HSDM Theses and Dissertations

Background: Platelet rich plasma (PRP) has been utilized as an adjunct to various periodontal procedures with the goal of potentializing the regenerative outcome of surgical therapy. The biopolymer gelatin hydroxylphenylpropionic acid (Gtn-HPA) has recently been developed and proposed to be used as an injectable scaffold for tissue engineering. The aim of the present study was to characterize the influence of the Gtn-HPA hydrogel incorporating PRP on osteoprogenitor cells behavior and growth factor release in vitro. Methods: Platelet rich plasma was obtained from 6 human donors followed by platelet counting. The mRNA levels of osteoblast differentiation markers including OCN and COL1A1 plus the growth factors receptors FGFr, EGFr and PDGFr were analyzed in 2D cultures, hydrogel only, 10% PRP, 20% PRP, 10% FBS and 20% FBS in a 2% Gtn-HPA hydrogel (n=3) and seeded with osteoprogenitor cells. Furthermore, protein quantification for PDGF, TGF-B, EGF, FGF and IL-1B (n=5) were analyzed in the PRP lysate samples. Results: Platelet concentration increased between 5 to 9-fold in PRP compared to whole blood counts. Protein quantification was consistently greater for TGFB, PDGF and EGF. Gene expression analysis showed a tendency of upregulation of the osteoblast differentiation markers and growth factor receptor genes in the PRP groups, however a great variability between the donors was noticed. Conclusion: Within the limitations of this study, the use of human PRP in a Gtn-HPA hydrogel reveals this biopolymer as a promising and suitable carrier/scaffold with optimal growth factor release and enhanced gene expression favoring tissue regeneration.

Background: Conventional bone augmentation for dental implants typically involves grafting materials, with or without barrier membranes, to preserve ridge dimensions. This study compares the changes in horizontal ridge width in patients undergoing alveolar ridge preservation (ARP) with a xenogenic (bovine) collagen plug, with and without a collagen membrane. Methods: A 6-month randomized clinical trial was conducted with 20 participants requiring ARP after tooth extraction. Participants were randomly assigned to two groups: Group I, which received a xenogenic plug alone, and Group II, which received a xenogeneic plug with a non-crosslinked collagen membrane. Both groups were allowed to heal by secondary intention. The outcomes of interest included clinical, radiographic, and ultrasonographic changes in hard and soft tissues, as well as histologic and histomorphometric evidence of new bone formation. Results: All enrolled participants completed the study, with a mean age of 54.25 ± 17.08 years, including 17 premolars and 3 . Baseline buccal bone thickness was similar between the groups (Group 1: 1.02 ± 0.44 mm; Group 2: 0.93 ± 0.45 mm; p = 0.662). At 6 months, no significant differences were observed between groups for keratinized tissue width (Group 1: 4.80 ± 1.48 mm; Group 2: 4.80 ± 1.14 mm; p = 0.999), gingival thickness (Group 1: 1.22 ± 0.25 mm; Group 2: 1.20 ± 0.35 mm; p = 0.650), horizontal bone width at 3 mm from the crest (Group 1: 9.02 ± 2.09 mm; Group 2: 8.41 ± 1.45 mm; p = 0.464), and change in vertical bone at the buccal aspect (Group 1: -0.78 ± 0.59 mm; Group 2: -1.31 ± 1.20 mm; p = 0.244). Ultrasound evaluations were consistent with clinical measurements. Histologic analysis showed similar new bone formation between groups (Group 1: 25.80 ± 15.48%; Group 2: 27.89 ± 18.21%; p = 0.803). Conclusion: The xenogeneic collagen plug alone yielded comparable ARP outcomes to its combination with a collagen membrane, simplifying the treatment process by reducing costs, time, and the need for additional materials.

