Expression of high-affinity receptors for TGF-beta during rat cranial suture fusion.

The etiology of craniosynostosis is unknown. The elucidation of the biological pathways responsible for this disorder has been hampered by an inability to evaluate cranial sutures before, during, and after cranial suture fusion. The programmed fusion of the rat posterofrontal (PF) suture postnatally provides an excellent model to study the molecular events that occur during cranial suture fusion. Previous experiments have implicated transforming growth factor beta (TGF-beta) growth factors in the regulation of PF suture fusion. The purpose of these experiments was to localize the expression of high-affinity receptors for these growth factors during cranial suture fusion. Four rats were sacrificed on postnatal days 8, 12, 17, and 40 (N = 16). The PF and sagittal sutures were harvested and prepared for immunohistochemical localization of TGF-beta receptor 1 and receptor 2 (Tbeta-RI, Tbeta-RII) protein. Results indicate that immunostaining for Tbeta-RI and Tbeta-RII is markedly increased in the dura mater and osteoblasts of the sutural margin of the PF suture during active suture fusion (on postnatal days 12, 17, and 40) compared with the osteoblasts and dura mater underlying the patent sagittal suture. These results, in combination with the authors' previous findings as well as studies supporting a role for TGF-beta molecules in the regulation of osteogenesis, implicate TGF-beta signaling in the regulation of suture fusion. The possible mechanisms of ligand-receptor interaction are discussed.     

Regional differentiation of cranial suture-associated dura mater in vivo and in vitro: implications for suture fusion and patency.

Despite its prevalence, the etiopathogenesis of craniosynostosis is poorly understood. To better understand the biomolecular events that occur when normal craniofacial growth development goes awry, we must first investigate the mechanisms of normal suture fusion. Murine models in which the posterior frontal (PF) suture undergoes programmed sutural fusion shortly after birth provide an ideal model to study these mechanisms. In previous studies, our group and others have shown that sutural fate (i.e., fusion vs. patency) is regulated by the dura mater (DM) directly underlying a cranial suture. These studies have led to the hypothesis that calvarial DM is regionally differentiated and that this differentiation guides the development of the overlying suture. To test this hypothesis, we evaluated the messenger RNA (mRNA) expression of osteogenic cytokines (transforming growth factor beta1 [TGF-beta1] and TGF-beta3) and bone-associated extracellular matrix (ECM) molecules (collagen I, collagen III, osteocalcin, and alkaline phosphatase) in freshly isolated, rat dural tissues associated with the PF (programmed to fuse) or sagittal (SAG; remains patent) sutures before histological evidence of sutural fusion (postnatal day 6 [N6]). In addition, osteocalcin protein expression and cellular proliferation were localized using immunohistochemical staining and 5-bromo-2'deoxyuridine (BrdU) incorporation, respectively. We showed that the expression of osteogenic cytokines and bone-associated ECM molecules is potently up-regulated in the DM associated with the PF suture. In addition, we showed that cellular proliferation in the DM associated with the fusing PF suture is significantly less than that found in the patent SAG suture just before the initiation of sutural fusion N6. Interestingly, no differences in cellular proliferation rates were noted in younger animals (embryonic day 18 [E18] and N2). To further analyze regional differentiation of cranial suture-associated dural cells, we established dural cell cultures from fusing and patent rat cranial sutures in N6 rats and evaluated the expression of osteogenic cytokines (TGF-beta1 and fibroblast growth factor 2 [FGF-2]) and collagen I. In addition, we analyzed cellular production of proliferating cell nuclear antigen (PCNA). These studies confirmed our in vivo findings and showed that dural cell cultures derived from the fusing PF suture expressed significantly greater amounts of TGF-beta1, FGF-2, and collagen I. In addition, similar to our in vivo findings, we showed that PF suture-derived dural cells produced significantly less PCNA than SAG suture-derived dural cells. Finally, coculture of dural cells with fetal rat calvarial osteoblastic cells (FRCs) revealed a statistically significant increase in proliferation (*p < 0.001) in FRCs cocultured with SAG suture-derived dural cells as compared with FRCs cocultured alone or with PF suture-derived dural cells. Taken together, these data strongly support the hypothesis that the calvarial DM is regionally differentiated resulting in the up-regulation of osteogenic cytokines and bone ECM molecules in the dural tissues underlying fusing but not patent cranial sutures. Alterations in cytokine expression may govern osteoblastic differentiation and ECM molecule deposition, thus regulating sutural fate. Elucidation of the biomolecular events that occur before normal cranial suture fusion in the rat may increase our understanding of the events that lead to premature cranial suture fusion.     

