FUTURE WATCH

H-D Nah

Authors’ affiliations: Hyun-Duck Nah, Department of Biochemistry, University of Pennsylvania School of Dental Medicine, and Department of Orthodontics, Temple University School of Dental Medicine, Philadelphia, PA Correspondence to: Hyun-Duck Nah, DMD, PhD Assistant Professor Department of Biochemistry University of Pennsylvania School of Dental Medicine 4001 Spruce Street Philadelphia, PA 19104 USA Tel: +1 215 573 6081 Fax: + 1 215 898 3695 E-mail: [email protected]

Suture biology: Lessons from molecular genetics of craniosynostosis syndromes Abstract: Sutures are critical growth sites in the developing craniofacial skeleton, and equally importantly, they are the target tissues for certain orthodontic therapeutic modalities. For these reasons, suture biology has long been an area of significant interest in orthodontics. In fact, much of our knowledge on sutures, such as structure, ontogeny, growth and development, pathology, and tissue response to biomechanical forces, is based on investigations conducted by orthodontic researchers in the 1950 – 1970s. Despite their significant impact in the establishment of the current paradigm in suture biology, these earlier studies have not provided much information on the molecular and cellular mechanisms of suture development. Only in the last 5 or 6 years, identification of genes involved in genetic disorders of premature suture closure (syndromes of craniosynostosis) has uncovered the first clues to some key molecules and their specific roles in this process. The list, which is expected to get longer, includes fibroblast growth factor receptors (FGFRs), MSX2, and

TWIST. FGFRs belong to a family of cell membrane receptor kinases and bind to their principal ligands, fibroblast growth factors (FGFs). Upon binding to FGFs, FGFRs transduce intracellular signaling cascades and ultimately modify cellular behavior, such as cell proliferation and differentiation. MSX2 and TWIST are transcription factors that bind to regulatory regions of target effector genes and determine their expression. In addition to the above-mentioned molecules, recent tissue recombination and biochemical studies have added FGFs and transforming growth factors to the list of key molecules in suture biogenesis. The purpose of this article is not to reiterate the earlier classical studies, which have been extensively reviewed by others, but to present the new and exciting developments in the field of suture research at the To cite this article: Clin. Orthod. Res. 3, 2000; 37 – 45 Nah H-D: Suture biology: Lessons from molecular genetics of craniosynostosis syndromes

genetic and cellular levels. These new perspectives and what

Copyright © Munksgaard 2000

FGFR; growth and development; growth modification; MSX2 ;

ISSN 1397-5927

syndromes; TWIST

lies ahead as future challenges in this field are discussed. Key words: cranial sutures; craniosynostosis; facial sutures;

Nah. Biology of sutural synostosis

Introduction Sutures are a rigid type of articulation, where the approximating margins of cranial and facial bony plates are joined by a thin layer of fibrous tissue. They are found only in the skull. Although some histological differences exist between cranial and facial sutures in early morphogenesis, at maturity they both eventually form a three-layered structure: bony margin – fibrous mesenchyme–bony margin (1). This simple-looking structure is not only a meeting place for the two bony margins, but it also acts as a shock absorber. Most importantly for the clinician, it plays an essential role in the growth and development of the craniofacial skeleton. Appositional growth at the suture accounts for a major portion of the total craniofacial skeletal growth. In this capacity, they serve as major growth sites in developing children. Unlike synchondroses in the cranial base, however, sutures do not grow in an autonomous manner (2). Instead, bony edges in sutures grow passively to compensate the growth of the soft-tissue envelope, brain, and airway. Sutures show remarkable plasticity along their bony margins in response to biomechanical forces (3). Earlier studies have repeatedly confirmed that compressive forces applied across sutures reduce bone deposition and induce bone resorption, while tensional forces increase bone deposition. This response characteristic makes sutures important target areas for orthodontic/orthopedic appliances designed to control vertical and transverse growth of the maxilla, such as palatal expander and cervical, high-pull and protraction headgears. Thus, it comes as no surprise that suture biology has long been a major field of interest to orthodontists. The pioneering studies published in the 1950–1970s by orthodontic researchers reflect their fascination with these tissues. Their reports form our basic knowledge of suture structure, ontogeny, growth and development, pathology, and tissue responses to biomechanical forces. These classical studies have been extensively reviewed by others (2 – 5). As much as these early studies make up the basis of what most of us know about sutures today, they do not explain the molecular components and mechanisms involved in the regulation of their development. After more than a decade of a semi-dormant period in the 1980s and early 1990s, concerted efforts from clinical geneticists and molecular biologists, facilitated by major advances and breakthroughs in biomedical sciences, have resulted in the 38 2Clin Orthod Res 3, 2000/37–45

