Human Alveolar Bone Research Articles

Primary Human Alveolar Bone Cells Isolated from Tissue Samples Acquired at Periodontal Surgeries Exhibit Sustained Proliferation and Retain Osteogenic Phenotype during In Vitro Expansion

ObjectivesBone tissue regeneration requires a source of viable, proliferative cells with osteogenic differentiation capacity. Periodontal surgeries represent an opportunity to procure small amounts of autologous tissues for primary cell isolation. Our objective was to assess the potential of human alveolar bone as a source of autologous osteogenic cells for tissue engineering and biomaterials and drug testing studies.Materials and MethodsAlveolar bone tissue was obtained from 37 patients undergoing routine periodontal surgery. Tissue harvesting and cell isolation procedures were optimized to isolate viable cells. Primary cells were subcultured and characterized with respect to their growth characteristics, gene expression of osteogenic markers, alkaline phosphatase activity and matrix mineralization, under osteogenic stimulation.ResultsAlveolar bone cells were successfully isolated from 28 of the 30 samples harvested with bone forceps, and from 2 of the 5 samples obtained by bone drilling. The yield of cells in primary cultures was variable between the individual samples, but was not related to the site of tissue harvesting and the patient age. In 80% of samples (n = 5), the primary cells proliferated steadily for eight subsequent passages, reaching cumulative numbers over 1010 cells. Analyses confirmed stable gene expression of alkaline phosphatase, osteopontin and osteocalcin in early and late cell passages. In osteogenic medium, the cells from late passages increased alkaline phosphatase activity and accumulated mineralized matrix, indicating a mature osteoblastic phenotype.ConclusionsPrimary alveolar bone cells exhibited robust proliferation and retained osteogenic phenotype during in vitro expansion, suggesting that they can be used as an autologous cell source for bone regenerative therapies and various in vitro studies.

The effect of epigallocatechin-3-gallate (EGCG) on human alveolar bone cells both in vitro and in vivo

ObjectiveThe effects of epigallocatechin-3-gallate (EGCG), a major catechin in green tea, on human and mouse osteoblasts remain controversial. This study investigated the direct effects of EGCG on human alveolar bone-derived cells (hABCs) both in vitro and in vivo. DesignhABCs which were collected from eight children (aged 7–9 years, seven males and one female) were treated with EGCG at various concentrations (1, 5, 10, 25, and 50μM), and a proliferation assay, flow cytometric analysis for apoptosis evaluation, migration assay, and in vitro osteogenic differentiation were performed. hABCs that were pretreated with 10μM EGCG and mixed with calcium phosphate carrier combined with EGCG (0.1, 0.5, or 1.5mg) in vivo were transplanted into immunodeficient mouse. Histological staining, quantitative gene expressions, and alkaline phosphatase activity were evaluated in the retrieved transplants. ResultsThe proliferation and migration were decreased when EGCG was present at over 25μM. The osteogenic differentiation increased slightly when EGCG was present at up to 10μM, and clearly decreased for higher concentrations of EGCG. In vivo, the potential for hard-tissue formation was slightly higher for the group with 0.1mg of EGCG than for the control group, and decreased sharply for higher concentrations of EGCG. ConclusionThe present observations suggest that EGCG at a low concentration can slightly enhance the osteogenic effect in vivo, whereas at a higher concentration it can prevent the osteogenic differentiation of hABCs both in vitro and in vivo.

Differences in morphogenesis of 3D cultured primary human osteoblasts under static and microfluidic growth conditions

As information on osteoblast mechanosensitivity response to biomechanical cues in three-dimensional (3D) in vitro microenvironments is sparse, the present study compared morphogenesis of primary human alveolar bone osteoblasts (PHABO) under microchip-based 3D-static conditions, and 3D-fluid flow-mediated biomechanical stimulation in perfusion bioreactors. Discrimination of the respective microenvironment by differential morphogenesis was evident from fluid flow-induced PHABO reorganization into rotund bony microtissue, comprising more densely packed multicellular 3D-aggregates, while viability of microtissues was flow rate dependent. Time-lapse microscopy and simple modeling of biomechanical conditions revealed that physiologically relevant fluid flow-mediated PHABO stimulation was associated with formation of mulberry-like PHABO aggregates within the first 24 h. Differential extracellular matrix deposition patterns and gene expression modulation in PHABO aggregates at day 7 further indicates progressive osteoblast differentiation exclusively in perfusion culture-developed bony microtissues. The results of our study strongly suggest PHABO morphogenesis as discriminator of microenvironmental growth conditions, which in case of the microfluidic 3D microchip-bioreactor are substantiated by triggering in vitro bone microtissue formation concomitant with progressive osteoblastic differentiation. Such microtissue outcomes provide unique insight for mechanobiological studies in response to biomechanical fluid flow cues, and clinically appear promising for in vitro PHABO preconditioning, enabling innovative bone augmentation procedures.

