Differential expression of fibromodulin, a transforming growth factor-beta modulator, in fetal skin development and scarless repair.

Transforming growth factor-beta (TGF-beta1, -beta2, and -beta3) has been implicated in the ontogenetic transition from scarless fetal repair to adult repair with scar. Generally, TGF-beta exerts its effects through type I and II receptors; however, TGF-beta modulators such as latent TGF-beta binding protein-1 (LTBP-1), decorin, biglycan, and fibromodulin can bind and potentially inhibit TGF-beta activity. To more fully explore the role of TGF-beta ligands, receptors, and potential modulators during skin development and wound healing, we have used a rat model that transitions from scarless fetal-type repair to adult-type repair with scar between days 16 and 18 of gestation. We showed that TGF-beta ligand and receptor mRNA levels did not increase during the transition to adult-type repair in fetal skin, whereas LTBP-1 and fibromodulin expression decreased. In addition, TGF-beta1 and -beta3; type I, II, and III receptors; as well as LTBP-1, decorin, and biglycan were up-regulated during adult wound healing. In marked contrast, fibromodulin expression was initially down-regulated in adult repair. Immunostaining demonstrated significant fibromodulin induction 36 hours after injury in gestation day 16, but not day 19, fetal wounds. This inverse relationship between fibromodulin expression and scarring in both fetal and adult rat wound repair suggests that fibromodulin may be a biologically relevant modulator of TGF-beta activity during scar formation.

"Pumping the regenerate": an evaluation of oscillating distraction osteogenesis in the rodent mandible.

Mandibular distraction osteogenesis (DO) has become an important technique to lengthen the hypoplastic mandible and to reconstruct osseous defects after ablative surgery. The hallmark of successful DO is the creation of new bone within the distraction gap. Several anecdotal reports have described alternating compressing and lengthening protocols (i.e., "pumping the regenerate") to augment regenerate bone formation. The purpose of this experiment was to analyze formally the effects of an alternating compression/distraction protocol with a traditional distraction protocol. Ten adult male rats underwent unilateral mandibular osteotomy with placement of a custom distractor. After a latency period of 5 days, distraction was initiated at a rate of 0.25 mm twice daily. Animals in the control group (N = 5) were distracted to a length of 5.0 mm for 10 days at a rate of 0.25 mm twice daily. In contrast, animals in the experimental group (N = 5) were distracted to a length of 2.5 mm (at a rate of 0.25 mm twice daily) for 5 days, then compressed 1.0 mm for a 2-day period, and redistracted to a length of 5.0 mm. Regenerate cross-sectional area was evaluated by computed tomography performed after 5 weeks of consolidation. Gross examination and histological analysis were performed by a panel of experienced reviewers. Radiological as well as histological analysis of regenerate cross-sectional area demonstrated no significant differences between experimental (i.e., "pumped") and control groups. Both groups demonstrated excellent regenerate bone formation with no evidence of fibrous union. This study represents the first attempt to investigate the anecdotal technique of pumping the mandibular regenerate. The authors have demonstrated that pumping the regenerate leads to no substantial differences in radiological or histological appearance of regenerate bone formation.

Controlled multiplanar distraction of the mandible. Part III: Laboratory studies of sagittal (anteroposterior) and horizontal (mediolateral) movements.

