The effects of mineral trioxide aggregate and second-generation autologous growth factor on pulpotomy via TNF-α and NF-kß/p65 pathways.

This study aims to investigate the effect of Mineral Trioxide Aggregate (MTA), a bioactive endodontic cement, and Concentrated Growth Factor (CGF), a second-generation autologous growth factor, on pulpotomy-induced pulp inflammation. The study utilized the maxillary anterior central teeth of thirty-six young male Sprague Dawley rats. Forty-eight teeth were randomly assigned to two groups (12 rats/group; 24 teeth/group) based on the capping material (MTA or CGF). Subsequently, two subgroups (MTAG and CGFG) were formed per group (12 teeth/group) based on the time following pulpotomy (2-weeks and 4-weeks). The central teeth of the 12 animals assigned to the control group (CG) were not manipulated in any way, both in the 2-week group and in the 4-week group. Tissue samples extracted from rats at the end of the experiment were stained with H&E for histopathological analysis. For immunohistochemical analysis, primary antibodies for TNF-α and NF-kß/65 were incubated. Data obtained from semi-quantitative analysis were assessed for normal distribution using Skewness-Kurtosis values, Q-Q plot, Levene's test, and the Shapiro-Wilk test on statistical software. A P value < 0.05 was considered significant. When compared with the control group, both MTAG and CGFG showed increased edematous and inflammatory areas. In MTAG, edematous and inflammatory areas decreased significantly from the 2nd week (2(2-2), 2(1-2)) to the 4th week (1(1-1), 1(0-1)), while in CGFG, edematous areas decreased (2(2-3), 1.5(1-2)), and inflammatory areas increased significantly (2(2-3), 3(2-2.5)). When compared with the control group, TNF-α and NF-kß/p65 positivity were higher in both MTAG and CGFG. In MTAG, TNF-α [2(1.5-2)] and NF-kß/p65 [1.5(1-2)] positivity decreased significantly from the 2nd week to the 4th week [TNF-α: 1(1-1), NF-kß/p65: 1(1-2)], while no significant change was observed in CGFG. In conclusion, this study revealed a reduction in cells showing TNF-α and NF-kß/p65 positivity in the MTA treatment group compared to the CGF group. Although MTA demonstrated more favorable results than CGF in mitigating pulpal inflammation within the scope of this study, further experimental and clinical investigations are warranted to obtain comprehensive data regarding CGF.

Biocompatibility, bioactivity and immunomodulatory properties of three calcium silicate-based sealers: an in vitro study on hPDLSCs.

OBJECTIVES: To assess the biocompatibility, bioactivity, and immunomodulatory properties of three new calcium silicate cement-based sealers: Ceraseal (CS), Totalfill BC Sealer (TFbc) and WellRoot ST (WR-ST) on human periodontal ligament stem cells (hPDLSCs). MATERIALS AND METHODS: HPDLSCs were isolated from extracted third molars from healthy patients. Eluates (1:1, 1:2, and 1:4 ratio) and sample discs of CS, TFbc and WR-ST after setting were prepared. A series of assays were performed: cell characterization, cell metabolic activity (MTT assay) cell attachment and morphology (SEM assay), cell migration (wound-healing assay), cytoskeleton organization (phaloidin-based assay); IL-6 and IL-8 release (ELISA); differentiation marker expression (RT-qPCR assay), and cell mineralization (Alizarin Red S staining). HPDLSCs cultured in unconditioned (negative control) or osteogenic (positive control) culture media were used as a comparison. Statistical significance was established at p < 0.05. RESULTS: All the tested sealers exhibited similar results in the cytocompatibility assays (cell metabolic activity, migration, attachment, morphology, and cytoskeleton organization) compared with a negative control group. CS and TFbc exhibited an upregulation of at least one osteo/cementogenic marker compared to the negative and positive control groups. CS and TFbc also showed a significantly higher calcified nodule formation than the negative and positive control groups. Both the marker expression and calcified nodule formation were significantly higher in CS-treated cells than TFbc treated cells. WR-ST exhibited similar results to the control group. CS and TFbc-treated cells exhibited a significant downregulation of IL-6 after 72 h of culture compared to the negative control group (p < 0.05). CONCLUSION: All the tested sealers exhibited an adequate cytocompatibility. CS significantly enhances cell differentiation by upregulating the expression of key genes associated with bone and cementum formation. Additionally, CS was observed to facilitate the mineralization of the extracellular matrix effectively. In contrast, the effects of TFbc and WR-ST on these processes were less pronounced compared to CS. Furthermore, both CS and TFbc exhibited an anti-inflammatory potential, contributing to their potential therapeutic benefits in regenerative endodontics. CLINICAL RELEVANCE: This is the first study to compare the biological properties and immunomodulatory potential of Ceraseal, Totalfill BC Sealer, and WellRoot ST. The results act as supporting evidence for their use in root canal treatment.