Real-time quantitative polymerase chain reaction and flow cytometric analyses of cell adhesion molecules expressed in human cell–multilayered periosteal sheets in vitro

Background aimsCultured human periosteal sheets more effectively function as an osteogenic grafting material at implantation sites than do dispersed periosteal cells. Because adherent cell growth and differentiation are regulated by cell-cell and cell–extracellular matrix contacts, we hypothesized that this advantage is a result of the unique cell adhesion pattern formed by their multiple cell layers and abundant extracellular matrix. To test this hypothesis, we prepared three distinct forms of periosteal cell cultures: three-dimensional cell-multilayered periosteal sheets, two-dimensional dispersed cell cultures, and three-dimensional hybrid mock-ups of cells dispersed onto collagen sponges. MethodsPeriosteal cells were obtained from human alveolar bone. Cell adhesion and extracellular matrix molecules were quantitatively determined at the messenger RNA and protein levels by means of real-time quantitative polymerase chain reaction and flow cytometry, respectively. ResultsReal-time quantitative polymerase chain reaction analysis demonstrated that regardless of culture media α1 integrin, vascular cell adhesion molecule-1, fibronectin and collagen type 1 were substantially upregulated, whereas CD44 was strongly downregulated in periosteal sheets compared with dispersed cell monolayers. With increased thickness, stem cell medium upregulated several integrins (β1, α1 and α4), CD146, vascular cell adhesion molecule-1, fibronectin and collagen type 1 in the periosteal sheets. Flow cytometric analysis revealed that the active configuration of β1 integrin was substantially downregulated in the stem cell medium–expanded cell cultures. The cell adhesion pattern found in the mock-up cultures was almost identical to that of genuine periosteal sheets. ConclusionsIntegrin α1β1 and CD44 function as the main cell adhesion molecule in highly cell-multilayered periosteal sheets and dispersed cells, respectively. This difference may account for the more potent osteogenic activity shown by the thicker periosteal sheets.

低出力超音波パルスを照射したヒト歯槽骨骨膜由来細胞 スフェロイドの組織学的変化

我々は，イヌの実験的骨欠損部に低出力超音波パルス（LIPUS）を照射すると，歯周組織の治癒が促進されることを報告した。しかし，その詳細なメカニズムは明らかでないため，今回は，ヒト歯槽骨骨膜由来細胞（HABPCs）の細胞凝集塊(スフェロイド)を作製して LIPUS を照射し，スフェロイド内の細胞形態および骨基質タンパク発現に与える LIPUS の影響を経時的に検索した。1 日，3，7，および 14 日間，LIPUS を照射したスフェロイドに対し，トルイジンブルーを用いた組織学的検索と，抗オステオポンチン（OP)，抗オステオカルシン（OC) を用いた免疫組織化学的検索を行ったところ，1 日目から細胞の走行などにより区別された層構造がスフェロイドに認められたが，照射近位部，照射遠位部，コントロールの順で経時的に層構造は不明瞭になっていった。また，全てのスフェロイドに深層から中層，表層の順で OP と OC 免疫陽性反応が認められた。経時的には，照射近位部，照射遠位部，コントロールの順に，OP，OC 免疫陽性反応が認められ，照射近位部では 1 日目よりスフェロイドの表層から深層へ向けて索状構造が観察された。これらの結果から，LIPUS 照射の影響が大きい照射近位部で早期に OP，OC の発現と索状構造が認められたため，LIPUS 照射により，スフェロイド内の HABPCs の細胞分化の促進と細胞の方向性の変化が起こることが示唆された。 日本歯周病学会会誌（日歯周誌）55(4)：312-325，2013

The Influence of β‐Tricalcium Phosphate Blocks Containing Extracellular Matrix on Osteogenic Differentiation of Rat Bone Marrow Stromal Cells