Distraction osteogenesis has proven to be an effective technique for the correction of mandibular deficiencies. However, problems have been encountered in achieving a final, idealized form of the mandible when using distraction devices capable of moving the bone segments in only one dimension (uniplanar). Specifically, occlusal irregularities and deficiencies in lower facial contour have been seen following uniplanar distraction. To address these problems, a distraction device capable of independent movements in three planes (multiplanar) was developed. Previously reported studies in a canine model have demonstrated that this device can successfully distract the mandible along both the sagittal axis (anteroposterior or z-axis) and the vertical axis (superoinferior or y-axis). This study examines the ability of the multiplanar device to distract along the sagittal and horizontal axes (mediolateral or x-axis). A total of 12 dogs were included in the study. All animals underwent unilateral or bilateral mandibular distraction using an external multiplanar device. After a latency period of 5 days, primary distraction along the anteroposterior axis at a rate of 1 mm/day for 10 days (10 mm total) was performed. During the following 10 days, along with an additional 11 mm to 20 mm of anteroposterior axis distraction, concomitant secondary distraction was performed along the horizontal (mediolateral) axis at a rate of 5 degrees/day (50 degrees total). Cephalometric radiographs were obtained preoperatively and at the conclusion of both anteroposterior and combined anteroposterior-mediolateral distraction. Computed tomography (CT) scans were obtained preoperatively and at the end of consolidation (28 days), after which all animals were sacrificed and the dry skulls examined. In all animals, distraction along the mediolateral or x-axis was found to change the anteroposterior projection of the mandible. Varus angulation of the device with respect to the midline of the mandible caused compression of the distracted segments and reduced the anteroposterior thrust of the mandible. In contrast, valgus positioning of the device, with respect to the midline of the mandible, created the opposite effect, increasing the distracted length in the anteroposterior direction. The bone (mandibular) segments being distracted assumed the orientation of the device only for valgus positioning of the device (producing a decrease in the bigonial distance). Conversely, there was no effect from the mediolateral angulation on the distracted segments during varus positioning of the device. A possible explanation for this finding may be a greater resistance to an increase in the bigonial distance (varus positioning of the device) posed by obstruction of lateral movement of the condyle. This stands in contrast to a decrease in the bigonial distance observed following valgus positioning of the device. These findings confirm the clinical impression that distraction along the anteroposterior or sagittal axis remains the critical or keystone therapeutic maneuver in distraction of the mandible. Mediolateral or horizontal axis distraction is best used only as a supplementary movement; in essence, it only affects the anteroposterior dimension with little impact on clinically relevant changes to the bigonial distance.

Immunolocalization of basic fibroblast growth factor and fibroblast growth factor receptor-1 and receptor-2 in rat cranial sutures.

Craniosynostosis is a common disorder with an unknown etiology. Recent genetic mapping studies have demonstrated a strong linkage between several familial craniosynostotic syndromes and mutations in fibroblast growth factor receptor 1 (FGF-R1) and 2 (FGF-R2). The purpose of this experiment was to investigate by immunohistochemistry the protein production of these receptors as well as of their most prevalent ligand, basic fibroblast growth factor (bFGF), before, during, and after sutural fusion in rat cranial sutures. The posterior frontal (normally fuses between postnatal days 12 and 22) and sagittal (remains patent) sutures of embryonic day 20 and neonatal days 6, 12, 17, 22, and 62 (n = 3 per group) were harvested, fixed, and decalcified. Five-micrometer sections were stained with polyclonal antibodies against bFGF, FGF-R1, and FGF-R2, and patterns of immunohistochemical staining were assessed by independent reviewers. Our results indicate that increased bFGF production correlates temporally with suture fusion, with increased staining of the dura underneath the fusing suture prior to fusion followed by increased staining within osteoblasts and sutural cells during fusion. FGF-R1 and, to a lesser extent FGF-R2 immunostaining revealed a different pattern of localization with increased immunostaining within the patent sagittal suture at these time points. These results implicate bFGF in the regulation of sutural fusion and may imply autoregulatory mechanisms in fibroblast growth factor receptor expression.

Rat mandibular distraction osteogenesis: Part I. Histologic and radiographic analysis.

The application of distraction osteogenesis to craniofacial surgery has altered the approach and treatment of congenital and acquired craniofacial defects. Although the histologic and ultrastructural changes associated with distraction osteogenesis have been described extensively, relatively little is known about the molecular regulation of this process. The elucidation of the molecular mechanisms of distraction osteogenesis has important clinical implications because it may facilitate the use of recombinant proteins or gene therapy to accelerate bone regeneration. Molecular analysis of distraction osteogenesis has been hindered by the use of large animal models in which only limited genetic information is available. In this study, a rat model of mandibular distraction osteogenesis is described. This report includes a pilot study (n = 50) to develop an appropriate distraction device and to determine the optimal placement of the osteotomy. The study subsequently included 80 animals, 35 of which were distracted at a rate of 0.25 mm per day for 6 days (1.5 mm total) and 35 that were distracted at a rate of 0.25 mm twice per day (3.0 mm total). These animals were killed at various time points (after latency and during the distraction and consolidation periods) and displayed histologic and radiographic findings of membranous bone distraction osteogenesis that were consistent with those in large ,animal and clinical models. In addition, five animals each were acutely lengthened 1.5 mm and 3.0 mm and demonstrated a fibrous nonunion. Furthermore, the utility of this model is demonstrated in the analysis of the molecular mechanisms of distraction osteogenesis by applying the polymerase chain reaction to total cellular RNA isolated from normal and distracted rat mandibles. In conclusion, it is believed that the rat model of distraction osteogenesis has significant advantages over traditional models, including decreased costs and facilitation of molecular analysis.