Microfocal CT: a method for evaluating murine cranial sutures in situ

INTRODUCTION: The murine model is a well-established surrogate for studying human cranial suture biology. In mice, all sutures with the exception of the posterior frontal (PF) suture remain patent throughout life. Histology is regarded as the gold standard for analyzing sutures. On this basis, PF suture fusion begins on day of life 25 and is complete by day 45. Cranial suture histology, however, requires sacrifice of the animal to obtain tissue for analysis. As a result, knowledge of the kinetics of cranial suture fusion is based on a patchwork analysis of many sutures from many different animals. The behavior of a single suture through time is unknown. Our goal is to develop a noninvasive means to repeatedly image mouse cranial sutures in vivo. As a first step, the present study was performed to evaluate microfocal computer tomography (micro-CT) technology for the use of capturing images of a mouse cranium in situ. METHODS: The micro-CT system consists of a microfocal X-ray source and a large format CCD camera optically coupled to a high-resolution X-ray image intensifier, digitally linked to a computer. The PF and sagittal sutures lie in continuity along the midline of the skull. Holes were drilled in the calvaria on both sides of the PF and sagittal sutures of a 45-day-old euthanized mouse. A micro-CT scan of this animal was performed and hundreds of cross-sectional images were generated for the cranium. These images were used to reconstruct three-dimensional volumetric images of the entire cranium. Comparisons were made between (1). the gross specimen and the three dimensional reconstructions; (2). two-dimensional coronal images obtained by micro-CT and those obtained by histology. RESULTS: Analysis of PF and sagittal sutures demonstrated the following: (1). The drilled holes were accurately rendered by micro-CT, when compared to both the gross specimen and the histology. (2). The sagittal suture was found to be patent by both micro-CT and histology. (3). The PF suture is fused by histology, but unexpectedly, the PF suture appears incompletely fused by micro-CT. By micro-CT, however, the anterior and endocranial regions appear more extensively fused than the remainder of the PF suture, a finding consistent with published histologic analysis. CONCLUSIONS: We successfully imaged 45-day-old mouse cranial sutures in situ using micro-CT technology. Precise correlation between histologic sections and radiologic images is difficult, but convincing similarities exist between the gross specimen and images from micro-CT and histology. PF suture fusion in a 45-day-old animal appears different by micro-CT than by histology. One possible explanation for this apparent discrepancy is that suture fusion in histology is determined based on the appearance of bone morphology and not tissue density, as the specimens are necessarily decalcified to section the bone. Micro-CT, on the other hand, distinguishes tissues on the basis of density. Newly forming bone may require bone matrix formation prior to complete calcification; PF suture in 45-day-old mice may be morphologically complete but incompletely ossified. Studies correlating histologic and micro-CT assessment of suture development are underway. Micro-CT appears to be a promising method for noninvasive imaging of mouse cranial suture.     

胰岛素生长因子1在颅缝闭合中调节作用的实验研究

目的：研究胰岛素生长因子1对大鼠颅缝细胞的骨诱导作用和体外调节小鼠矢状缝闭合的作用。方法：获取新出生的SD大鼠的矢状缝细胞进行培养和出生8d的CD1小鼠矢状缝进行体外无血清器官培养基培养，加入胰岛素生长因子1（insulin lilce growth factor 1 IGF1），浓度分别为10ng/ml和40ng/ml，并设立不加IGF1者为对照组，应用RT－PCR检测颅缝细胞的成骨细胞表型碱性磷酸酶、骨钙素和骨桥蛋白mRNA表达，ELISA法检测培养液I型胶原的分泌，光镜观察小鼠矢状缝闭合的情况。结果：加入IGF1的矢状缝细胞的成骨细胞表型碱性磷酸酶、骨钙素和骨桥蛋白mRNA表达以及培养液I型胶原分泌量较对照组明显增高。有IGF1干预的矢状缝移植体培养8d时，颅缝内侧面骨板开始接近，培养20d时，颅缝小部分闭合，培养30d时颅缝大部分闭合，无IGF1干预的对照组培养30d，颅缝未发生闭合。结论：胰岛素生长因子1通过增强颅缝细胞的骨诱导促进颅缝的闭合。     

