F-spondin negatively regulates dental follicle differentiation through the inhibition of TGF-β activity

ObjectiveF-spondin is an extracellular matrix (ECM) protein that belongs to the thrombospondin type I repeat superfamily and is a negative regulator of bone mass. We have previously shown that f-spondin is specifically expressed in the dental follicle (DF), which gives rise to the periodontal ligament (PDL) during the tooth root formation stage. To investigate the molecular mechanism of PDL formation, we investigated the function of f-spondin in DF differentiation.DesignThe expression patterning of f-spondin in the developing tooth germ was compared with that of periodontal ligament-related genes, including runx2, type I collagen and periostin, by in situ hybridization analysis. To investigate the function of f-spondin during periodontal ligament formation, an f-spondin adenovirus was infected into the bell stage of the developing tooth germ, and the effect on dental differentiation was analyzed.ResultsF-spondin was specifically expressed in the DF of the developing tooth germ; by contrast, type I collagen, runx2 and periostin were expressed in the DF and in the alveolar bone. F-spondin-overexpresssing tooth germ exhibited a reduction in gene expression of periostin and type I collagen in the DF. By contrast, the knockdown of f-spondin in primary DF cells increased the expression of these genes. Treatment with recombinant f-spondin protein functionally inhibited periostin expression induced by transforming growth factor-β (TGF-β).ConclusionOur data indicated that f-spondin inhibits the differentiation of DF cells into periodontal ligament cells by inhibiting TGF-β. These data suggested that f-spondin negatively regulates PDL differentiation which may play an important role in the immature phenotype of DF.     

Immunoexpression of TNF-α and TGF-β in central and peripheral giant cell lesions of the jaws

J Oral Pathol Med (2012) 41 : 194–199 Background: Peripheral giant cell lesion (PGCL) is a reactive process associated with a local irritating factor that shows low recurrence after treatment, especially if the irritating factor is eliminated. On the other hand, central giant cell lesion (CGCL) presents a variable clinical behavior ranging from slow and asymptomatic growth without recurrence to rapid, painful and recurrent growth. Our aim was to compare the immunoexpression of tumor necrosis factor‐alpha (TNF‐α) and transforming growth factor‐beta (TGF‐β) in CGCL and PGCL. Methods: Twenty CGCL and 20 PGCL were selected for analysis of the immunoexpression of TNF‐α and TGF‐β in multinucleated giant cells (MGC) and mononucleated cells (MC). Results: The PGCL showed lightly higher expression of TNF‐α than CGCL. In comparison with PGCL, the CGCL showed higher expression of TGF‐β in MC and MGC (P < 0.05) and in total cells (P < 0.05). Significant positive correlation was found between expressions of TGF‐β and TNF‐α in CGCL (P < 0.05). Conclusions: Our results suggest that, in CGCL, coordinated interactions between TGF‐β and TNF‐α may be important for osteoclastogenesis and bone resorption. PGCL occasionally cause bone resorption but to a lower extent, a fact that might be explained by the lower expression of TGF‐β in these lesions.     

Comparison of different autografts for aural cartilage in aesthetic rhinoplasty: is the tragal cartilage graft a viable alternative?

Auricular cartilage is an important source of grafts for various reconstructive procedures such as aesthetic rhinoplasty. The purpose of this investigation was to compare tragal cartilage with auricular cartilage harvested from the concha and scapha, and describe its clinical viability, indications, and morbidity in rhinoplasty. A total of 150 augmentation rhinoplasties with a total of 170 grafts were included. The donor sites were tragus (n = 136), concha (n = 26), and scapha (n = 8). The time needed to harvest the grafts, the donor site morbidity, and the indications for operation were recorded. The anthropometric changes to 4 auricular variables after the cartilage had been harvested were analysed and compared with those on the opposite side in 48 patients using Student's paired t-test. Intraobserver reliability was assessed using Pearson's intraclass correlation. The mean (SD) harvesting time was 27 (8) min for the concha, 4.5 (1.4) min for the tragus, and 5.7 (1.6) min for the scapha. The largest graft was taken from the concha (28 × 19 mm), followed by the tragus (20 × 12 mm), and the scapha (18 × 6 mm). The grafts were placed at the following sites: tip grafts (n = 123), columella struts (n = 80), shield (n = 20), rim (n = 17), and dorsal onlay (n = 15). Harvesting tragal cartilage is safe, simple, fast, and has a low morbidity, but it can affect the patient's ability to wear earphones. Tragal cartilage is a good alternative for nasal reconstruction if a graft of no longer than 20 mm is required.     

