Richard T. Kao

Platelet-Derived Growth Factor Promotes Periodontal Regeneration in Localized Osseous Defects: 36-Month Extension Results From a Randomized, Controlled, Double-Masked Clinical Trial

BACKGROUND Recombinant human platelet-derived growth factor (rhPDGF) is safe and effective for the treatment of periodontal defects in short-term studies up to 6 months in duration. We now provide results from a 36-month extension study of a multicenter, randomized, controlled clinical trial evaluating the effect and long-term stability of PDGF-BB treatment in patients with localized severe periodontal osseous defects. METHODS A total of 135 participants were enrolled from six clinical centers for an extension trial. Eighty-three individuals completed the study at 36 months and were included in the analysis. The study investigated the local application of β-tricalcium phosphate scaffold matrix with or without two different dose levels of PDGF (0.3 or 1.0 mg/mL PDGF-BB) in patients possessing one localized periodontal osseous defect. Composite analysis for clinical and radiographic evidence of treatment success was defined as percentage of cases with clinical attachment level (CAL) ≥2.7 mm and linear bone growth (LBG) ≥1.1 mm. RESULTS The participants exceeding this composite outcome benchmark in the 0.3 mg/mL rhPDGF-BB group went from 62.2% at 12 months, 75.9% at 24 months, to 87.0% at 36 months compared with 39.5%, 48.3%, and 53.8%, respectively, in the scaffold control group at these same time points (P <0.05). Although there were no significant increases in CAL and LBG at 36 months among all groups, there were continued increases in CAL gain, LBG, and percentage bone fill over time, suggesting overall stability of the regenerative response. CONCLUSION PDGF-BB in a synthetic scaffold matrix promotes long-term stable clinical and radiographic improvements as measured by composite outcomes for CAL gain and LBG for patients possessing localized periodontal defects ( ClinicalTrials.gov no. CT01530126).

Periodontal regeneration - intrabony defects: a systematic review from the AAP Regeneration Workshop.

BACKGROUND Previous systematic reviews of periodontal regeneration with bone replacement grafts and guided tissue regeneration (GTR) were defined as state of the art for clinical periodontal regeneration as of 2002. METHODS The purpose of this systematic review is to update those consensus reports by reviewing periodontal regeneration approaches developed for the correction of intrabony defects with the focus on patient-, tooth-, and site-centered factors, surgical approaches, surgical determinants, and biologics. This review adheres to the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines for systematic reviews. A computerized search of the PubMed and Cochrane databases was performed to evaluate the clinically available regenerative approaches for intrabony defects. The search included screening of original reports, review articles, and reference lists of retrieved articles and hand searches of selected journals. All searches were focused on clinically available regenerative approaches with histologic evidence of periodontal regeneration in humans published in English. For topics in which the literature is lacking, non-randomized observational and experimental animal model studies were used. Therapeutic endpoints examined included changes in clinical attachment level, changes in bone level/fill, and probing depth. For purposes of analysis, change in bone fill was used as the primary outcome measure, except in cases in which this information was not available. The SORT (Strength of Recommendation Taxonomy) grading scale was used in evaluating the body of knowledge. RESULTS 1) Fifty-eight studies provided data on patient, tooth, and surgical-site considerations in the treatment of intrabony defects. 2) Forty-five controlled studies provided outcome analysis on the use of biologics for the treatment of intrabony defects. CONCLUSIONS 1) Biologics (enamel matrix derivative and recombinant human platelet-derived growth factor-BB plus β-tricalcium phosphate) are generally comparable with demineralized freeze-dried bone allograft and GTR and superior to open flap debridement procedures in improving clinical parameters in the treatment of intrabony defects. 2) Histologic evidence of regeneration has been demonstrated with laser therapy; however, data are limited on clinical predictability and effectiveness. 3) Clinical outcomes appear most appreciably influenced by patient behaviors and surgical approach rather than by tooth and defect characteristics. 4) Long-term studies indicate that improvements in clinical parameters are maintainable up to 10 years, even in severely compromised teeth, consistent with a favorable/good long-term prognosis.

