Roles of SP/KLF transcription factors in odontoblast differentiation: From development to diseases.

OBJECTIVES: A zinc-finger transcription factor family comprising specificity proteins (SPs) and Krüppel-like factor proteins (KLFs) plays an important role in dentin development and regeneration. However, a systematic regulatory network involving SPs/KLFs in odontoblast differentiation has not yet been described. This review examined the expression patterns of SP/KLF gene family members and their current known functions and mechanisms in odontoblast differentiation, and discussed prospective research directions for further exploration of mechanisms involving the SP/KLF gene family in dentin development. MATERIALS AND METHODS: Relevant literature on SP/KLF gene family members and dentin development was acquired from PubMed and Web of Science. RESULTS: We discuss the expression patterns, functions, and related mechanisms of eight members of the SP/KLF gene family in dentin development and genetic disorders with dental problems. We also summarize current knowledge about their complementary or synergistic actions. Finally, we propose future research directions for investigating the mechanisms of dentin development. CONCLUSIONS: The SP/KLF gene family plays a vital role in tooth development. Studying the complex complementary or synergistic interactions between SPs/KLFs is helpful for understanding the process of odontoblast differentiation. Applications of single-cell and spatial multi-omics may provide a more complete investigation of the mechanism involved in dentin development.

Effect of photobiomodulation therapy on TGF-ß release from dentin, migration and viability of dental pulp stem cells in regenerative endodontics treatment: An ex vivo study.

BACKGROUND AND AIM: Regenerative endodontic procedures (REPs) are oriented by the principles of tissue engineering, incorporating dental pulp stem cells (DPSC), crucial growth factors like Transforming growth factor-ß (TGF-ß1), and scaffolds to facilitate the regeneration of dental pulp tissues. The present study aimed to investigate the effect of photobiomodulation (PBM) therapy, using an 808 nm diode laser on cellular modulation mechanisms in REPs. METHOD AND MATERIAL: A total of 108 human dentin discs obtained from intact single root teeth were randomly assigned into six groups (n = 8): 1. Positive control (EDTA), 2. PBM-1 (3 J/cm2), 3. PBM-2 (5 J/cm2), 4. EDTA+PBM-1, 5. EDTA+PBM-2, and 6. Negative control (NaOCl). Then, an extract solution was prepared from each disc and the concentration of released TGF-ß1 from the discs was measured using enzyme-linked immunosorbent assay (ELISA). Moreover, the extract solution was added to DPSC culture medium to evaluate cell viability and migration through MTT assay and scratch test, respectively. RESULT: The group exposed to PBM-1 showed the highest cell viability, while treatment with EDTA and EDTA+PBM-2 decreased cellular viability. Also, the PBM-treated groups showed significantly higher release of TGF-ß1 compared to the negative control. EDTA and EDTA+PBM-1 showed the highest release among all the groups. No significant difference was found between EDTA and EDTA+PBM-1, as well as between PBM-1 and PBM-2. Moreover, the PBM-1 group exhibited the highest migration after 24 h, which was significantly greater than other groups, except for the PBM-2 group. CONCLUSION: According to the obtained data, 808 nm mediated-PBM (3 J/cm2), both independently and in conjunction with EDTA, enhanced the release of TGF-ß1 from dentin and improved cell viability and migration of DPSCs. It seems that, PBM under the specific parameters employed in this study, could be an effective adjunctive therapy in REPs.

Sequential Application of Calcium Phosphate and ε-Polylysine Show Antibacterial and Dentin Tubule Occluding Effects In Vitro.