Academic Training in Oral and Maxillofacial Surgery – when and how to enter the pathway

Entering into surgical academia can seem a daunting prospect for an oral and maxillofacial surgery (OMFS) trainee. However, the streamlining of academic training by the NIHR to create the integrated academic training (IAT) pathway has simplified academic training and more clearly defined academic positions and entry points for trainees. In this article we review the current NIHR IAT pathway and the various grades and entry points available to OMF surgeons, both pre- and post-doctoral. We highlight the unique challenges facing OMF trainees and provide advice and insight from both junior and senior OMFS academics. Finally, we focus on the planning and application for a doctoral research fellowship – discussing funding streams available to OMF surgeons.     

The Role of TGF-β Signaling in Cranial Neural Crest Cells during Mandibular and Tooth Development

To investigate the role of transforming growth factor-β (TGF-β) in regulating the fate of cranial neural crest cells (CNC) during mandibular and tooth development, we generated mutant mice with tissue-specific Tgfbr2 gene ablation using a Cre/loxP recombination system exclusively in the cranial neural crest lineage (Tgfbr2^fl/fl; Wntl-Cre). Tgfbr2^fl/fl; Wntl-Cre mice show mandible defects including the abnormal shape of Meckel's cartilage, a small mandibular bone, and perturbed chondrogenesis in the proximal region of the mandible. TGF-β signaling stimulates the proliferative activity of chondrocytes in Meckel's cartilage through Ctgf and of osteoblasts in mandibular bone through Msxl expression. Interestingly, in the proximal region of Tgfbr2^fl/fl; Wntl-Cre mice, cartilage was replaced by bone formation as a result of accelerated osteoblast differentiation due to elevated Runx2 and Dlx5 expression in osteo-chondroprogenitor cells. Additionally, the deletion of Dlx5 in Tgfbr2^fl/fl; Wntl-Cre mice resulted in the rescue of cartilage formation in the angular processes. Regarding tooth development, Tgfbr2^fl/fl; Wntl-Cre mice showed abnormal dentin formation. Following kidney capsule transplantation, Tgbr2 mutant tooth germs exhibited defects, with a decreased dentin thickness and absent dentinal tubules. The expression of Coll and Dsp was reduced. In addition, the expression of the intermediate filament nestin was decreased in Tgfbr2^fl/fl; Wntl-Cre mouse samples. This suggests that TGF-β signaling controls odontoblast maturation and dentin formation during tooth development. In this review, we reveal these mandibular and tooth phenotypes in Tgfbr2^fl/fl; Wntl-Cre, and discuss the intrinsic requirement of TGF-β signaling during craniofacial development.     

PERIOSTIN regulates MMP-2 expression via the αvβ3 integrin/ERK pathway in human periodontal ligament cells

ObjectiveDuring orthodontic tooth movement, activation of the vascular system in the compressed periodontal ligament (PDL), which becomes hypoxic, is essential for periodontal tissue remodelling. PERIOSTIN, an extracellular matrix protein, is expressed in PDL and its concentration is increased on the compressive side during orthodontic tooth movement. PERIOSTIN promotes angiogenesis through upregulation of matrix metalloproteinase (MMP)-2, which has been shown to be expressed via αvβ3 integrin/extracellular signal-related kinase (ERK) signalling pathway and vascular endothelial growth factor (VEGF). Therefore, we hypothesized that hypoxia-induced PERIOSTIN promotes MMP-2 expression via αvβ3 integrin/ERK signalling and VEGF in PDL cells.MethodsHuman PDL cells were cultured in condition medium containing desferrioxamine (DFO) to mimic hypoxia. The total RNA, cell lysates or supernatant were collected, and MMP2 and VEGF expression, PERIOSTIN expression and ERK phosphorylation, and MMP-2 activity were analysed by real-time RT-PCR, western blot analysis, and zymography, respectively. A recombinant human PERIOSTIN or PERIOSTIN siRNA was applied to the cells, then the total RNA was extracted to measure MMP-2 and VEGF expression. The cells were treated with αvβ3 integrin-blocking antibody or ERK inhibitor followed by PERIOSTIN stimulation. MMP-2 expression was measured by real-time RT-PCR.ResultsPERIOSTIN was upregulated in a time-dependent manner in human PDL cells treated with DFO, a chemical hypoxia mimic. MMP-2 and VEGF expression, and MMP-2 activity were increased by DFO or PERIOSTIN treatment, and decreased by PERIOSTIN silencing. PERIOSTIN treatment also induced ERK phosphorylation, and PERIOSTIN-induced MMP-2 was reduced by αvβ3 integrin-blocking antibody or ERK inhibitor.ConclusionThese data suggest that PERIOSTIN upregulates MMP-2 expression via the αvβ3 integrin/ERK signalling pathway and VEGF expression in human PDL cells.     