Background: Peri-implant disease management continues to evolve, with increasing emphasis on early detection and monitoring to prevent and treat peri-implant disease. Conventional periodontal probing is the gold standard for diagnosis, however, high-frequency dental ultrasonography has emerged as an innovative diagnostic tool capable of assessing peri-implant lesions and tissue perfusion associated with inflammation. In addition, peri-implant crevicular fluid (PICF) biomarker analysis is another new diagnostic tool for peri-implant disease, shedding light onto the host inflammatory response to pathogenic bacteria. The aim of this longitudinal split-mouth study was to assess whether ultrasonographic and biomarker analysis can aid in early diagnosis of peri-implant disease and predict the outcomes of peri-implant mucositis (PM) treatment over a 1-year period. Methods: Twenty patients treated at the Harvard Dental Center with two non-adjacent implants (one healthy and one diagnosed with PM) were enrolled. Ultrasonographic and clinical parameters were assessed at baseline (T0), four weeks post-nonsurgical therapy (T1), and one year post-therapy (T2). Ultrasonographic measurements at midfacial and interproximal surfaces included tissue perfusion (via Doppler analysis), presence of a “lesion” (hypoechoic supracrestal area [HSA]), mucosal thickness (MT), buccal bone distance (BBD), buccal bone thickness (BBT), supracrestal tissue height (STH), and mesial and distal papilla height (PH). Clinical parameters included probing depth (PD), bleeding on probing (BOP), suppuration, keratinized mucosa width (KMW), plaque accumulation, and peri-implant soft tissue dehiscence (PSTD). Peri-implant crevicular fluid samples were collected from healthy and PM sites at T0 and T1 to quantify biomarkers associated with PM (bFGF, IL-1β, IL-6, IL-17, MMP-8, MMP-9, TNF-α, and VEGF). Patient reported outcome measures (PROMs) assessing esthetic satisfaction and discomfort during brushing (0–10 scale) were recorded at all time points. Non-surgical therapy was delivered to PM implants at T0, and oral hygiene instructions were reinforced. Patients were followed by their primary providers at three and six months. Fourteen of twenty participants completed the one-year follow-up. Clinical, ultrasonographic, and PROM assessments were repeated at T2.

Results: At T1, ten of the twenty (50%) diseased implant sites exhibited complete resolution of inflammation. At T2, five of fourteen sites (35.7%) initially categorized as PM were diagnosed as healthy. The initial round of non-surgical therapy at diseased implants resulted in a significant reduction in coronal MT (p.05) and BOP (p.01). At T0, 75% of PM implants presented BOP at the mesiobuccal aspect, 65% at the midbuccal aspect, 65% at the distobuccal aspect, and 68.3% across all three buccal sites. At T1, relative BOP reduction was 46.7%, 38.5%, and 53.8% at the mesiobuccal aspect, midbuccal aspect, and distobuccal aspect, respectively (p.01 for all comparisons). From T0 to T2, relative BOP reduction was 36%, 29%, and 44% at the mesiobuccal aspect, midbuccal aspect, and distobuccal aspect, respectively (p.05 for all comparisons). Implant recession increased by a mean of 0.51 mm from T0 to T1 and 0.38 mm from T0 to T2. At T0, 19 healthy implant sites (95%) did not show ultrasonographic presence of an HSA (95%). The proportion of healthy sites without HSA was 80% at T1 and 78.6% at T2. In contrast, 90% of PM implants exhibited a detectable HSA at T0, while 80% and 64.3% of PM sites displayed an HSA at T1 and T2, respectively. The mean vertical extension of the HSA was 5.75 mm at T0, 3.2 mm at T1, and 3.9 mm at T2. The mean HSA area was 4.94 mm² at T0, 2.61 mm² at T1, and 3.11 mm² at T2. Color Doppler velocity and power Doppler intensity were significantly higher in PM sites compared to healthy sites at all time points (p.01) and correlated with HSA area (p.01), but not with other clinical parameters. Multilinear regression analysis revealed that baseline HSA area (p.01) and baseline power Doppler intensity (p.05) were significant predictors of PM resolution at one year. Baseline clinical parameters and biomarkers did not significantly correlate with resolution at T2. No significant differences in PROMs were observed between healthy and diseased sites at any time point. Conclusions: Echo-Doppler ultrasonography represents a valuable non-invasive tool for characterizing healthy and PM implant sites. Ultrasonographic parameters, including the presence of an HSA and Doppler-measured tissue perfusion, were associated with the clinical diagnosis of PM and showed potential predictive value for 1-year resolution following non-surgical therapy. In this cohort, non-surgical therapy resulted in modest resolution rates, highlighting variability in treatment response. Longer-term studies are required to validate these preliminary findings.