In this study, ε-polylysine and calcium phosphate precipitation (CPP) methods were employed to induce antibacterial effects and dentin tubule occlusion. Antibacterial effects of ε-polylysine were evaluated with broth dilution assay against P. gingivalis. CPP solution from MCPM, DCPD, and TTCP was prepared. Four concentrations of ε-polylysine(ε-PL) solutions (0.125%, 0.25%, 0.5%, 1%) were prepared. Dentin discs were prepared from recently extracted human third molars. Dentin discs were incubated with P. gingivalis (ATCC 33277) bacterial suspension (ca. 105 bacteria) containing Brain Heart Infusion medium supplemented with 0.1 g/mL Vitamin K, 0.5 mg/mL hemin, 0.4 g/mL L-cysteine in anaerobic jars (37 °C) for 7 days to allow for biofilm formation. P. g-infected dentin specimens were randomly divided into four groups: CPP + 0.125% ε-PL, CPP + 0.25% ε-PL, CPP + 0.5% ε-PL, CPP + 1% ε-PL. On each dentin specimen, CPP solution was applied followed by polylysine solution with microbrush and immersed in artificial saliva. Precipitate formation, antibacterial effects, and occlusion of dentinal tubules were characterized in vitro over up to 72 h using scanning electron microscopy. ε-PL showed 34.97% to 61.19% growth inhibition levels against P. gingivalis (P. g) after 24 h of incubation. On P. g-infected dentin specimens, DCPD + 0.25% ε-PL, and DCPD + 0.5% ε-PL groups showed complete bacterial inhibition and 78.6% and 98.1% dentin tubule occlusion, respectively (p < 0.001). The longitudinal analysis on fractured dentin samples in DCPD and TTCP groups revealed deeply penetrated hydroxyapatite-like crystal formations in dentinal tubules after 72 h of incubation in artificial saliva.

Influence of Surface Conditioning on the Repair Strength of Bioactive Restorative Material.

OBJECTIVE: To evaluate the effects of surface treatment and repair material on the repair shear bond strength (SBS) of the bioactive restorative material. METHODS: A total of 240 Activa BioActive Restorative (Activa) discs were prepared, aged, and polished, and divided randomly into eight groups (n = 30). Groups 1-4 discs were repaired with bulk-fill flowable resin-based composite (Bulk-RBC), and Groups 5-8 discs with Activa. Surface treatment used for each repair material type were air abrasion with silica-coated 30-m Al2O3 particles (air abrasion) (Groups 2 & 6), Air abrasion with universal primer (Groups 3 & 7), and Air abrasion with universal adhesive (Groups 4 & 8). Groups 1 and 5 were controls without surface treatment. SBS test was performed, and the failure mode and surface topography were assessed. RESULTS: Surface treatment with air abrasion significantly improved the SBS for repair using both Activa and Bulk-RBC. Repair SBS using Activa was significantly higher compared with Bulk-RBC. Cohesive failure in substrate and mixed failures were most common in the surface-treated groups (2-4, 6-8). Air abrasion produced prominent surface topography changes compared with polishing. CONCLUSION: Air abrasion enhances the repair SBS of aged bioactive restorative material. The use of the same material (Activa) for repair affords a higher bond strength compared with the use bulk-RBC.

Combined effects of calcium hydroxide and photobiomodulation therapy on apexogenesis of immature permanent teeth in dogs.

Treatment of high-risk traumatic immature teeth due to incomplete root development is challenging. Apexogenesis is currently the ideal treatment option that allows normal root development. The purpose of this study was to evaluate the apexogenesis process of immature permanent teeth of dogs when co-administered with calcium hydroxide and photobiomodulation therapy (PBMT). A total of 36 immature permanent anterior and premolar teeth were selected from three 4-6-month-old dogs of Iranian mixed generation. The teeth were categorized into two groups, calcium hydroxide with laser irradiation (CHL) and calcium hydroxide without laser irradiation (CH). All the selected teeth received calcium hydroxide pulpotomy. After restoring teeth with amalgam, the CHL group received gallium­aluminum-arsenide (GaAlAs) diode laser (810 nm, 4.2 J/cm2, 0.3 W, 9 s,CW) on apical one-third of both buccal and lingual roots. The irradiation was repeated every 48 h for fourteen days. Intravenous tetracycline was used to observe newly formed dentin in the first, third, seventh, and fourteenth days. The distance between tetracycline lines (DTL) was examined by Fluorescence microscopy. Generalized estimating equations (GEE) were used for data analysis. In all assessments, the mean DTL were greater in the CHL group. However, the two groups had no significant differences in the amount of deposited dentin between the first and third, third and seventh, and first and seventh lines. Meanwhile, there was a significant difference between the two groups in terms of the distances between lines 7 and 14, 1 and 14 and also 3 to 14 (P < .001). In other words, from the 7th day onwards, there was a significant difference between the two groups. Within the limitation of this study, the combination therapy of PBMT and pulpotomy with calcium hydroxide accelerated apexogenesis in immature permanent dogs' teeth.

