NP-12 peptide functionalized nanoparticles counteract the effect of bacterial lipopolysaccharide on cultured osteoblasts.

OBJECTIVE: To evaluate whether nanoparticles (NPs) functionalized with Tideglusib (TDg, NP-12), and deposited on titanium surfaces, would counteract the effect of bacterial lipopolysaccharide (LPS) on osteoblasts. METHODS: Experimental groups were: (a) Titanium discs (TiD), (b) TiD covered with undoped NPs (Un-NPs) and (c) TiD covered with TDg-doped NPs (TDg-NPs). Human primary osteoblasts were cultured onto these discs, in the presence or absence of bacterial LPS. Cell proliferation was assessed by MTT-assay and differentiation by measuring the alkaline phosphatase activity. Mineral nodule formation was assessed by the alizarin red test. Real-time quantitative polymerase chain reaction was used to study the expression of Runx-2, OSX, ALP, OSC, OPG, RANKL, Col-I, BMP-2, BMP-7, TGF-ß1, VEGF, TGF-ßR1, TGF-ßR2, and TGF-ßR3 genes. Osteoblasts morphology was studied by Scanning Electron Microscopy. One-way ANOVA or Kruskal-Wallis and Bonferroni multiple comparisons tests were carried out (p < 0.05). RESULTS: TDg-NPs enhanced osteoblasts proliferation. Similarly, this group increased ALP production and mineral nodules formation. TDg-NPs on titanium discs resulted in overexpression of the proliferative genes, OSC and OSX, regardless of LPS activity. In the absence of LPS, TDg-NPs up-regulated Runx2, COL-I, ALP, BMP2 and BMP7 genes. OPG/RANKL gene ratios were increased about 2500 and 4,000-fold by TDg-NPs, when LPS was added or not, respectively. In contact with the TDg-NPs osteoblasts demonstrated an elongated spindle-shaped morphology with extracellular matrix production. SIGNIFICANCE: TDg-NPs on titanium discs counteracted the detrimental effect of LPS by preventing the decrease on osteoblasts proliferation and mineralization, and produced an overexpression of proliferative and bone-promoting genes on human primary osteoblasts.

Tunable polymer-peptide hybrids for dentin tissue repair.

OBJECTIVES: This study targets to assess the remineralization capability of conditioned dentin infiltrated with polymeric nanoparticles (NPs) doped with tideglusib (TDg) (TDg-NPs). METHODS: Dentin conditioned surfaces were infiltrated with NPs and TDg-NPs. Bonded interfaces were created, stored for 24 h and submitted to mechanical and thermal challenging. Resin-dentin interfaces were evaluated through nanohardness, Masson's trichrome staining microscopy, and Raman analysis. RESULTS: Dentin surfaces treated with TDg-NPs and load cycled produced higher nanohardness than the rest of the groups at the hybrid layer. At the bottom of the hybrid layer, all samples treated with TDg-NPs showed higher nanohardness than the rest of the groups. Active remineralization underneath the hybrid layer was detected in all groups after TDg application and load cycling, inducting new dentinal tubuli formation. After thermocycling, remineralization at the hybrid layer was not evidenced in the absence of NPs. Raman analysis showed increase mineralization, enriched carbonate apatite formation, and improved crosslinking and scaffolding of the collagen. CONCLUSIONS: Mechanical loading on the specimens obtained after TDg-NPs dentin infiltration inducts an increase of mineralization at the resin/dentin interface, indicating remineralization of peritubular and intertubular dentin with augmented crystallographic maturity in crystals. Enriched collagen quality was produced, generating an adequate matrix organization to promote apatite nucleation, after tideglusib infiltration. CLINICAL SIGNIFICANCE: At the present research, it has been proved the creation of reparative dentin, at the resin-dentin interface, after tideglusib dentin infiltration. Chemical stability, to favor integrity of the resin-dentin interface, is warranted in the presence of the TDg-NPs in the demineralized dentin collagen.

Effect of functionalized titanium particles with dexamethasone-loaded nanospheres on macrophage polarization and activity.

