Yasushi Furuichi

FGF-2 Stimulates Periodontal Regeneration Results of a Multi-center Randomized Clinical Trial

The efficacy of the local application of recombinant human fibroblast growth factor-2 (FGF-2) in periodontal regeneration has been investigated. In this study, a randomized, double-blind, placebo-controlled clinical trial was conducted in 253 adult patients with periodontitis. Modified Widman periodontal surgery was performed, during which 200 µL of the investigational formulation containing 0% (vehicle alone), 0.2%, 0.3%, or 0.4% FGF-2 was administered to 2- or 3-walled vertical bone defects. Each dose of FGF-2 showed significant superiority over vehicle alone (p < 0.01) for the percentage of bone fill at 36 wks after administration, and the percentage peaked in the 0.3% FGF-2 group. No significant differences among groups were observed in clinical attachment regained, scoring approximately 2 mm. No clinical safety problems, including an abnormal increase in alveolar bone or ankylosis, were identified. These results strongly suggest that topical application of FGF-2 can be efficacious in the regeneration of human periodontal tissue that has been destroyed by periodontitis.

Endocannabinoid, anandamide in gingival tissue regulates the periodontal inflammation through NF-κB pathway inhibition

Anandamide (AEA) exhibits anti‐inflammatory effects. However, its role in the periodontal field remains unknown. Here, we found that gingival crevicular fluid contained a detectable level of AEA. The cannabinoid receptors CB1 and CB2 were expressed by human gingival fibroblasts (HGFs), and markedly upregulated under pathological conditions. AEA significantly reduced the production of pro‐inflammatory mediators (IL‐6, IL‐8 and MCP‐1) induced by Porphyromonas gingivalis LPS in HGFs, and this effect was attenuated by AM251 and SR144528, selective antagonists of CB1 and CB2, respectively. Moreover, AEA completely blocked LPS‐triggered NF‐κB activation, implying that AEA may regulate hyperinflammatory reactions in periodontitis.

Injectable Bone Tissue Engineering Using Expanded Mesenchymal Stem Cells

Patients suffering from bone defects are often treated with autologous bone transplants, but this therapy can cause many complications. New approaches are therefore needed to improve treatment for bone defects, and stem cell therapy presents an exciting alternative approach. Although extensive evidence from basic studies using stem cells has been reported, few clinical applications using stem cells for bone tissue engineering have been developed. We investigated whether injectable tissue‐engineered bone (TEB) composed of mesenchymal stem cells (MSCs) and platelet‐rich plasma was able to regenerate functional bone in alveolar deficiencies. We performed these studies in animals and subsequently carried out large‐scale clinical studies in patients with long‐term follow‐up; these showed good bone formation using minimally invasive MSC transplantation. All patients exhibited significantly improved bone volume with no side effects. Newly formed bone areas at 3 months were significantly increased over the preoperation baseline (p < .001) and reached levels equivalent to that of native bone. No significant bone resorption occurred during long‐term follow‐up. Injectable TEB restored masticatory function in patients. This novel clinical approach represents an effective therapeutic utilization of bone tissue engineering. STEM CELLS2013;31:572–580

FGF-2 induces proliferation of human periodontal ligament cells and maintains differentiation potentials of STRO-1 + /CD146 + periodontal ligament cells

The presence of human STRO-1(+)/CD146(+) periodontal ligament (PDL) cells has been reported, but obtaining a large amount of these cells is difficult. The purpose of this study was to evaluate the percentages of STRO-1(+)/CD146(+) cells in PDL cells and determine the effects of FGF-2 on the proliferation and multilineage differentiation potency of these cells. Human PDL (HPDL) cells were individually prepared from 15 extracted teeth. HPDL cells were cultured with or without FGF-2, and the percentages of STRO-1(+)/CD146(+) cells in each HPDL cell culture was examined using FACSAria™. The STRO-1(+)/CD146(+) cells were sorted with FACSAria™, and the mRNA expression and differentiation potency of the sorted cells were subsequently examined. The numbers of the STRO-1(+)/CD146(+) cells in the FGF-2 cultures were significantly higher than those cultured in the absence of FGF-2. The sorted STRO-1(+)/CD146(+) cells expressed mRNA of PDL markers and differentiated into adipocytes and osteoblast-like cells. The present study shows that FGF-2 augmented the proliferation of the STRO-1(+)/CD146(+) cells in the HPDL cultures whilst retaining adipogenic and osteogenic differentiation potentials. Thus, it may be useful to culture HPDL cells with FGF-2 for the application of the human STRO-1(+)/CD146(+) PDL cells in periodontal tissue regeneration.