Antifungal and Synergistic Effects of the Ethyl Acetate Extract of Tanacetum vulgare (L.) Against Candida albicans.

BACKGROUND With the continued demand for new, effective, and safe endodontic therapies, the aim of this study was assessment of efficiency of the ethyl acetate (EthOAc) extract of Tanacetum vulgare (L.) against Candida albicans. MATERIAL AND METHODS The antifungal effectiveness of the EthOAc extract of T. vulgare was determined using the agar disk diffusion method. The inhibition zones induced by the EthOAc extract were compared after 5 minutes, 60 minutes, and 24 hours to those induced by standard solutions (2% chlorhexidine, saturated calcium hydroxide, and 2% sodium hypochlorite). Statistical analysis of the results was performed using the Kruskal-Wallis test and one-way ANOVA. RESULTS The inhibition zone of chlorhexidine against C. albicans was 30.3-19.3 mm, but in combination with EthOAc extract (100 mg/mL) of T. vulgare, this inhibition was from 32.7-30 mm, indicating that this combination exerted a marked synergistic effect against C. albicans. The inhibition zone of sodium hypochlorite (69.7-65 mm) was higher than the inhibition zone of EthOAc extract and chlorhexidine. The combination of EthOAc extract with sodium hypochlorite resulted in a loss of antifungal activity. Furthermore, the activity of the EthOAc extract against C. albicans was decreased after mixing the extract with dentine at a concentration of 25 mg/50 µL (30.3-20.7 mm). The EthOAc extract did not show a genotoxic effect on lymphocyte cells. CONCLUSIONS The EthOAc extract of T. vulgare may be a useful tool to discover natural bioactive agents that have antifungal activity against C. albicans and could be used as endodontic therapies.

Efficacy of phototherapy in the adhesive bonding of different dental posts to root dentin: A systematic review.

BACKGROUND: The aim of the current systematic review was to evaluate the efficacy of phototherapy in the adhesive bonding of different dental posts to root dentin. METHODS: The outline of this systematic review followed the PRISMA (Preferred Reporting Items for Systematic Review and Meta-Analysis) guidelines. Searches in both electronic and manual literature were performed in the main databases 'MEDLINE' and 'EMBASE' up to March 2019 using the following terms: (Phototherapy) AND (push out test) AND (bond strength) AND (post) OR (fibre) OR (metal) AND (smear layer). RESULTS: Six in-vitro studies were included and processed for data extraction. All studies incorporated the use of fiber posts. The mean shear bond strength for test group ranged from 2.23 to 15.17 MPa while mean shear bond strength for control group ranged from 2.93 to 9.38 MPa. The wavelengths of diode lasers ranged from 660 to 2940 nm (nm). Power was reported in 5 studies ranging from 0.075 W to 3 W. All studies compared shear bond strength of phototherapy in the adhesive bonding of different dental posts to root dentin. In all the included studies, greater bond strength achieved by phototherapy compared to nonirradiated group. CONCLUSION: Within the limits, this study shows that the dentin to post bond strength was significantly enhanced by phototherapy.

Influence of Er: Cr: YSGG laser on adhesive strength and microleakage of dentin bonded to resin composite. In-vitro study.

