Growth plate closure and therapeutic interventions
Article information
Abstract
Height gains result from longitudinal bone growth, which is largely dependent on chondrocyte differentiation and proliferation within the growth plates of long bones. The growth plate, that is, the epiphyseal plate, is divided into resting, proliferative, and hypertrophic zones according to chondrocyte characteristics. The differentiation potential of progenitor cells in the resting zone, continuous capacity for chondrocyte differentiation and proliferation within the proliferative zone, timely replacement by osteocytes, and calcification in the hypertrophic zone are the 3 main factors controlling longitudinal bone growth. Upon adequate longitudinal bone growth, growth plate senescence limits human body height. During growth plate senescence, progenitor cells within the resting zone are depleted, proliferative chondrocyte numbers decrease, and hypertrophic chondrocyte number and size decrease. After senescence, hypertrophic chondrocytes are replaced by osteocytes, the extracellular matrix is calcified and vascularized, the growth plate is closed, and longitudinal bone growth is complete. To date, gonadotropin-releasing hormone analogs, aromatase inhibitors, C-type natriuretic peptide analogs, and fibroblast growth factor receptor 3 inhibitors have been studied or used as therapeutic interventions to delay growth plate closure. Complex networks of cellular, genetic, paracrine, and endocrine signals are involved in growth plate closure. However, the detailed mechanisms of this process remain unclear. Further elucidation of these mechanisms will enable the development of new therapeutic modalities for the treatment of short stature, precocious puberty, and skeletal dysplasia.
Key message
Height gains result from longitudinal bone growth. Upon adequate growth, growth plate closure limits longitudinal bone growth. To date, gonadotropin-releasing hormone analogs, aromatase inhibitors, C-type natriuretic peptide analogs, and fibroblast growth factor receptor 3 inhibitors have been studied or used as therapeutic interventions to delay growth plate closure and increase human height. The development of more effective therapeutic modalities for short stature, precocious puberty, and skeletal dysplasia is anticipated.
Graphical abstract. Growth plate closure mechanism and therapeutic interventions used to delay its senescence to increase human height. GH, growth hormone; IGF-1, insulin-like growth factor-1; GnRHa, gonadotropin-releasing hormone analog; AI, aromatase inhibitor; FGFR-3, FGF receptor-3; CNP, C-type natriuretic peptide; FGF, fibroblast growth factor.
Introduction
Longitudinal bone growth results from chondrogenesis at the growth plate and endochondral ossification [1-5]. Chondrocyte hypertrophy, differentiation, and proliferation as well as extracellular matrix secretion continue during growth plate maturation [6]. Once adequate growth plate maturation is achieved,unknown signals cause growth plate senescence and closure. At that time, progenitor cells within the resting zone become depleted, proliferative chondrocyte numbers decrease, and hypertrophic chondrocyte numbers and size decrease. Subsequently, hypertrophic chondrocytes are replaced by osteocytes, the extracellular matrix becomes calcified and vascularized, and longitudinal bone growth ends [7].
Although the detailed mechanisms of these processes remain unclear, many pathways associated with genetic, intracellular, extracellular, autocrine, paracrine, and endocrine factors are involved [8,9]. Thus, the elucidation of the mechanisms underlying growth plate maturation, senescence, and closure may provide an opportunity to develop new treatment strategies for various growth disorders. This review details evidence-based reports on the mechanisms of and therapeutic trials of treatments to delay growth plate closure.
Growth plate structure, function, and closure
The growth plate, located between the epiphysis and metaphysis of the long bone, is divided into 3 zones according to the characteristics of the chondrocytes within each. The resting zone comprises small chondrocytes that act as progenitor cells with slow replication rates. The proliferative zone contains flat chondrocytes that line the long axis of the bone and replicate quickly. The hypertrophic zone is a layer of chondrocytes undergoing terminal differentiation that features increased thickness, surrounds a calcified matrix, and attracts factors for bone and vessel formation [1-5] (Fig. 1A).
During linear growth, growth plate thickening and maturation, chondrocyte proliferation and differentiation, extracellular matrix secretion, hypertrophic zone calcification, osteoblast invasion and differentiation, and blood vessel formation processes repeat continuously [4-9]. However, at the end of puberty, these processes cease and longitudinal bone growth is completed via the growth plate senescence and closure processes [8,9] (Fig. 1B).
1. Resting zone
Progenitor cells within the resting zone can continuously differentiate and proliferate into chondrocytes during growth plate maturation but lose this capacity during growth plate senescence [10-12]. Therefore, postnatal skeletal growth appears to be driven by the epiphyseal stem cell niche [13-15].
There are 2 hypotheses regarding programmed senescence of the growth plate: cell counting and a biological timing mechanism [10,12]. In a growth plate transplantation experiment, the growth rate of the transplanted growth plate was dependent on donor animal, but not recipient animal, age. This experiment showed the finite proliferative capacity of the growth plate similar to the cell-counting mechanism [16]. In addition, in a murine study, loss of telomerase activity had no major effects on skeletal growth, indicating that telomere shortening is not the primary mechanism limiting chondrocyte proliferation [17]. However, in vitro studies have shown that epigenetic changes in the methylation of genomic DNA may limit chondrocyte replication [18]. The cellular programmed factors related to these theories have yet to be clearly elucidated.