Impaired posterior frontal sutural fusion in the biglycan/decorin double deficient mice

Biglycan (Bgn) and decorin (Dcn) are highly expressed in numerous tissues in the craniofacial complex. However, their expression and function in the cranial sutures are unknown. In order to study this, we first examined the expression of biglycan and decorin in the posterior frontal suture (PFS), which predictably fuses between 21 and 45 days post-natal and in the non-fusing sagittal (S) suture from wild-type (Wt) mice. Our data showed that Bgn and Dcn were expressed in both cranial sutures. We then characterized the cranial suture phenotype in Bgn deficient, Dcn deficient, Bgn/Dcn double deficient, and Wt mice. At embryonic day 18.5, alizarin red/alcian blue staining showed that the Bgn/Dcn double deficient mice had hypomineralization of the frontal and parietal craniofacial bones. Histological analysis of adult mice (45-60 days post-natal) showed that the Bgn or Dcn deficient mice had no cranial suture abnormalities and immunohistochemistry staining showed increased production of Dcn in the PFS from Bgn deficient mice. To test possible compensation of Dcn in the Bgn deficient sutures, we examined the Bgn/Dcn double deficient mice and found that they had impaired fusion of the PFS. Semi-quantitative RT-PCR analysis of RNA from 35 day-old mice revealed increased expression of Bmp-4 and Dlx-5 in the PFS compared to their non-fusing S suture in Wt tissues and decreased expression of Dlx-5 in both PF and S sutures in the Bgn/Dcn double deficient mice compared to the Wt mice. Failure of PFS fusion and hypomineralization of the calvaria in the Bgn/Dcn double deficient mice demonstrates that these extracellular matrix proteoglycans could have a role in controlling the formation and growth of the cranial vault.     

Cranial deformation in craniosynostosis. A new explanation.

Skull growth after premature fusion of a single suture was described by Virchow in 1851. He observed that growth was restricted in a plane perpendicular to a fused suture. However, he failed to predict the compensatory growth patterns that produce many of the deformities recognized as features of individual craniosynostosis syndromes. The deformities resulting from premature closure of a coronal, sagittal, metopic, or lambdoid suture can be predicted by the following observations: (1) cranial vault bones that are prematurely fused act as a single bone plate with decreased growth potential; (2) asymmetrical bone deposition occurs mainly at perimeter sutures, with increased bone deposition directed away from the bone plate; (3) sutures adjacent to the stenotic suture compensate in growth more than those sutures not contiguous with the closed suture; and (4) enhanced bone deposition occurs along both sides of a nonperimeter suture that is a continuation of the prematurely closed suture. These four rules were derived by critically examining the clinical deformities observed with each form of craniosynostosis. These rules assume that cranial sutures have the capacity to compensate by depositing bone asymmetrically along their edges. Unequal growth patterns have been demonstrated in the frontonasal suture of rabbits by Selman and Sarnat. In addition, unequal bone deposition has also been demonstrated along the parieto-interparietal suture in albino rats by Baer. Human studies to determine if asymmetrical bone deposition actively occurs along cranial vault sutures in response to a stenotic suture have not been performed, however. It is also unclear whether these four guidelines apply to cranial base abnormalities observed with craniosynostosis. As new radiologic techniques develop to define the configuration of the skull in intricate detail, a skull pattern of growth explaining the pathogenesis of all deformities created by premature fusion of a cranial vault suture may become apparent.     