Free gingival grafts (FGG) are the gold standard for peri-implant phenotype augmentation in sites with inadequate keratinized mucosa (KM 2mm) and adhered mucosal width (AMW 1mm). However, FGGs are historically associated with postoperative morbidity due to palatal harvesting and esthetic shortcomings. Previous studies demonstrate Buccal Strip Gingival Graft (BSGG) combined with a xenogeneic collagen matrix (XCM) as a minimally invasive, patient-centered alternative to achieve phenotype modification and improved esthetic results. To compare the clinical, ultrasonographic, and patient-reported outcome measures (PROMs) of both methods, a randomized control trial with 12 months of follow up is presented under a non-inferiority framework. Thirty-two participants (38 implants) were randomized (1:1) to receive either BSGG + XCM (n = 16) or FGG (n = 16). The primary success criteria were successful phenotype modification defined as KM ≥ 2 mm with adherent, non-movable tissue (AMW ≥ 1mm) at 12 months. Secondary endpoints included KM gain, AWM gain, vestibular depth, mucosal thickness, volumetric changes, and peri-implant soft-tissue stability. Ultrasonographic biomarkers and measurements such as soft tissue thickness gain, perfusion index (PI), elasticity, and echo intensity (EI) were evaluated as a surrogate marker of graft vascularization and collagen maturation. Peri-implant crevicular fluid (PICF) biomarkers were analyzed via ELISA to explore early wound-healing dynamics and their predictive value for final outcomes. PROMs, including postoperative pain, recovery time, esthetic satisfaction, oral health-related quality of life, and willingness to retreat, were recorded through standardized questionnaires and analyzed under a superiority framework. Mixed-effects regression models and Fisher’s exact tests were used for continuous and categorical analyses, respectively. Correlations between ultrasonographic perfusion parameters, clinical healing indices, and PROMs were also examined. Non-inferiority of BSGG + XCM relative to FGG was demonstrated when comparing the primary outcome of achieving KM ≥ 2mm and AMW ≥ 1mm (BSGG + XCM = 88.2%; FGG = 89.5%). KM gain, PD, and recession changes between both groups were statistically equivalent (p > 0.05). While FGG produced greater increased vestibular depth (p = 0.04), BSGG + XCM demonstrated significantly lower postoperative pain (p 0.01), faster functional recovery, higher esthetic satisfaction, quicker recovery time, and greater willingness to retreat (all p 0.05). Comparable soft tissue maturity between groups at 12 months were confirmed by ultrasonographic analysis. Early PICF biomarker expression patterns differed between groups, suggesting distinct biologic healing pathways; exploratory analyses indicated potential associations between early angiogenic and inflammatory mediator levels and final tissue outcomes. The BSGG + XCM approach achieves non-inferior clinical, ultrasonographic, and biomarker outcomes compared with FGG. The BSGG + XCM PROMs reported greater patient comfort, recovery, and esthetic satisfaction. FGG may yield greater dimensional soft tissue gains and vestibular depth, but it is associated with increased postoperative morbidity. BSGG + XCM provides a minimally invasive soft tissue alternative for peri-implant phenotype modification.

Background: Periodontitis and type II diabetes mellitus (T2DM) are both chronic inflammatory diseases and have been shown to have a bidirectional relationship. Periodontal inflammation can worsen glycemic control, while hyperglycemia can raise baseline inflammatory load and worsen periodontal tissue breakdown.

Aim: To evaluate whether an electronic toothbrush with a smartphone application, used as an adjunct to scaling and root planing (ScRP), improves glycemic control and periodontal outcomes, compared to a manual toothbrush.

Methods: Adults ≥35-year-old with stage II-III periodontitis and T2DM with HbA1c ≥7% were included for the study. Patients received ScRP and oral hygiene instruction, and frequent periodontal maintenance. Patients were randomized into either (1) manual toothbrush (control) or (2) electronic toothbrush with smartphone application (test) group. Patients receives periodontal maintenances at follow-ups visits at 3, 6, 9, and 12 months. Oral hygiene instruction and periodontal maintenance was provided at follow-up visits at 3, 6, 9, and 12 months. The primary outcome was HbA1c changes from baseline. Secondary outcomes included changes in probing pocket depth (PPD), residual deep pocket, bleeding on probing (BOP), plaque index (PI), and inflammatory biomarkers collected from gingival crevicular fluid (GCF).