OBJECTIVE: The aim of this study was to determine the effect of titanium micro particles (TiP) previously functionalized with nanoparticles doped with dexamethasone (Dex) and doxycycline (Dox), on macrophage polarization and activity. METHODS: Macrophages RAW264.7 were cultured in the presence TiP loaded with dexamethasone -NPs (Dex)- and doxycycline -NPs (Dox)-, and as control, TiP with or without doped NPs. Cells were tested with and without previous bacterial lipopolysaccharide endotoxin (LPS) stimulation. Their morphology, proliferation, cytotoxicity, phenotypic change, and cytokines release were assessed by LIVE/DEAD, DNA release, metabolic activity, brightfield and scanning electron microscopy. The test Kruskall-Wallis was used for comparisons, while the cytokine expression profiles were examined by hierarchical clustering (p < 0.05). RESULTS: Upon exposure with TiP macrophages were activated and polarized to M1, but without depicting cytotoxic effects. The particles were phagocytised, and vacuolized. When exposed to functionalised TiP with NPs(Dex) and NPs(Dox), the ratio M1/M2 was up to forty times lower compared to TiP alone. When exposed to LPS, TiP reduced cell viability in half. Functionalised TiP with NPs(Dex) inhibited the cytokine release exerted by TiP on macrophages. When macrophages were exposed to functionalised TiPs with NPs(Dex) with and without LPS, the effect of TiP on cytokine secretion was inhibited. SIGNIFICANCE: Functionalised TiPs with NPs(Dex) and NPs(Dox) may potentially have beneficial effects on modulating titanium and LPS-related inflammatory reactions.

Dentin remineralization using a stimuli-responsive engineered small molecule GSK3 antagonists-functionalized adhesive.

OBJECTIVES: Tideglusib has shown great performance in terms of dentin regenerative properties. This study aims to evaluate bonding ability, of demineralized dentin infiltrated with polymeric nanoparticles (NPs) doped with tideglusib (TG) (TG-NPs). METHODS: Dentin conditioned surfaces were infiltrated with NPs and TG-NPs. Bonded interfaces were created and stored for 24 h and then submitted to mechanical, chemical and thermal challenging. The resin-dentin interface was evaluated through a doubled dye fluorescent technique and a calcium chelator fluorophore under a confocal laser scanning microscopy, and by field emission scanning electron microscopy. RESULTS: Dentin surfaces treated with TG-NPs and load cycled produced higher bond strength than the rest of the groups. Immersion of dentin specimens treated with undoped-NPs in collagenase solution attained the lowest microtensile bond strength (MTBS) values. Both porosity and nanoleakage decreased when dentin was infiltrated with TG-NPs, that revealed strong signals of xylenol orange stain at both hybrid layer and dentinal tubules. The presence of NPs, in general, inducted the presence of mineralized interfaces after mechanical loading and thermocycling. CONCLUSIONS: Nanoparticles doped with tideglusib promoted the highest dentin bonding efficacy among groups, as they facilitated the maximum bond strength values with creation of mineral deposits at the hybrid layer and dentinal walls. Tideglusib enabled scarce porosity, nanoleakage and advanced sealing among dentin groups. SIGNIFICANCE: Doping hydrophilic polymeric NPs with tideglusib, infiltrated in etched dentin represents a reproducible technique to create reparative dentin at the resin-dentin interface, by inducing therapeutic bioactivity.

Effects of an MDP-based surface cleaner on dentin structure, morphology and nanomechanical properties.

OBJECTIVE: To analyze the effect of Katana™ Cleaner (KC) in nanomechanical and triboscopic properties of etched dentin. METHODS: Dentin disks from human third molars were prepared. Two main groups of study were established in function of the etching conditioning, phosphoric acid (PA) and Clearfil SE Bond primer (CSEB). Four subgroups were tested within each group: i) untreated dentin (UD), ii) etched dentin (ED) [(PAED/CSEB)], iii) etched dentin contaminated with saliva (PAED+S)/(CSEB+S), and iv) etched and contaminated dentin treated with KC (PAED+S+KC)/(CSEB+S+KC). Nano-DMA testing and imaging, atomic force microscopy (AFM) analysis and nanoroughness (SRa) measurements were obtained. Field emission scanning electron microscopy (FESEM) images were also acquired. RESULTS: Phosphoric acid etched dentin samples and those specimens contaminated with saliva (PAED+S) attained the highest SRa values, that decreased after Katana™ Cleaner application (PAED+S+KC). In the group of dentin treated with CSEB primer, all subgroups performed similar, except in CSEB+S that attained the highest SRa values. The treatment with KC restored the original values of complex modulus of the untreated dentin. KC application produced the lowest and the highest tan delta values on PAED and CSEB groups, respectively. CONCLUSION: Katana™ Cleaner provided equally mature dentin surfaces after any of the etching methods. Tan delta increased when Katana™ Cleaner was applied on the dentin surface previously etched and contaminated with saliva, regardless the kind of etchant, thus facilitating the dissipation of energy for elastic recoil during loading. CLINICAL SIGNIFICANCE: Katana™ Cleaner application after saliva contamination originated similar low roughness levels, regardless the type of etching method. Both complex and storage moduli were similar, after Katana™ Cleaner application, in any case.

