Hiroki Sugito

A distinct cohort of progenitor cells participates in synovial joint and articular cartilage formation during mouse limb skeletogenesis

The origin, roles and fate of progenitor cells forming synovial joints during limb skeletogenesis remain largely unclear. Here we produced prenatal and postnatal genetic cell fate-maps by mating ROSA-LacZ-reporter mice with mice expressing Cre-recombinase at prospective joint sites under the control of Gdf5 regulatory sequences (Gdf5-Cre). Reporter-expressing cells initially constituted the interzone, a compact mesenchymal structure representing the first overt sign of joint formation, and displayed a gradient-like distribution along the ventral-to-dorsal axis. The cells expressed genes such as Wnt9a, Erg and collagen IIA, remained predominant in the joint-forming sites over time, gave rise to articular cartilage, synovial lining and other joint tissues, but contributed little if any to underlying growth plate cartilage and shaft. To study their developmental properties more directly, we isolated the joint-forming cells from prospective autopod joint sites using a novel microsurgical procedure and tested them in vitro. The cells displayed a propensity to undergo chondrogenesis that was enhanced by treatment with exogenous rGdf5 but blocked by Wnt9a over-expression. To test roles for such Wnt-mediated anti-chondrogenic capacity in vivo, we created conditional mutants deficient in Wnt/beta-catenin signaling using Col2-Cre or Gdf5-Cre. Synovial joints did form in both mutants; however, the joints displayed a defective flat cell layer normally abutting the synovial cavity and expressed markedly reduced levels of lubricin. In sum, our data indicate that cells present at prospective joint sites and expressing Gdf5 constitute a distinct cohort of progenitor cells responsible for limb joint formation. The cells appear to be patterned along specific limb symmetry axes and rely on local signaling tools to make distinct contributions to joint formation.

Regulation of CCN2 gene expression and possible roles in developing tooth germs

CCN proteins are extracellular and cell-associated molecules involved in several developmental processes, but their expression patterns and regulation in tooth development remain unclear. Here we first determined the expression patterns of CCN genes in mouse tooth germs. We found that at early stages CCN2 was detected in dental lamina, dental mesenchyme, and primary enamel knot, while other CCN family members were expressed broadly. By the bell stage, all members were expressed in differentiating odontoblasts and ameloblasts, but CCN1 and CCN2 transcripts were conspicuous in differentiating osteoblasts in dental follicle. Next, we asked what signalling molecules regulate CCN2 expression and what roles CCN2 may have. We found that upon surgical removal of dental epithelium CCN2 was not longer expressed in dental mesenchyme in cultured bud stage germs. Implantation of beads pre-coated with BMPs and FGFs onto E12-13 mandibular explants induced CCN2 expression in dental mesenchyme. There was a dose-dependent effect of BMP-4 on CCN2 induction; a concentration of 100 ng/μl was able to induce strong CCN2 expression while a minimum concentration of 25 ng/μl was needed to elicit appreciable expression. Importantly, Noggin treatment inhibited endogenous and BMP-induced CCN2 expression, verifying that CCN2 expression in developing tooth germs requires BMP signalling. Lastly, we found that rCCN2 stimulated proliferation in dental mesenchyme in a dose-dependent manner. Together, the data indicate that expression of CCN genes is spatio-temporally regulated in developing tooth germs. CCN2 expression appears to depend on epithelial and mesenchymal-derived signalling factors, and CCN2 can elicit strong proliferation in dental mesenchyme.

Point-of-care detection of Tannerella forsythia using an antigen–antibody assisted dielectrophoretic impedance measurement method

UNLABELLED The importance of periodontal treatment planning based on diagnosis with clinical detection of periodontal pathogens has been well recognized. However, reliable detection and quantification methods that can be conveniently used at chair-side have yet to be developed. This study aimed to evaluate the clinical use of a novel apparatus which uses an antigen-antibody reaction assisted dielectrophoretic impedance measurement (AA-DEPIM) for the detection of a prominent periodontal pathogen, Tannerella forsythia. A total of 15 patients with a clinical diagnosis of chronic periodontitis, three periodontally healthy volunteers and two with gingivitis were subjected to clinical and microbiological examinations. Saliva samples were analyzed for the presence of T. forsythia using AA-DEPIM, PCR-Invader and real-time PCR methods. The measurement values for total bacteria and T. forsythia using the prototype AA-DEPIM apparatus were significantly greater in periodontitis group than those in healthy/gingivitis group. Using the AA-DEPIM apparatus with tentative cut-off values, T. forsythia was detected for 14 (12 with periodontitis and 2 either healthy or with gingivitis) out of 20 individuals. The measurement for the detection of T. forsythia by the AA-DEPIM method showed a significant positive correlation with the detection by PCR-Invader (r = 0.541, p = 0.01) and the real-time PCR method (r = 0.834, p = 0.01). When the PCR-Invader method was used as a reference, the sensitivity and specificity of the AA-DEPIM method were 76.5% and 100%, respectively. The results suggested that the AA-DEPIM method has potential to be used for clinically evaluating salivary presence of T. forsythia at chair-side. TRIAL REGISTRATION UMIN Clinical Trials Registry (UMIN-CTR) UMIN000012181.

