Craniofacial biology: Animal surgical experimentation and clinical practice

Studies are presented on the growth of the mandible in the pig, growth of the frontonasal suture and snout in the rabbit, and the development of the face and jaws in a human patient with anodontia. Growth of the snout after extirpation of the frontonasal suture is contrasted with its growth following resection of the cartilaginous nasal septum. The results of the studies have clinical applications in surgery and dentistry.     

Anatomy of the nasal hump

Histologic studies were performed on the cartilaginous nasal hump in 10 cadavers (newborn to age 80) and 10 clinical patients. In all sections, the upper two-thirds of the nasal cartilaginous hump is a fused unit between the upper lateral cartilages and the septum. This fused unit is permanently altered when the hump is removed.     

Surgical correction of the severely deviated nose by extramucosal excision of the osseocartilaginous septum and replacement as a free graft

Twenty-five patients with severe internal and external deviation of the nose characterized by deviation of the septum in several planes and almost total obstruction of the airway on one or both sides were operated on. The entire bony and cartilaginous septum was removed in each of these patients, preserving the mucoperichondrial and mucoperiosteal flaps. The extramucosal technique of septal dissection was used. A support graft was fashioned from cartilaginous remnants of the septal cartilage and placed between the mucoperichondrial flaps as a free graft. All patients were followed for a minimum of 1 year. The longest follow-up is 15 years. Aesthetic improvement of the nose was obtained in all patients. All patients experienced varying degrees of improvement in nasal blockade.     

A comparative radioautographic study of chondrocytic proliferation in nasal septal cartilage of the 5-day-old rat, rabbit, guinea pig and beagle.

Growth sites within the cartilaginous nasal septa of four different species of animals (5-day-old rats, rabbits, guinea pigs and beagles) were identified by monitoring cellular proliferation radioautographically. A statistical analysis (MANOVA) was employed. It showed that, of the six combinations compared (rat-beagle, rat-guinea pig, rat-rabbit, beagle-guinea pig, beagle-rabbit, and guinea pig-rabbit), in only one (beagle-guinea pig) was there any similarity in growth pattern. The other five combinations all were significantly different. Since no particular areas emerged, with any consistency, as common growth sites within any of the four kinds of septa, it was concluded that the nasal septum might well play a passive role in midfacial growth, rather than an active role as previously thought.     

Surgical correction of the nose and midface in maxillonasal dysplasia (Binder's syndrome)

In 48 patients with maxillonasal dysplasia the retruded nasal base was corrected with onlay cancellous bone grafts after subperiosteal dissection using an oral vestibular approach. Support for the nasal dorsum was achieved in 39 patients with an L-shaped bone graft from the iliac crest introduced through the same approach. The advancement of the nose was found stable on lateral cephalograms; i.e., resorption did not occur. However, the grafts showed considerable remodeling. Half the patients found the stiffness of the nose to be disturbing. In nine patients, the cartilaginous septum was used instead as a support for the nasal dorsum and tip. At operation, the entire cartilaginous septum was mobilized after subperichondrial dissection and rotated forward either pedicled at the nasal dorsum or completely released. Cartilage regenerated in the periochondrial pocket left behind the advanced septum. The anterior transfer of the nose was 6 to 10 mm. The use of septal advancement is preferred over bone implants in the correction of maxillonasal dysplasia in patients in whom the bony nasal dorsum is of adequate height because it results in a soft and flexible nose and the risk of traumatic fracture and resorption is eliminated. The technique has been used in adolescents with promising results.     

A different rasp for rhinoplasty

A new rasp for rhinoplasty is presented. It can be dismantled and has a renewable blade that looks like the one you find in a plane. The main advantage of this rasp is that it rasps the cartilaginous septum as well as the bony part of the hump.     

Dorsal hump morphology in pink salmon (Oncorhynchus gorbuscha)

Mature male Pacific salmon (Genus Oncorhynchus) develop a dorsal hump, as a secondary male sexual characteristic, during the spawning period. Previous gross anatomical studies have indicated that the dorsal humps of salmon are mainly composed of cartilaginous tissue (Davidson [1935] J Morphol 57:169–183.) However, the histological and biochemical characteristics of such humps are poorly understood. In this study, the detailed microstructures and components of the dorsal humps of pink salmon were analyzed using histochemical techniques and electrophoresis. In mature males, free interneural spines and neural spines were located in a line near to the median septum of the dorsal hump. No cartilaginous tissue was detected within the dorsal hump. Fibrous and mucous connective tissues were mainly found in three regions of the dorsal hump: i) the median septum, ii) the distal region, and iii) the crescent‐shaped region. Both the median septum and distal region consisted of connective tissue with a high water content, which contained elastic fibers and hyaluronic acid. It was also demonstrated that the lipid content of the dorsal hump connective tissue was markedly decreased in the mature males compared with the immature and maturing males. Although, the crescent‐shaped region of the hump consisted of connective tissue, it did not contain elastic fibers, hyaluronic acid, or lipids. In an ultrastructural examination, it was found that all of the connective tissues in the dorsal hump were composed of collagen fibers. Gel electrophoresis of collagen extracts from these tissues found that the collagen in the dorsal hump is composed of Type I collagen, as is the case in salmon skin. These results indicate that in male pink salmon the dorsal hump is formed as a result of an increase in the amount of connective tissue, rather than cartilage, and the growth of free interneural spines and neural spines. J. Morphol. 275:514–527, 2014. © 2013 Wiley Periodicals, Inc.     