The combination of endoscopy and distraction osteogenesis in the development of a canine midface advancement model.

The requirements for reconstruction in patients with midface hypoplasia can be formidable: a bicoronal scalp incision, Le Fort III or monobloc skeletal advancement, harvesting and insertion of bone grafts, application of rigid (and occasionally intermaxillary) fixation, blood transfusions, and prolonged operative time and hospitalization. The introduction of the endoscope offers the possibility of minimally invasive surgery with improved visualization of the osteotomy sites. The development of distraction osteogenesis as a surgical technique allows controlled and gradual advancement of the osteotomized skeletal segment and associated soft tissue. The purpose of this study was to develop a canine model of an endoscopically assisted Le Fort III osteotomy with attendant midface distraction. Four mongrels (20 kg in weight) were study subjects. Three 2-cm skin incisions were made (two perpendicular to the zygomaticomaxillary suture and one perpendicular to the nasofrontal suture). The soft tissue and periosteum were evaluated bluntly. Retractors specially designed for the project created a space for endoscopic visualization. Bilateral zygomatic, nasofrontal, and medial orbital wall osteotomies, corticotomies, or both were performed under endoscopic visualization using a reciprocating saw; the medial orbital wall sectioning was specifically not completed (i.e., corticotomy) to avoid laceration of the mucosa and attendant bleeding. The pterygomaxillary osteotomy was completed with an osteotome and mallet. Finally, the nasal septum was only partially divided with an osteotome to avoid excessive blood loss. Four distraction devices were placed across the above-noted osteotomies (two across the nasofrontal osteotomy and one across each lateral osteotomy). The animals were distracted 1 mm per day for 16 to 40 days after surgery (16-40 mm of linear distraction). Cephalograms and computed tomography scans were obtained before and after distraction. The animals were killed after remaining in fixation for 4 to 6 weeks after distraction. All soft tissue was removed and the skull was examined. Photos were obtained throughout the experiment for documentation. The study demonstrated that Le Fort III osteotomies can be performed successfully via small incisions with endoscopic assistance in canine subjects with excellent visualization and minimal bleeding. The advancement of the midface segment can be achieved by activation of an external distraction device.

Scarless healing. The fetal wound.

Fetal wounds heal without a scar early in gestation, and may hold the key to scarless repair. Several important concepts central to the fetal wound-healing response have been determined. The fetal fibroblast modulates the wound-healing response through collagen deposition, extracellular matrix deposition, and growth factor secretion. Fetal repair is both gestational-age and wound-size dependent, with a transition from scarless to scarring repair occurring during fetal life. Fetal fibroblasts manifest a decreased ability to induce dermal appendage formation from fetal epithelium at the same time that scarring in the fetus begins, suggesting that epithelial-mesenchymal interactions play an important role in scarless fetal repair. The fetal immune response during wound healing differs from the adult response, with a primarily mononuclear cell infiltrate and decreased activity and presence of polymorphonuclear leukocytes, whereas the cytokine profile of the fetal wound differs markedly from that of the adult wound. Patterning genes (homeobox genes) involved in organogenesis may prove integral to fetal healing, and are emerging as an active area of research. Once the biology of fetal wound healing is fully determined, attempts to manipulate the adult wound undoubtedly will progress rapidly, and scarless repair may become a clinical reality in children and adults.

Intracapsular foldable posterior chamber lens implantation in eyes with posterior capsule tears or zonular fiber instability.

A surgical technique is described for foldable posterior chamber intraocular lens implantation in the capsular bag in the presence of a posterior capsule tear or weakened zonular fiber support. Haptics are compressed by suturing before endocapsular insertion, minimizing capsular and zonular fiber stress.