Anti-inflammatory effects of EMD in the presence of biomechanical loading and interleukin-1β in vitro

Enamel matrix derivative (EMD) used to promote periodontal regeneration has been shown to exert anti-inflammatory effects. This in vitro study was performed to investigate if the anti-inflammatory actions of EMD are modulated by the local cellular environment, such as inflammation or occlusal, i.e., biomechanical, loading. Human periodontal ligament cells were seeded on BioFlex plates and incubated with EMD under normal, inflammatory, and biomechanical loading conditions for 1 and 6 days. In order to mimic inflammatory and biomechanical loading conditions in vitro, cells were stimulated with interleukin (IL)-1β and exposed to dynamic tensile strain, respectively. The gene expression of IL-1β, IL-1 receptor antagonist (IL-1RN), IL-6, IL-8, IL-10, and cyclooxygenase (COX)-2 was analyzed by real-time RT-PCR and the IL-6 protein synthesis by enzyme-linked immunoassay. For statistical analysis, Student's t test, ANOVA, and post-hoc comparison tests were applied (p < 0.05). EMD downregulated significantly the expression of IL-1β and COX-2 at 1 day and of IL-6, IL-8, and COX-2 at 6 days in normal condition. In an inflammatory environment, the anti-inflammatory actions of EMD were significantly enhanced at 6 days. In the presence of low biomechanical loading, EMD caused a downregulation of IL-1β and IL-8, whereas high biomechanical loading significantly abrogated the anti-inflammatory effects of EMD at both days. Neither IL-1RN nor IL-10 was upregulated by EMD. These data suggest that high occlusal forces may abrogate anti-inflammatory effects of EMD and should, therefore, be avoided immediately after the application of EMD to achieve best healing results.     

Inhibition of the receptor for advanced glycation promotes proliferation and repair of human periodontal ligament fibroblasts in response to high glucose via the NF-κB signaling pathway

ObjectiveTo observe if inhibition of the receptor for advanced glycation endproducts (RAGE) promotes proliferation and repair of human periodontal ligament fibroblasts (hPDLFs) stimulated by high glucose. In addition, we also discuss the effects of the NF-κB signaling pathway in relation to this process.MethodsPrimary cultured hPDLFs were exposed to either low glucose (5.5 mmol/L) or high glucose (25 mmol/L), and RAGE expression was measured by Western blot analysis. Cells were cultured in high glucose with different concentrations of the RAGE inhibitor, FPS-ZM1. We measured cell proliferation using the Cell Counting Kit-8 and expression of collagen type 1 and fibronectin by real-time PCR and ELISA, respectively. The relative protein expression levels of NF-κB p65 and phosphorylated p65 were measured by Western blot analysis.ResultsHigh glucose enhanced RAGE expression and suppressed cell growth. While FPS-ZM1 increased proliferation and expression of repair-related factors in high glucose, there was a concurrent decline in the phosphorylation level of NF-κB p65.ConclusionFPS-ZM1 rescued the proliferative capacity and repair capability of hPDLFs via the RAGE-NF-κB signaling pathway in response to high glucose.     

Leukocyte-platelet-rich plasma (L-PRP) impairs the osteoconductive capacity of the autograft associated to changes in the immunolocalization of TGF-β1 and its co-expression with Wnt10b and CD34 cells

BackgroundThis study evaluated the osteoconductive effect of an autograft, in the presence or absence of the L-PRP, using histomorphometric analysis of the bone formed, and we compared the results in the presence of TGF-β1, Wnt10b and CD34 detected by immunohistochemistry.Materials and methodsTwo bone defects were produced in the calvaria of 20 rabbits. The defects were treated with autograft and autograft combined with L-PRP. The animals were euthanized at 15 and 40 days post-surgery. Data were analyzed by Student-Newman–Keuls (p ≤ 0.05) test for histomorphometric and immunohistochemical interpretation.ResultsThe results revealed that the presence of bone matrix was significantly less in the defects treated with L-PRP. These results coincided with changes of the immunolocalization of the TGF-β1. In the L-PRP-free groups the TGF-β1 was restricted to bone matrix while the CD34 was scarce and the Wnt10b occurred in peritrabecular cells. In contrast, in defects that received L-PRP the presence of TGF-β1 occurred in cells, which occupied whole area of defect. These TGF-β1+ cells also were co-expressed to Wnt10b and CD34.ConclusionThese results suggest that L-PRP induces a cross-reaction between TGF-β1 and Wnt10b, which stimulates the self-renewal and maintenance of CD34+ stem cells immunophenotype, impairing the osteoconductivity properties of the autograft.     