BACKGROUND: The aim of this study was to evaluate the surface treatment of dentin with phototherapy (ER-CR-YSGG laser) in the presence of different bonding systems on their shear bond strength and microleakage. MATERIALS AND METHODS: Eighty intact human third molars were divided into two groups. Forty teeth were treated with conventional flat wheel diamond bur and the remaining forty were treated with Er Cr YSGG laser (phototherapy). The two groups i.e. treated with laser (l) and group treated with abrasive bur (NL) were further divided into two subgroups of twenty samples each. Subgroup 1 was surface treated with etch and rinse (ER) and other 20 remaining samples were treated with self- etch (SE) bonding regime. Specimens from all the groups were assessed for shear bond strength and microleakage scores. Ten samples from all groups were immersed in 2% methylene blue for 24 h and assessed under a digital microscope for microleakage. Data was assessed using analysis of variance and tukey multiple comparisons test. RESULTS: The lowest bond strength was achieved in laser prepared phototherapy group bonded with self-etch (LSE-11.87 ± 1.21). The maximum bond strength score was observed in non-laser-etch and rinse group (NLER-23.66 ± 2.56). The highest mean microleakage was observed among laser bonded with self-etch (LSE) specimens (1.0 ± 0.13). The lowest microleakage scores were seen in group treated with non-phototherapy etch and rinse (NLER-0.4 ± 0.11) regime. CONCLUSION: Use of etch and rinse dentin bonding regime in combination with ER-CR-YSGG phototherapy dentin treatment has the potential for clinical application in comparison to conventional conditioning technique.

Cell Homing for Pulp Tissue Engineering with Endogenous Dentin Matrix Proteins.

INTRODUCTION: Compelling evidence pinpoints that pulp tissue engineering after the transplantation of stem cells is possible. Although intriguing, severe problems regarding clinical feasibility remain. Cell homing has been proposed as a viable alternative in which dentin-derived growth factors in a conducive scaffold may attract resident cells to form pulplike tissue. In this study, an ectopic animal model for in situ dental pulp tissue engineering was developed to evaluate whether pulplike tissue formation in empty root canals after the attraction of stem cells was possible and whether this could be enhanced by dentin-derived growth factors. METHODS: Three types of fibrin (custom-made fibrin, fibrin sealant, and plasma rich in growth factors [PRGF]) as well as a self-assembling peptide were evaluated in vivo in a modified tooth root model using human teeth. Root canal dentin was conditioned with EDTA, tooth roots were filled with growth factor-laden scaffolds, and dental pulp stem cells in collagen were placed at the root tip. Constructs were implanted into immunocompromised mice for 4 weeks and subsequently analyzed histologically. Differential interference contrast and second harmonic generation imaging were performed for selected sections. RESULTS: For custom-made fibrin and fibrin sealant with dentin matrix proteins, migration into the roots and the formation of a pulplike tissue were observed, whereas the peptide-based scaffold appeared less suitable. PRGF supported tissue formation regardless of the addition of dentin matrix proteins. In the test groups with dentin matrix proteins and EDTA conditioning, differentiated odontoblastlike cells extended cellular processes into the dentinal tubules, which coincided with the deposition of the newly formed collagenous dentin matrix. CONCLUSIONS: This new cell homing model provides evidence that fibrin derivatives make applicable scaffolds and that dentin-derived proteins induce chemotaxis and pulplike tissue formation.

Improved reactive nanoparticles to treat dentin hypersensitivity.

The aim of this study was to evaluate the effectiveness of different nanoparticles-based solutions for dentin permeability reduction and to determine the viscoelastic performance of cervical dentin after their application. Four experimental nanoparticle solutions based on zinc, calcium or doxycycline-loaded polymeric nanoparticles (NPs) were applied on citric acid etched dentin, to facilitate the occlusion and the reduction of the fluid flow at the dentinal tubules. After 24 h and 7 d of storage, cervical dentin was evaluated for fluid filtration. Field emission scanning electron microscopy, energy dispersive analysis, AFM and Nano-DMA analysis were also performed. Complex, storage, loss modulus and tan delta (δ) were assessed. Doxycycline-loaded NPs impaired tubule occlusion and fluid flow reduction trough dentin. Tubules were 100% occluded in dentin treated with calcium-loaded NPs or zinc-loaded NPs, analyzed at 7 d. Dentin treated with both zinc-NPs and calcium-NPs attained the highest reduction of dentinal fluid flow. Moreover, when treating dentin with zinc-NPs, complex modulus values attained at intertubular and peritubular dentin were higher than those obtained after applying calcium-NPs. Zinc-NPs are then supposed to fasten active dentin remodeling, with increased maturity and high mechanical properties. Zinc-based nanoparticles are then proposed for effective dentin remineralization and tubular occlusion. Further research to finally prove for clinical benefits in patients with dentin hypersensitivity using Zn-doped nanoparticles is encouraged. STATEMENT OF SIGNIFICANCE: Erosion from acids provokes dentin hypersensitivity (DH) which presents with intense pain of short duration. Open dentinal tubules and demineralization favor DH. Nanogels based on Ca-nanoparticles and Zn-nanoparticles produced an efficient reduction of fluid flow. Dentinal tubules were filled by precipitation of induced calcium-phosphate deposits. When treating dentin with Zn-nanoparticles, complex modulus values attained at intertubular and peritubular dentin were higher than those obtained after applying Ca-nanoparticles. Zn-nanoparticles are then supposed to fasten active dentin remodeling, with increased maturity and high mechanical properties. Zinc-based nanogels are, therefore, proposed for effective dentin remineralization and tubular occlusion. Further research to finally prove for clinical benefits in patients with dentin hypersensitivity using Zn-doped nanogels is encouraged.

