Rat Hard Palate Research Articles

Functional Validation of a New Alginate-based Hydrogel Scaffold Combined with Mesenchymal Stem Cells in a Rat Hard Palate Cleft Model.

Background:One of the major difficulties in cleft palate repair is the requirement for several surgical procedures and autologous bone grafting to form a bony bridge across the cleft defect. Engineered tissue, composed of a biomaterial scaffold and multipotent stem cells, may be a useful alternative for minimizing the non-negligible risk of donor site morbidity. The present study was designed to confirm the healing and osteogenic properties of a novel alginate-based hydrogel in palate repair.Methods:Matrix constructs, seeded with allogeneic bone marrow–derived mesenchymal stem cells (BM-MSCs) or not, were incorporated into a surgically created, critical-sized cleft palate defect in the rat. Control with no scaffold was also tested. Bone formation was assessed using microcomputed tomography at weeks 2, 4, 8, and 12 and a histologic analysis at week 12.Results:At 12 weeks, the proportion of bone filling associated with the use of hydrogel scaffold alone did not differ significantly from the values observed in the scaffold-free experiment (61.01% ± 5.288% versus 36.91% ± 5.132%; p = 0.1620). The addition of BM-MSCs stimulated bone formation not only at the margin of the defect but also in the center of the implant.Conclusions:In a relevant in vivo model of cleft palate in the rat, we confirmed the alginate-based hydrogel’s biocompatibility and real advantages for tissue healing. Addition of BM-MSCs stimulated bone formation in the center of the implant, demonstrating the new biomaterial’s potential for use as a bone substitute grafting material for cleft palate repair.

Development of an Animal Model for Aggregatibacter Actinomycetemcomitans Biofilm‐Mediated Oral Osteolytic Infection: A Preliminary Study

Biofilm-induced inflammatory osteolytic oral infections, such as periodontitis and peri-implantitis, have complex etiology and pathogenesis. A significant obstacle to research has been the lack of appropriate animal models where the inflammatory response to biofilms can be investigated. The aim of this study is to develop a novel animal model to study the host response to Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans)-biofilm colonizing titanium implants. Titanium implants were inoculated in vitro with A. actinomycetemcomitans, establishing a biofilm for 1 to 3 days. Biofilm-inoculated and control implants were transmucosally placed into rat hard palate or alveolar ridge. Analysis included documentation of clinical inflammation, polymerase chain reaction, and culture detection of A. actinomycetemcomitans and microcomputed tomography quantitation of peri-implant bone volume. Viable A. actinomycetemcomitans biofilm was successfully established on titanium implants in vitro, detected by confocal laser scanning microscopy. An inflammatory response characterized by clinical inflammation, bleeding, ulceration, hyperplasia, and necrosis was observed around biofilm-inoculated implants. A. actinomycetemcomitans was detected by polymerase chain reaction and culture analysis on 100% of biofilm-inoculated implants for up to 3 weeks and 25% for up to 6 weeks. Microcomputed tomography analysis demonstrated significantly lower bone volume (P <0.05) around biofilm-inoculated implants (29.6% ± 7.6%) compared to non-inoculated implants (50.5% ± 9.6%) after 6 weeks. These results describe a novel animal model where A. actinomycetemcomitans biofilm was established in vitro on titanium implants before placement in rat oral cavity, leading to an inflammatory response, osteolysis, and tissue destruction. This model may have potential use for investigation of host responses to biofilm pathogens and antibiofilm therapy.

Effects of ovariectomy and/or dietary calcium deficiency on bone dynamics in the rat hard palate, mandible and proximal tibia

The effects of ovariectomy and/or dietary calcium deficiency on bone dynamics were examined by comparing the histomorphometric changes in these bones. Five groups of rats were studied, (1) unoperated basal controls; (2) sham-operated, fed on a normal-calcium diet; (3) ovariectomized, fed on a normal-calcium diet; (4) sham-operated, fed on a calcium-deficient diet; (5) ovariectomized, fed on a calcium-deficient diet. The basal controls were killed at 6 weeks of age, and the remaining groups were killed at 18 weeks of age. The hard palate, mandible and proximal tibia were processed undemineralized for quantitative bone histomorphometry. Bone volume, eroded surface, osteoid surface and bone-formation ratio were calculated. A significant age-related increase in bone volume and a significant decrease in bone formation were observed in the hard palate and mandible, whereas no significant age-related increase in bone volume could be found in the tibia. In the hard palate, ovariectomy neither inhibited age-related increases in bone volume nor affected bone dynamics, while both the combined ovariectomy and dietary calcium deficiency and dietary calcium deficiency alone led to bone loss and increased bone turnover. In contrast, in the mandible and proximal tibia, ovariectomy alone as well as dietary calcium deficiency led to bone loss and increased bone turnover. Ovariectomy, therefore, produced no significant changes in the hard palate, but affected bone dynamics in the mandible and tibia. However, dietary calcium deficiency induced bone loss and increased bone turnover in the hard palate, mandible and proximal tibia, independently of ovariectomy.

Innervation of the hard palate in the rat studied by anterograde transport of horseradish peroxidase conjugates.

The innervation of the rat hard palate and the bordering part of the soft palate was studied after anterograde transport of horseradish peroxidase conjugated to wheat germ agglutinin (WGA-HRP) and to choleragenoid (B-HRP) in separate experiments. WGA-HRP labeling showed leakage from several types of nerve endings, whereas B-HRP did not. Both conjugates gave rise to heavy labeling of a variety of nerve endings. Intragemmal and, especially, perigemmal fibers were labeled in chemosensory corpuscles, which were most common in the medial wall of the incisive canal and in the most anterior part of the soft palate. Ruffini endings of different sizes were labeled in the incisive papilla. Other subepithelial endings forming elongated expanded profiles with medium- to large-caliber source fibers were most common in protruding parts of the palate. Labeled intraepithelial endings included Merkel endings, which were most frequent in the incisive papilla and the rugae. Other labeled profiles were medium-caliber afferents giving rise to irregular, beaded, and sometimes branched endings often located far superficially in the epithelium. Such endings were present both within and between protruding parts of the palate. Fine-caliber intraepithelial endings were labeled almost exclusively in WGA-HRP experiments.