Deciphering the Cell-Specific Effect of Osteoblast-Macrophage Crosstalk in Periodontitis.

In periodontitis, the bone remodeling process is disrupted by the prevalent involvement of bacteria-induced proinflammatory macrophage cells and their interaction with osteoblast cells residing within the infected bone tissue. The complex interaction between the cells needs to be deciphered to understand the dominant player in tipping the balance from osteogenesis to osteoclastogenesis. Yet, only a few studies have examined the crosstalk interaction between osteoblasts and macrophages using biomimetic three-dimensional (3D) tissue-like matrices. In this study, we created a cell-laden 3D tissue analog to study indirect crosstalk between these two cell types and their direct synergistic effect when cultured on a 3D scaffold. The cell-specific role of osteoclast differentiation was investigated through osteoblast- and proinflammatory macrophage-specific feedback studies. The results suggested that when macrophages were exposed to osteoblasts-derived conditioned media from the mineralized matrix, the M1 macrophages tended to maintain their proinflammatory phenotype. Further, when osteoblasts were exposed to secretions from proinflammatory macrophages, they demonstrated elevated receptor activator of nuclear factor-κB ligand (RANKL) expression and decreased alkaline phosphate (ALP) activities compared to osteoblasts exposed to only osteogenic media. In addition, the upregulation of tumor necrosis factor-alpha (TNF-α) and c-Fos in proinflammatory macrophages within the 3D matrix indirectly increased the RANKL expression and reduced the ALP activity of osteoblasts, promoting osteoclastogenesis. The contact coculturing with osteoblast and proinflammatory macrophages within the 3D matrix demonstrated that the proinflammatory markers (TNF-α and interleukin-1ß) expressions were upregulated. In contrast, anti-inflammatory markers (c-c motif chemokine ligand 18 [CCL18]) were downregulated, and osteoclastogenic markers (TNF receptor associated factor 6 [TRAF6] and acid phosphatase 5, tartrate resistant [ACP5]) were unchanged. The data suggested that the osteoblasts curbed the osteoclastogenic differentiation of macrophages while macrophages still preserved their proinflammatory lineages. The osteoblast within the 3D coculture demonstrated increased ALP activity and did not express RANKL significantly different than the osteoblast cultured within a 3D collagen matrix without macrophages. Contact coculturing has an anabolic effect on bone tissue in a bacteria-derived inflammatory environment.

Dexamethasone and doxycycline functionalized nanoparticles enhance osteogenic properties of titanium surfaces.

OBJECTIVES: To evaluate the effect of doxycycline and dexamethasone doped nanoparticles covering titanium surfaces, on osteoblasts proliferation and differentiation. METHODS: Doxycycline and dexamethasone doped polymeric nanoparticles were applied on titanium discs (Ti-DoxNPs and Ti-DexNPs). Undoped NPs and uncovered Ti discs were used as control. Human MG-63 osteoblast-like cells were cultured. Osteoblasts proliferation was tested by MTT assay. Alkaline phosphatase activity was analyzed. Differentiation gene expression was assessed by real-time quantitative polymerase chain reaction. Scanning Electron Microscopy was performed to assess osteoblasts morphology. Mean comparisons were conducted by ANOVA and Wilcoxon or Tukey tests (p < 0.05). RESULTS: No differences in osteoblasts proliferation were found. Osteoblasts grown on Ti-DoxNPs significantly increased alkaline phosphatase activity. Doxycycline and dexamethasone nanoparticles produced an over-expression of the main osteogenic proliferative genes (TGF-ß1, TGF-ßR1 and TGF-ßR2). The expression of Runx-2 was up-regulated. The osteogenic proteins (AP, OSX and OPG) were also overexpressed on osteoblasts cultured on Ti-DoxNPs and Ti-DexNPs. The OPG/RANKL ratio was the highest when DoxNPs were present (75-fold increase with respect to the control group). DexNPs also produced a significantly higher OPG/RANKL ratio with respect to the control (20 times higher). Osteoblasts grown on titanium discs were mainly flat and polygonal in shape, with inter-cellular connections. In contrast, osteoblasts cultured on Ti-DoxNPs or Ti-DexNPs were found to be spindle-shaped and had abundant secretions on their surfaces. SIGNIFICANCE: DoxNPs and DexNPs were able to stimulate osteoblasts differentiation when applied on titanium surfaces, being considered potential inducers of osteogenic environment when performing regenerative procedures around titanium dental implants.