identification of several genes that are key players in suture formation. These and subsequent investigations during the last 5–6 years have uncovered the first clues to the complex molecular mechanisms underlying the normal and abnormal suture formation, providing us with a new understanding of suture biology at the genetic and cellular levels. These recent and exciting developments will be the focus of this review. Since the new knowledge of suture biology comes from studies of the genes linked to inherited diseases affecting the normal suture development, especially the timing of suture closure, they will be discussed first. It will be followed by studies in other molecules implicated in suture formation, in particular growth factors. Finally, new perspectives in suture biology and what lies ahead as future challenges in this field will be discussed.

Craniosynostosis: genes and mechanisms Craniosynostosis, which refers to premature fusion of calvarial bones at the sutures, is a relatively common birth defect and occurs about 1 in 2000–3000 individuals (6). It is frequently found as part of syndromic features, but it also appears in an isolated non-syndromic form. The clinical consequences of prematurely fused cranial sutures are cranial and facial asymmetry, tower-shaped head, midface deficiency, hypertelorism with exophthalmia, and occasional neurological complications, depending on the location of affected sutures and the timing of the fusion (Fig. 1). The list of the genes that have been identified in association with craniosynostosis syndromes includes fibroblast growth factor receptors (FGFRs), MSX2, and TWIST. FGFRs

To date, four craniosynostosis syndromes, known as Apert, Pfeiffer, Jackson–Weiss, and Crouzon syndromes, have been linked to mutations in the FGFR genes (7, 8) These syndromes are inherited as an autosomal dominant trait or found occasionally in a sporadic pattern due to newly acquired mutations. Because similar craniofacial features result from premature suture fusion, clinical distinctions among these syndromes are usually made based on the severity of non-craniofacial phenotypes, i.e., foot and hand phenotypes characterized by soft-tissue and bony tissue syndactyly (mitten-like hands and feet) and/or

Nah. Biology of sutural synostosis

Fig. 1. A patient with Apert’s syndrome. A) The front facial view of the patient features hypertelorism, ptosis, and high forehead. B) The lateral demonstrates the high tower-shaped head, exophthalmosis, and midface deficiency. C,D) Non-craniofacial phenotypes includes hand and foot anormalies characterized by soft-tissue and bony tissue syndactyly (mitten-like hands and feet).

broadened first digits (Fig. 1)(7). The most severe of these syndromes is Apert’s syndrome, followed by Pfeiffer’s and Jackson–Weiss syndromes. Crouzon’s syndrome displays the mildest phenotype, featuring only the craniofacial anomaly. The first successful linkage of FGFR gene mutations to craniosynostosis syndromes was the identification of a mutation in the fibroblast growth factor receptor-1 gene (FGFR1 ) in a family with Pfeiffer’s syndrome. Since then, many other mutations in FGFR2 and FGFR3 have been linked to the above syndromes, implicating crucial roles played by FGFRs in the normal development as well as patency of cranial sutures. What are the molecular nature and the functions of FGFRs? As the name implies, they are the cell surface receptors for fibroblast growth factors (FGFs), which elicit a variety of cellular functions in different cell types, including cell proliferation, differentiation, migration, and pattern formation (9). FGFRs belong to a family of proteins known as transmembrane tyrosine kinases. So far, four FGFRs (FGFR1 – 4) have been identified. They all consist of an extracellular ligand-binding domain, a transmembrane domain, and intracellular kinase domains (Fig. 2). Notably, these receptors differ in their ligand specificity (10) and tissue distribution (11, 12). For example, in calvarial bones, FGFR1 is mainly expressed in