大鼠颅骨体外培养矢状缝牵引成骨模型的建立

目的探讨建直幼年SD大鼠顿骨器官体外培养矢状缝牵引成骨模型。方法取19日龄的SD大鼠颅顶骨矢状缝组织块．随机分为对照组和实验组进行体外器官培养。实验组加力约3．92×10^3N（0．4g）力Hr照组不加力。培养24h结束时,进行大体观察及倒置显微镜下观察;并经苏木精-伊红染色后进行组织学观察。结果大体观察／受倒置显微镜下观察发现,实验组骨缝逐渐明显加宽。至加力24h,所有标本欠状缝未见断裂。组织学观察发现,缝两侧区域为成骨活跃部位,两侧骨化前沿交错排列。缝内可见纤维结缔纠织、成骨细胞、成纤维细胞及毛细血管,缝内组织与硬脑膜相连。对照组骨缝保持正常发育,未见明显变化,以成骨细胞为主。结论大鼠颅骨骨缝器官可以在体外培养中成功存活并继续生长。     

Interrelationship of Cranial Suture Fusion, Basicranial Development, and Resynostosis Following Suturectomy in Twist1+/? Mice, a Murine Model of Saethre-Chotzen Syndrome

The interrelationships among suture fusion, basicranial development, and subsequent resynostosis in syndromic craniosynostosis have yet to be examined. The objectives of this study were to determine the potential relationship between suture fusion and cranial base development in a model of syndromic craniosynostosis and to assess the effects of the syndrome on resynostosis following suturectomy. To do this, posterior frontal and coronal suture fusion, postnatal development of sphenooccipital synchondrosis, and resynostosis in Twist1(+/+) (WT) and Twist1(+/-) litter-matched mice (a model for Saethre-Chotzen syndrome) were quantified by evaluating μCT images with advanced image-processing algorithms. The coronal suture in Twist(+/-) mice developed, fused, and mineralized at a faster rate than that in normal littermates at postnatal days 6-30. Moreover, premature fusion of the coronal suture in Twist1(+/-) mice preceded alterations in cranial base development. Analysis of synchondrosis showed faster mineralization in Twist(+/-) mice at postnatal days 25-30. In a rapid resynostosis model, there was an inability to fuse both the midline posterior frontal suture and craniotomy defects in 21-day-old Twist(+/-) mice, despite having accelerated mineralization in the posterior frontal suture and defects. This study showed that dissimilarities between Twist1(+/+) and Twist1(+/-) mice are not limited to a fused coronal suture but include differences in fusion of other sutures, the regenerative capacity of the cranial vault, and the development of the cranial base.     

SD大鼠颅缝细胞不同时期GPC3基因表达变化的研究

目的 探索GPC3基因在颅缝组织和细胞中不同时期的表达情况,为后续的疾病模型研究提供参照。方法研究并观察1 d、3 d、7 d的SD大鼠颅缝细胞组织,应用组织切片免疫荧光染色、RT-q PCR、Western Blot等方法,检测其在不同年龄SD大鼠中的表达水平,判断其与颅骨成骨及颅缝闭合的关系。结果 免疫荧光显示,GPC3在大鼠未闭合颅缝组织不同阶段胞内、细胞表面及胞外均有表达;RT-q PCR、Western Blotting结果显示,GPC3基因表达随着大鼠年龄增长呈现下降趋势。结论 SD大鼠中,GPC3与颅缝闭合密切相关。     

An analysis of the growth of the cranial vault in rabbits by vital staining with lead acetate

The mode and the rate of formation of individual bones in the cranium were studied by vital staining with lead acetate in 13 New Zealand white rabbits beginning at 35 days of age. These evaluations were compared with⁴⁵Ca uptake at bilaterally comparable sites.The growth patterns differed from one suture to another. Elongation of the cranial vault in the anterior direction in the snout area was rapid. In the sagittal suture complex, active growth was observed on the intrasutural surfaces of the internasal and metopic sutures, while much less growth was taking place in the sagittal suture. Active growth also was taking place at the intrasutural surface of the nasal bone at the naso-premaxillary suture while resorption was found on the surface of the parietal bone in the squamosal suture.Increase in thickness of the cranial vault appeared to be the result of accretion on the ectocranial and to a lesser extent on the endocranial surfaces of the frontal and parietal bones.Good correlations between the linear measurement with the lead lines and⁴⁵Ca uptake were observed in the bones of the cranial vault, especially when⁴⁵Ca was administered 3 or 6 days before sacrifice.Fellowship F05-TW-1029.     