Doped Electrospinned Material-Guides High Efficiency Regional Bone Regeneration.

The main target of bone tissue engineering is to design biomaterials that support bone regeneration and vascularization. Nanostructured membranes of (MMA)1-co-(HEMA)1/(MA)3-co-(HEA)2 loaded with 5% wt of SiO2-nanoparticles (Si-M) were doped with zinc (Zn-Si-M) or doxycycline (Dox-Si-M). Critical bone defects were effectuated on six New Zealand-bred rabbit skulls and then they were covered with the membranes. After six weeks, a histological analysis (toluidine blue technique) was employed to determine bone cell population as osteoblasts, osteoclasts, osteocytes, M1 and M2 macrophages and vasculature. Membranes covering the bone defect determined a higher count of bone cells and blood vessels than in the sham group at the top regions of the defect. Pro-inflammatory M1 appeared in a higher number in the top regions than in the bottom regions, when Si-M and Dox-Si-M were used. Samples treated with Dox-Si-M showed a higher amount of anti-inflammatory and pro-regenerative M2 macrophages. The M1/M2 ratio obtained its lowest value in the absence of membranes. On the top regions, osteoblasts were more abundant when using Si-M and Zn-Si-M. Osteoclasts were equally distributed at the central and lateral regions. The sham group and samples treated with Zn-Si-M attained a higher number of osteocytes at the top regions. A preferential osteoconductive, osteoinductive and angiogenic clinical environment was created in the vicinity of the membrane placed on critical bone defects.

Dexamethasone-doped nanoparticles improve mineralization, crystallinity and collagen structure of human dentin.

OBJECTIVES: Bioactive materials have been used for functionalization of adhesives to promote dentin remineralization. This study aims to evaluate bonding ability and both mechanical and chemical behavior of demineralized dentin infiltrated with polymeric nanoparticles doped with dexamethasone (Dex-NPs). METHODS: Dentin conditioned surfaces were infiltrated with NPs, Dex-NPs or Dex-Zn-NPs. Bonded interfaces were also created and stored for 24 h or 21d, and then submitted to microtensile bond strength testing. Dentin remineralization was analyzed by Nanohardness, Young's modulus and Raman analysis. RESULTS: At 21d of storage, dentin treated with undoped-NPs attained the lowest nanohardness and Young's modulus. Dex-NPs and Zn-Dex-NPs increased dentin nanohardness and Young's modulus after 21d Raman analysis showed high remineralization, crystallinity, crosslinking and better structure of collagen when functionalized Dex-NPs were present at the dentin interface. CONCLUSIONS: Infiltration of dentin with Dex-NPs promoted functional remineralization as proved by nanomechanical and morpho-chemical evaluation tests. Dexamethasone in dentin facilitated crystallographic maturity, crystallinity and improved maturity and secondary structure of dentin collagen. CLINICAL SIGNIFICANCE: Using dexamethasone-functionalized NPs before resin infiltration is a clear option to obtain dentin remineralization, as these NPs produce the reinforcement of the dentin structure, which will lead to the improvement of the longevity of resin restorations.

Dexamethasone and zinc loaded polymeric nanoparticles reinforce and remineralize coronal dentin. A morpho-histological and dynamic-biomechanical study.

OBJECTIVE: To investigate the effect of novel polymeric nanoparticles (NPs) doped with dexamethasone (Dex) on viscoelasticity, crystallinity and ultra-nanostructure of the formed hydroxyapatite after NPs dentin infiltration. METHODS: Undoped-NPs, Dex-doped NPs (Dex-NPs) and zinc-doped-Dex-NPs (Zn-Dex-NPs) were tested at dentin, after 24 h and 21 d. A control group without NPs was included. Coronal dentin surfaces were studied by nano-dynamic mechanical analysis measurements, atomic force microscopy, X-ray diffraction and transmission electron microscopy. Mean and standard deviation were analyzed by ANOVA and Student-Newman-Keuls multiple comparisons (p < 0.05). RESULTS: At 21 d of storage time, both groups doped with Dex exhibited the highest complex, storage and loss moduli among groups. Zn-Dex-NPs and Dex-NPs promoted the highest and lowest tan delta values, respectively. Dex-NPs contributed to increase the fibril diameters of dentin collagen over time. Dentin surfaces treated with Zn-Dex-NPs attained the lowest nano-roughness values, provoked the highest crystallinity, and produced the longest and shortest crystallite and grain size. These new crystals organized with randomly oriented lattices. Dex-NPs induced the highest microstrain. Crystalline and amorphous matter was present in the mineral precipitates of all groups, but Zn and Dex loaded NPs helped to increase crystallinity. SIGNIFICANCE: Dentin treated with Zn-Dex-NPs improved crystallographic and atomic order, providing structural stability, high mechanical performance and tissue maturation. Amorphous content was also present, so high hydroxyapatite solubility, bioactivity and remineralizing activity due to the high ion-rich environment took place in the infiltrated dentin.