differentiated osteoblasts while FGFR2 is expressed in preosteoblasts. On the other hand, FGFR3 is found mostly in cartilagenous tissues. The expression of FGFR4 has not been determined in craniofacial bones. The principal function of FGFRs is to transduce extracellular FGF signals to the intracellular compartment, where a cascade of specific biochemical events lead to activation or repression of target genes and proteins. Ultimately, cellular behavior, such as proliferation and differentiation, is modified (9). At the molecular level, FGFR-mediated signal transduction is triggered by binding of FGFs to the extracellular domains of the FGFRs, followed by dimerization of FGFRs, which in turn induces trans-phosphorylation of the intracellular kinase domain of one receptor molecule by the other in the dimer (Fig. 2). The phosphorylated FGFR kinases then selectively phosphorylate intracellular molecules switching on appropriate signaling pathways. The importance of FGFR functions in normal suture development is amply demonstrated (7, 8) from molecular genetic studies of craniosynostosis syndromes associated with FGFR mutations. What, then, are the normal functions of FGFRs in suture development, and how do mutations affect normal functions of FGFRs and eventually development of the suture? FGFR mutations linked to craniosynostosis are mostly single amino acid substitution mutations. Most of them induce dimerization of FGFRs in the absence of FGF ligands, leading to subsequent activation of the FGFR kinase and signaling pathways (Fig. 2). Thus, mutant FGFRs are expected to be constitutively active. This is in contrast to physiological FGF/FGFR signaling, in which availability of specific FGF ligands is the key factor for controlled activation of FGFRs. A recent study investigated the effect of one of the Apert’s mutations in FGFR2 on the calvarial bone cell phenotype (13). The histological analysis of calvarial bone from these patients showed accelerated ossification and an increased thickness of subperiosteal bone compared to normal calvarial bone. Also, analyses of calvarial bone cells from the same patients for the alkaline phosphatase activity and expression of other osteogenic markers confirmed a correlation between Apert’s genotype and increased osteogenic commitment and maturation. Thus, it is quite conceivable that the constitutively activated FGFR2 function by the Apert’s mutation is the direct cause of increased osteogenic activity in cells at the calvarial bony margins that lead to premature suture closure. It is not known, however, whether or not increased osteogenic activity is comClin Orthod Res 3, 2000/37–452

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Fig. 2. A diagram illustrating the FGF signaling mediated by normal and mutant FGFRs in calvarial osteoblasts. FGFRs consist of extracellular ligand-binding, transmembrane, and intracellular kinase domains. Upon binding to the FGF and heparan sulfate complex, normal FGFRs dimerize, autophosphorylate the intracellular kinase domain, and transduce the intracellular signaling. The FGF signaling modulates gene expression in calvarial bone cells, influencing cellular events associated with proliferation and differentiation. In contrast, mutant FGFRs harboring craniosynostosis dimerize independent of FGFs, autophosphotylate the intracellular kinase domain, and perpetually activate the intracellular signaling pathway.

mon in synostotic calvarial bone from patients with other types of FGFR mutations. Further studies are needed to establish this cause-and-effect relationship. MSX2

The first molecular delineation of craniosynostosis syndrome came from the identification of the mutation in the MSX2 gene in Boston-type craniosynostosis (14). Bostontype craniosynostosis is also an autosomal dominant disorder and is characterized by typical craniofacial features (resulting from premature suture fusion), but without any other clinical phenotype. Thus, it represents an isolated form of craniosynostosis. The MSX2 gene encodes for a transcription factor that belongs to a family of homeobox-containing genes. The 40 2Clin Orthod Res 3, 2000/37–45

homeobox refers to a highly conserved DNA-binding domain found in a family of transcription factors involved in pattern formation during development. The homeobox domain of the MSX2 is also expected to bind regulatory DNA sequences in specific target genes and determines if and when they should be expressed. The Boston-type craniosynostosis is a single-base mutation in the homeobox domain. Interestingly, this mutation enhances DNAbinding affinity of the MSX2 homeobox domain without altering target gene specificity (15). In this case, the mutant MSX2 is likely to stay bound to target genes for a prolonged period, resulting in an enhanced MSX2 activity in cells. Therefore, the gain of function could be the underlying molecular basis for the Boston-type craniosynostosis phenotype. This contention is further supported by a transgenic animal model in which