The aim of the present study is to assess the osteogenic differentiation of rat bone marrow stromal cells (RBMCs) in β-tricalcium phosphate (β-TCP) blocks containing extracellular matrix (ECM) produced by human alveolar bone periosteal cells (HABPCs). HABPCs were cultured in β-TCP blocks for 1 week (group 1) and 2 weeks (group 2). β-TCP blocks containing ECM were then created by drying the blocks for 3 days. RBMCs were cultured in the blocks containing ECM for 2 weeks. In the control group, RBMCs were cultured in β-TCP blocks alone for 2 weeks. HABPCs and RBMCs in the blocks were examined by histologic and immunohistochemical analyses. Histology revealed a significantly higher number of HABPCs in the group 2 blocks than in the group 1 blocks. HABPCs produced several bone matrix proteins in the blocks, and these positive reactions in group 2 were significantly higher than in group 1. Both groups showed a significantly higher number of RBMCs than the control group. RBMCs produced osteopontin and osteocalcin in the blocks, and these positive reactions were significantly higher in both test groups than in the control group. The number of osteocalcin-positive reactions was higher in group 2 than in group 1. Osteogenic differentiation of RBMCs cultured for 2 weeks in β-TCP blocks containing ECM was significantly higher than that of cells cultured for 1 week or without ECM.

Effect of Emdogain enamel matrix derivative and BMP-2 on the gene expression and mineralized nodule formation of alveolar bone proper-derived stem/progenitor cells

The objective of this study was to evaluate the effect of Emdogain (Enamel Matrix Derivative, EMD) and Bone Morphogenetic Protein-2 (BMP-2), either solely or in combination, on the gene expression and mineralized nodule formation of alveolar bone proper-derived stem/progenitor cells.Stem/progenitor cells were isolated from human alveolar bone proper, magnetically sorted using STRO-1 antibodies, characterized flowcytometrically for their surface markers' expression, and examined for colony formation and multilineage differentiation potential. Subsequently, cells were treated over three weeks with 100 μg/ml Emdogain (EMD-Group), or 100 ng/ml BMP-2 (BMP-Group), or a combination of 100 ng/ml BMP-2 and 100 μg/ml Emdogain (BMP/EMD-Group). Unstimulated stem/progenitor cells (MACS+-Group) and osteoblasts (OB-Group) served as controls. Osteogenic gene expression was analyzed using RTq-PCR after 1, 2 and 3 weeks (N = 3/group). Mineralized nodule formation was evaluated by Alizarin-Red staining.BMP and EMD up-regulated the osteogenic gene expression. The BMP Group showed significantly higher expression of Collagen-I, III, and V, Alkaline phosphatase and Osteonectin compared to MACS+- and OB-Group (p < 0.05; Two-way ANOVA/Bonferroni) with no mineralized nodule formation.Under in-vitro conditions, Emdogain and BMP-2 up-regulate the osteogenic gene expression of stem/progenitor cells. The combination of BMP-2 and Emdogain showed no additive effect and would not be recommended for a combined clinical stimulation.

Human mesenchymal progenitor cells derived from alveolar bone and human bone marrow stromal cells: a comparative study

The aim of the present study was to evaluate the potential of intraoral harvested alveolar bone as an alternative source of multipotent mesenchymal stromal cells for future applications in oral and maxillofacial tissue engineering. Explant cultures were established from 20 alveolar bone samples harvested from the oblique line immediately before wisdom tooth removal. Morphology and proliferation characteristics of the in vitro expanded cells, referred to as human alveolar bone-derived cells (hABDCs), were studied using phase-contrast microscopy. Immunocytochemical analysis of their surface marker expression was conducted using monoclonal antibodies defining mesenchymal stromal cells. To evaluate their multilineage differentiation potential, hABDCs were induced to differentiate along the osteogenic, adipogenic, and chondrogenic lineage and compared to bone marrow mesenchymal stromal cells (hBMSCs) on mRNA and protein levels applying RT-PCR and cytochemical staining methods. hABDCs showed typical morphological characteristics comparable to those of hBMSCs such as being mononuclear, fibroblast-like, spindle-shaped, and plastic adherent. Immunophenotypically, cells were positive for CD105, CD90, and CD73 while negative for CD45, CD34, CD14, CD79α, and HLA-DR surface molecules, indicating an antigen expression pattern considered typical for multipotent mesenchymal stromal cells. As evidenced by RT-PCR and cytochemistry, hABDCs showed multilineage differentiation and similar chondrogenic and osteogenic differentiation potentials when compared to hBMSCs. Our findings demonstrate that human alveolar bone contains mesenchymal progenitor cells that can be isolated and expanded in vitro and are capable of trilineage differentiation, providing a reservoir of multipotent mesenchymal cells from an easily accessible tissue source.