Doxycycline-doped collagen membranes accelerate in vitro osteoblast proliferation and differentiation.

OBJECTIVE: The aim of the study was to evaluate the effect of doxycycline- and dexamethasone-doped collagen membranes on the proliferation and differentiation of osteoblasts. BACKGROUND: Collagen barrier membranes are frequently used to promote bone regeneration and to boost this biological activity their functionalization with antibacterial and immunomodulatory substances has been suggested. METHODS: The design included commercially available collagen membranes doped with doxycycline (Dox-Col-M) or dexamethasone (Dex-Col-M), as well as undoped membranes (Col-M) as controls, which were placed in contact with cultured MG63 osteoblast-like cells (ATCC). Cell proliferation was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assay and differentiation by measuring the alkaline phosphatase (ALP) activity using spectrophotometry. Real-time quantitative polymerase chain reaction was used to study the expression of the genes: Runx-2, OSX, ALP, OSC, OPG, RANKL, Col-I, BMP-2, BMP-7, TGF-ß1, VEGF, TGF-ßR1, TGF-ßR2, and TGF-ßR3. Scanning electron microscopy was used to study osteoblast morphology. Data were assessed using one-way analysis of variance or Kruskal-Wallis tests, once their distribution normality was assessed by Kolmogorov-Smirnov tests (p > .05). Bonferroni for multiple comparisons were carried out (p < .05). RESULTS: Osteoblast proliferation was significantly enhanced in the functionalized membranes as follows: (Col-M < Dex-Col-M < Dox-Col-M). ALP activity was significantly higher on cultured osteoblasts on Dox-Col-M. Runx-2, OSX, ALP, OSC, BMP-2, BMP-7, TGF-ß1, VEGF, TGF-ßR1, TGF-ßR2, and TGF-ßR3 were overexpressed, and RANKL was down-regulated in osteoblasts cultured on Dox-Col-M. The osteoblasts cultured in contact with the functionalized membranes demonstrated an elongated spindle-shaped morphology. CONCLUSION: The functionalization of collagen membranes with Dox promoted an increase in the proliferation and differentiation of osteoblasts.

Biomimetic Collagen Membranes as Drug Carriers of Geranylgeraniol to Counteract the Effect of Zoledronate.

To counteract the effect of zoledronate and decrease the risk of osteonecrosis of the jaw (BRONJ) development in patients undergoing guided bone regeneration surgery, the use of geranylgeraniol (GGOH) has been proposed. Collagen membranes may act as biomimetical drug carriers. The objective of this study was to determine the capacity of collagen-based membranes doped with GGOH to revert the negative impact of zoledronate on the growth and differentiation of human osteoblasts. MG-63 cells were cultured on collagen membranes. Two groups were established: (1) undoped membranes and (2) membranes doped with geranylgeraniol. Osteoblasts were cultured with or without zoledronate (50 µM). Cell proliferation was evaluated at 48 h using the MTT colorimetric method. Differentiation was tested by staining mineralization nodules with alizarin red and by gene expression analysis of bone morphogenetic proteins 2 and 7, alkaline phosphatase (ALP), bone morphogenetic proteins 2 and 7 (BMP-2 and BMP-7), type I collagen (Col-I), osterix (OSX), osteocalcin (OSC), osteoprotegerin (OPG), receptor for RANK (RANKL), runt-related transcription factor 2 (Runx-2), TGF-ß1 and TGF-ß receptors (TGF-ßR1, TGF-ßR2, and TGF-ßR3), and vascular endothelial growth factor (VEGF) with real-time PCR. One-way ANOVA or Kruskal-Wallis and post hoc Bonferroni tests were applied (p < 0.05). Scanning electron microscopy (SEM) observations were also performed. Treatment of osteoblasts with 50 µM zoledronate produced a significant decrease in cell proliferation, mineralization capacity, and gene expression of several differentiation markers if compared to the control (p < 0.001). When osteoblasts were treated with zoledronate and cultured on GGOH-doped membranes, these variables were, in general, similar to the control group (p > 0.05). GGOH applied on collagen membranes is able to reverse the negative impact of zoledronate on the proliferation, differentiation, and gene expression of different osteoblasts' markers.