Results: 32 participants were enrolled; 23 were included in the final analysis (11 control, 12 test). The mean age was 62-year-old for both groups. HbA1c declined initially for both groups and then increased again at the end of 12-month follow-up. In the control group, the greatest mean HbA1c reduction of 0.51% was at 6 months. In the test group the greatest mean HbA1c reduction of 0.85% was at 3 months. Overall, HbA1c level changes did not demonstrate a statistically significant difference between the two groups over time. For periodontal clinical parameters, PPD reduction were significant across all time points for both control and test groups, however, no between-group differences were observed. End point parameter of residual deep pockets showed significant differences across all time points within-group and a significant difference at 9 months between two groups, favoring the test group. BOP showed statistically significant within-group reduction across all time points for test group but only significant at 2 and 6 months for control group. Between-group difference was statistically significant at 9 months, favoring the test group. PI showed no statistical significance between the two groups. GCF biomarker analyses were completed for 14 inflammatory biomarkers. Statistically significant between-group differences at the p .05 threshold were limited to TNFα, IL-2, IL-4, IL-5, and VEGF.

Conclusions: Electronic toothbrush with smartphone application when used as adjunct to nonsurgical periodontal therapy with regular maintenance enhanced periodontal clinical outcomes. However, the effects on HbA1c are comparable between two groups.

Background: Conventional antibacterial treatment fails to eradicate biofilms associated with dental caries and periodontal disease. Biofilm targeted-photodynamic therapy (PDT) could be a solution to these problems. Objective: to identify broad-spectrum bacterial binding ligands from one-bead one-compound (OBOC) libraries to develop targeting ligands against biofilm thorough phototherapy.
Material & Methods: Biofilm bacteria of S. mutans and E.coli were created on tissue flasks. After thorough washing to remove the free bacteria, the biofilm bacteria were scraped with a sterile blade. The scraped bacteria were ultra-sonicated and were incubated with OBOC libraries. The beads with S. mutans binding were sorted. After removing the bacteria from beads, the S. mutans binding beads were then re-incubated with E.coli. The sequences were determined by edman chemistry. The focused library was generated according to the mortif. The bacteria of S. mutans, P.gingivitis, E.coli, P. aeruginosa, Klebsiella sp, E. faecalis, citrabacter sp., and S. aureus were screened with focused library one by one. The compound beads with the property of binding to all of above bacteria were isolated and then incubated with human oral epithelial cells as well as human endothelial cells. The beads that bind to bacteria, but not to human oral epithelial cells and endothelial cells were finally identified. The chemical structures of identified compounds were determined by using peptide sequencing. The bacterial binding compounds were then conjugated with biotin through lysine and challenged with biofilm to testing their binding efficacy through streptavidin 488. The “all in one” targeting nanoparticles with hydrophobic core of photosensitizers were developed by decorating these bacterial binding ligands with porphyrins through click chemistry. The phototoxic efficacy of targeting nanoparticles on biofilm was evaluated by using viable account.
Results: four bacterial compounds were identified for possessing the property of specifically binding to all of the above bacteria, but not binding to normal human oral cells and endothelial cells. MTT assays indicated that these four compounds have no cytotoxicity to human epithelial cells and endothelial cells at up to 500 uM. Zonal inhibition assay indicated these four ligands do not have bactericidal effects. Compared with the porphyrin nanoparticles control, targeting nanoparticles caused a 100-fold reduction in the number of mix E. coli and S.mutans in the biofilm, whereas control group phototherapy had no effect on the number of E. coli.
Conclusion: A high throughput bacterial binding compound screening method was established and four novel bacterial binding compounds were identified. The use of a compounds photosensitizer conjugate is a more selective method of delivery thermal and dynamic therapy to biofilm. Work is currently under way to evaluate the phototoxic efficacy of these targeted nanoparticles on S. mutans, P. gingivitis, E. coli, P. aeruginosa, Klebsiella sp, E. faecalis, citrabacter sp., and S.aureus biofilm to identify the optimal parameters to fight biofilm.