Dentine matrix proteins: isolation and effects on human pulp cells.

AIM: To establish a simplified and efficient protocol for the isolation and concentration of matrix proteins from human dentine, and to assess the effects of extracted dentine matrix proteins (eDMP) on the behaviour of human pulp cells. METHODOLOGY: Matrix proteins were isolated from human dentine, purified, concentrated and characterized with protein and enzyme-linked immunosorbent assays (ELISA). Culture media were supplemented with eDMP in different concentrations, referred to as eDMP 1-10 000, to assess viability and proliferation of human pulp cells by DNA and MTT assays; apoptotic events were quantified by flow cytometry. Chemotactic effects of eDMP were assessed in a modified Boyden chamber assay. Expression levels of odontoblastic marker genes in pulp cells cultured with eDMPs were determined by real-time quantitative PCR, and the ability to induce mineralization was demonstrated by alizarin red staining. Nonparametric statistical analysis was performed to pairwise compare different groups at all time-points (Mann-Whitney U-test, α = 0.05). RESULTS: High concentrations of eDMP exhibited significant antiproliferative effects (P ≤ 0.023) after 5 (eDMP 1000) and 7 days (eDMP 500) without affecting cell viability. Apoptosis was barely influenced (P ≥ 0.089). eDMP exerted a concentration-dependent chemotactic stimulus on dental pulp cells with statistical significance already at low dosage (P = 0.006 at eDMP 10). Changes in gene expression indicated a differentiation into odontoblast-like cells, which was corroborated by findings of mineral nodule formation. CONCLUSIONS: A novel, effective and time-saving protocol for isolation and concentration of dentine matrix proteins is presented. As eDMP stimulates chemotaxis, differentiation and mineralization without affecting viability, endogenous dentine matrix proteins might be valuable for approaches to regenerate or engineer dental pulp.

Microtensile Bond Strength of Resin-Modified Glass Ionomer Cement to Sound and Artificial Caries-Affected Root Dentin With Different Conditioning.

In this laboratory study, the microtensile bond strengths (µTBS) of resin-modified glass ionomer cement (RM-GIC) to sound and artificial caries-affected bovine root dentin (ACAD) using three different conditioning agents were evaluated after 24 hours and three months. The fractured interface was examined with a scanning electron microscope (SEM). Specimens were created on bovine root dentin that was embedded in epoxy resin. For the ACAD specimens, artificial carious lesions were created. The RM-GIC (Fuji II LC) was applied either directly (no treatment), after application of self conditioner, cavity conditioner, or 17% ethylenediamine tetraacetic acid (EDTA) applied for 60 seconds, on sound dentin and ACAD, then light cured. They were stored in artificial saliva for 24 hours or three months. Following this, the specimens were cut into sticks for the µTBS test, and the failure mode of the debonded specimens was examined by using SEM. Pretest failures were excluded from the statistical analysis of the µTBS values because of their high incidence in some groups. Results showed that the µTBS values were significantly affected by the dentin substrate as well as the conditioning agent. Self conditioner provided the highest and most stable µTBS values, while cavity conditioner showed stable µTBS values on sound dentin. Both self conditioner and cavity conditioner had significantly higher µTBS values than the no treatment groups. EDTA conditioning reduced the µTBS after three months to sound dentin, while it showed 100% pretest failure with ACAD for both storage periods.

