Osteoclast formation from mouse bone marrow cells on micro/nano-scale patterned surfaces

ObjectivesOsteoclasts can sense the surface topography of materials. However, it is difficult to identify the structural factors that affect osteoclast formation and its function. Furthermore, we hypothesized that the type of osteoclast precursor cells also affects osteoclastogenesis in the materials. In this study, we investigated the effects of defined micro/nanoscale patterns on osteoclastogenesis from bone marrow cells (BMCs).MethodsVarious cyclo-olefin polymer (COP) patterns were prepared using nanoimprinting. The effects of shape, size, and height of the patterns, and the wettability of the patterned surfaces on osteoclastogenesis from BMCs were evaluated in vitro.ResultsOsteoclast formation was promoted on pillars (diameter, 1 μm or 500 nm; height, 500 nm). Notably, osteoclastogenesis from BMCs was better promoted on hydrophobic pillars than on hydrophilic pillars. In contrast, decreased osteoclast formation was observed on the nanopillars (diameter, 100 nm; height, 200 nm).ConclusionsWe demonstrated the promotion of osteoclast formation from BMCs on hydrophobic pillars with diameters of 1 μm and 500 nm. Some cellular behaviors in the patterns were dependent on the type of osteoclast precursor cells. The designed patterns are useful for designing the surface of dental implants or bone replacement materials with a controllable balance between osteoblast and osteoclast activities.     

Characterization of verruciform xanthoma by in situ hybridization and immunohistochemistry

Verruciform xanthoma (VX) is a rare, benign lesion, mainly found in the oral mucosa. Histologically and ultrastructurally. the lesion is characteristic and well defined. However, the etiology of the lesion remains unclear. The purpose of the present study was to elaborate upon the pathogenesis of VX by evaluation of an additional series of oral examples for human papillomaviruses (HPV). using immunohistochemistry and in situ hybridization, and to further characterize the cellular components of VX immunohistochemically. Twelve specimens diagnosed as VX were retrospectively collected. One of the twelve specimens was positive for HPV types 6/11 by in situ hybridization. None of the twelve specimens demonstrated the presence of HPV antigen by immunohistochemistry. By immunohis-tochemical studies, the predominant cells in the inflammatory infiltrate were T cells. The foam cells were of monocyte/macrophage lineage. S-100-positive (Langerhans) cells were occasionally found in the suprabasal layer of the epithelium. HLA-DR-positive keratinocytes were noted at the intense inflammatory sites. Taken together, these findings suggest that an immune response may play a role, at leas! in part, in VX pathogenesis.     

Advances in skeletal tissue engineering with hydrogels

OBJECTIVES: Tissue engineering has the potential to make a significant impact on improving tissue repair in the craniofacial system. The general strategy for tissue engineering includes seeding cells on a biomaterial scaffold. The number of scaffold and cell choices for tissue engineering systems is continually increasing and will be reviewed. DESIGN: Multilayered hydrogel systems were developed to coculture different cell types and develop osteochondral tissues for applications including the temporomandibular joint. EXPERIMENTAL VARIABLE: Hydrogels are one form of scaffold that can be applied to cartilage and bone repair using fully differentiated cells, adult and embryonic stem cells. OUTCOME MEASURE: Case studies represent an overview of our laboratory's investigations. RESULTS: Bilayered scaffolds to promote tissue development and the formation of more complex osteochondral tissues were developed and proved to be effective. CONCLUSION: Tissue engineering provides a venue to investigate tissue development of mutant or diseased cells and potential therapeutics.     

Cytokeratin phenotypes at the dento-gingival junction in relative health and inflammation, in smokers and non-smokers

OBJECTIVES: The cells of the junctional epithelium (JE) provide and maintain the epithelial attachment, and remain morphologically and phenotypically distinct from oral sulcular (OSE) and external oral epithelia (EOE), from which they may be regenerated de novo. Expression of cytokeratins (CK) in human epithelia has been shown to be highly site-specific, implying a functional role. The aims of this study were to differentiate between the cyto-keratin profiles of JE, OSE, EOE and pocket epithelia (PE) in health and disease, in smokers and non-smokers.
MATERIALS AND METHODS: The cytokeratin profiles of 40 samples of healthy and clinically inflamed human gingival tissue taken from 15 smokers and 25 non-smokers were studied by immunocytochemistry. Cryostat sections of fresh frozen gingival tissues were stained with a panel of monoclonal antibodies (mAb) and visualised by a biotin-Streptavidin-peroxidase complex technique.
RESULTS: JE and PE expressed an identical range of cytokeratins irrespective of the inflammatory or smoking status, with the exception of CK4 expression, which tended to be increased in smokers. The OSE and EOE expressed non-cornifying and cornifying differentiation cytokeratins respectively, but in the presence of inflammation, both these epithelia showed increased expression of CK19 at a basal level in association with expression of one or more of the simple cytokeratins. JE/PE expressed CK17 in external layers only, approximating the tooth surface. All epithelia expressed CK6,16 the markers of high cell turnover.
CONCLUSIONS: CK19 was a consistent differentiation marker for JE and PE.Expression of CK8,18 was enhanced by inflammation. CK4 expression increased in association with smoking. Markers of differentiation were not always co-expressed equally within a pair. Pairs were not always completely mutually exclusive with frequent co-localisation.     