Elastin degradation by proteases from cultured human breast cancer cells

Summary Potent elastinolytic activity was elaborated into the media of several cultured human breast cancer cell lines. Cultured stromal cells from normal and tumor-bearing human breast tissue contained no such activity. This elastase-like activity was resolved into three major peaks of activity by DEAE-cellulose column chromatography. Two of the three activities were separated from the collegenolytic activity also produced by these cells. Inhibition profiles of the elastinolytic activities indicated that two were serine proteases and one was a metalloprotease. Each of these three proteases were in an inactive form and required trypsin pretreatment for activity. The desmoplastic reaction to human breast cancer contains both collagen and elastin. The present findings indicate that the breast tumor cells themselves may modulate the turnover of these stromal proteins.

Ascorbic acid stimulates production of glycosaminoglycans in cultured fibroblasts

The effect of ascorbic acid on collagen synthesis is well characterized. Proteoglycans and their attached glycosaminoglycans are components of the extracellular matrix closely associated with collagen fibers. We examined whether ascorbic acid also plays a role in glycosaminoglycan production. Synthesis and deposition of glycosaminoglycans into the extracellular matrix and secretion into the media were followed in human skin fibroblasts cultured in the presence and absence of ascorbic acid. Specific glycosaminoglycans were identified and quantitated by differential enzyme digestion, ion-exchange column chromatography, and cellulose-acetate electrophoresis. No major qualitative changes in glycosaminoglycans were observed. However, quantitatively, synthesis of glycosaminoglycans increased 30 to 90%, and deposition into the extracellular matrix increased 80% in the presence of ascorbic acid. This effect was only in part secondary to decreased levels of collagen, and the diminished capacity of underhydroxylated collagen to bind proteoglycans. The effect of ascorbic acid on extracellular macromolecules is thus more pervasive than previously assumed.

Collagen and elastin synthesis in human stroma and breast carcinoma cell lines: modulation by the extracellular matrix.

The desmoplastic response to breast carcinoma is being studied. The stimulation of stromal cell proliferation by a preformed breast tumor matrix was shown. An additional mechanism for stimulating scleroprotein deposition is described here. On a per-cell basis, the synthesis of collagen and elastin was increased by 50% and 70%, respectively, in fibroblasts grown on the preformed breast tumor matrix compared to the same cells grown on plastic or on their own preformed matrix. Breast tumor cells themselves synthesized small amounts of collagen and elastin compared to fibroblasts. These levels were unchanged when breast tumor cells were grown on the preformed matrix of fibroblasts. Addition of steroid hormones to cultured cells grown on plastic or on preformed matrices in various combinations, did not change the levels of either collagen or elastin synthesis. The matrix of human breast tumor cells exerts a dual effect; it is mitogenic for fibroblasts, and also stimulates the level of collagen and elastin synthesis, events that could contribute to the formation of the desmoplastic response to human breast cancer in situ.

Periodontal Regeneration – Intrabony Defects: A Consensus Report From the AAP Regeneration Workshop

BACKGROUND Treatment of intrabony defects is an important therapeutic goal of periodontal therapy. The goal of this consensus report was to critically appraise the evidence for the available approaches for promoting periodontal regeneration in intrabony defects. In addition to evaluating the effectiveness of new regenerative approaches for intrabony defects, recommendations for future research were defined for this area. METHODS A systematic review was conducted using computerized searches of PubMed and Cochrane databases, supplemented with screening of references in original reports, review articles, and a hand search in selected journals. All searches were focused on regenerative approaches with histologic evidence of periodontal regeneration (proof of principle), clinical trials, and case reports. For purposes of analysis, change in intrabony defect fill was considered the primary outcome variable, with change in clinical attachment as a secondary outcome. The SORT (Strength of Recommendation Taxonomy) grade was used to evaluate the quality and strength of the evidence. During the consensus meeting, the group agreed on the outcomes of the systematic review, pertinent sources of evidence, clinical recommendations, and areas requiring future research. RESULTS The systematic review, which was conducted for the consensus conference, evaluated the effectiveness of the use of biologics for the treatment of intrabony defects. Enamel matrix derivative (EMD) and recombinant human platelet-derived growth factor-BB (rhPDGF-BB) with β-tricalcium phosphate were shown to be efficacious in regenerating intrabony defects. The level of evidence is supported by multiple studies documenting effectiveness. The clinical application of biologics supports improvements in clinical parameters comparable with selected bone replacement grafts and guided tissue regeneration (GTR). Factors negatively affecting regeneration included smoking and excessive tooth mobility. CONCLUSIONS Periodontal regeneration in intrabony defects is possible on previously diseased root surfaces, as evidenced by a gain in clinical attachment, decreased pocket probing depth, gain in radiographic bone height, and overall improvement in periodontal health. These clinical findings are consistent with available histologic evidence. Clinical improvements can be maintained over long periods (>10 years). Although bone replacement grafts have been the most commonly investigated modality, GTR, biologics, and combination therapies have also been shown to be effective. Future research should emphasize patient-reported outcomes, individual response differences, and emerging technologies to enhance treatment results. CLINICAL RECOMMENDATIONS Early management of intrabony defects with regenerative therapies offers the greatest potential for successful periodontal regeneration. The clinical selection and application of a regenerative therapy or combination of therapies for periodontal regeneration should be based on the clinicians experiences and understanding of the regenerative biology and technology. This decision-making process should take into consideration the potential adverse influence of factors, such as smoking, poor oral hygiene, tooth mobility, and defect morphology, on regeneration. Management should be coupled with an effective maintenance program for long-term success.