Micro-Computed Tomography Assessment of Human Alveolar Bone: Bone Density and Three-Dimensional Micro-Architecture

Micro-computed tomography (micro-CT) is a valuable means to evaluate and secure information related to bone density and quality in human necropsy samples and small live animals. The aim of this study was to assess the bone density of the alveolar jaw bones in human cadaver, using micro-CT. The correlation between bone density and three-dimensional micro architecture of trabecular bone was evaluated. Thirty-four human cadaver jaw bone specimens were harvested. Each specimen was scanned with micro-CT at resolution of 10.5 μm. The bone volume fraction (BV/TV) and the bone mineral density (BMD) value within a volume of interest were measured. The three-dimensional micro architecture of trabecular bone was assessed. All the parameters in the maxilla and the mandible were subject to comparison. The variables for the bone density and the three-dimensional micro architecture were analyzed for nonparametric correlation using Spearman's rho at the significance level of p < .05. A wide range of bone density was observed. There was a significant difference between the maxilla and mandible. All micro architecture parameters were consistently higher in the mandible, up to 3.3 times greater than those in the maxilla. The most linear correlation was observed between BV/TV and BMD, with Spearman's rho = 0.99 (p = .01). Both BV/TV and BMD were highly correlated with all micro architecture parameters with Spearman's rho above 0.74 (p = .01). Two aspects of bone density using micro-CT, the BV/TV and BMD, are highly correlated with three-dimensional micro architecture parameters, which represent the quality of trabecular bone. This noninvasive method may adequately enhance evaluation of the alveolar bone.

Distinct Cell Functions of Osteoblasts on UV-Functionalized Titanium- and Zirconia-Based Implant Materials Are Modulated by Surface Topography

Though recent studies report decisive positive effects on cells, elicited by ultraviolet (UV)-induced bioactivation of biomaterial implant surfaces, they frequently employ cells other than of human origin or cells not representing oral implant targets. Therefore, the present study aims at exploring distinct cell functions of primary human alveolar bone osteoblasts (PHABO) in response to bioactivated microstructured titanium and zirconia implant surfaces with matched controls. UV-treatment significantly reduced surface carbon, while concomitantly increasing wettability. In case of titanium or zirconia biomaterial source of equal roughness, bioactivation did not significantly improve cell functions, including initial cell attachment, morphogenesis, proliferation, and gene expression of osteogenic biomarkers osteocalcin, alkaline phosphatase and collagen type I. However, cell functions discriminated surface roughness by either comparing titanium and zirconia or interindividual zirconia surfaces. While rough surfaces primarily favored primary adhesion, proliferation appeared improved on smooth surfaces, and gene expression seemed to be stronger modulated on the smoothest biomaterial. Our results show for the first time that bioactivation appears to be not the main causative for the observed modulation of the distinct cell functions analyzed in PHABO, but add to the body of evidence that they were more governed by surface architecture rather than by bioactivation.

Microstructural Observation with MicroCT and Histological Analysis of Human Alveolar Bone Biopsy from a Planned Implant Site: A Case Report

The subject was a 53-year-old male. An alveolar bone sample was obtained from the site of the lower left first molar, before dental implant placement. Although the details of the trabecular structure were not visible with conventional computed tomography, micro-computed tomography (microCT) three-dimensional images of the alveolar bone biopsy sample showed several plate-like trabeculae extending from the lingual cortical bone. Histological observations of the bone sample revealed trabeculae, cuboidal osteoblasts, osteoclasts and hematopoietic cells existing in the bone tissue at the implantation site. Bone metabolic markers and calcaneal bone density were all within normal ranges, indicating no acceleration of the patient’s bone metabolism.Using microCT, and histological and histomorphometrical techniques, a great deal of valuable information about the bone tissue was obtained from a biopsy sample extracted from the patient’s planned implant site.