VCAM-1 and ICAM-1 are expressed by Langerhans cells, macrophages and endothelial cells in oral lichen planus

Expression of intercellular adhesion molecule-1 (ICAAM-1, CD54) and vascular cell adhesion molecule-1 (VCAM-l, CD106) was examined in oral lichen planus (OLP) and normal oral mucosa (NOM). Immunoperoxidase staining showed ICAM-1 expression by vascular endothelium in all biopsies of OLP and NOM whereas endothelial VCAM-l staining was found in 2/7 NOM and 8/9 OLP. In the lamina propria of NOM occasional cells were ICAM-1 or VCAM-l positive, and virtually no staining of intraepilhelial dendritic cells was seen for either marker. Intraepithelial dendritic cells stained for ICAM-1 in 7/9 and VCAM-1 in 4/9 OLP biopsies. Double immunofluorescence showed dual labelling of Langerhans cells (LC) with CD1a and VCAM-l in a further 5/12 cases of OLP, but there was no such staining in four NOM. This is the first report of LC staining with VCAAM-l. Induction of ICAM-1 and VCAM-l on LC and macrophages in OLP suggests these cells are activated and may contribute to the pathogenesis of OLP by presenting antigen to infiltrating lymphocytes.     

不同代数人牙周膜细胞成骨特征稳定性的实验研究

目的：观察培养代数牙周膜细胞的成骨细胞表型特征的影响。方法：利用4、6、8、10代细胞进行以形态学、细胞生长曲线、碱性磷酸酶（ALP）活性测定以及矿化能力和面积的观察。结果：在细胞形态和超微结构、细胞生长曲线、碱性磷酸酶活性和矿化能力方面，第4、6、8、10代无明显差别；10代以后的细胞各方面指标减弱，提示细胞进入衰老期。结论：利用10代以内牙周膜细胞进行实验或移植其结果可靠。     

牙体预备对牙龈结合上皮损伤后的超微结构观察

目的 :研究烤瓷修复体牙体预备对牙龈结合上皮损伤后 ,牙龈结合上皮在透射电镜下形态学变化的特点。方法 :将 16只家兔的 3 2颗上前牙分别用金刚砂车针和手术刀破坏左侧上前牙的牙龈结合上皮 (JE) ,以右侧同名牙为对照 ,分别于 1、2周取材 ,利用透射电镜观察结合上皮的组织学改变 ,并对 2实验组进行对比观察。结果 :金刚砂车针处理组和手术刀处理组 2周后牙龈结合上皮附着恢复 ,两处理组无明显差别 ,与未经手术处理的JE比较 ,处理组主要超微结构变化是线粒体空泡化 ,基膜不完整。结论 :牙体预备对牙龈结合上皮的损伤 ,可导致结合上皮的超微结构改变     

PAF levels in saliva are regulated by inflammatory cells

Platelet activating factor (PAF), a powerful inflammatory phospholipid mediator, has been detected in normal human saliva and found to be increased in periodontitis. The cellular source of PAF in saliva is controversial although several data suggest an origin related to the presence of inflammatory cells. PAF levels in biological fluids are regulated by PAF-producing cells and by the PAF-degrading acetylhydrolase. Although in normal human saliva acetylhydrolase activity is very low, no information is available on the levels of this enzyme in inflammatory conditions of the mouth. The aim of our study was to assess the contribution of inflammatory cells to the levels of PAF in saliva in normal subjects and in patients with periodontitis. PAF was measured by radioimmunoassay (RIA) in mixed uncentrifuged saliva and in cell-free saliva from healthy subjects, before and after tooth brushing, and in patients with periodontitis. In healthy subjects PAF levels were significantly higher in whole saliva than in centifuged saliva (1.51 ± 0.22 vs. 0.92 ± 0.04 ng/ml, p<0.0039). A significant increase in the amount of PAF was detected in whole saliva, but not in centifuged saliva, 2 h after tooth brushing. In patients with periodontitis PAF levels were not different from those of healthy individuals when using centrifuged saliva but were significantly higher when using whole, uncentrifuged saliva. Exogenous radiolabelled PAF was degraded much more rapidly by the saliva of periodontitis patients than by that of normal subjects. In conclusion, our study shows that inflammatory cells regulate the levels of PAF in saliva contributing to its production and degradation. The differential degradation of PAF in normal and inflammatory saliva highlights the absolute need of a series of methodological precautions when performing studies on salivary PAF.     

Behaviour of bone marrow osteoblast-like cells on mineral trioxide aggregate: morphology and expression of type I collagen and bone-related protein mRNAs

AIM: To investigate the in vitro behaviour of rat bone marrow cells (RBM) on mineral trioxide aggregate (MTA) (ProRoot, MTA Root Canal Repair Material; Dentsply Tulsa, Tulsa, OK, USA) compared with intermediate restorative materials (IRM) (Dentsply Caulk, Milford, DE, USA). METHODOLOGY: RBM were obtained from rat femur and were primary cultured and then subcultured. Cells were then seeded on three dishes of each material, and cultured for 3 days, after which they were evaluated morphologically using scanning (SEM) and transmission (TEM) electron microscopy. Furthermore, the calcium released from hydrated material, the cell proliferation ratio and alkaline phosphatase (ALP) activity were analysed, and the expression of type I collagen and bone-related protein mRNAs were evaluated. The data were averaged and analysed via one-way analysis of variance (anova) and were then compared by the Scheffe's test. RESULTS: SEM showed that RBM attached to MTA and had a flattened appearance without nuclear protrusions and microspikes. TEM showed that the cells attached in the same manner as the control group, but gaps larger than 2 microm were frequently seen. The calcium released from hydrated MTA was about 130 ppm after 3 days of immersion in saline. The ALP activity was similar to the control group. Cell proliferation and expression of type I collagen mRNA was significantly lower, while the expression of osteopontin mRNA was significantly higher than the control group at the third day of culture. In IRM groups, a few rounded cells were observed on the material but no living cells were seen. CONCLUSIONS: MTA is a material of low toxicity which does not inhibit cell growth, but does suppress the differentiation of osteoblast-like cells.