Objectives: The first specific aim of this thesis was to define the presence, assess the distribution, and characterize PRX1 expressing cells within the periodontium. The second specific aim was to assess the contribution of PRX1 expressing cells to the periodontal formation, post-natal periodontal hemostasis, and periodontal regeneration. Methods: To assess the significance of the expression of PRX1 in human periodontium, we tested the expression of the Prx1 gene in samples of human bone, periodontal ligament (PDL), and periodontal stem cells (PDLSCs). The presence of PRX1 expressing cells (PRX1+ cells) in the mouse periodontium was explored by means of in vivo imaging. The distribution of PRX1+ cells in mouse periodontium was investigated in inducible lineage tracing analysis performed postnatally (Prx1-CreER-EGFP+/-;Rosa26-tdTOMATO+/-). PRX1+ cells also were isolated from periodontal ligament of mouse mandibular incisors and their gene expression pattern was explored. The contribution of PRX1+ cells to the formation of the mouse periodontium was investigated by constitutive lineage tracing analysis performed during embryogenesis (Prx1-cre+/-;Rosa26-tdTOMATO+/-). By means of diphtheria toxin (DTA)-mediated conditional ablation of PRX1+ cells, we also explored their contribution to periodontal homeostasis by assessing the width of PDL in Prx1-CreER-EGFP+/-;Rosa26-DTA+/- mice (ablation mouse model) (n=5) and their non-ablating littermates. The contribution of PRX1+ cells to periodontal regeneration was assessed by creating sub-critical size periodontal fenestration defects by the mandibular incisor and first molar teeth. The healing of defects was compared in ablation mouse model (test group) and non-ablation mouse model (control group) (n=5).
Results: Gene expression analysis of human samples confirmed that PRX1 is significantly expressed in human PDL and PDLSCs. The gene expression signature of PRX1+ cells demonstrated the expression mesenchymal stem cells markers, up-regulation of Notch signaling, and down-regulation of Wnt signaling. Lineage tracing analyses showed that PRX1+ cell are present in high abundance within the PDL of mouse incisor teeth, and PRX1-lineage cells contribute to the formation of the mouse incisor’s periodontium. Post-natal ablation of PRX1+ cells resulted in a significant enlargement of the periodontal ligament space and lack of periodontal regeneration in sub-critical size periodontal defects in the continuously regenerating periodontal ligament of mouse incisor. Conclusions: PRX1 expressing cells are present in high abundance within the continuously regenerating periodontal ligament of mouse incisor teeth and at such location they contribute to periodontal formation, periodontal homeostasis, and periodontal regeneration. This study, for the first time, reports the presence of PRX1 expressing cells within human periodontal ligament. Thus, studying mouse periodontal PRX1+ cells may provide significant information for the development of novel and more effective therapeutic approaches to periodontal regeneration in humans.

Periodontitis and peri-implantitis are chronic inflammatory diseases in which dysregulated host-microbial homeostasis compromises the oral mucosal barrier and leads to progressive soft-tissue destruction and bone loss. Although epithelial and stromal compartments are positioned to regulate barrier integrity and immune infiltration, how these non-immune cells differentially mediate inflammation in periodontitis versus peri-implantitis remains incompletely understood. The present study tested the hypothesis that distinct inflammatory profiles in periodontitis and peri-implantitis are associated with condition-specific changes in epithelial and stromal cellular states and their interactions with immune cells. Soft-tissue biopsies were collected from adults with periodontal health (n = 5 sites), periodontitis (n = 7 sites), and peri-implantitis (n = 8 sites). Single-nucleus RNA sequencing generated a cell-resolved atlas across epithelial, stromal, and immune compartments. Differential abundance, gene set enrichment, and pseudotime analyses assessed condition-specific shifts in cell proportions, biological pathways, and epithelial differentiation trajectories. Spatial transcriptomics and multiplexed immunofluorescence were used to validate candidate epithelial and endothelial niches and to visualize proximity-based interactions in situ. An in vitro endothelial assay tested cytokine inducibility of MHC class II under pro-inflammatory stimulation. The epithelial compartment resolved into seven differentiation states spanning a pseudotemporal continuum from basal repair-associated cells to suprabasal barrier-forming cells. Periodontitis was characterized by expansion of least differentiated, repair-skewed epithelial states and loss of suprabasal barrier-forming states. Peri-implantitis retained suprabasal differentiation programs but showed enrichment of stress keratins (KRT6A/B/C and KRT16). CXCL14 expression localized predominantly to suprabasal epithelial cells and showed the strongest spatial association with immune proximity in periodontitis. In the stromal compartment, a rare HLA-DR/MHC II-positive endothelial subset was enriched in disease and localized adjacent to T cells. This phenotype was recapitulated in cultured human endothelial cells under pro-inflammatory stimulation. These results support a model in which epithelial differentiation state and endothelial immune adaptation contribute in distinct ways to inflammatory niche organization in periodontitis and peri-implantitis. By integrating high-resolution single-cell and spatial data, this work delineates cellular niches and regulatory signatures that distinguish these diseases and identifies barrier-associated and vascular programs relevant to future host-modulatory strategies.