A Genome-wide Association Study of Periodontitis in a Japanese Population

Periodontitis is a multifactorial disease in which bacterial, lifestyle, and genetic factors are involved. Although previous genetic association studies identified several susceptibility genes for periodontitis in European populations, there is little information for Asian populations. Here, we conducted a genome-wide association study and a replication study consisting of 2,760 Japanese periodontitis patients and 15,158 Japanese controls. Although single-nucleotide polymorphisms that surpassed a stringent genome-wide significance threshold (P < 5 × 10−8) were not identified, we found 2 suggestive loci for periodontitis: KCNQ5 on chromosome 6q13 (rs9446777, P = 4.83 × 10−6, odds ratio = 0.82) and GPR141-NME8 at chromosome 7p14.1 (rs2392510, P = 4.17 × 10−6, odds ratio = 0.87). A stratified analysis indicated that the GPR141-NME8 locus had a strong genetic effect on the susceptibility to generalized periodontitis in Japanese individuals with a history of smoking. In conclusion, this study identified 2 suggestive loci for periodontitis in a Japanese population. This study should contribute to a further understanding of genetic factors for enhanced susceptibility to periodontitis.

Effectable application of vascular endothelial growth factor to critical sized rat calvaria defects.

OBJECTIVE An early vascular response for angiogenesis is essential for the normal progression of bone defect healing. Vascular endothelial growth factor (VEGF) is a potent inducer of angiogenesis. The aim of this study was to evaluate the effects of a poly (L,D-lactic-co-glycolic acid) (PLGA) membrane with VEGF encapsulated into PLGA microspheres on bone regeneration at bone defects in rat calvaria. STUDY DESIGN Microspheres of PLGA incorporating VEGF(165) (VEGF microspheres) were prepared, and critical-size bone defects were created in rat calvaria. The VEGF microspheres, PLGA microspheres, or VEGF microspheres plus PLGA membrane were applied to the defects. Bone regeneration was evaluated using image analysis based on soft radiographic and histologic examination. RESULTS Mature thick bone regeneration was observed in selected sites at bone defects that had been applied with VEGF microspheres/PLGA membrane compared with those that had been applied with the other treatments. CONCLUSION A combination of VEGF microspheres and a PLGA membrane effectively enhances bone regeneration.

Randomized Placebo-Controlled and Controlled Non-Inferiority Phase III Trials Comparing Trafermin, a Recombinant Human Fibroblast Growth Factor 2, and Enamel Matrix Derivative in Periodontal Regeneration in Intrabony Defects

We investigated the efficacy, safety, and clinical significance of trafermin, a recombinant human fibroblast growth factor (rhFGF)‐2, for periodontal regeneration in intrabony defects in Phase III trials. Study A, a multicenter, randomized, double‐blind, placebo‐controlled study, was conducted at 24 centers. Patients with periodontitis with 4‐mm and 3‐mm or deeper probing pocket depth and intrabony defects, respectively, were included. A total of 328 patients were randomly assigned (2:1) to receive 0.3% rhFGF‐2 or placebo, and 323 patients received the assigned investigational drug during flap surgery. One of the co‐primary endpoints, the percentage of bone fill at 36 weeks after drug administration, was significantly greater in the rhFGF‐2 group at 37.131% (95% confidence interval [CI], 32.7502 to 41.5123; n = 208) than it was in the placebo group at 21.579% (95% CI, 16.3571 to 26.8011; n = 100; p < 0.001). The other endpoint, the clinical attachment level regained at 36 weeks, was not significantly different between groups. Study B, a multicenter, randomized, blinded (patients and evaluators of radiographs), and active‐controlled study was conducted at 15 centers to clarify the clinical significance of rhFGF‐2. Patients with 6‐mm and 4‐mm or deeper probing pocket depth and intrabony defects, respectively, were included. A total of 274 patients were randomly assigned (5:5:2) to receive rhFGF‐2, enamel matrix derivative (EMD), or flap surgery alone. A total of 267 patients received the assigned treatment during flap surgery. The primary endpoint, the linear alveolar bone growth at 36 weeks, was 1.927 mm (95% CI, 1.6615 to 2.1920; n = 108) in the rhFGF‐2 group and 1.359 mm (95% CI, 1.0683 to 1.6495; n = 109) in the EMD group, showing non‐inferiority (a prespecified margin of 0.3 mm) and superiority of rhFGF‐2 to EMD. Safety problems were not identified in either study. Therefore, trafermin is an effective and safe treatment for periodontal regeneration in intrabony defect, and its efficacy was superior in rhFGF‐2 compared to EMD treatments.