Analysing the bioactive makeup of demineralised dentine matrix on bone marrow mesenchymal stem cells for enhanced bone repair.

Dentine matrix has proposed roles for directing mineralised tissue repair in dentine and bone; however, the range of bioactive components in dentine and specific biological effects on bone-derived mesenchymal stem cells (MSCs) in humans are less well understood. The aims of this study were to further elucidate the biological response of MSCs to demineralised dentine matrix (DDM) in enhancing wound repair responses and ascertain key contributing components. Dentine was obtained from human teeth and DDM proteins solubilised with ethylenediaminetetraacetic acid (EDTA). Bone marrow derived MSCs were commercially obtained. Cells with a more immature phenotype were then selected by preferential fibronectin adhesion (FN-BMMSCs) for use in subsequent in vitro assays. DDM at 10 µg/mL reduced cell expansion, attenuated apoptosis and was the minimal concentration capable of inducing osteoblastic differentiation. Enzyme-linked immunosorbent assay (ELISA) quantification of growth factors indicated physiological levels produced the above responses; transforming growth factor ß (TGF-ß1) was predominant (15.6 ng/mg DDM), with relatively lower concentrations of BMP-2, FGF, VEGF and PDGF (6.2-4.7 ng/mg DDM). Fractionation of growth factors from other DDM components by heparin affinity chromatography diminished osteogenic responses. Depletion of biglycan from DDM also attenuated osteogenic potency, which was partially rescued by the isolated biglycan. Decorin depletion from DDM had no influence on osteogenic potency. Collectively, these results demonstrate the potential of DDM for the delivery of physiological levels of growth factors for bone repair processes, and substantiate a role for biglycan as an additional adjuvant for driving osteogenic pathways.

Ultrasonic activation of irrigants increases growth factor release from human dentine.

OBJECTIVES: Bioactive proteins are sequestered in human dentine and play a decisive role in dental pulp regeneration and repair. They can be released and exposed on the dentine surface by acids, but also chelators, such as ethylenediaminetetraacetic acid (EDTA). The objectives of this study were (i) to evaluate whether ultrasonic activation of irrigants in the root canal will promote growth factor release from dentine and (ii) to collect bioactive proteins in a physiological solution. MATERIALS AND METHODS: Human dentine disks underwent irrigation with and without ultrasonic activation. The protocols included treatment by either a single or two consecutive steps with 10 % EDTA and phosphate-buffered saline (PBS), where each sample was treated three times. To mimic clinical conditions, selected irrigation regimens were applied to root canals of extracted human teeth after preparation. Amounts of transforming growth factor ß1 (TGF-ß1) in solution were quantified using enzyme-linked immunosorbent assays. Nonparametric statistical analysis was performed to compare different groups as well as repetitions within a group (Mann-Whitney U test, α = 0.05). Additionally, morphological changes of dentine surfaces were visualized by scanning electron microscopy (SEM). RESULTS: TGF-ß1 was not detectable after irrigation of dentine with PBS, neither with nor without ultrasonic activation. Irrigation with EDTA released TGF-ß1, and ultrasonic activation of EDTA enhanced this effect. However, preceding EDTA conditioning enabled the release of bioactive proteins into PBS solution. Similar results were observed in dentine disks and root canals. Visualization of dentine surfaces after different treatment revealed superficial erosion after ultrasonic activation irrespective of the irrigant solution, but different degrees of exposure of organic substance. CONCLUSIONS: Ultrasonic activation enhances growth factor release from human dentine. Bioactive proteins can be isolated in physiological solvents and may act as autologous supplements for regenerative endodontic treatment or pulp tissue engineering. CLINICAL RELEVANCE: Autologous growth factors from human dentine can advance treatment strategies in dental pulp tissue engineering.

The Great Irish Famine: Identifying Starvation in the Tissues of Victims Using Stable Isotope Analysis of Bone and Incremental Dentine Collagen.