Introduction: One of the greatest differences between natural teeth and dental implants lies in the attachment apparatus. Collagen fiber bundles connect the tooth to the gingiva in the form of connective tissue attachment which inserts perpendicularly to the root surface. On the contrary, bundles of connective tissue fibers run parallel to the implant surface at the transmucosal level which make for a vulnerable seal. The goal of the present study is to develop a nano-engineered titanium surface with perpendicularly attached collagen that stimulate platelet activation for the restoration of periodontium-like connective tissue around dental implants. Materials & Methods: The titanium surface modification is executed in two stages. First, TiO2 nanotube array is fabricated via anodization. Diameters and depths of TiO2 nanotubes are controlled by applied voltage and duration. Subsequently, an electrophoretic fusion (EPF) method is applied to fuse type I collagen (Col-I) into nanotube arrays on the titanium surface. Surface morphology of the collagen-modified titanium surface was observed using scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR) and atomic force microscopy (AFM). Next, platelet-rich plasma (PRP) is prepared and samples (tests and control) were incubated with PRP for 30 minutes, one hour and 3 hours on a see-saw table at 37 °C. At the end of the incubation period, the samples were rinsed and fixed for observation under SEM while supernatants were subjected to enzyme-linked immunosorbent assay (ELISA). To verify the integration of fibroblast-produced Col-I to the nanoengineered surface, samples (tests and control) were co-cultured with fibroblasts using a cell culture insert with mesh size of 3 µm. At the end of the incubation period, samples were either dried in serial alcohol dehydration for SEM observation or stained with a FITC-conjugated anti-Col-I antibody for confocal fluorescence microscopy. In another culture system, fibroblasts between passaged 10 ~20 were seeded directly onto sample (test and control) surfaces. 50 mg/mL of L-Ascorbic acid was supplemented every other day for 8 days. At the end of the incubation period, samples were fixed with 4% PFA for 20 minutes and store in phosphate buffered solution (PBS). The samples were subjected to observation with SEM, multiphoton laser scanning microscopy and RAMAN spectroscopy. Results: SEM with EDX observation revealed that a uniform array of nanotubes measuring 67 nm in diameter was obtained by anodization at 30 volts for 3 hours. Electrophoresis of type-I collagen through native PAGE gel into Ti nanotube surface resulted in Col-I depositions in a perpendicular fashion. SEM images showed a uniform array of perpendicular collagen fibrils deposited into and around the nanotubes while the presence of an amide C=O absorption peak at 1550 cm-1 in the spectrum was confirmed with FTIR. The perpendicularly attached Col-I to the nanotubes demonstrated a significantly more robust binding (compared to other methods of Col-I deposition), resistant to high-power sonication. When platelet-rich plasma (PRP) was applied onto titanium, platelets aggregated on perpendicular collagen-fused nanotube surface while there were none or few for pure titanium surfaces at an early time point. Furthermore, confocal fluorescence microscopy revealed an increased amount of collagen on the modified Ti surfaces in a co-culture model with fibroblasts, confirming that fibroblast-derived Col-I fused with the engineered Col-I.
Polarized Raman spectroscopy revealed perpendicularly oriented collagen fibers extruding out from the nano-engineered collagen on nanotube-titanium surface. Conclusions: In this study, we hypothesized that our modified TiO2 surface with perpendicular collagen-fused nanotube array would facilitate biomimetic restoration of peri-implant soft and hard tissue. The key innovation is the orientation of the fused Col-I. The nanotubes support the perpendicular insertion of Col-I monomers and these monomer projections, in terms, serve as the priming site for activation of in vivo healing process. In this report, we have shown that the nano-engineered surface with perpendicular collagen coating promoted fibroblasts attachment and induced platelet activation. An in vitro culture model revealed that fibroblasts secreted collagen fibrils attached to the engineered surface in a perpendicular fashion. We will move toward in vitro 3D culture model as well as in vivo mice animal model to further investigate the establishment of a direct connective tissue attachment to the modified titanium surface.