Nah. Biology of sutural synostosis

over-expression of the MSX2 gene at the suture area in mice is correlated with premature suture closure and ectopic membranous bone formation (16). Boston-type craniosynostosis and the associated mutation in the MSX2 gene have been reported only in one family so far. Importantly, its expression in the osteogenic front of the developing calvarial bone at the suture (14) and successful induction of craniosynostosis by over-expression of the MSX2 gene in transgenic mice (16) confirm that the MSX2 gene function is necessary for normal development of sutures. The exact role of the MSX2 in suture development, however, is not known. Similar to other homeobox-containing genes, the MSX2 gene plays an important role in pattern formation in the developing embryo. It is also involved in other important developmental events, such as programmed cell death (apoptosis) (17, 18) and tissue interaction (19). Therefore, it is quite plausible that the mutated MSX2 gene product might either alter apoptotic rate in suture cells or interaction between sutures and adjacent tissues, or both, to promote the abnormal development of sutures in Bostontype craniosynostosis. TWIST

Mutations in the TWIST gene have been identified in a subset of Saethre – Chotzen syndrome, which is probably the most common autosomal dominant disorder of craniosynostosis in man (1 in 25000) (20). This syndrome is characterized by craniofacial and limb anomalies similar to the craniosynostosis syndrome associated with FGFR mutations (20, 21). The most common dismorphic facial features in these patients are an abnormal head shape resulting from premature cranial suture closure, hypertelorism, and midface deficiency. Other less frequently found craniofacial phenotypes include cleft palate, malocclusion, enamel hypoplasia, deviated nasal septum, low-set hairline, a high forehead, ptosis, and defective hearing. The typical limb abnormalities seen in these patients are brachydactyly and soft-tissue syndactyly. The protein product of the TWIST gene is a transcription factor containing a highly conserved DNA and protein-binding basic helix-loop-helix (b-HLH) motif. Similar to MSX2, TWIST regulates the expression of a number of genes by binding to regulatory regions of the target genes. In mice, it is expressed in the developing embryonic head, mesenchymal, and limb bud cells, and later in neonatal calvarial osteoblasts (22, 23). This gene expres-

sion pattern correlates well with craniofacial and limb phenotypes observed in patients with Saethre–Chotzen syndrome. The expression analysis of the TWIST gene has not yet been done in humans, but it is expected to be similar to that in mice. The TWIST mutations associated with Saethre– Chotzen syndrome either truncate or disrupt the DNAbinding domain of the TWIST. Consequently, the TWIST mutants fail to interact with target genes, suggesting loss of function as the underlying molecular basis for Saethre– Chotzen syndrome. This view is further supported by knock-out mouse studies (24). Heterozygous knock-out mice, containing only one functional TWIST allele, produce one-half of the amount of TWIST protein compared to normal mice (haploinsufficiency). Thus, the autosomal dominant nature of the Saethre–Crotzen mutation is recreated in these mice. The phenotype of heterozygous mice is quite similar to the Saethre–Crotzen phenotype, displaying skull and limb defects. Meanwhile, homozygous knock-out mice, completely lacking TWIST protein expression from both alleles, die early in utero, explaining why patients with recessive TWIST mutations have not been found. It can be seen, unlike FGFRs, normal development of the suture is sensitive to the level of TWIST expression. One of the most exciting findings of the TWIST gene studies is the first clue to the regulatory cascades of the molecular components involved in suture development. Studies in Drosophilae have provided several lines of evidence that TWIST may function as an upstream regulator of FGFRs. It was shown that activation of the DFR1 gene (a homologue of FGFRs in Drosophila) requires TWIST (25), and these two genes are expressed in similar regions. Additionally, the close resemblance of phenotypes among patients with the FGFR mutation and the TWIST mutation further strengthens the evidence for the same developmental pathways shared by FGFRs and TWIST.

Growth factors: TGF-b and FGF Molecular genetic studies linking specific mutations to craniosynostosis syndromes have established FGFRs, MSX2, and TWIST as key factors in suture formation. In most cases of craniosynostosis, however, the etiology remains unknown. Clearly, one can foresee that many more molecules are yet to be identified before the list of components involved in suture development is complete. Clin Orthod Res 3, 2000/37–452