The major components of human diet both past and present may be estimated by measuring the carbon and nitrogen isotope ratios (δ13C and δ15N) of the collagenous proteins in bone and tooth dentine. However, the results from these two tissues differ substantially: bone collagen records a multi-year average whilst primary dentine records and retains time-bound isotope ratios deriving from the period of tooth development. Recent studies harnessing a sub-annual temporal sampling resolution have shed new light on the individual dietary histories of our ancestors by identifying unexpected radical short-term dietary changes, the duration of breastfeeding and migration where dietary change occurs, and by raising questions regarding factors other than diet that may impact on δ13C and δ15N values. Here we show that the dentine δ13C and δ15N profiles of workhouse inmates dating from the Great Irish Famine of the 19th century not only record the expected dietary change from C3 potatoes to C4 maize, but when used together they also document prolonged nutritional and other physiological stress resulting from insufficient sustenance. In the adults, the influence of the maize-based diet is seen in the δ13C difference between dentine (formed in childhood) and rib (representing an average from the last few years of life). The demonstrated effects of stress on the δ13C and δ15N values will have an impact on the interpretations of diet in past populations even in slow-turnover tissues such as compact bone. This technique also has applicability in the investigation of modern children subject to nutritional distress where hair and nails are unavailable or do not record an adequate period of time.

Comparison of the ability of chondroitin sulfate derived from bovine, fish and pigs to suppress human osteoclast activity in vitro.

Chondroitin sulfate (CS) compounds are commonly used to manage OA symptoms. Recent literature has indicated that abnormal subchondral bone metabolism may have a role in the pathogenesis of OA. The aim of this study was to access the effects of chondroitin sulfate obtained from bovine, fish and porcine sources on human osteoclast formation and activity in vitro. Human osteoclasts were generated from blood mononuclear cells. Cells were cultured over 17 days with the addition of macrophage colony stimulating factor (M-CSF) and then stimulated with receptor activator of nuclear factor kappa B ligand from day 7. Cells were treated with the CS commencing from day 7 onwards. To assess effects on osteoclasts, tartrate resistant acid phosphatate (TRAP) expression and resorption of whale dentine assays were used. Bovine-derived CS consistently suppressed osteoclast activity at concentrations as low as 1 µg/ml. Fish and porcine CS was less consistent in their effects varying with different donor cells. All CS compounds had little effect on TRAP activity. mRNA analysis using real-time PCR of bovine CS treated cells indicated that the inhibition of activity was not due to inhibition of the late stage NFATc1 transcription factor (p > 0.05). These results are consistent with CS inhibition of mature osteoclast activity rather than the formation of mature osteoclasts. It would appear that there are differences in activity of the different CS compounds with bovine-derived CS being the most consistently effective inhibitor of osteoclast resorption, but the results need to be confirmed.

The efficacy of gaseous ozone on some cariogenic bacteria.

The aim of this study is to analyze ozone impact on some cariogenic bacteria in ex vivo and in vitro conditions. The in vitro part of study inoculated dentine with strains of Streptococcus mutans ATCC 33402 and Lactobacillus paracasei ATCC 11974 bacteria. Samples of dentine before and after 40s ozone treatment were collected and anaerobically incubated. Samples of cariogenic dentine (N = 24) were collected from permanent molars within the ex vivo segment of the study, prior and after 40s ozone treatment and a number of colonies were counted after incubation. For the in vitro part of study, results have shown a statistically significant average value of reduction of Streptococcus mutans ATCC 33402 and Lactobacillus paracasei ATCC 11974 prior and after ozone treatment (p < 0.001). The ex vivo segment of the study has also demonstrated a statistically significant difference in the number of bacteria prior and after ozone implementation (p < 0.001). Gaseous ozone demonstrated a strong antimicrobial effect on cariogenic bacteria in both in vitro and ex vivo conditions and it can be used as an adjuvant in caries therapy.

Short-term PTH administration increases dentine apposition and microhardness in mice.