Multipotency of clonal cells derived from swine periodontal ligament and differential regulation by fibroblast growth factor and bone morphogenetic protein

BACKGROUND AND OBJECTIVE A blood supply is indispensable for the regeneration of damaged or lost periodontal ligament (PDL) tissue. Mesenchymal stem cell-like activity of cells derived from the PDL has been identified by their capacity to form fibrous and osseous tissue and cementum. However, it remains to be clarified whether the cells have an ability to build the capillary network of blood vessels. This study evaluated the potential of cells derived from the PDL to construct a blood vessel-like structure and examined how growth factors controlled the multipotency of the cells. MATERIAL AND METHODS The ability of a swine PDL fibroblast cell line, TesPDL3, to construct a blood vessel-like structure was evaluated on and in the self-assembling peptide scaffold, PuraMatrix(TM). In addition, the ability of the cells to form mineralized nodules was evaluated on type I collagen-coated plastic plates. In some cases, fibroblast growth factor (FGF)-2 and bone morphogenetic protein (BMP)-2 were added to these cultures. The status of the expression of vascular and osteoblastic cell-specific markers in the cells was evaluated using reverse transcriptase-polymerase chain reaction (RT-PCR), Western blotting and immunofluorescence analyses. RESULTS The TesPDL3 cells not only formed mineralized nodules in response to BMP-2 stimulation but also constructed tube-like structures in response to FGF-2 stimulation. Intriguingly, FGF-2 inhibited the BMP-2-induced formation of mineralized nodules. Conversely, BMP-2 inhibited the FGF-2-induced formation of tube-like structures. CONCLUSION Periodontal ligament fibroblasts have the potential to differentiate not only into osteoblastic but also into vascular cell lineages. The destiny of the cells was reciprocally regulated by BMP-2 and FGF-2.

Prevalence and risk factors for peri-implant diseases in Japanese adult dental patients

We investigated the prevalences and risk factors for peri-implant diseases in Japanese adult dental patients attending a follow-up visit at dental hospitals or clinics as part of their maintenance program. This cross-sectional multicenter study enrolled patients with dental implants who attended regular check-ups as part of a periodontal maintenance program during the period from October 2012 through September 2013. Patients with implants with at least 3 years of loading time were included in the study. The condition of peri-implant tissue was examined and classified into the following categories: healthy, peri-implant mucositis, and peri-implantitis. Patients were also evaluated for implant risk factors. A total of 267 patients (110 men, 157 women; mean age: 62.5 ± 10.7 years) were analyzed. The prevalence of patient-based peri-implant mucositis was 33.3% (n = 89), and the prevalence of peri-implantitis was 9.7% (n = 26). Poor oral hygiene and a history of periodontitis were strong risk factors for peri-implant disease. The present prevalences were lower than those previously reported. The quality of periodontal therapy before and after implant installation and patient compliance and motivation, as indicated by plaque control level, appear to be important in maintaining peri-implant tissue health.

E-Selectin Mediates Porphyromonas gingivalis Adherence to Human Endothelial Cells

ABSTRACT Porphyromonas gingivalis, a major periodontal pathogen, may contribute to atherogenesis and other inflammatory cardiovascular diseases. However, little is known about interactions between P. gingivalis and endothelial cells. E-selectin is a membrane protein on endothelial cells that initiates recruitment of leukocytes to inflamed tissue, and it may also play a role in pathogen attachment. In the present study, we examined the role of E-selectin in P. gingivalis adherence to endothelial cells. Human umbilical vein endothelial cells (HUVECs) were stimulated with tumor necrosis factor alpha (TNF-α) to induce E-selectin expression. Adherence of P. gingivalis to HUVECs was measured by fluorescence microscopy. TNF-α increased adherence of wild-type P. gingivalis to HUVECs. Antibodies to E-selectin and sialyl Lewis X suppressed P. gingivalis adherence to stimulated HUVECs. P. gingivalis mutants lacking OmpA-like proteins Pgm6 and -7 had reduced adherence to stimulated HUVECs, but fimbria-deficient mutants were not affected. E-selectin-mediated P. gingivalis adherence activated endothelial exocytosis. These results suggest that the interaction between host E-selectin and pathogen Pgm6/7 mediates P. gingivalis adherence to endothelial cells and may trigger vascular inflammation.