Three Years After Transplants in Human Mandibles, Histological and In-Line Holotomography Revealed That Stem Cells Regenerated a Compact Rather Than a Spongy Bone: Biological and Clinical Implications

Mesenchymal stem cells deriving from dental pulp differentiate into osteoblasts capable of producing bone. In previous studies, we extensively demonstrated that, when seeded on collagen I scaffolds, these cells can be conveniently used for the repair of human mandible defects. Here, we assess the stability and quality of the regenerated bone and vessel network 3 years after the grafting intervention, with conventional procedures and in-line holotomography, an advanced phase-imaging method using synchrotron radiation that offers improved sensitivity toward low-absorbing structures. We found that the regenerated tissue from the graft sites was composed of a fully compact bone with a higher matrix density than control human alveolar spongy bone from the same patient. Thus, the regenerated bone, being entirely compact, is completely different from normal alveolar bone. Although the bone regenerated at the graft sites is not of the proper type found in the mandible, it does seem to have a positive clinical impact. In fact, it creates steadier mandibles, may well increase implant stability, and, additionally, may improve resistance to mechanical, physical, chemical, and pharmacological agents.

Micro-architecture and mineralization of the human alveolar bone obtained with microCT

ObjectivesThe primary dental implant stability depends on the location of the implant in the jaw. This study analysed the architecture and mineralization of the trabecular bone at different jaw locations and thereby identified potential prognostic factors for implant failure. It has checked the hypotheses: (1) the mandible contains more compact and less mineralized trabecular bone than the maxilla and (2) within the mandible the trabecular bone is more compact and less mineralized in the anterior region. MethodsAlveolar bone specimens were produced from the cadavers of ten humans (7 males and 3 females; mean age: 73.7±12.5 years) and scanned with a high-resolution microCT system. Volumes of interest were chosen next to the roots of molars and incisors in both the maxilla and mandible. Several morphological parameters as well as the tissue mineral density were determined. ResultsThe alveolar bone specimens had a very high bone volume fraction (mean=0.31) with large differences (SD=0.17) between and within subjects. Yet several significant differences were found between the maxilla and the mandible. The bone volume fraction and trabecular thickness were significantly higher in the mandible than in the maxilla (p<0.01). But the tissue mineral density was not significantly different. ConclusionsA higher primary implant stability coincides with a higher bone volume fraction and degree of anisotropy Although local differences remain more important for implantology, the results suggest that the micro-structure also affects the implant stability. The tissue mineral density seems to have no predictive value.

Cytotoxicity Testing of Methyl and Ethyl 2-Cyanoacrylate Using Direct Contact Assay on Osteoblast Cell Cultures

Cyanoacrylate has been used as a commercial tissue adhesive. Recently, ethyl 2-cyanoacrylate has been suggested for the fixation of onlay autogenous bone graft. However, ethyl 2-cyanoacrylate must be biocompatible with bone tissue. This study evaluated the cytotoxicity of cyanoacrylate adhesives using a direct contact assay on human oral osteoblast cells. Osteoblastic cells derived from human alveolar bone of the mandible were cultured with or without cyanoacrylate. The CA1 group contained methyl 2-cyanoacrylate, the CA2 group contained ethyl 2-cyanoacrylate, and the CA3 group did not contain cyanoacrylate (control). This study investigated cell morphology, which included the inhibition zone, and cytotoxicity was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, which was measured as optical density. Data from the MTT assay were tested statistically using SigmaStat 3.5. Dead cells found around the CA1- and CA2-treated cells constituted inhibitory zones that varied from 200 to 500 μm. There was no inhibitory zone in the CA3 group. Cell viability evaluated by the MTT assay showed that the CA2 and CA3 optical densities were not significantly different. The CA1 optical densities differed significantly from the CA3 optical densities. Within the limits of this study, the MTT method supported the conclusion that ethyl 2-cyanoacrylate is biocompatible according to a direct contact assay on human osteoblast cell cultures and suggests its usefulness in bone graft fixation.