Deformation of nasal septal cartilage during mastication

The cartilaginous nasal septum plays a major role in structural integrity and growth of the face, but its internal location has made physiologic study difficult. By surgically implanting transducers in 10 miniature pigs (Sus scrofa), we recorded in vivo strains generated in the nasal septum during mastication and masseter stimulation. The goals were (1) to determine whether the cartilage should be considered as a vertical strut supporting the nasal cavity and preventing its collapse, or as a damper of stresses generated during mastication and (2) to shed light on the overall pattern of snout deformation during mastication. Strains were recorded simultaneously at the septo‐ethmoid junction and nasofrontal suture during mastication. A third location in the anterior part of the cartilage was added during masseter stimulation and manipulation. Contraction of jaw closing muscles during mastication was accompanied by anteroposterior compressive strains (around ?1,000 με) in the septo‐ethmoid junction. Both the orientation and the magnitude of the strain suggest that the septum does not act as a vertical strut but may act in absorbing loads generated during mastication. The results from masseter stimulation and manipulation further suggest that the masticatory strain pattern arises from a combination of dorsal bending and/or shearing and anteroposterior compression of the snout. J. Morphol., 2009. © 2009 Wiley‐Liss, Inc.     

Formation of cartilage in the heart of the Spanish terrapin,Mauremys leprosa (Reptilia,Chelonia)

Cartilaginous deposits are regularly present in the heart of several reptilian, avian, and mammalian species. The formation of these extraskeletal cartilages has been studied in birds and mammals, but not in reptiles. The aim here was to elucidate this question in the Spanish terrapin. Hearts from 23 embryos belonging to Yntema (1968) developmental stages 17 to 26 and eight terrapins age 3 months to 10 years were examined using histological, histochemical, and immunohistochemical techniques. In the heart of the Spanish terrapin (Mauremys leprosa), chondrogenesis can start during embryonic life. Cartilaginous tissue develops from a mesenchymal cellular condensation that extends along the aorticopulmonary septum and the incipient pars fibrosa of the ventricular horizontal septum. This cellular condensation, which is smooth muscle alpha-actin (SMalpha-actin)-negative and type II collagen-negative during stages 17 to 22, acts as a prechondrogenic condensation. In stage 23, production of type II collagen begins in the central core of the condensation and gradually spreads toward its periphery. The type II collagen-positive (chondrogenic) cellular condensation remains devoid of perichondrium prior to birth. Thereafter, it converts into hyaline cartilage that extends along the proximal part of the aorticopulmonary septum and the pars fibrosa of the horizontal septum. Our findings are consistent with the assumption that, as in birds and mammals, the precursors of the cardiac chondrocytes in chelonians are neural crest-derived cells of nonmuscular nature. In addition, they point to the possibility that cells from the neural crest populate the embryonic pars fibrosa of the horizontal septum, thereby contributing to its alignment with the aorticopulmonary septum. In the present species, a second cartilaginous deposit of a hyaline nature extends along the sinus wall of the right semilunar valve of the right aorta, penetrating the fibrous cushion that constitutes the proximal support of the corresponding valve leaflet. This cartilage develops after birth, between the third and eighteenth month of life; its morphogenetic origin is unclear. The cartilaginous foci occurring in hearts of Spanish terrapin appear to act as pivots resisting mechanical tensions generated during the cardiac cycle. In the specimens examined there was no sign of replacement of the cardiac cartilages by bone tissue.     

Morphology of the interorbital region of Saimiri sciureus

The skull of the platyrrhine primate Saimiri sciureus is distinguished by a large interorbital fenestra. Juvenile skulls still show a bony interorbital septum with some small gaps. A morphogenetic study was undertaken to better understand the structures of the interorbital region, which represents a linkage between the base of the braincase and the nasal skeleton. Already in early ontogenetic stages a reduction of the posterior portion of the nasal capsule and of the cartilaginous interorbital septum are observed, resulting in the formation of a primary interorbital fenestra. A bony interorbital septum is mainly formed in perinatal age stages by ossification of the presphenoid and by medial fusion of the frontals; the primary interorbital fenestra is retained as a small opening. It only occurs in late juvenile stages when the definitive interorbital fenestra develops by by secondary transformation of bone into a membrane of dense connective tissue; this process is most probably caused by mechanical friction of the very closely approximated eyes of both sides.     