Short versus standard implants at sinus augmented sites: a systematic review and meta-analysis.

BACKGROUND: Short implants are proposed as a less invasive alternative with fewer complications than standard implants in combination with sinus lift. The aim of this systematic review and meta-analysis was to state the efficacy of placing short implants (≤ 6 mm) compared to standard-length implants (≥ 8 mm) performing sinus lift techniques in patients with edentulous posterior atrophic jaws. Efficacy will be evaluated through analyzing implant survival (IS) and maintenance of peri-implant bone (MBL). METHODS: Screening process was done using the National Library of Medicine (MEDLINE by PubMed), EMBASE, the Cochrane Oral Health, and Web of Science (WOS). The articles included were randomized controlled trials. Risk of bias was evaluated according to The Cochrane Collaboration's tool. Weighted means were calculated. Heterogeneity was determined using Higgins (I2). A random-effects model was applied. Secondary outcomes such as surgical time, patient satisfaction, mucositis and peri-implantitis, pain, and swelling were analyzed. RESULTS: Fourteen studies (597 patients and 901 implants) were evaluated. IS was 1.02 risk ratio, ranging from 1.00 to 1.05 (CI 95%) (p = 0.09), suggesting that IS was similar when both techniques were used. MBL was higher in patients with standard-length implants plus sinus lift elevation (p = 0.03). MBL was 0.11 (0.01-0.20) mm (p = 0.03) and 0.23 (0.07-0.39) mm (p = 0.005) before and after 1 year of follow-up, respectively, indicating that the marginal bone loss is greater for standard-length implants. DISCUSSION: Within the limitations of the present study, as relatively small sample size, short dental implants can be used as an alternative to standard-length implants plus sinus elevation in cases of atrophic posterior maxilla. Higher MBL was observed in the groups where standard-length implants were used, but implant survival was similar in both groups. Moreover, with short implants, it was observed a reduced postoperative discomfort, minimal invasiveness, shorter treatment time, and reduced costs. CLINICAL CLINICAL RELEVANCE: The low MBL promoted by short implants does contribute to a paradigm shift from sinus grafting with long implants to short implants. Further high-quality long-term studies are required to confirm these findings.

Dexamethasone and Doxycycline Doped Nanoparticles Increase the Differentiation Potential of Human Bone Marrow Stem Cells.

Non-resorbable polymeric nanoparticles (NPs) are proposed as an adjunctive treatment for bone regenerative strategies. The present in vitro investigation aimed to evaluate the effect of the different prototypes of bioactive NPs loaded with zinc (Zn-NPs), doxycycline (Dox-NPs) or dexamethasone (Dex-NPs) on the viability, morphology, migration, adhesion, osteoblastic differentiation, and mineralization potential of human bone marrow stem cells (hBMMSCs). Cell viability, proliferation, and differentiation were assessed using a resaruzin-based assay, cell cycle analysis, cell migration evaluation, cell cytoskeleton staining analysis, Alizarin Red S staining, and expression of the osteogenic-related genes by a real-time quantitative polymerase chain reaction (RT-qPCR). One-Way ANOVA and Tukey's test were employed. The resazurin assay showed adequate cell viability considering all concentrations and types of NPs at 24, 48, and 72 h of culture. The cell cycle analysis revealed a regular cell cycle profile at 0.1, 1, and 10 µg/mL, whereas 100 µg/mL produced an arrest of cells in the S phase. Cells cultured with 0.1 and 1 µg/mL NP concentrations showed a similar migration capacity to the untreated group. After 21 days, mineralization was increased by all the NPs prototypes. Dox-NPs and Dex-NPs produced a generalized up-regulation of the osteogenic-related genes. Dex-NPs and Dox-NPs exhibited excellent osteogenic potential and promoted hBMMSC differentiation. Future investigations, both in vitro and in vivo, are required to confirm the suitability of these NPs for their clinical application.

Biomimetic Remineralization of an Extracellular Matrix Collagen Membrane for Bone Regeneration.

Natural extracellular matrix (ECM) collagen membranes are frequently used for bone regeneration procedures. Some disadvantages, such as rapid degradation and questionable mechanical properties, limit their clinical use. These membranes have a heterologous origin and may proceed from different tissues. Biomineralization is a process in which hydroxyapatite deposits mainly in collagen fibrils of the matrices. However, when this deposition occurs on the ECM, its mechanical properties are increased, facilitating bone regeneration. The objective of the present research is to ascertain if different membranes from distinct origins may undergo biomineralization. Nanomechanical properties, scanning electron (SEM) and multiphoton (MP) microscopy imaging were performed in three commercially available ECMs before and after immersion in simulated body fluid solution for 7 and 21 d. The matrices coming from porcine dermis increased their nanomechanical properties and they showed considerable mineralization after 21 d, as observed in structural changes detected through SEM and MP microscopy. It is hypothesized that the more abundant crosslinking and the presence of elastin fibers within this membrane explains the encountered favorable behavior.