Effects of Growth Factors on Mucosal Scar Fibroblasts in Culture — a Possible Role of Growth Factors in Scar Formation

Various growth factors are known to play important roles in wound healing, especially in an early inflammatory phase. However, their roles in subsequent scar formation phase are relatively unexplored. The aim of this study is to investigate the mechanisms of regulation in scar formation by these factors.Scar fibroblasts (SF) were obtained from immature scar tissue made at rat hard palate 1 month after excision and normal fibroblasts (NF) were obtained from the palatal mucosa of untreated control animals. SF showed a longer doubling time, and increased level of protein synthesis when compared to NF. Platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) stimulated [3H]-thymidine uptake less effectively in SF than in NF. In both cells, transforming growth factor-β1 (TGF-β1) inhibited EGF-induced stimulation of [3H]-thymidine uptake, but had no effects when it was added alone. TGF-β1 increased collagen synthesis more effectively in SF than in NF.These data indicate that the growth factors may play key roles in regulating proliferation and metabolic activity of fibroblasts during scar formation.

Substance-P-Containing Fibers in the Incisive Papillae of the Rat Hard Palate

Substance P (SP)-containing fibers in the incisive papillae of rat hard palates, which include various components of sensory receptors, i.e. mechanoreceptors, free nerve endings and chemosensory corpuscles (taste buds), were examined using immunoperoxidase techniques and light and electron microscopes. Immunolabeled fibers were consistently distributed in the medial part of the orifice of the incisive canals, i.e. in the taste-bud-enriched region. Dense immunolabeled fibers were found in subgemmal regions and in the lamina propria papillae. Some fine fibers entered and ascended the taste buds or occasionally the epithelium outside the taste buds. In addition, a rich innervation by SP-containing fibers close to blood capillaries was clearly identified. Electron microscopy revealed no specialized synaptic contact between the immunolabeled fibers and taste bud cells. Synaptic-like images could be found only between nonimmunolabeled nerve endings and the underlying taste bud cells. In the lamina propria papillae, mechanoreceptors observed in the present study contained no immunoperoxidase end products, whereas free nerve endings with an immunolabeled small-diameter axon (630-730 nm in diameter) were frequent. Similar axons were located at the adventitia of the blood capillaries. The possible functional role of SP-containing fibers in the incisive papillae was given attention.

Sensory nerve endings of the incisive papilla of rat hard palate studied by peroxidase cytochemical methods.

This study utilized peroxidase cytochemical methods to survey and analyze the distribution, morphology, and functional relationship of the various components of sensory nerve endings in a focal region of the anterior hard palate (incisive papilla) of the rat. The studies included an anatomical survey of this relatively unknown oral structure, confirmation of the trigeminal origin (labeled by retrograde axonal transport of WGA-HRP injected into the incisive papilla) of the sensory nerve endings, and a complete analysis of the distribution of these nerve endings (labeled by anterograde axonal transport of HRP or WGA-HRP injected into the trigeminal ganglion). Three methods of fixation and two methods of cytochemical reaction were used for selection of an optimal technique for these studies. The results showed distinct patterns of peroxidase-labeled sensory nerve endings at three regions of the incisive papilla: a) dome region (ventral), where labeled nerve endings formed three parallel channels in association with three surface ridges; b) chemosensory corpuscle enriched region (medial to incisive canal), where 82% of the 30-40 chemosensory corpuscles were labeled; and c) lateral labium (lateral to incisive canal), where labeled nerve endings formed a circumscribed network guarding the orifice of incisive canal. The discrete organization of multiple sensory nerve endings in the incisive papilla of the rat may provide an easily accessible model system for various studies in sensory physiology.

口こう粘膜の創傷治ゆに関する実験的研究

The rate of epithelialization on intraoral operative wounds was studied with special reference to the influence of blood supply disturbance on mucosa surrounding wound and of the nature of wound surface. A standardized wound was made on the rat hard palate and after various surgical treatments the epithelial extension from the wound edge was observed.Results1) Epithelial extension depended mainly on migration of the epithelial cell.2) The disturbance of blood supply to the surrounding mucosa prolonged the lag period, then the extension started. This factor did not infleunce the rate of epithelialization thereafter.3) In the groups in which the deep portion of the lamina propria was preserved on the wound surface, the rate of epithelialization was 0.19-0.21 mm/day. This value was similar to that reported previously about open skin wounds.Saliva eased dehydration of the connective tissue surface of wounds but did not accelerate epithelialization.4) In the groups in which the palatal bone was exposed to the wound surface, epithelialization was very slow because proliferation of the connective tissue was needed for the epithelial extension. But once the wound surface was covered with a new connective tissue, rate increased to half the level of the groups with the intact deep portion of the lamina propria preserved.5) When the deep portion of the lamina propria, which was suffering from blood supply disturbance, was left on the wound surface, the rate of epithelialization was half the level of groups with the intact deep portion of the lamina propria preserved.6) Edema and inflammation in the wound connective tissue decreased the rate of epithelialization.7) From these findings, it can be stated that the nature of the wound surface is a very important factor which influences epithelialization of intraoral operative wounds. To increase the rate of epithelialization, it is effective to preserve the intact lamina propria in the wound and secondarily to accelerate proliferation of the granulation tissue. It is also effective to prevent inflammation of the connective tissue in the wound.