In vitro growth of the nasal septal cartilage of the rat in a serum-free culture medium. [3H]-thymidine incorporation studies

To evaluate the capacity for growth of cartilage in vitro, the cartilaginous nasal septum was cultured in a serum-free medium in short- and long-term incubation periods. The proliferative activity was assessed by relating counts per minute (cpm) obtained in the liquid scintillater after [3H]-thymidine incorporation with the DNA content of the tissue under investigation. Cell proliferation was interpreted as being synonymous with the DNA synthesizing activity or the cpm/microgram DNA ratio. Short-term incubation (24 h) tended to show the same proliferative activity which we have previously found in vivo and as autotransplants, showing that the presence of serum seems to be of little importance under these conditions. Long-term incubation in a serum-free medium tended to show a gradual loss of proliferative activity, emphasizing the need for either serum or other growth-promoting factors as necessary for continued normal proliferation of cartilage in vitro.     

Anatomy of the nasal cartilages of the unilateral complete cleft lip nose

The purpose of this study was to disclose the relationship between the anomaly of the cartilaginous framework and the nasal deformity of cleft lip. The noses of six stillborn infants with unilateral complete cleft lip were carefully dissected. The size and weight of the lower lateral cartilages were measured to determine whether there was a significant difference between the normal and involved sides. The position of the nasal cartilages was observed, and the distance between them was measured to determine whether they were normal. The surgical dissection revealed that the lower lateral cartilages from both sides were asymmetrical in three dimensions, indicating the displacement of the lower lateral cartilage on the involved side. There was displacement of the cartilaginous septum and the upper lateral cartilage. The statistical evaluation did not demonstrate a significant difference between weight and size of the two sides. One of the major causative factors of nasal deformity is displacement of the nasal cartilages. There is no hypoplasia of nasal cartilage in newborn infants with cleft lip.     

Relationship of Nandinia binotata (Gray) to the Superfamily Feloidea (Mammalia, Carnivora)

The phylogenetic relationship between Nandinia binotata and Feloidea is analysed by the cladistic method, based on a literature review of osteological characters used in systematic works on carnivores for more than a century. The reduced or lost postglenoid foramen is a synapomorphy that define Nandinia and Feloidea as a monophyletic group. Nandinia does not have an ectotympanic septum in the bulla nor a paroccipital process nested with the posterior wall of the bulla, which are autapomorphies for the Feloidea. Thus it is hypothesized that Nandinia binotata has a sister-group relation to Feloidea. The cartilaginous caudal entotympanic is an autapomorphy for Nandinia.     

Specific effects of retinoic acid on the skeletal morphogenesis of the 11-day mouse embryo forelimb bud in vitro

Using our improved method for culturing 11-day mouse forelimb buds in vitro, we have investigated the effects of a local application of all-trans-retinoic acid (RA) on growth, cartilaginous differentiation and skeletal patterning in the mammalian limb bud. Carrier implants of catgut impregnated with DMSO or various doses of RA in DMSO were inserted at the apex of the buds in the proximo-distal axis just beneath the apical ectodermal ridge. After 6 days of culture, cartilaginous skeletons were stained and explants were processed for morphological analysis and quantitative study using computerized optical image analysis. Buds treated with low doses of RA exhibited stimulated growth and chondrogenesis. Moreover, hypertrophied and fused metacarpals were seen within explants treated with the lowest dose. High doses strongly inhibited growth and skeletal morphogenesis. An intermediate dose sustained cartilaginous differentiation at the same level as low doses, but concomitantly disturbed the skeletal pattern. These results are discussed considering reported RA effects on other experimental systems including avian limb bud as an in vivo model or cell cultures as an in vitro simplified model.     

Use of cartilaginous autografts in nasal surgery: 8 years of experience

Although the cartilaginous autografts are one of the tissues more utilized in nasal surgery, a comparative study does not exist to determine which are better options and their precise indications. It is for this reason that a histopathologic analysis was carried out comparing the characteristics and properties of the four principal cartilages that are utilized in aesthetic functional surgery of the nose. Considering these particularities, the precise indications for the employment of the different cartilage as nasal autografts were determined. Of 1120 aesthetic functional rhinoplasties during a period of 8 years, 930 (83 percent) required cartilaginous autografts, 86 percent were primary, 11 percent were secondary, and 3 percent had two or more surgeries. Eighty-three percent of the grafts used were from nasal septum, 12 percent from the auricle, 3 percent from alar cartilages, and 2 percent from the rib. The anatomic sites in which they were employed consisted of the following: 64 percent between the medial crura, 28 percent as in Sheen's graft, 19 percent in the nasal dorsum, 8 percent as spreader grafts, 8 percent as in Peck's graft, and 3 percent in the rim to improve alar collapse. We followed at all times the previous indications for obtaining and placing the nasal autografts. Eighty-four percent of the patients were totally satisfied and only 8 percent required a second surgical procedure to achieve the results desired. Based on this study, it is recommended to utilize the cartilaginous autografts in nasal surgery considering three parameters: the physical and histologic characteristics of each cartilage, the anatomic site in which they are to be placed, and the effect desired with their application.     