OBJECTIVE: The purpose of this study is to investigate the effects of intermittent parathyroid hormone (PTH) administration on the apposition rate and structural features of dentine from mouse incisors. METHODS: Young male A/J Unib mice were treated daily for 6 and 10 days with 40 µg/kg of hPTH 1-34 or a vehicle. Dentine apposition rates measured by fluorescent labels (tetracycline and calcein) and alkaline phosphatase (ALP) plasma levels were evaluated after 6 days of treatment. Knoop microhardness testing and element content measurements in at.% of calcium (Ca), phosphorus (P), oxygen (O), and magnesium (Mg) in the peritubular and intertubular dentine were performed by Energy Dispersive X-ray (EDX) microanalysis via Scanning Electron Microscopy (SEM) after 10 days of treatment. RESULTS: Histometric analysis revealed an increase of 5% in the apposition rate of dentine and 25% in the ALP plasma levels in the PTH treated group. In addition, knoop microhardness testing revealed that the animals treated with PTH had a greater microhardness (11%). EDX microanalysis showed that PTH treatment led to increases in P (23%) and Ca (53%) at.% content, as well as the Ca/P ratio (24%) in peritubular dentine. The chemical composition of intertubular dentine did not vary between the groups. CONCLUSIONS: These findings indicate that intermittent administration of hPTH (1-34) increases apposition and mineralization of the dentine during young mice incisor formation.

An in vitro model for the study of chemical exchange between glass ionomer restorations and partially demineralized dentin using a minimally invasive restorative technique.

OBJECTIVES: This study was designed to validate an in vitro model of the Atraumatic Restorative Technique (ART). This model allowed in depth analyses of the migration of apatite forming Sr and F, from a glass-ionomer, Fuji IXGP (GC Corporation, Tokyo, Japan), into partly demineralized dentine. The second objective was to study the effects of the levels of mineral loss on such ionic exchange. METHODS: Artificial lesions were created in thirty third molars, which were divided into three groups based on time of exposure to the demineralizing solution (7, 14 and 21 days). These were restored with Fuji IXGP and left for 21 days. The method of analysis was electron probe microanalysis (EPMA) with wavelength dispersive spectrometry (WDS). This combination can yield both qualitative identification of elements as well as quantitative compositional information, with a minimum detection limit of 0.01%. RESULTS: The results indicated that there was no significant difference in the levels of Ca and P from the control and test sides for all three groups of demineralized dentine. The differences in the depth of penetration and the amount of Sr and F between control and test were significant. CONCLUSIONS: The validity of the in vitro model was confirmed. There was migration of both strontium and fluorine to the artificially demineralized dentine in a pattern consistent with remineralization and similar to what was observed in a previous in vivo study.

Long-term stability of dentin matrix following treatment with various natural collagen cross-linkers.

OBJECTIVES: Collagen disorganization is one of the main degradation patterns found in unsuccessful adhesive restorations. The hypothesis of this study was that pretreatment using natural collagen cross-linking agents rich in proanthocyanidin (PA) would improve mechanical properties and stability over time of the dentin collagen and, thus, confer a more resistant and lasting substrate for adhesive restorations. METHODS: PA-based extracts, from grape seed (GSE), cocoa seed (CSE), cranberry (CRE), cinnamon (CNE) and açaí berry (ACE) were applied over the demineralized dentin. The apparent elastic modulus (E) of the treated dentin collagen was analyzed over a 12 month period. Specimens were immersed in the respective solution and E values were obtained by a micro-flexural test at baseline, 10, 30, 60, 120 and 240 min. Samples were stored in artificial saliva and re-tested after 3, 6 and 12 months. Data was analyzed using ANOVA and Tukey test. RESULTS: GSE and CSE extracts showed a time-dependent effect and were able to improve [240 min (MPa): GSE = 108.96 ± 56.08;CSE = 59.21 ± 24.87] and stabilize the E of the organic matrix [12 months (MPa): GSE = 40.91 ± 19.69;CSE = 42.11 ± 13.46]. CRE and CNE extracts were able to maintain the E of collagen matrices constant over 12 months [CRE = 11.17 ± 7.22;CNE = 9,96 ± 6.11;MPa]. ACE (2.64 ± 1.22 MPa) and control groups immersed in neat distilled water (1.37 ± 0.69 MPa) and ethanol-water (0.95 ± 0.33 MPa) showed no effect over dentin organic matrix and enable their degradation and reduction of mechanical properties. SIGNIFICANCE: Some PA-based extracts were capable of improving and stabilizing collagen matrices through exogenous cross-links induction.