Biological roles of KGF,CTGF and TGF-β in cyclosporine-A- and phenytoin- induced gingival overgrowth: A comparative experimental animal study

ObjectiveTo identify the possible biological roles of keratinocyte growth factor (KGF), connective tissue growth factor (CTGF) and transforming growth factor-β (TGF-β) in cyclosporine-A (CsA) and phenytoin (PNT)-induced gingival overgrowth (GO) and to correlate them with each other.MethodsSixty adult male albino rats were selected and divided into 3 equal groups. Group I rats received no treatment. Group II rats were administrated CsA for 7 weeks. Group III were administrated PNT for the same period. Rats were euthanized at the end of the experiment and routine tissue processing was carried out. The obtained specimens were stained with H&E, KGF, CTGF and TGF-β antibodies.ResultsOne-way MANOVA test for KGF, CTGF and TGF-β revealed an overall significant difference between the different groups (P < 0.001). LSD post hoc test for multiple comparisons revealed a significant difference between each two groups. Two-tailed Pearson correlation for group II revealed non-significant weak positive correlations between KGF & CTGF and between CTGF & TGF-β. Non-significant weak negative correlation was found between KGF & TGF-β. Meanwhile, group III revealed non-significant weak positive correlation between KGF & TGF-β and between CTGF & TGF-β. Significant moderate positive correlation was found between KGF & CTGF.ConclusionThe findings of the present study indicated that KGF, CTGF and TGF-β have biological roles in progression of CsA- and PNT- induced GO. KGF plays a greater role in CsA- induced GO than in PNT- induced GO. Meanwhile, CTGF and TGF-β play a role in PNT- induced GO greater than in CsA- induced GO.     

Tumor necrosis factor-α and interleukin-1β expression pathway induced by Streptococcus mutans in macrophage cell line RAW 264.7

Streptococcus mutans, a major etiological agent of dental caries, frequently causes systemic disease, such as subacute bacterial endocarditis, if it enters the bloodstream. In this study, the production pathways of the proinflammatory cytokines, tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), induced by S. mutans in mouse macrophage were examined using a quantitative real-time polymerase chain reaction and an enzyme-linked immunosorbent assay. The S. mutans stimulated the expression of TNF-α and IL-1β mRNA at a multiplicity of infection of 1 : 100, which increased at 2 and 4 h, respectively, to 24 h. It also induced the production of high levels of the TNF-α and IL-1β proteins, which increased at 2 h and reached a peak at 4 and 24 h, respectively. Nuclear factor-κB (NF-κB) was activated and reached a maximum level 30 min after the S. mutans treatment. The expression of TNF-α and IL-1β mRNA and protein was suppressed by the treatment with pyrrolidine dithiocarbamate, an NF-κB inhibitor. The S. mutans-induced TNF-α expression was suppressed by the presence of SB203580, a p38 mitogen-activated protein (MAP) kinase inhibitor, or SP600125, a Jun N-terminal kinase (JNK) MAP kinase inhibitor. On the other hand, IL-1β expression was inhibited by extracellular signal-regulated kinase (ERK)/p38/JNK MAP kinase inhibitor pretreatment. In addition, TNF-α production was suppressed more in the Toll-like receptor 2(-/-) (TLR2(-/-)) macrophages than in the TLR4(-/-) macrophages, whereas IL-1β production was suppressed more in the TLR4(-/-) macrophages than in the TLR2(-/-) macrophages. These results show that S. mutans stimulates the production of TNF-α and IL-1β in the mouse macrophage cell line, RAW 264.7, by activating ERK/p38/JNK, and NF-κB through TLR2 and TLR4, respectively.     

What is the role of biofilms in severe head and neck infections?

Most infections of the head and neck and virtually all of those encountered in the practice of dentistry are caused by bacteria that are organized into biofilms. A biofilm is a complex, usually multispecies, highly communicative community of bacteria that is surrounded by a polymeric matrix. Treatment of these types of infections with traditional antibiotics alone is ineffective, and surgical removal of diseased tissue is still necessary.     

Malocclusion and caries prevalence: is there a connection in the primary and mixed dentitions?

The purpose of this epidemiological cross-sectional study was to determine the prevalence of malocclusion and caries in children and to investigate whether a relationship exists between prevalence of caries and studied malocclusion. The study consisted of 8,864 preschool and schoolchildren with primary dentitions (mean age 4.5 years) and mixed dentitions (mean age 8.9 years). 1997 WHO dental caries criteria were applied to both groups. The existence of an increased caries risk was deducted from the dmft and DMFT indices related to age. Malocclusion in primary and mixed dentitions was classified into seven types. Fifty-seven percent of all children had some form of malocclusion. Prevalence of malocclusion increased and was significantly greater in the mixed dentition sample (p<0.001) than in the primary dentition sample. Seventy-four percent of children with primary dentitions and 23% of children with mixed dentitions had zero dmft and DMFT scores. Mean dmft indices in subjects with primary and mixed dentitions were 1.02 and 1.53, respectively. No positive correlation between prevalence of caries and malocclusion could be established in the sub sample with primary teeth only. However, statistically significant parallelism in prevalence of malocclusion and caries were found for posterior cross-bite (p=0.050) and mandibular overjet (p=0.013) in children with mixed dentitions.