Immunohistochemical Localization of Bone Morphogenetic Proteins (BMPs) and their Receptors in Solitary and Multiple Human Osteochondromas

The expression of bone morphogenetic proteins (BMPs) and their cognate receptors (BMPRs) in osteochondromas has not been investigated. We determined the immunohistochemical localization and distribution of BMP-2/4, -6 and -7; BMP receptors BMPR-1A, BMPR-1B and BMPR-2; signal transducing proteins phosphorylated Smad1/5/8; and BMP antagonist noggin in the cartilaginous cap of solitary (SO) and multiple (MO) human osteochondromas and compared these with bovine growth plate and articular cartilage. The distribution and localization patterns for BMP-6, BMP-7, BMPR-1A and BMPR-2 were similar between the cartilaginous cap and the growth plate. BMP-2/4 and BMPR-1B were present throughout the growth plate. However, BMP-2/4 and phosphorylated Smad1/5/8 were mainly detected in proliferating chondrocytes of the cartilaginous cap. Also, BMPR-1B was found in hypertrophic chondrocytes of SO and proliferating chondrocytes of MO. Noggin was observed in resting chondrocytes and, to a lesser extent, in clustered proliferating chondrocytes in SO. On the other hand, noggin in MO was observed in proliferating chondrocytes. Since BMPs can stimulate proliferation and hypertrophic differentiation of chondrocytes, these findings suggest that there is an imbalance of BMP-2/4 and noggin interactions that may lead to abnormal regulation of chondrocyte proliferation and differentiation in the cartilaginous cap of human osteochondromas.     

Specific effects of retinoic acid on the skeletal morphogenesis of the 11-day mouse embryo forelimb bud in vitro

Using our improved method for culturing 11-day mouse forelimb buds in vitro, we have investigated the effects of a local application of all-trans-retinoic acid (RA) on growth, cartilaginous differentiation and skeletal patterning in the mammalian limb bud. Carrier implants of catgut impregnated with DMSO or various doses of RA in DMSO were inserted at the apex of the buds in the proximo-distal axis just beneath the apical ectodermal ridge. After 6 days of culture, cartilaginous skeletons were stained and explants were processed for morphological analysis and quantitative study using computerized optical image analysis. Buds treated with low doses of RA exhibited stimulated growth and chondrogenesis. Moreover, hypertrophied and fused metacarpals were seen within explants treated with the lowest dose. High doses strongly inhibited growth and skeletal morphogenesis. An intermediate dose sustained cartilaginous differentiation at the same level as low doses, but concomitantly disturbed the skeletal pattern. These results are discussed considering reported RA effects on other experimental systems including avian limb bud as an in vivo model or cell cultures as an in vitro simplified model.     

Localization and activity of the calcineurin catalytic and regulatory subunit complex at the septum is essential for hyphal elongation and proper septation in Aspergillus fumigatus

Calcineurin, a heterodimer composed of the catalytic (CnaA) and regulatory (CnaB) subunits, plays key roles in growth, virulence and stress responses of fungi. To investigate the contribution of CnaA and CnaB to hyphal growth and septation, ΔcnaB and ΔcnaAΔcnaB strains of Aspergillus fumigatus were constructed. CnaA colocalizes to the contractile actin ring early during septation and remains at the centre of the mature septum. While CnaB's septal localization is CnaA-dependent, CnaA's septal localization is CnaB-independent, but CnaB is required for CnaA's function at the septum. Catalytic null mutations in CnaA caused stunted growth despite septal localization of the calcineurin complex, indicating the requirement of calcineurin activity at the septum. Compared to the ΔcnaA and ΔcnaB strains, the ΔcnaAΔcnaB strain displayed more defective growth and aberrant septation. While three Ca(2+) -binding motifs in CnaB were sufficient for its association with CnaA at the septum, the amino-terminal arginine-rich domains (16-RRRR-19 and 44-RLRKR-48) are dispensable for septal localization, yet required for complete functionality. Mutation of the 51-KLDK-54 motif in CnaB causes its mislocalization from the septum to the nucleus, suggesting it is a nuclear export signal sequence. These findings confirm a cooperative role for the calcineurin complex in regulating hyphal growth and septation.