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That list might also include growth factors, such as transforming growth factor-b (TGF-b) and FGF. Earlier tissue recombination studies have demonstrated that cranial suture patency depends on underlying dura mater, the fibrous tissue interposed between brain and cranial bones (26–28). In these studies, fetal or neonatal rat coronal sutures were transplanted or co-cultured with or without dura mater. Sutures without dura mater were soon obliterated by bone, while sutures with dura mater resisted ossification, suggesting that tissue interaction of biochemical nature is required to maintain sutures open. Interestingly, this study also demonstrated that suture obliteration can be prevented when sutures stripped of dura mater were co-cultured with dura mater on the opposite side of a Millipore filter, which allows only low-molecular weight soluble molecules to travel across it. These studies concluded, therefore, that the influence of dura mater on suture patency is mediated by soluble factors rather than cell – cell or cell – matrix interactions. A later biochemical study has demonstrated that activities of the factors derived from dura mater are heparan sulfate dependent (27), suggesting that they could be TGF-b and/or FGF. Since the first characterization of FGF as a fibroblast mitogen, 17 members have been added to the FGF family to date. Diverse biological activities, such as differentiation, angiogenesis, embryonic development, and tumorogensis, are now associated with FGFs. Members of the FGF family bind to specific FGFRs with high affinity in the presence of heparan sulfate, a co-receptor (Fig. 2) (29). Evidence for the role of FGF signaling in suture formation is clearly supported by the identification of FGFR mutations in the craniosynostosis syndromes as mentioned above. Also, locally applied FGF2 to fetal mouse coronal suture in utero leads to ectopic bone formation at the suture area (30). Similarly, FGF4 delivered to the osteogenic front of calvarial bone in organ culture accelerates suture formation (30). Suture mesenchyme and underlying dural tissue produce FGF2, FGF9 (30), and possibly some other FGFs. Therefore, it is likely that signals generated by specific FGFs that are synthesized by dural cells regulate suture development via the FGFRs present in the advancing osteogenic front. TGF-b1-3 belongs to a family of related heparan-binding peptides. They are involved in diverse biological activities in normal growth and development of various organs, including regulation of osteoblast and chondrocyte differentiation, proliferation, and matrix gene expression. These activities are mediated by high affinity receptors that are 42 2Clin Orthod Res 3, 2000/37–45

associated with intracellular signaling. Compelling evidence for the involvement of TGF-b in suture formation is obtained from a series of expression studies. Continued expression of TGF-b1 and 2 is shown in association with obliterating sutures, while increased expression of TGF-b3 is found in non-fusing sutures (31). All three isoforms of TGF-b are expressed in dural tissues (31). Therefore, these studies, together with data from tissue recombination studies described above, suggest that different isoforms of TGF-b might determine calvarial bone growth at the sutures as well as their patency. Interestingly, however, no mutation in the TGF-b genes has been linked to craniosynostosis syndromes so far, indicating the complexity and functional redundancy of regulatory mechanisms that govern suture formation.

Perspectives and challenges Molecular genetic studies of naturally occurring mutations in craniosynostosis syndromes have identified several genes involved in regulation of suture development and yield new insights into both normal and abnormal suture formation. These studies have also been catalysts for the recent progress in the field of suture research. To date, more than 100 craniosynostosis syndromes have been described. Of those, six syndromes have been assigned to specific genes. The remarkable enthusiasm of academic and pharmaceutical researchers to identify the disease-causing mutations will rapidly bring about identification of additional genes involved in suture development. The real challenge, however, is the delineation of the molecular mechanisms by which these genes regulate suture formation. It is necessary to define the function of each molecular component and to determine how the interplay among these components contribute to suture formation. This will require concerted efforts from scientists in various disciplines, such as genetics, pathology, molecular biology, biochemistry, cell biology, developmental biology, mouse genetics, and the like. Sutures are fascinating structures in which a few cell layers of fibrous mesenchyme interposed between approximating bony edges resist mineralization. This property is critically important for normal growth and development of the craniofacial skeleton as is shown in craniosynostosis syndromes. An interesting aspect of craniosynostosis syndromes is that these mutated genes affect cranial sutures but not facial sutures. This demonstrates that despite