颅面多骨缝固定对颅颌形态影响的实验研究

目的通过固定幼兔颅骨缝，模拟颅骨多骨缝早闭，观察固定后颅颌面生长方式的变化及相互影响的关系。方法采用牙科釉质粘合剂固定２周龄幼兔冠状缝、矢状缝和额间缝，模拟颅骨多骨缝早闭。术后不同时期测量颅穹窿长度、高度、面中部高度、上颌骨长度、面角、腭角和颅底角。结果多骨缝早闭后，颅穹窿高度和长度明显减少，使颅骨变短且扁平。面中部高度和上颌骨长度增大，整个面中骨结构有整体向上移动趋势。面、腭和颅底角增大，这与颅穹窿变浅，面中部高度增大相对应。结论兔出生后１０周前是颅骨缝扩张性生长高峰期，在此期间，某些原因影响颅骨正常生长，必然导致颅面畸形。实验结果与临床颅骨骨缝早闭后表现类似，和主张早期手术矫正畸形观点一致     

A Pro253Arg mutation in fibroblast growth factor receptor 2 (Fgfr2) causes skeleton malformation mimicking human Apert syndrome by affecting both chondrogenesis and osteogenesis

Apert syndrome is one of the most severe craniosynostosis that is mainly caused by either a Ser252Trp(S252W) or Pro253Arg(P253R) mutation in fibroblast growth factor receptor 2 (FGFR2). As an autosomal dominant disorder, Apert syndrome is mainly characterized by skull malformation resulting from premature fusion of craniofacial sutures, as well as syndactyly, etc. A P253R mutation of FGFR2 results in nearly one-thirds of the cases of Apert syndrome. The pathogenesis of Apert syndrome resulting from P253R mutation of FGFR2 is still not fully understood. Here we reported a knock-in mouse model carrying P253R mutation in Fgfr2. The mutant mice exhibit smaller body size and brachycephaly. Analysis of the mutant skulls and long bones revealed premature fusion of coronal suture, shortened cranial base and growth plates of long bones. In vitro organ culture studies further revealed that, compared with wild-type littermates, the mutant mice have prematurely fused coronal sutures and retarded long bone growth. Treatment of the cultured calvaria and femur with PD98059, an Erk1/2 inhibitor, resulted in partially alleviated coronal suture fusion and growth retardation of femur respectively. Our data indicated that the P253R mutation in Fgfr2 directly affect intramembranous and endochondral ossification, which resulted in the premature closure of coronal sutures and growth retardation of long bones and cranial base. And the Erk1/2 signaling pathway partially mediated the effects of P253R mutation of Fgfr2 on cranial sutures and long bones.     

Craniofacial growth following experimental craniosynostosis and craniectomy in rabbits.

Premature fusion of the coronal suture was produced in 9-day-old rabbits by immobilization of the suture area bilaterally with methyl-cyanoacrylate adhesive. The effects of suture fusion and its surgical release on suture growth and on skull morphology were evaluated by radiographic cephalometry. Immobilization resulted in significant changes in the angular dimensions in the vault toward an anteroposterior shortening. No permanent deformity was observed in the angular relationship between the cranial base and the facial skeleton. Craniectomy at 30 days, when a skull deformity had been established, resulted in rapid separation of the bones at the suture site which returned the deformed skull to a normal configuration by 90 days of age. Surgical removal of a normal suture in a control group also resulted in accelerated separation of the bones at the excised suture site, but it was less than after removal of an immobilized suture. The experimental data indicate that premature fusion of rapidly growing sutures results in consistent skull deformity. Early release of the fusion, when this is the primary abnormality, will result in spontaneous correction of the deformity.     