Histomorphometric Analysis of Differential Regional Bone Regeneration Induced by Distinct Doped Membranes.

Our objective is to evaluate the regional regenerative potential of calvarial bone in critical-sized defects in a rabbit model using novel nanostructured silica-loaded membranes doped with zinc or doxycycline. Nanostructured membranes of (MMA)1-co-(HEMA)1/(MA)3-co-(HEA)2 loaded with 5 wt% of SiO2 nanoparticles (HOOC-Si-Membranes) were doped with zinc (Zn-HOOC-Si-Membrane) or doxycycline (Dox-HOOC-Si-Membrane). Critical bone defects were created on six New-Zealand-breed rabbit skulls and covered with the membranes. A sham defect without a membrane was used as the control. After six weeks, a histological analysis (toluidine blue technique) was employed to determine the area percentages of newly formed bone, osteoid bone, and soft tissue. The measurements were performed by dividing the total defect area into top (close to the membrane) and bottom (close to the dura mater) regions, or peripheral (adjacent to the old bone) and central (the sum of the remaining zones) regions. The peripheral regions of the defects showed higher osteogenic capacity than the central areas when the membranes were present. The proportion of new bone adjacent to the dura was similar to that adjacent to the membrane only when the HOOC-Si-Membranes and Zn-HOOC-Si-Membranes were used, indicating a direct osteoinductive effect of the membranes.

Treating Gingival Recessions Using Coronally Advanced Flap or Tunnel Techniques with Autografts or Polymeric Substitutes: A Systematic Review and Meta-Analysis.

Gingival recessions are a prevalent oral mucosa alteration. To solve this pathology, palatal mucosa or polymeric soft tissue substitutes are used when performing coronal advanced flap (CAF) or tunnel (TUN) surgical techniques. To evaluate which is the most successful approach, a literature review and meta-analysis were conducted. For the electronic search the National Library of Medicine, the Cochrane Oral Health Group Trials Register, EMBASE and WOS were used. Pooled data for the percentage of root coverage was collected and weighted means were calculated. Heterogeneity was determined using the Higgins (I2) statistic and a random-effects model was applied. Thirteen studies were included in the systematic review (12 randomized and 1 controlled clinical trials) in which both techniques (394 patients) were compared with a follow-up of 4 to 12 months. Galbraith and Baujat plots were used to control for the presence of potential outliers. After performing the meta-analysis (11 studies), the mean root coverage was similar when using the TUN or CAF techniques (p = 0.49). The only differences between the two were found for single recessions, where CAF offered a higher percentage of root coverage (mean difference = 4.98%; p = 0.006). There were no differences when applying an autograft or a polymeric substitute with either of the two tested surgical techniques (p = 0.445).

An in-vitro investigation of the bond strength of experimental ion-releasing dental adhesives to caries-affected dentine after 1 year of water storage.

OBJECTIVE: The aim of this study was to evaluate the bonding performance after 1 year storage of an experimental dental adhesive containing analogues of phosphoproteins and fluoride-doped bioglass (EXP), applied in self-etching (SE) or etch & rinse (ER) mode, to caries-affected dentine after selective caries removal. MATERIALS AND METHODS: Fifty human molars with dentine carious lesions were excavated selectively using CarisolvTM gel and then connected to simulated pulpal pressure system. Teeth were divided randomly into five groups based on the tested materials: EXP-SE, EXP-ER, a resin-modified glass-ionomer cement (RMGIC), a three-step adhesive system (OPT) and a universal adhesive applied in SE mode (UA-SE). The specimens were submitted to different analytical tests (µTBS, SEM fractographic analysis and dye-enhanced confocal microscopy) at baseline (T0) and after 1 year (T1). RESULTS: At T0 there was no difference in bond strength between the tested materials (p>0.05). At T1, EXP-SE and EXP-ER were the only materials to show no significant reduction in bond strength (p<0.05). The SEM showed a clear presence of minerals deposited on the dentine surface after bonding in the EXP-SE and EXP-ER groups. The specimens restored with RMGIC showed no exposure of the dentine surface after failure. The OPT and UA-SE specimens showed clear signs of degradation at the interface. Confocal microscopy imaged mineral precipitation at the interface of the EXP groups. CONCLUSION: Conventional adhesives may have inadequate bonding performance with evident degradation at the dentine-bonded interface over time. Although the RMGIC may present a drop in bond strength after prolonged storage, the bonding interface may result less affected by degradation over time. Innovative ion-releasing adhesives may remineralise the caries-affected dentine and achieve a stable bond over time. CLINICAL SIGNIFICANCE: GIC-based materials may represent an appropriate dentine replacement material after selective chemo-mechanical caries removal rather than conventional adhesive systems.