Several transcription factors are considered involved in progenitor cell differentiation (Table 1). The conversion of progenitor cells in the mesenchymal condensation into the chondrocyte lineage is controlled by SRY-Box transcription factor 9 (SOX9) expression. The subsequent chondrocyte differentiation is influenced by SOX9, SOX5, and SOX6 [19].
Genetic and paracrine factors controlling the growth plate function
Bone morphogenic protein (BMP) is a paracrine factor that promotes progenitor cell differentiation into proliferative chondrocytes [20].
2. Proliferative zone
Chondrocytes continue to proliferate and differentiate into prehypertrophic and hypertrophic chondrocytes before growth plate senescence under the influence of complex interactions among paracrine, endocrine, and transcription factors [1-6] (Fig. 2).
Molecular networks involved in chondrocyte differentiation, proliferation, and hypertrophy. SOX9, SRY-box transcription factor 9; BMP, bone morphogenic protein; PTHrP, parathyroid hormone-related peptide; IHH, Indian hedgehog; Runx2, runt-related transcription factor 2; FGF, fibroblast growth factor; CNP, C-type natriuretic peptide; HIF-1, hypoxia-inducible factor-1; VEGF, vascular endothelial growth factor; WNT, Wingless-type mouse mammary tumor virus integration site signaling; CC, calcified cartilage. Modified from Ağırdil et al. EFORT Open Rev 2020;5:498-507 with permission [5].
BMP is a paracrine signal that promotes progressive differentiation from the resting state through proliferation to hypertrophic chondrocytes. BMP is involved in perichondrium, periosteum, and osteoblast development [21-23].
Parathyroid hormone-related peptide (PTHrP), which is secreted from periarticular chondrocytes, diffuses across the growth plate cartilage to maintain chondrocyte proliferation [24].
Indian hedgehog (IHH), secreted by prehypertrophic and hypertrophic chondrocytes, stimulates PTHrP secretion and promotes the differentiation of proliferative chondrocytes into prehypertrophic chondrocyte [25-27].
Fibroblast growth factor (FGF) is also important for perichondria, periosteum, and osteoblast development. In vivo studies have indicated that FGF receptor-1 and FGF receptor-3 (FGFR-3) have growth-inhibiting effects and FGF receptor-2 has growth-promoting effects on longitudinal bone growth [28-29]. Runt-related transcription factor 2 (Runx2) is a transcription factor that is required for further chondrocyte differentiation and hypertrophy [20].
Various endocrine factors are involved in chondrocyte proliferation and differentiation (Table 2) [30]. Growth hormones (GHs) positively regulate insulin-like growth factor-1 (IGF-1) synthesis and secretion in the liver and growth plates. Longitudinal bone growth is stimulated by GH, circulating IGF-1, and locally secreted IGF-1 in the growth plate. Among them, IGF-1 produced locally from chondrocytes is especially important for chondrocyte differentiation, proliferation, and hypertrophy; moreover, it stimulates extracellular matrix production and ossification within the growth plate [31].
Endocrine factors control growth plate function
The binding of estrogen to its receptor (estrogen receptor [ER]) stimulates the GH-IGF-1 axis, especially during the pubertal growth spurt. Both receptor subtypes (ERα and ERβ) are related with the augmentation of GH secretion and expressed in the resting, proliferative, and hypertrophic zones of the growth plate [32-35]. There is evidence that growth plate closure occurs when the proliferative capacity of progenitor cells within the resting zone becomes exhausted. Estrogen can accelerate the exhaustion of proliferative potential and advance growth plate senescence. The binding of estrogen to each ER subtype is considered related to growth plate closure. ERα appears to be the dominant mediator of estrogen actions. ERβ has some repressive functions within the long bones in mice; however, its definite roles in the long bones of humans remain unclear. Estrogen also promotes bone formation and remodeling by stimulating osteoblastogenesis and inhibiting osteoclastogenesis [36-39].
Androgens with aromatized estrogen contribute to growth plate maturation and senescence through direct or indirect interactions with growth plate chondrocytes. In an organ culture study, testosterone stimulated chondrocyte proliferation with increased local IGF-1 production, while dihydrotestosterone promoted chondrocyte proliferation and proteoglycan synthesis within the growth plates. In growth plate closure, the function of androgen is attributed to the aromatization of androgen to estrogen in various peripheral tissues, including the growth plate cartilage [40,41].
Thyroid hormones stimulate chondrocyte proliferation, hypertrophy, and growth plate maturation [8].
Glucocorticoids inhibit longitudinal bone growth by inhibiting the GH-IGF-1 axis and chondrocyte proliferation as well as stimulating apoptosis of hypertrophic chondrocytes.It also delays growth plate senescence [8].