Nah. Biology of sutural synostosis

the morphological similarity between cranial and facial sutures, differences in biochemical and/or molecular nature exist between them. This could be because cranial suture formation requires interaction with underlying dural tissue while the facial suture develops in the absence of such tissues. Growth factors produced by the dural tissue could significantly influence the molecular and biochemical makeup of overlying cranial suture cells. It could also be reasoned that cells in facial and cranial sutures have different embryonic origins. It is possible that the mechanisms for keeping the facial suture open may not be same as that for the cranial suture. Currently, there are no data available that compare the differences between facial and cranial sutures at the molecular level. What keeps the suture from closing? Based on the known functions of genes expressed in the sutures and genes associated with craniosynostosis syndromes, it is clear that suture patency as well as development are regulated by complex mechanisms, including tissue interaction, cell signaling, and activities of specific transcription factors. As we add more names to the list of key players in suture development, regulation of this process will likely turn out to be more complex than is currently believed. A complete picture of molecular and cellular mechanisms for suture development will not be within sight in the very near future. Yet, a detailed delineation of the regulatory pathways governed by several key factors may unveil important common pathways shared by many molecules. Only then will we be able to influence suture development to our advantage by preventing or intervening in sutural pathogenesis. One can also envision that future orthodontists will be able to manage maxillary skeletal growth anomalies by modifying specific gene activities or signaling pathways in circummaxillary sutural cells to facilitate growth modification in the maxilla as well as adjacent bones. Such a remedy could even include gene therapy.

Abstrakt Suturen sind kritische Wachstumszonen des sich entwickelnden Gesichtsscha¨dels und gleichfalls bedeutend, weil sie der Gewebeangriffspunkt fu¨r bestimmte kieferorthopa¨dische Behandlungsmaßnahmen sind. Aus diesen beiden Gru¨nden war die Biologie der Suturen schon lange ein Gebiet besonderen Interesses in der Kieferorthopa¨die. So basiert unser Wissen u¨ber Suturen, wie zum Beispiel Struktur, Entstehung, Wachstum und Entwicklung, Pathologie und Reaktion auf biomechanische Kra¨fte, vielfach auf Untersuchungen, die von kieferorthopa¨dischen Wissenschaftlern zwischen 1950 und 1970 durchgefu¨hrt wurden. Trotz ihres bedeutenden Einflusses auf die Entstehung des gegenwa¨rtigen Paradigmas der Suturenbiologie haben diese fru¨heren Studien nicht viel Information

u¨ber den molekularen und zellula¨ren Mechanismus der Suturenentwicklung geliefert. Erst in den letzten 5 oder 6 Jahren hat die Identifizierung der Gene, die bei Erbkrankheiten mit vorzeitigem Suturenverschluß (Syndrome der Kraniosynostosen) eine Rolle spielen, erste Hinweise aufgedeckt, daß es fu¨r den Suturenverschluß einige Schlu¨sselmoleku¨le gibt und welche Rolle sie spielen. Die Aufza¨hlung dieser Schlu¨sselmoleku¨le, die vermutlich la¨nger werden wird, umfaßt die beiden Rezeptoren der Wachstumsfaktoren von Fibroblasten (FGFRs = fibroblast growth factor receptors), MSX2 und TWIST. FGFRs geho¨ren zur Familie der Zellmembran-Rezeptorkinasen, die sich mit ihren Hauptliganden, den FibroblastenWachstumsfaktoren (FGFs =fibroblast growth factors), verbinden. Bei der Bindung an FGFs u¨bertragen FGFRs eine Kaskade intrazellula¨rer Signale, die letzten Endes das Zellverhalten wie beispielsweise die Proliferation und Differenzierung vera¨ndern. MSX2 und TWIST sind Transkriptionsfaktoren, die sich an die Regulationsgebiete von Zieleffektorgenen binden und dadurch deren Wirkung bestimmen. Ju¨ngere Geweberekombinationen und biochemische Untersuchungen haben dazu gefu¨hrt, FGFs und TGF-b (transforming growth factor) zusa¨tzlich zu den oben erwa¨hnten Moleku¨len in die Liste der Schlu¨sselmoleku¨le fu¨r die Suturenbiogenese aufzunehmen. Der Sinn dieses Artikels ist es nicht, fru¨here klassische Studien zu wiederholen, die durch andere ausreichend bewertet wurden, sondern neue und spannende Entwicklungen im Gebiet der Suturenforschung auf genetischer und zellula¨rer Ebene darzustellen. Diese neuen Ausblicke werden ebenso diskutiert wie das, was an zuku¨nftigen Herausforderungen in diesem Gebiet vor uns liegt.