Cranial vault growth in craniosynostosis

Skull growth after single suture closure was described in 1851 by Virchow, who noted that growth in the plane perpendicular to a fused suture was restricted. However, this observation failed to predict compensatory growth patterns that produce many of the deformities recognized as features of individual syndromes. The deformities resulting from premature closure of a coronal, sagittal, metopic, or lambdoid suture can be predicted on the basis of the following observations: 1) cranial vault bones that are prematurely fused secondary to single suture closure act as a single bone plate with decreased growth potential; 2) asymmetrical bone deposition occurs mainly at perimeter sutures, with increased bone deposition directed away from the bone plate; 3) sutures adjacent to the prematurely fused suture compensate in growth more than those sutures not contiguous with the closed suture; and 4) enhanced symmetrical bone deposition occurs along both sides of a non-perimeter suture that is a continuation of the prematurely closed suture. These observations regarding growth in craniosynostosis are illustrated with clinical material in this report.     

Toward an understanding of nonsyndromic craniosynostosis: altered patterns of TGF-beta receptor and FGF receptor expression induced by intrauterine head constraint

Although the etiology of nonsyndromic forms of craniosynostosis remains uncertain, recent experiments from our laboratory have demonstrated that fetal head constraint induces cranial suture fusion in mice through a process associated with altered patterns of transforming growth factor beta (TGF-beta) isoform expression. Other recent studies have highlighted the role of secreted signaling molecules, including members of the TGF-beta superfamily and the fibroblast growth factors (FGFs), as well as their receptors, in regulating suture development and fusion. The purpose of these experiments was to examine the potential role of TGF-beta receptors and FGF receptor 2 (FGFR2) in nonsyndromic craniosynostosis by determining their temporospatial patterns of expression during development complicated by intrauterine head constraint. This study consisted of two groups of C57BI/6J mice: an experimental group subjected to intrauterine constraint and a control unconstrained group. Fetal head constraint was induced by performing uterine cerclage on day 17.5 of gestation and allowing intrauterine fetal growth to continue 24 and 48 hours beyond the normal gestational period. Control animals underwent hysterotomy on day 17.5 and the nonconstrained pups were allowed to continue intra-abdominal fetal growth 48 hours beyond normal gestation. Expression of TGF-beta receptor types I and II, and FGFR2 in the calvarial tissue was determined by immunohistochemical analysis. In the unconstrained control animals, there was minimal immunoreactivity for both TGF-beta receptors and FGFR2 within the coronal suture. After 24 hours of constraint, however, there was a marked increase in immunoreactivity of TGF-beta receptors and FGFR2 in the osteoblasts along the osteogenic fronts and in the dural cells. After 48 hours, there was continued expression of both type I and type II receptors and FGFR2 within the midsutural mesenchyme of the coronal suture, in the osteoblasts, and in the dura. The authors demonstrated substantial upregulation of TGF-beta receptor types I and II and FGFR2 in coronal sutures subjected to in utero constraint. These results suggest an important role for TGF-beta/TGF-beta receptor, and FGF/FGFR signaling in the pathogenesis of constraint-induced craniosynostosis.     

Infantile craniosynostosis: Clinical,radiological, and surgical considerations based on 100 surgically treated cases

Summary One hundred children affected by craniosynostosis were operated on from January 1952 to February 1977.Forty-six patients were operated on within the first year of life, and only 9 after six years of age.In 27 cases only one cranial suture was synostotic (mostly the coronal: 15 cases). In 36 cases two sutures were involved (mostly coronal plus sagittal: 25 cases). In 28 cases all the cranial sutures were involved. Also included in this series are seven cases of Crouzon's, one of Apert's, and one of Carpenter's syndromes. The clinical, radiological, and surgical features of this condition are discussed, and the long-term results are reported.     

A case of pycnodysostosis

Pycnodysostosis is a term introduced by Maroteaux and Lamy in 1962 to describe characteristic features such as dwarfism, separated cranial sutures, open fontanelles, hypoplasia and absence of mandibular angle, partial aplasia of the terminal phalanges of the fingers and toes, generalized condensation of bone shadow, bone fragility, and autosomal recessive inheritance. We reported a such case in a 13-years old boy associated with an epidural hematoma. Four weeks after removal of the hematoma, lumbar puncture was performed with the initial pressure of 200mm H2O in an awake state and the plateau waves were observed several times during sleep. Such increased intracranial pressure may have caused the open fontanelle and cranial sutures in addition to dysostosis. Thus such patients require careful attention in order to prevent major bleeding from the superior sagittal sinus directly beneath the separated sagittal suture.     