3. Hypertrophic zone
1) Replacement of chondrocytes by osteocytes
Chondrocytes in the hypertrophic zone are replaced by osteocytes during growth plate maturation and senescence. To the best of our knowledge,there are 4 theories (apoptosis, autophagy, hypoxia, and transdifferentiation) regarding the mechanism of cartilage replacement in bone tissue [11,42-46].
2) Blood vessel formation
Blood vessel formation and vascular invasion are critical for substituting avascular cartilage with vascular bone and marrow tissues. Vessel formation is mediated by hypoxia-inducible factor-1 (HIF-1), vascular endothelial growth factor (VEGF), Runx2, FGF, BMP, transforming growth factor, IGF, and platelet-derived growth factor within the growth plate [47-50].
3) Osteoblast differentiation and ossification
Five osteoblastic lineage differentiation steps (preosteoblasts, mature osteoblasts, osteoid osteocytes, early osteocytes, and mature osteocytes) are necessary to achieve the final development of bone tissue within the growth plate. Wingless-type mouse mammary tumor virus integration site/β-catenin signaling, Runx2, Osterix, BMP, IHH, and IGF are the required factors for differentiation [51].
During this process, bones can deposit minerals from the extracellular matrix that are rich in type I collagen, completing ossification and growth plate closure [52].
Therapeutic interventions for delaying growth plate closure
1. Gonadotropin-releasing hormone analog
Various preparations of gonadotropin-releasing hormone analogs (GnRHa), such as leuprolide acetate, triptorelin phosphate, and histrelin acetate, have been used as major treatment modalities for idiopathic central precocious puberty. Their effects are mediated by the desensitization of pituitary gonadotrophs occupying GnRH receptors, resulting in the suppression of gonadal sex steroid secretion and postponement of growth plate closure. The effects on adult height increase varied from 2 to 10 cm in children with precocious puberty and from 1.7 to 6.7 cm in adolescents with idiopathic short stature or growth-limiting syndromes. However, adverse effects on bone mineral density have been reported in adolescents with idiopathic short stature who do not undergo early-onset puberty [53-56].
2. Aromatase inhibitors
Aromatase inhibitors (AIs) inhibit aromatase, which converts androgen into estrogen. Letrozole, anastrozole, and exemestane, representative selective AI, were initially used to treat breast cancer in women and gynecomastia in men. A few reports have described their positive effects on adult height after their off-label use to delay growth plate closure. However, their general use to increase height in humans is not widely accepted because of the possibility of adverse effects on bone mineral density and vertebral deformity [57].
3. C-type natriuretic peptide analog
C-type natriuretic peptide (CNP) binds natriuretic peptide receptor-B and results in the transformation of guanosine 5 -triphosphate to cyclic guanosine monophosphate and the inhibition of mitogen-activated protein kinase (MAPK) and FGF signaling. These signaling pathways stimulate the latter steps of chondrocyte hypertrophy within the growth plate and promote growth plate closure. CNP analogs have been studied as treatment modalities in patients with achondroplasia, a disorder characterized by the activation mutation of FGFR-3, which results in earlier growth plate closure and a severely short stature. Two types of CNP analogs, a recombinant CNP analog (vosoritide) and transiently conjugated CNP (navepegritide), can inhibit MAPK and FGFR signaling and are new treatment methods for achondroplasia [58,59].
4. FGFR-3 inhibitor
FGF binds to FGFR-3 and stimulates Ras-MAPK signaling, inhibiting chondrocyte differentiation and proliferation. FGFR3 inhibitors have been studied as treatment modalities for achondroplasia because they block the Ras-MAPK pathway and stimulate chondrocyte differentiation and proliferation. Such inhibitors include the soluble FGFR3 decoy (recifercept), anti-FGFR3 monoclonal antibody (vofatamab), and FGFR3-selective tyrosine kinase inhibitor (infigratinib) [58,60].
Conclusion
Cellular and genetic factors related to the aging of progenitor cells within the resting zone and the transdifferentiation of chondrocytes into osteocytes within the hypertrophic zone are considered important in limiting height growth; however, their exact mechanisms of action remain unclear. Many paracrine and genetic factors are also involved in chondrocyte differentiation, proliferation, and hypertrophy within the cartilage as well as vascularization and ossification in the extracellular matrix of the growth plate; however, further studies of their interaction mechanisms are needed. Among endocrine factors, estrogen and the GH-IGF-I axis are important in chondrocyte differentiation, proliferation, and hypertrophy, but the exact mechanisms by which estrogen accelerates growth plate senescence remain unknown.
The complex interactions between the cellular, genetic, paracrine, and endocrine systems within the growth plates are closely related to the mechanisms of growth plate maturation, senescence, and closure. Until now, GnRHa, AIs, CNP analogs, and FGFR-3 inhibitors have been studied or used as the therapeutic modalities for delaying growth plate closure. Elucidating the detailed processes of growth plate physiology will increase our ability to explain the molecular mechanisms responsible for idiopathic short stature, central precocious puberty, and skeletal dysplasia as well as facilitate the development of more effective therapeutic modalities for these diseases.
Notes
Conflicts of interest
No potential conflict of interest relevant to this article was reported.
Funding
This study received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.