Introduction and Objective: Actinomyces israelii is an early colonizer of the dental biofilm and a recognized causative agent of actinomycosis that has been implicated as a conditional pathogen in chronic periodontal inflammation. Clinical studies have demonstrated that A. israelii is more frequently isolated from diseased periodontal pockets than from healthy sites, yet its antimicrobial susceptibility profile and adaptive responses to commonly prescribed periodontal antibiotics remain poorly characterized. This study investigated the phenotypic, transcriptional, and biofilm-level responses of A. israelii to amoxicillin and metronidazole through a three-phase experimental design. Methods: In Phase 1, minimum inhibitory concentrations (MICs) of amoxicillin and metronidazole were determined for four Actinomyces species (A. gerencseriae, A. israelii, A. naeslundii, and A. oris) using the E-test method. In Phase 2, transcriptomic profiling was performed on planktonic A. israelii cultures exposed to amoxicillin (0.04 µg/mL) and metronidazole (13 µg/mL) for 2 hours, with differential gene expression analysis conducted using DESeq2. In Phase 3, a 40-species multispecies biofilm model grown on hydroxyapatite-coated pegs (MBEC Assay® system) was exposed to both antibiotics and assessed by scanning electron microscopy (SEM) and the triphenyltetrazolium chloride (TTC) metabolic assay at 3- and 7-day timepoints. Results: All Actinomyces species demonstrated marked susceptibility to amoxicillin (MICs ≤0.058 µg/mL) and intrinsic resistance to metronidazole (MICs ≥37 µg/mL), with A. israelii showing MIC .016 and >256 µg/mL, respectively. Transcriptomic analysis revealed antibiotic-specific gene expression programs: amoxicillin selectively upregulated resistance genes aph(3')-Ia (log₂FC = 4.70) and tet(C) (log₂FC = 2.81), while metronidazole predominantly upregulated virulence and stress response genes including uvrA, recA, and dnaK. Biofilm metabolic activity was transiently reduced at day 3 across treatment groups but recovered by day 7, with amoxicillin-treated biofilms exceeding control levels — suggesting compensatory metabolic responses within the multispecies community. SEM imaging confirmed that amoxicillin markedly disrupted biofilm architecture at day 7, whereas metronidazole had minimal structural impact consistent with the intrinsic resistance profile. Conclusions: A. israelii mounts distinct, antibiotic-specific adaptive programs in response to amoxicillin and metronidazole. Multispecies biofilm resilience is not predicted by planktonic susceptibility testing, highlighting the need for biofilm-based assessments to inform more targeted periodontal antimicrobial strategies.

Plasma technology is an emerging treatment with potential for implant surface decontamination and surface modification. Our in vitro study evaluates the effects of plasma treatment on fibroblast and osteoblast adhesion, proliferation, and differentiation on titanium surfaces. Plasma treatment was applied to machined and rough-surface titanium discs, and surface characterization was conducted using scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), and hydrophilicity tests. Cell adhesion and proliferation were assessed through SEM imaging, viable cell assays, and immunohistologic staining. Statistical comparisons between control (no plasma) and test (plasma) groups were performed using an independent two-sample t-test (α = 0.05). While SEM imaging showed no significant changes, XPS analysis revealed a substantial reduction (p .001) in carbon content, indicating decreased hydrocarbon buildup. Plasma treatment significantly increased surface hydrophilicity in both machined (p 0.0001) and rough (p 0.0001) surfaces, suggesting potential benefits for implant osseointegration. SEM and IHC imaging of fibroblast and osteoblasts showed a higher prevalence of cells in the test group. Cell viability assays indicated significantly greater live cell attachment in plasma-treated groups for both osteoblasts and fibroblasts at early time points, though the differences were not observed after 12 hours. In conclusion, plasma treatment of titanium surfaces, 1) reduce hydrocarbon buildup, 2) enhances surface hydrophilicity, and 3) improve early-stage fibroblast and osteoblast attachment to titanium surfaces. Further research is needed to explore whether these findings are clinically significant through clinical trials, and whether the technology can be used in the treatment of peri-implant disease.