Regulation of bone morphogenetic protein signalling and cranial osteogenesis by Gpc1 and Gpc3

From birth, the vault of the skull grows at a prodigious rate, driven by the activity of osteoblastic cells at the fibrous joints (sutures) that separate the bony calvarial plates. One in 2500 children is born with a medical condition known as craniosynostosis because of premature bony fusion of the calvarial plates and a cessation of bone growth at the sutures. Bone morphogenetic proteins (BMPs) are potent growth factors that promote bone formation. Previously, we found that Glypican-1 (GPC1) and Glypican-3 (GPC3) are expressed in cranial sutures and are decreased during premature suture fusion in children. Although glypicans are known to regulate BMP signalling, a mechanistic link between GPC1, GPC3 and BMPs and osteogenesis has not yet been investigated. We now report that human primary suture mesenchymal cells coexpress GPC1 and GPC3 on the cell surface and release them into the media. We show that they inhibit BMP2, BMP4 and BMP7 activities, which both physically interact with BMP2 and that immunoblockade of endogenous GPC1 and GPC3 potentiates BMP2 activity. In contrast, increased levels of GPC1 and GPC3 as a result of overexpression or the addition of recombinant protein, inhibit BMP2 signalling and BMP2-mediated osteogenesis. We demonstrate that BMP signalling in suture mesenchymal cells is mediated by both SMAD-dependent and SMAD-independent pathways and that GPC1 and GPC3 inhibit both pathways. GPC3 inhibition of BMP2 activity is independent of attachment of the glypican on the cell surface and post-translational glycanation, and thus appears to be mediated by the core glypican protein. The discovery that GPC1 and GPC3 regulate BMP2-mediated osteogenesis, and that inhibition of endogenous GPC1 and GPC3 potentiates BMP2 responsiveness of human suture mesenchymal cells, indicates how downregulation of glypican expression could lead to the bony suture fusion that characterizes craniosynostosis.     

Significance of differences in patency among cranial sutures

OBJECTIVE: To evaluate the gross external characteristics of the coronal, lambdoid, and sagittal sutures in human cadaver skulls and determine if a difference exists in terms of patency, sex, and age. METHODS: The coronal, lambdoid, and sagittal sutures were described using a modified grading scale to quantify sutural patency. An open suture was graded as 0, a fused suture as 1, and an obliterated suture as 2, 3, or 4, depending on the extent of obliteration. RESULTS: Thirty-six skulls were examined, including 17 female and 19 male (age range, 56-101 y). When compared with the sagittal suture, the lambdoid suture was significantly more likely to be patent and least likely to be obliterated. No significant difference in suture grades was found between female and male skulls, and no significant difference was found between age and suture grade. CONCLUSION: The prolonged patency of the lambdoid suture may be due to external forces, such as the greater number of muscles affecting the lambdoid suture when compared with the sagittal suture.     

子宫内局部生物力学改变与胚胎兔颅缝早闭相关性的实验研究

目的研究兔子宫内作用于胚胎颅骨局部生物力在兔颅缝闭合过程中的调控作用。方法通过对妊娠25d大白兔宫内胚胎，硬脑膜外的颅骨矢状缝用5—0可吸收线缝合，改变其矢状缝局部生物作用力，建立子宫内胚胎颅骨局部生物力学改变的模型。采用胚胎外科技术、组织化学技术，研究局部生物动力学改变后，颅缝组织中的骨钙素（Osteocalcin，OCN）及骨桥蛋白（Osteopontin，OPN）的分泌情况。结果在胚胎兔出生后9d，其被缝合的矢状缝组织中OCN及OPN的含量明显增加。与对照组之间有明显差异（p〈0．05）。结论宫内局部生物力学改变，促进兔单条颅缝组织中OCN及OPN的分泌。