Cyclic AMP signaling in bone marrow stromal cells has reciprocal effects on the ability of mesenchymal stem cells to differentiate into mature osteoblasts versus mature adipocytes

Stimulatory G protein-mediated cAMP signaling is intimately involved in skeletal homeostasis. However, limited information is available on the role of the cAMP signaling in regulating the differentiation of mesenchymal stem cells into mature osteoblasts and adipocytes. To investigate this, we treated primary mouse bone marrow stromal cells (BMSCs) with forskolin to stimulate cAMP signaling and determined the effect on osteoblast and adipocyte differentiation. Exposure of differentiating osteoblasts to forskolin markedly inhibited progression to the late stages of osteoblast differentiation, and this effect was replicated by continuous exposure to PTH. Strikingly, forskolin activation of cAMP signaling in BMSCs conditioned mesenchymal stem cells (MSCs) to undergo increased osteogenic differentiation and decreased adipogenic differentiation. PTH treatment of BMSCs also enhanced subsequent osteogenesis, but promoted an increased adipogenesis as well. Thus, activation of cAMP signaling alters the lineage commitment of MSCs, favoring osteogenesis at the expense of adipogenesis.

Effect of demineralized bone powder on osteoblast-like cells in culture: A potential rapid quality control assay

Demineralized bone powder (DBP) has been shown to induce osteogenesis in a variety of bone defects and extra-osseous sites. Previous investigations have been carried out in animal models which are time-consuming and expensive. We studied the effect of DBP on well-established populations of osteoblast and non-osteoblast-like cells in culture to establish an inexpensive, efficient and reliable assay for bone induction. DBP and BP (non-demineralized powder), of particle size 38-53 microns, were prepared from rat long bones. ROS (rat osteosarcoma) 17/2.8 and ROS 24/1 cell lines were subcultured weekly. For both 17/2.8 (well differentiated) and 24/1 (poorly differentiated) cells, proliferation, i.e. cell count, was significantly greater in DBP enriched medium when compared with control or medium with BP. Cell counts for wells with BP were no different from controls. The increased cell count in DBP-enriched medium was significant on days 2-5 (peak effect 2-3 days). Alkaline phosphatase production reached peak levels after day 3 when proliferation was beginning to taper off. In this study a consistent increase in osteoblast proliferation and alkaline phosphatase production under the influence of DBP was demonstrated. The tissue culture assay for proliferation must now be correlated with bone induction in vivo. In future, the method may be useful for investigating the mechanism of bone induction.

Mitogenic Response of Cells in Culture to Demineralized Bone Matrix

In this study the mitogenic effect of demineralized bone powder (DBP) on cells in tissue culture was evaluated. The cells studied have osteoblastic properties. At a DBP concentration of 100 micrograms/well, rat osteosarcoma cells (ROS) 17/2.8 cells, ROS 24/1 cells, and periosteal fibroblasts (POF) all showed a significant increase in proliferation in comparison to controls. The peak effect was on day two for ROS 17/2.8 (125% of control), on day three for ROS 24/1 (139% of control), and on day four for POF (145% of control). The results indicate that this in vitro system may be useful for evaluating the osteoinductive activity of DBP preparations.

Tissue Engineering Approach to Implant Site Development

One of the greatest challenges for the surgical placement of dental implants is the lack of adequate bone volume. Many techniques are available but they are both technique sensitive, and the results are not always predictable. Tissue engineering through the use of biologics provides a strategy for enhancing bone regeneration. Though this is a relatively new strategy, this chapter focuses on technologies that are available for clinicians to utilize at this time.