Abstracto Las suturas son a´reas de crecimiento indispensables para el desarrollo del esqueleto craniofacial. Igualmente importantes, son tejidos claves para ciertas modalidades terape´uticas ortodonticas. Por estas razones, la biologı´a de sutura siempre ha sido un a´rea de intere´s significativo en la ortodoncia. De hecho, mucho de lo que se conoce acerca de las suturas como la estructura, la ontogenia, el crecimiento y desarrollo, la patologı´a y la respuesta del tejido a fuerzas biomeca´nicas, es basado en estudios conducidos por investigadores ortodonticos desde la de´cada del 50 hasta la del 70. A pesar del impacto significativo en el establecimiento del paradigma actual en la biologı´a de sutura, estos estudios anteriores no han proveı´do mucha informacio´n acerca de los mecanismos tanto moleculares como celulares en el desarrollo de las suturas. Recientemente, en los u´ltimos cinco o seis an˜os, la identificacio´n de los genes envueltos en desordenes gene´ticos de cierre prematuro de sutura (sı´ndrome de craniosinostosis) han descubierto las primeras pistas para algunas mole´culas claves y su desempen˜o especı´fico en este proceso. La lista, la cual se espera que aumente en taman˜o, incluye receptores de factor de crecimiento de fibroblasto (FGFRs), MSX2 y TWIST. FGFRs pertenecen a una familia de receptores kinasas de la membrana celular y se enlazan a sus ligandos principales, factores de crecimiento de fibroblasto (FGFs). Luego de enlazarse al FGFs, el FGFRs transduce cascadas intracelulares de sen˜alamiento, y finalmente modifica el comportamiento celular tales como la proliferacio´n celular y la diferenciacio´n. MSX2 y TWIST son factores de transcripcio´n que se enlazan a regiones reguladoras de genes efectores claves y determinan su expresio´n. Adema´s de las mole´culas recientemente mencionadas, recombinaciones recientes de tejidos y estudios bioquı´micos han an˜adido FGFs y sectores de crecimiento de transformacio´n (TGF-b) a la lista de las mole´culas claves para la biogenesis de sutura. El propo´sito de este artı´culo no es el de reiterar los estudios cla´sicos anteriores, los cuales han sido revaluados extensamente por otros, sino presentar el desarrollo nuevo y excitante en el campo de las investigaciones de las suturas a nivel celular y gene´tico. Estas nuevas perspectivas y lo previsto como un reto en el futuro, es discutido en este estudio. Clin Orthod Res 3, 2000/37–452

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References 1. Prichards JJ, Scott JH, Girgis FS. The structure and development of cranial and facial sutures. J Anat 1956;90:73– 86. 2. Kokich VG. The biology of sutures. In: Cohen MM, editor. Craniosynostosis: Diagnosis, Evaluation, and Management. New York: Raven Press; 1986. pp. 81 – 103. 3. Wagemans PAH, van der Velde J, Kuijpers-Jagtman AM. Sutures and forces: a review. Am J Orthod Dentofac Orthop 1988;94:129– 41. 4. Persson M. The role of sutures in normal and abnormal craniofacial growth. Acta Odontol Scand 1995;53:152– 61. 5. Cohen MM. Suture biology and correlates of craniosynostosis. Am J Med Genet 1993;47:581– 616. 6. Wilkie AO. Craniosynostosis: genes and mechanisms. Hum Mol Genet 1997;6:1647– 56. 7. Muenke M, Schell U. Fibroblast-growth-factor receptor mutations in human skeletal disorders. Trends Genet 1995;11:308–13. 8. de Moerlooze L, Dickson C. Skeletal disorders associated with fibroblasts growth factor receptor mutations. Curr Op Genet Dev 1997. 9. Wilkie AOM, Morriskay GM, Jones EY, Heath JK. Functions of fibroblast growth factors and their receptors. Curr Biol 1995;5:500– 7. 10. Ornitz DM et al. Receptor specificity of the fibroblast growth factor family. J Biol Chem 1996;271:15292– 7. 11. Patstone G, Pasquale EB, Maher PA. Different members of the fibroblast growth factor receptor family are specific to distinct cell types in the developing chicken embryo. Dev Biol (Orlando) 1993;155:107– 23. 12. Wilke TA, Gubbels S, Schwartz J, Richman JM. Expression of fibroblast growth factor receptors (FGFR1, FGFR2, FGFR3) in the developing head and face. Dev Dyn 1997;210:41– 52. 13. Lomri A et al. Increased calvaria cell differentiation and bone matrix formation induced by fibroblast growth factor receptor 2 mutations in Apert syndrome. J Clin Invest 1998;101:1310–7. 14. Jabs EW et al. A mutation in the homeodomain of the human MSX2 gene in a family affected with autosomal dominant craniosynostosis. Cell 1993;75:443– 50. 15. Ma L, Golden S, Wu L, Maxson R. The molecular basis of Boston-type craniosynostosis: the Pro148-- \His mutation in the N-terminal arm of the MSX2 homeodomain stabilizes DNA binding without altering nucleotide sequence preferences. Hum Mol Genet 1996;5:1915– 20. 16. Liu YH et al. Premature suture closure and ectopic cranial bone in mice expressing Msx2 transgenes in the developing skull. Proc Natl Acad Sci USA 1995;92:6137– 41. 17. Marazzi G, Wang Y, Sassoon D. Msx2 is a transcriptional regulator in the BMP4-mediated programmed cell death pathway. Dev Biol (Orlando) 1997;186:127– 38. 18. Graham A, Koentges G, Lumsden A. Neural crest apoptosis and the establishment of craniofacial pattern: an honorable death. Mol Cell Neurosci 1996;8:76– 83. 19. Maas R, Bei M. The genetic control of early tooth development. Crit Rev Oral Biol Med 1997;8:4– 39. 20. Howard TD et al. Mutations in TWIST, a basic helix-loop-helix transcription factor, in Saethre – Chotzen syndrome. Nat Genet 1997;15:36– 41. 21. el Ghouzzi V et al. Mutations of the TWIST gene in the Saethre– Chotzen syndrome. Nature Genet 1997;15:42– 6.