Enhanced Osteogenesis of Human Alveolar Bone-Derived Mesenchymal Stem Cells for Tooth Tissue Engineering Using Fluid Shear Stress in a Rocking Culture Method

This study instituted a simple approach to stimulate alveolar bone regeneration for tooth tissue engineering by controlling effects of low fluid dynamic shear stress (LFDSS) on growth and differentiation in vitro. Human alveolar bone-derived mesenchymal stem cells (hABMSCs) harvested from human mandibular alveolar bone were cultured with LFDSS to generate cultures containing bone-like formations. To distinguish between osteodifferentiation and bone-like formation, cells were cultured either with or without fluid shear stress. The calcium content and alkaline phosphatase (ALP) activity of hABMSCs were used as indicators of osteogenesis. Cell viability and proliferation after stimulating with LFDSS for 10-60 min/day were higher than with longer stimulations. Mineralized nodules formed when osteoblasts were cultured with an induction medium, a marker of osteogenic differentiation. ALP activity tended to increase after 10 and 60 min/day of stimulation. In addition, LFDSS conditions also increased gene expression of IBSP, RUNX2, COL-I, ALP, OCN, and OPN, as shown by reverse transcriptase-polymerase chain reaction. From the results of a proteomics array, LFDSS groups were intensely expressed with several factors (EGF, HGF, IGF, TGF, and PDGF). Furthermore, CD146 and Stro-1 expression increased in cells treated with 30 min/day and decreased in cells treated with 120 min/day, as determined by cell surface antigen analysis by fluorescence-activated cell-sorting analysis. These results strongly showed that LFDSS at the proper intensity and time enhanced the differentiation and maturation of hABMSCs. In conclusion, an appropriate level of LFDSS can potently and positively modulate proliferation and differentiation in hABMSCs.

Crosstalk on Cell Behavior in Interactive Cocultures of hMSCs with Various Oral Cell Types

When prospectively applied for regenerative therapies, human bone-marrow-derived mesenchymal stem cells (hMSCs) interact with the locally residing host cells. With respect to the developmentally particular origin of oral cells, little is known about the putatively discriminative behavioral responses of hMSCs in interaction with various oral cell types, including human alveolar bone osteoblasts (hOAs), periodontal ligament fibroblasts (hPDLs), and gingival fibroblasts (hGFs). To assess the crosstalk between hMSCs and oral cells, interactive cocultures were established by combining well-characterized hMSCs with hOAs, hPDLs, or hGFs, and the behavioral hMSC aspects, that is, proliferation and gene expression, were measured by employing a 5-bromo-2'-deoxyuridine assay and real-time polymerase chain reaction, while apoptosis was quantified by in situ cell death detection kit. hMSCs expressed the typical antigen spectrum lacking CD34, CD45, CD14, CD19, and HLA-DR, while expressing CD73, CD90, and CD105, and could successfully be transformed into adipocytes, osteocytes, and chondrocytes. Monocultured control hMSCs proliferated readily, whereas a general reduction of BrdU-labeled cells was observed in cocultures. Globally, upon extending time periods, interactive coculture combinations of hMSCs with hOAs reduced both osteogenic gene and stem cell marker transcription in hMSCs, a phenomenon appearing less pronounced by combining hMSCs with hPDLs, such that the observed effects in terms of proliferation and gene expression followed the same ranking: hOAs>hGFs>hPDLs. Vice versa, in interactive hMSC cocultures, the cell survival rate was significantly increased, irrespective from the combined coculture cell counterpart. Our results show for the first time that behavior of hMSCs reflected by proliferation and gene expression was governed by interaction with various oral cells in a cell-type-discriminative manner. In addition, hMSC coculture restrains apoptosis, such that influences on cell behavior appear as a crosstalk. In summary, interactive cocultures render the basis for a prospective prediction of mutual cell behavior in hMSC-based oral tissue regeneration disclosing that oral cells shift hMSC behavior from proliferation to differentiation and apoptosis-repressing features.

Anthropometric Measurements in Toothed and Toothless Maxillaries and its Consequences in Human Alveolar Bone Resorption

Este estudio propuso medir y comparar las medidas antropometricas entre los maxilares con y sin dientes. Fueron utilizados 26 craneos humanos, 13 maxilares con dientes y 13 sin dientes. Las mediciones fueron realizadas a ambos lados por dos evaluadores entre las distancias del reborde alveolar (RA): a la espina nasal anterior (ENA), al canal incisivo (CI) y a lo foraamenes palatinos mayor (FPMA) y menor (FPME). Los datos mostraron que la media de las mediciones fueron proporcionalmente mayores en las maxilas con dientes, para todas las mediciones en cualquiera de los evaluadores. El RA-ENA (p = 0,001), AR-CI (p = 0,006), RA-APMA derecho (p = 0,001) y RA-APMA izquierdo (p <0,001) y RA-APME en ambos lados (p = 0,001), comprobando la existencia de diferencias significativas entre los dos tipos de maxila. Fue verificado que a partir de la perdida dental ocurren irreversibles cambios anatomicos en el maxilar, que deben ser considerados en el diseno y fabricacion de protesis e implantes osteointegrados.

