagen type 1 chains 1 and 2 are the cause of osteogenesis imperfecta in the majority of cases [
], while osteonectin is a collagen -binding glycoprotein, which is critical for bone mass maintenance and normal bone remodeling. Homozygosity for missense variants in the osteonectin gene also causes osteogenesis imperfecta type XVII, whereas in vivo osteonectin-null mice show decrease osteoblast number and bone formation rate [
], which as pluripotent cells can be differentiated into osteoblasts, chondrocytes, adipocytes and myoblasts in vitro. In this study VPA suppressed mesenchymal cell proliferation in the presence of extracellular matrix proteins such as fibronectin and type I collagen, which are known inducers of mesenchymal cell proliferation [
], showing that it may rearrange the cytoskeleton of various cell types. In a microarray analysis of somatic tissue from mouse embryos, VPA was shown to alter the microtubule cytoskeleton and actin filament and thus could be associated with malformations in these embryos' axis skeleton [
In another study with MSCs cultures, VPA enhanced the migration of cord blood mesenchymal stromal cells to injured tissues through increasing the expression of stromal cell-derived factor receptors, CXCR4 and CXCR7, without, however, affecting their ability to differentiate to osteocytes and chondrocytes [
Based on current evidence it appears that while VPA may exert positive effects on osteoblasts proliferation and differentiation, it suppresses the activity of mature osteoblasts and this observation may be the molecular link with the increased skeletal fragility and fracture risk that is shown in clinical studies (Fig. 2). Although osteocytes are of critical importance in skeletal integrity and bone strength, there are no data available for a direct effect of VPA on osteocytes’ viability or gene expression. Similarly, there are no data available on a direct VPA effect on osteoclasts.
One of the major contributing factors of VPA effect in bone is VPA-induced osteomalacia, which is considered to be mediated through accelerated catabolism of 1,25 (OH)2 vitamin D.
Vitamin D is a fat-soluble vitamin, obtained from food such as fish, liver, milk and eggs, or de novo synthesized from cholesterol in the liver. The active metabolite of vitamin D displays two sequential hydroxylations. The first 25-hydroxylation takes place in the liver and the second 1a-hydroxylation, which is considered to be the rate limiting step of active form of vitamin D synthesis, in the kidney.
The active form of vitamin D, 1,25 (OH)2 vitamin D, binds with high affinity and selectivity to its specific vitamin D receptor (VDR), which belongs to the nuclear receptor family of transcription factors. Upon activation by vitamin D, VDR forms a heterodimer with the retinoid-X receptor and binds to the DNA responsive elements of the target cells. The interaction of 1,25(OH)2D with the VDR initiates a complex cascade of molecular events, regulating the transcription of specific genes or gene networks. Non-genomic actions of 1,25(OH)2D, mediating rapid and non–transcriptional dependent actions of vitamin D, have also been described [
]. Interestingly, both studies have shown that this effect was not mediated solely through VPA action as a HDAC1 inhibitor, but also included activation of extracellular signal-regulated kinase and direct activation of androstane receptor and pregnane- X receptor.
5.3 Indirect effect on bone metabolism through endocrine complications
It is controversial whether the endocrine dysfunction in epilepsy patients is caused by the epilepsy itself, the antiepileptic therapy, or both. Treatment with VPA has been associated with hypogonadism, hypothyroidism, hyponatremia, and mild hypercortisolemia, which can also contribute to bone loss seen in these patients.
Chronic administration of VPA affects the function of the hypothalamic-pituitary-gonadal axis primarily in males [
]. The underlying mechanisms of VPA effects on reproductive function are not fully elucidated but several theories have been proposed. It has been suggested that the increase in male androgens could be associated with the inductive effect of VPA on liver enzymes responsible for the production of sex hormone binding globulin (SHBG) [
]. In addition, it has been shown that VPA blocks androgens and progesterone receptors within the therapeutic levels and that especially the blockade of androgens receptor could contribute to the reported impaired reproductive functions during VPA treatment [
]. Finally, it has been reported that VPA treatment induces carnitine deficiency by inhibiting its biosynthesis through decreasing the concentration of alpha-ketoglutarate [
]. Carnitine is an amino acid derivative, which is an essential cofactor in the beta-oxidation of fatty acids and is considered an indicator of epididymal functioning. Through carnitine deficiency, VPA affects indirectly the semen parameters, such as sperm motility [
L: -carnitine fumarate and isovaleryl-L: -carnitine fumarate accelerate the recovery of bone volume/total volume ratio after experimetally induced osteoporosis in pregnant mice.
] supplementation with carnitine has been shown to reverse decreased BMD.
Chronic VPA treatment also produces an elevation in plasma cortisol concentrations without any concomitant rise in ACTH concentrations. It is possible that valproate produces a direct effect at the level of the adrenal cortex, or perhaps increases production of an adrenocortical-stimulating hormone other than ACTH.
Severe symptomatic hyponatremia and syndrome of inappropriate antidiuretic hormone secretion have also been reported in relation to VPA exposure in adults [
Syndrome of inappropriate antidiuretic hormone secretion (SIADH) or hyponatraemia associated with valproic Acid: four case reports from the Netherlands and a case/non-case analysis of vigibase.
Assessment of low-density lipoprotein oxidation: paraoxonase activity, and arterial distensibility in epileptic children who were treated with anti-epileptic drugs.
]. Elevated thyroid-stimulating hormone (TSH) was also found in children (mean age 3.7) receiving VPA monotherapy for 9 months compared with pre-treatment levels [
]. These adverse effects on thyroid function were shown to be largely reversible in both men and women upon discontinuation of VPA monotherapy in one study [
VPA is established worldwide as one of the most widely used AEDs in the treatment of both generalized and partial seizures in adults and children.
The broad spectrum of antiepileptic efficacy of VPA is reflected in preclinical both in vivo and in vitro models, including a variety of animal models of seizures or epilepsy. VPA may have many adverse effects that require careful monitoring during the chronic treatment. In particular, its use is not recommended in patients with some pre-existing conditions e.g. hepatic and pancreatic insufficiency as well as in obese patients at risk of developing metabolic syndrome and in female pubertal patients because VPA can be expected to induce polycystic ovaries. In recent years, chronic use of VPA has been shown to exert variable effects on bone metabolism. There is no single mechanism of VPA action that can account for all the numerous effects of the drug on bone but the overall effect is an increased fracture rate. The main effects of VPA in bone include direct effects on bone cells through a decrease in osteoblast proliferation, changes in collagen synthesis and an induction of vitamin D catabolism. Furthermore, experimental and clinical observations show that many endocrine side effects, such as hypogonadism, hypothyroidism, hypercortisolemia and carnitine deficiency induced by VPA treatment, may also indirectly contribute to the increased bone fragility.
Further basic and controlled clinical trials are necessary to promote our understanding of the mechanisms of action of this broad spectrum antiepileptic drug on bone.
References
Burton B.S.
On the propyl derivatives and decomposition products of ethylacetate.
Fish liver oil and propolis as protective natural products against the effect of the anti-epileptic drug valproate on immunological markers of bone formation in rats.
L: -carnitine fumarate and isovaleryl-L: -carnitine fumarate accelerate the recovery of bone volume/total volume ratio after experimetally induced osteoporosis in pregnant mice.
Syndrome of inappropriate antidiuretic hormone secretion (SIADH) or hyponatraemia associated with valproic Acid: four case reports from the Netherlands and a case/non-case analysis of vigibase.
Assessment of low-density lipoprotein oxidation: paraoxonase activity, and arterial distensibility in epileptic children who were treated with anti-epileptic drugs.
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