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.

Cranial sutures function as bone growth centers while themselves remaining unossified. Rat frontonasal sutures become obliterated by neonatal day 21 (N21), while coronal sutures do not fuse over the life of the animal. Coronal sutures induced to undergo osseous obliteration in vitro after removal of the dura mater were found to require soluble, heparin-binding factors present in dura mater to resist osseous obliteration. Transforming growth factor beta 1 (TGF-beta 1), beta 2, and beta 3, heparin-binding factors known to regulate bone cell proliferation and differentiation, were considered likely candidates. The presence and distribution of these factors in calvarial tissues both in vivo and in vitro were established by immunohistochemical analysis, while reverse transcription followed by polymerase chain reaction (RT/PCR) was employed to determine the presence of transcripts for these factors in mRNA isolated from microdissected dura mater. Results indicated that the presence of TGF-beta 1 and TGF-beta 2 were associated with developing coronal and frontonasal sutures, and that the continued presence of these factors was associated with osseous obliteration of the frontonasal suture. However, increased TGF-beta 3 immunoreactivity was associated with the coronal suture remaining unossified. RT/PCR demonstrated the presence of transcripts for TGF-beta 1, beta 2, and beta 3 in dural tissues isolated from rat calvaria. These data support the notion of a role for TGF-beta s in regulating cranial suture morphogenesis and establish the in vitro model as a valid system for examining mechanisms by which growth factors regulate both suture morphogenesis and bone growth at the suture site.

Dura mater maintains rat cranial sutures in vitro by regulating suture cell proliferation and collagen production.

Craniosynostosis, the premature osseous obliteration of cranial vault sutures, can result from mutations in genes encoding components of growth factor signaling systems or the extracellular matrix (ECM). Little is known of the capacity of osteoprogenitor cells of the cranial sutures to divide or to synthesize ECM in situ. Osteoblasts derived from patients with prematurely fused sutures were reported to express alkaline phosphatase and osteocalcin at elevated levels, while proliferating at a rate comparable to control cells [DePollack et al., JBMR, 1996]; however, the suture osteoprogenitors, the population most likely to show proliferative abnormalities, were not present in the fused sutures used for this study. A model in which rat coronal sutures and associated bones develop normally in vitro, but in which sutures can be induced to fuse in the absence of dura mater, was used to examine cell proliferation and total protein synthesis in unfused sutures cultured in the presence of dura mater or in sutures induced to fuse in the absence of dura mater. Significantly increased cell proliferation was seen in suture cells prior to sutural obliteration, which returned to control levels as sutural fusion proceeded. Collagen synthesis in fusing sutures was elevated compared to non-fusing sutures and comparable to that seen in bone. Results indicated that in the absence of intercellular signals provided by the dura mater, suture cell proliferation increased initially, followed by increased synthesis of collagenous ECM within the suture and subsequent osseous obliteration of the suture. Thus factors originating in the dura mater affected suture cell proliferation and ECM production and were required for the maintenance of suture patency.

Elevated levels of transforming growth factors beta 2 and beta 3 in lambdoid sutures from children with persistent plagiocephaly.

OBJECTIVE: To analyze the pertinent history and physical findings specific to the subset of patients with a progressive posterior skull deformity, requiring surgery to correct their deformity. PATIENTS: Since the Academy of Pediatrics issued its recommendation on supine positioning of infants to prevent sudden infant death syndrome (SIDS) in 1992, 73 children have presented to the University of Virginia Craniofacial Anomalies Clinic with posterior-skull deformities. The majority were successfully managed with conservative therapy, but in six patients, the deformity was severe and persistent, requiring surgical correction. All six children were older (7.5-12 mo), presenting with more severe morphologic appearances and a higher incidence of associated neurodevelopmental delay. Three had family backgrounds of isolated craniosynostosis. METHODS: Characteristics of these patients were examined to determine why they may have differed from those that responded to conservative management. Immunohistochemical staining of their lambdoid sutures was performed. RESULTS: Significantly increased staining for TGF-beta 2 and TGF-beta 3, potent stimulators of bone cell growth and differentiation, was seen in all 'affected' sutures from the flattened side of the skull, compared to unaffected sutures from the protruding side of the skull-a pattern similar to that seen during normal bony obliteration of calvarial sutures. CONCLUSION: The majority of patients with posterior plagiocephaly associated with positioning responded to conservative management, while a small subset of patients with persistent posterior skull deformation required surgical intervention. A genetic basis for the latter patients' persistent plagiocephaly, rather than positioning, cannot be ruled out. Genetics, prolonged external pressure against the sutures, or a combination of these factors may lead to permanently raised levels of growth factors in 'affected' sutures.