Controlled multiplanar distraction of the mandible, Part II: Laboratory studies of sagittal (anteroposterior) and vertical (superoinferior) movements.

The application of distraction osteogenesis in craniofacial surgery has significantly altered the treatment of congenital mandibular deficiencies. However, evaluation of results in both animal studies and clinical cases has revealed deficiencies, particularly in two areas. First, distraction using a uniplanar device in an anteroposterior direction can result in a persistent anterior open bite. Second, the lateralization of the distracted hemimandible was often limited, with insufficient incremental gain in the bigonial distance. To overcome these shortcomings, a multiplanar distraction device was developed and tested in the canine model. This report details canine studies addressing the first problem: combined anteroposterior or sagittal (z-axis) and superoinferior or vertical (y-axis) movements. Six dogs underwent bilateral mandibular distraction with an external (extraoral), multiplanar device and completed sagittal plus vertical distraction. Evaluation included clinical examination (facial form, jaw position, and occlusion), photography, cephalograms (posteroanterior, basilar, and lateral), three-dimensional computed tomography reconstructions, and examination of dry skulls. The dogs averaged 18.5 mm (range, 15-20 mm) of sagittal distraction and 41.0 degrees (range, 21-50 degrees) of vertical distraction. Marked anterior open bites were produced after vertical distraction secondary to premature contact of the maxillary and mandibular molars. Distraction in the vertical direction also had the additive effect of increasing the sagittal gains by approximately 5% to 10%. In conclusion, a multiplanar distraction device (with the potential for distraction in three planes) was effective in increasing mandibular anteroposterior thrust (sagittal distraction) and also in creating an anterior open bite (vertical or superoinferior distraction). Vertical distraction probably requires bilateral osteotomies to obtain optimal results. The preliminary gains in sagittal length are modified (reduced or increased) after distraction in a second plane (vertical and horizontal). Specifically, vertical distraction in the inferior direction (creating an open bite) also leads to isolated increases in the anteroposterior plane. Conversely, vertical distraction in the superior direction (closing an open bite), as seen in a human malocclusion, may lead to isolated decreases in the anteroposterior plane, but this question remains to be investigated in the laboratory.

Strength of clear corneal incisions in cadaver eyes.

PURPOSE: To determine the variation in strength of clear corneal incisions, as demonstrated by degree of incision leakage when challenged by increased intraocular pressure, in relationship to the incision width and length. SETTING: Mackool Eye Institute, Astoria, New York. METHODS: Clear corneal incisions 3.0 or 3.5 mm in width and from 1.0 to 3.5 mm in length (0.5 mm increments) were studied in human cadaver whole globes. Pressure was applied at the corneal apex or 8.0 mm posterior to the external wound margin to determine the strength of the incision with the application of external force. RESULTS: Clear corneal incisions of 3.0 or 3.5 mm in width and at least 2.0 mm in length demonstrated substantially greater resistance to incision failure than shorter incision lengths with both apical and posterior applied forces. CONCLUSION: Clear corneal incisions 2.0 mm or greater in length demonstrate resistance to leakage comparable to similarly constructed scleral tunnel incisions.

Effect of foldable intraocular lens insertion on incision width.

PURPOSE: To determine whether the surgical incision enlarges during insertion of foldable intraocular lenses (IOLs). SETTING: Mackool Eye Institute, Astoria, New York. METHODS: A variety of IOL insertion devices and foldable and injectable IOLs were inserted through 3.0 or 3.5 mm keratome incisions made in cadaver eyes. The external and internal incision widths were then measured. RESULTS: Each 3.0 mm incision was enlarged externally by 0.10 to 0.65 mm and internally by 0.50 to 0.75 mm by a variety of insertional devices and IOLs. One forceps-IOL combination required a 3.5 mm incision for lens insertion and resulted in a 0.4 mm enlargement of the internal incision. CONCLUSIONS: The use of a 3.5 mm incision and insertion devices that do not enlarge an incision of this size might be desirable.

Intracapsular posterior chamber intraocular lens insertion with posterior capsular tears or zonular instability.

A surgical technique is described for posterior chamber intraocular lens implantation within the capsular bag with a posterior capsular tear or weakened zonular support. Haptics are compressed before endocapsular insertion, minimizing capsular and zonular stress.