Comparison of Mitral Regurgitant Volume Assessment between Proximal Flow Convergence and Volumetric Methods in Patients with Significant Primary Mitral Regurgitation: An Echocardiographic and Cardiac Magnetic Resonance Imaging Study.

BACKGROUND: Discrepancies have been observed between transthoracic echocardiography (TTE) and cardiac magnetic resonance imaging (CMR) severity grading in primary mitral regurgitation (MR). OBJECTIVES: We sought to compare mitral regurgitant volume (RVol) determined by the TTE proximal flow convergence (proximal isovelocity surface area [PISA]) method and by volumetric methods (TTE and CMR) and to study the relationship between left ventricle (LV) size and RVol obtained by either the PISA or volumetric methods. METHODS: Two centers prospectively recruited 188 patients with at least moderate to severe primary MR due to prolapse in sinus rhythm who underwent TTE and CMR examinations. Regurgitant volume was estimated by either PISA (PISA-RVol) or volumetric methods (LV total stroke volume-systolic aortic forward outflow volume) using either CMR (CMR-RVol) or TTE (TTE-RVol). RESULTS: The PISA-RVol was weakly correlated with CMR-RVol and TTE-RVol (r = 0.29 and 0.30, respectively; P < .001 for both). On multivariable analysis, smaller CMR-left ventricular end-diastolic volume (LVEDV) and absence of mitral annular disjunction independently correlated with increased magnitude of RVol difference between PISA and volumetric methods. While PISA-RVol and LVEDV were unrelated, CMR-RVol and TTE-RVol moderately correlated with LVEDV (r = 0.66 and 0.68, respectively; P < .001 for both). In contrast, LVEDV and regurgitant fraction (RVol/LV total stroke volume), assessed with either TTE or CMR, were poorly correlated (r = 0.17, P = .02; and r = 0.12, P = .10, respectively). CONCLUSIONS: Mitral RVol values estimated by PISA and volumetric methods are not directly comparable. The expected proportional relationship between volumetric RVol and LV size, which was not observed with PISA-RVol, suggests that PISA-RVol would be inaccurate. Given that RVol assessed with volumetric methods depends on LV size, determination of a unique RVol threshold for severe MR is challenging. In contrast to RVol, calculating regurgitant fraction by volumetric methods allows the quantification of MR severity independently from LV size.

Antibacterial Effect of Functionalized Polymeric Nanoparticles on Titanium Surfaces Using an In Vitro Subgingival Biofilm Model.

This investigation aimed to evaluate the antibacterial effect of polymeric nanoparticles (NPs), functionalized with calcium, zinc, or doxycycline, using a subgingival biofilm model of six bacterial species (Streptococcus oralis,Actinomyces naeslundii, Veillonela parvula, Fusobacterium nucleatum, Porphyromonas gingivalis, and Aggregatibacter actinomycetemcomitans) on sandblasted, large grit, acid-etched titanium discs (TiDs). Undoped NPs (Un-NPs) or doped NPs with calcium (Ca-NPs), zinc (Zn-NPs), or doxycycline (Dox-NPs) were applied onto the TiD surfaces. Uncovered TiDs were used as negative controls. Discs were incubated under anaerobic conditions for 12, 24, 48, and 72 h. The obtained biofilm structure was studied by scanning electron microscopy (SEM) and its vitality and thickness by confocal laser scanning microscopy (CLSM). Quantitative polymerase chain reaction of samples was used to evaluate the bacterial load. Data were evaluated by analysis of variance (p < 0.05) and post hoc comparisons with Bonferroni adjustments (p < 0.01). As compared with uncovered TiDs, Dox-NPs induced higher biofilm mortality (47.21% and 85.87%, respectively) and reduced the bacterial load of the tested species, after 72 h. With SEM, scarce biofilm formation was observed in Dox-NPs TiDs. In summary, Dox-NPs on TiD reduced biofilm vitality, bacterial load, and altered biofilm formation dynamics.