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Nah. Biology of sutural synostosis

22. Murray SS, Glackin CA, Winters KA, Gazit D, Kahn AJ, Murray EJ. Expression of helix-loop-helix regulatory genes during differentiation of mouse osteoblastic cells. J Bone Miner Res 1992;7:1131– 8. 23. Stoetzel C, Weber B, Bourgeois P, Bolcato-Bellemin AL, PerrinSchmitt F. Dorso-ventral and rostro-caudal sequential expression of M-twist in the postimplantation murine embryo. Mech Dev 1995;51:251– 63. 24. Chen ZF, Behringer RR. Twist is required in head mesenchyme for cranial neural tube morphogenesis. Genes Dev 1995;9:686– 99. 25. Shishido E, Higashijima S, Emori Y, Saigo K. Two FGF-receptor homologues of Drosophila: one is expressed in mesodermal primordium in early embryos. Development 1993;117:751–61. 26. Opperman LA, Chhabra A, Nolen AA, Bao Y, Ogle RC. Dura mater maintains rat cranial sutures in vitro by regulating suture cell proliferation and collagen production. J Craniofac Genet Dev Biol 1998;18:150– 8. 27. Opperman LA, Passarelli RW, Morgan EP, Reintjes M, Ogle RC. Cranial sutures require tissue interactions with dura mater t

28.

29.

30.

31.

resist osseous obliteration in vitro. J Bone Miner Res 1995;10: 1978 – 87. Opperman LA, Sweeney TM, Redmon J, Persing JA, Ogle RC. Tissue interactions with underlying dura mater inhibit osseous obliteration of developing cranial sutures. Dev Dyn 1993;198:312– 22. Ornitz DM, Yayon A, Flanagan JG, Svahn CM, Levi E, Leder P. Heparin is required for cell-free binding of basic fibroblast growth factor to a soluble receptor and for mitogenesis in whole cells. Mol Cell Biol 1992;12:240– 7. Iseki S, Wilkie AO, Heath JK, Ishimaru T, Eto K, Morriss-Kay GM. Fgfr2 and osteopontin domains in the developing skull vault are mutually exclusive and can be altered by locally applied FGF2. Development 1997;124:3375– 84. Opperman LA, Nolen AA, Ogle RC. TGF-beta 1, TGF-beta 2, and TGF-beta 3 exhibit distinct patterns of expression during cranial suture formation and obliteration in vivo and in vitro. J Bone Miner Res 1997;12:301– 10.

Clin Orthod Res 3, 2000/37–452

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