Initial responses of osteoblasts derived from human alveolar bone to various compressive forces

Mechanical stress generated by orthodontic force is recognized as a major factor in the modulation of alveolar bone remodeling. During this process, osteoblasts play a crucial role, not only by participating in bone formation but also by promoting osteoclastogenesis. The aim of this study was to investigate how continuous compressive force (CF) affects human primary osteoblasts (HOBs) in terms of cell proliferation, apoptosis, and expression of interleukin-6 (IL-6) and chemokine CXC ligand 8 (CXCL8). Human primary osteoblasts, isolated from human mandibular bone pieces, were cultured with or without CF (1-4 g cm(-2)) for up to 72 h. Cell viability and proliferation were evaluated using the MTT assay. RT-PCR was used to determine the levels of expression of KI67 (a proliferation marker), BAX (a pro-apoptotic marker), BCL2 (an apoptotic inhibitor), IL6, and CXCL8 mRNAs, while a multiplexed bead immunoassay was used to measure the release of IL-6 and CXCL8. The results revealed that CF decreased cell viability and proliferation in a time- and force-dependent manner. After applying CF for 24 h, the mRNA expression of KI67 was markedly inhibited, whereas the mRNA expression of BAX and BCL2 was unaltered. In addition, CF enhanced the levels of IL6 and CXCL8 mRNAs in a force-dependent manner, whereas the levels of the corresponding proteins were reduced in the compressed HOBs.

The effect of enamel matrix derivative (Emdogain®) on gene expression profiles of human primary alveolar bone cells

Emdogain® is frequently used in regenerative periodontal treatment. Understanding its effect on gene expression of bone cells would enable new products and pathways promoting bone formation to be established. The aim of the study was to analyse the effect of Emdogain® on expression profiles of human-derived bone cells with the help of the micro-array, and subsequent validation. Bone was harvested from non-smoking patients during dental implant surgery. After outgrowth, cells were cultured until subconfluence, treated for 24 h with either Emdogain® (100 µg/ml) or control medium, and subsequently RNA was isolated and micro-array was performed. The most important genes demonstrated by micro-array data were confirmed by qPCR and ELISA tests. Emdogain tipped the balance between genes expressed for bone formation and bone resorption towards a more anabolic effect, by interaction of the PGE2 pathway and inhibition of IL-7 production. In addition the results of the present study indicate that Emdogain possibly has an effect on gene expression for extracellular matrix formation of human bone cells, in particular on bone matrix formation and on proliferation and differentiation. With the micro-array and the subsequent validation, the genes possibly involved in Emdogain action on bone cells were identified. These results can contribute to establishing new products and pathways promoting bone formation.

The influence of pore size on osteoblast phenotype expression in cultures grown on porous titanium

This study investigated the effect of pore size on osteoblastic phenotype development in cultures grown on porous titanium (Ti). Porous Ti discs with three different pore sizes, 312μm (Ti 312), 130μm (Ti 130) and 62μm (Ti 62) were fabricated using a powder metallurgy process. Osteoblastic cells obtained from human alveolar bone were cultured on porous Ti samples for periods of up to 14 days. Cell proliferation was affected by pore size at day 3 (p=0.0010), day 7 (p=0.0005) and day 10 (p=0.0090) in the following way: Ti 62<Ti 130<Ti 312. Gene expression of bone markers evaluated at 14 days was affected, RUNX2 (p=0.0153), ALP (p=0.0153), BSP (p=0.0156), COL (p=0.0156), and OPN (p=0.0156) by pore size as follows: Ti 312<Ti 130<Ti 62. Based on these results, the authors suggest that porous Ti surfaces with pore sizes near 62μm, compared with those of 312μm and 130μm, yield the highest expression of osteoblast phenotype as indicated by the lower cell proliferation rate and higher gene expression of bone markers.