Periodontist-Dental Hygienist Collaboration in Periodontal Care for Chronic Periodontitis: An 11-year Case Report

We report a case of severe chronic periodontitis treated and longitudinally maintained by a periodontist and dental hygienists. The patient was a 45-year-old woman who presented with the chief complaint of gingival bleeding and tooth mobility. An initial examination revealed generalized gingival inflammation and subgingival calculus in the premolar and molar regions. Premature contact was observed in #14 and 45. Clinical examination revealed 42% of sites with a probing depth (PD) of ≥4 mm and 44% of sites with bleeding on probing. Radiographic examination revealed vertical bone resorption in #35, 36, and 45, and horizontal bone resorption in other regions. Based on a clinical diagnosis of severe chronic periodontitis, initial periodontal therapy consisting of plaque control, scaling and root planing, and removal of an ill-fitting prosthesis was performed. Following suppression of inflammation, occlusal adjustment of premature contact sites was performed. Open flap debridement was performed for teeth with a PD of ≥5 mm. After confirming the stability of the periodontal tissue, final prostheses were placed on #16, 35-37, and 46. Following re-evaluation, the patient was placed on supportive periodontal therapy. It has been 11 years since the patients first visit, and the periodontal conditions have remained stable. Meticulous periodontal care maintained over a number of years by a periodontist and dental hygienist have yielded a clinically favorable outcome.

Periodontal Surgery Involving Modified Widman Flap Procedure and Connective Tissue Graft for Generalized Aggressive Periodontitis: A Case Report

We report a case of generalized aggressive periodontitis (AgP) requiring periodontal treatment including flap surgery and ridge augmentation. The patient was a 39-year-old woman who presented with the chief complaint of pus discharge from tooth #36. No other obvious signs of gingival inflammation were observed. Periodontal examination revealed multiple sites with a probing depth of ≥10 mm. Radiography showed pro-nounced bone defects in the maxillary incisors and molar region. Real-time PCR was used to detect Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, and Tannerella forsythia in subgingival plaque; all 3 pathogens were found. Based on a clinical diagnosis of generalized AgP, periodontal therapy was initiated, which resulted in an improvement in clinical and microbiological parameters. A modified Widman flap procedure was then performed on sites with residual periodontal pockets. Next, a connective tissue graft was performed for ridge augmentation at #22, which had shown evidence of ridge resorption. Postoperative reevaluation revealed a reduction in probing depth and an improvement in marginal bone levels. Oral function was then restored using a fixed bridge prosthesis and maintenance therapy initiated. The periodontal condition has remained stable over a 2.5-year period. In the present case of AgP, surgical intervention reduced periodontal pockets and periodontal pathogens and improved the architecture of both the hard and soft tissues, allowing subsequent care of the periodontium to be performed efficiently by the patient.

Treatment Capabilities of Postgraduate Dental Trainees at okyo Dental College Suidobashi Hospital Based on Dental Services Income

As of fiscal year 2006, it became compulsory for all newly licensed dentists in Japan to undergo a year of practical training at one of the designated training facilities found throughout the country as part of their postgraduate programs. The goal of this training is for the trainees to acquire diagnostic and therapeutic skills. While officially trainees, they are nonetheless regarded as members of staff. Clinical training emphasizes improving both technical skills and theoretical knowledge. However, taking on such trainees is sometimes considered unprofitable, as work productivity is claimed to be low. The purpose of this study was to compare work outcomes and income generated between trainees and part-time dentists working at the Tokyo Dental College Suidobashi Hospital. The part-time dentists comprised clinical instructors and dentists responsible for outpatients. Postgraduate dental trainees also generally conduct dental treatment for outpatients. Therefore, part-time dentists were considered the most suitable for a control group. No significant difference was observed in the total number of patients seen by either group by the final term of clinical training. Furthermore, no significant difference was observed in insurance-based dental treatment unit income (insured care unit income) per patient between the two groups from the mid-term period of training onwards. These results suggest that, although the trainees were less efficient in terms of time taken to provide a diagnosis and treatment, their performance was almost equal when viewed from the viewpoint of insured care provided. Taken together, this suggests that time-related care efficiency must be increased and specific training policies and strategies developed to improve the work productivity of dental trainees.