Background The bidirectional relationship between Metabolic Syndrome (MetS) and periodontal disease is well established. While periodontal therapy has been shown to improve certain MetS parameters, no studies to date have examined the impact of MetS on long-term periodontal treatment outcomes. This study investigated the effect of MetS and its individual components (hypertension, obesity, dyslipidemia, diabetes), as well as clinical and behavioral factors, on periodontal outcomes over five years. Methods In this retrospective longitudinal cohort study, 25,433 patient charts from the Harvard School of Dental Medicine were screened, and 320 met inclusion criteria. Patients were categorized into: Perio-Only (periodontitis without systemic comorbidities), Non-MetS (periodontitis with one or two systemic conditions), and MetS (periodontitis with ≥3 MetS components). Outcomes were evaluated at 6–8 weeks, and at 1, 3, and 5 years post-baseline. The primary outcome was number of residual sites with probing depth ≥5 mm. Results: At 1 year, patients with MetS had 4.75 more residual sites compared to those without MetS (p = 0.006). Differences were not statistically significant at 3 or 5 years, potentially reflecting reduced sample size at later follow-up time points. Dyslipidemia was associated with 3.41 additional residual sites at 1 year (p = 0.043), and obesity (BMI > 25) with 6.40 additional residual sites at 3 years (p = 0.01). Adherence to supportive periodontal therapy was strongly associated with improved outcomes, with compliant patients exhibiting 4.38, 16.62, and 13.66 fewer residual sites at 1, 3, and 5 years, respectively. Greater baseline disease severity was consistently associated with more residual sites over time. Conclusions: MetS and specific systemic components (dyslipidemia and obesity) negatively influence short- and mid-term periodontal treatment outcomes. Adherence to supportive care and baseline disease severity were also strong determinants of long-term results. These findings offer new clinically relevant insights into the oral–systemic connection and lay the groundwork for future prospective studies in this area.

Background and Object: Patient’s demands have progressively increased over the last decade that esthetic concerns related to anterior dental implants are not uncommon. Several techniques have been attempted for the treatment of peri-implant soft tissue dehiscences/deficiencies (PSTDs), mostly employing the use of autogenous connective tissue graft (CTG). The objective of this study was to assess the effectiveness of combining recombinant human platelet-derived growth factor (rhPDGF)-BB with a xenogeneic “volume-stable” collagen matrix collagen matrix (VCMX) compared to the gold standard CTG for the treatment of peri-implant soft tissue dehiscences PSTDs at anterior implant sites. Methods: Twenty-eight subjects with isolated PSTDs were enrolled and randomized to receive a prosthetic-surgical approach involving either VCMX + rhPDGF-BB or CTG. The treatment protocol involved a presurgical prosthetic phase, with the removal of the pre-existing crown and the fabrication of a temporary crown. After 1 month, the surgical procedure for the correction of the PSTD was performed via soft tissue augmentation (VCMX + rhPDGF-BB or CTG, based on the randomization). After a provisional phase with the creation of an adequate emergence profile, a new definitive crown was made. The primary outcomes of interest in this study were the mean PSTDs coverage between the two treatment groups at 1 year. Additional outcomes of interest included: i) volumetric changes; ii) assessment of modifications in the peri-implant soft tissue phenotype; iii) ultrasonographic tissue perfusion-related outcomes; iv) ultrasonographic tissue elasticity-related outcomes: v) assessment of patient-reported outcomes during the first post-operative month; vi) evaluation of patient-reported outcomes at the last study visit and their changes compared to baseline. Optical scanning and the superimposition of the digital models were employed to evaluate volumetric changes at the treated areas. High-frequency ultrasonography was executed at different time points over 12 months to assessed changes in the peri-implant soft tissue phenotype, as well as tissue perfusion, and strain elastography. Multilevel linear mixed models were used to assess the study outcomes, accounting for repeated measures with random effects for time and subject, and fixed effects for potential confounders. Results: The preliminary findings from the randomized clinical trial depicted a mean follow-up period of 7.6 months. No statistically significant differences were observed for the primary outcome (mean PSTD of 94% for CTG, and 87% for VCMX + rhPDGF-BB, p>0.005). CTG-treated sites showed significantly higher KMW changes (2.9 mm vs 0.6 mm, p.01), MT gain (1.6 mm vs 0.9 mm, p=0.02) and Vol gain (114 mm3 vs 81 mm3, p=0.04) compared to VCMX + rhPDGF. On the other hand, the patients allocated to CTG reported significantly greater post-operative morbidity (20.9 VAS vs 5.5 VAS, p.01) and time for recovery (8.3 days vs 4.1 days, p.01) than the subjects that received VCMX + rhPDGF-BB. No significant differences were found between the groups for patient-reported treatment satisfactions and esthetic assessment (p.05). Conclusion: Treatment of PSTD with CTG or VCMX + rhPDGF-BB resulted in similar mean PSTD coverage. CTG resulted in a significantly greater KMW, MT, and Vol gains, while it also caused significantly higher post-operative pain and time to recover than VCMX + rhPDGF