Novel L-(CaP-ZnP)/SA Nanocomposite Hydrogel with Dual Anti-Inflammatory and Mineralization Effects for Efficient Vital Pulp Therapy.

Background: Vital pulp therapy (VPT) is considered a conservative treatment for preserving pulp viability in caries and trauma-induced pulpitis. However, Mineral trioxide aggregate (MTA) as the most frequently used repair material, exhibits limited efficacy under inflammatory conditions. This study introduces an innovative nanocomposite hydrogel, tailored to simultaneously target anti-inflammation and dentin mineralization, aiming to efficiently preserve vital pulp tissue. Methods: The L-(CaP-ZnP)/SA nanocomposite hydrogel was designed by combining L-Arginine modified calcium phosphate/zinc phosphate nanoparticles (L-(CaP-ZnP) NPs) with sodium alginate (SA), and was characterized with TEM, SEM, FTIR, EDX, ICP-AES, and Zeta potential. In vitro, we evaluated the cytotoxicity and anti-inflammatory properties. Human dental pulp stem cells (hDPSCs) were cultured with lipopolysaccharide (LPS) to induce an inflammatory response, and the cell odontogenic differentiation was measured and possible signaling pathways were explored by alkaline phosphatase (ALP)/alizarin red S (ARS) staining, qRT-PCR, immunofluorescence staining, and Western blotting, respectively. In vivo, a pulpitis model was utilized to explore the potential of the L-(CaP-ZnP)/SA nanocomposite hydrogel in controlling pulp inflammation and enhancing dentin mineralization by Hematoxylin and eosin (HE) staining and immunohistochemistry staining. Results: In vitro experiments revealed that the nanocomposite hydrogel was synthesized successfully and presented desirable biocompatibility. Under inflammatory conditions, compared to MTA, the L-(CaP-ZnP)/SA nanocomposite hydrogel demonstrated superior anti-inflammatory and pro-odontogenesis effects. Furthermore, the nanocomposite hydrogel significantly augmented p38 phosphorylation, implicating the involvement of the p38 signaling pathway in pulp repair. Significantly, in a rat pulpitis model, the L-(CaP-ZnP)/SA nanocomposite hydrogel downregulated inflammatory markers while upregulating mineralization-related markers, thereby stimulating the formation of robust reparative dentin. Conclusion: The L-(CaP-ZnP)/SA nanocomposite hydrogel with good biocompatibility efficiently promoted inflammation resolution and enhanced dentin mineralization by activating p38 signal pathway, as a pulp-capping material, offering a promising and advanced solution for treatment of pulpitis.

Comparison of remineralization ability of tricalcium silicate and of glass ionomer cement on residual dentin: an in vitro study.

OBJECTIVE: This study aimed to compare the remineralization effects of a calcium silicate-based cement (Biodentine) and of a glass ionomer cement (GIC: Fuji IX) on artificially demineralized dentin. METHODS: Four standard cavities were prepared in dentin discs prepared from 34 extracted sound human third molars. In each disc, one cavity was covered with an acid-resistant varnish before demineralization (Group 1). The specimens were soaked in a chemical demineralization solution for 96 h to induce artificial carious lesions. Thereafter, one cavity each was filled with Biodentine (Group 2) and GIC (Group 3), respectively, and one carious lesion was left unrestored as a negative control (Group 4). Next, specimens were immersed in simulated body fluid (SBF) for 21 days. After cross-sectioning the specimens, the Ca/P ratio was calculated in each specimen by using scanning electron microscopy-energy-dispersive X-ray spectroscopy (SEM-EDX). Finally, data were analyzed using repeated-measures ANOVA with post-hoc Bonferroni correction. RESULTS: Both cement types induced dentin remineralization as compared to Group 4. The Ca/P ratio was significantly higher in Group 2 than in Group 3 (p < 0.05). CONCLUSION: The dentin lesion remineralization capability of Biodentine is higher than that of GIC, suggesting the usefulness of the former as a bioactive dentin replacement material. CLINICAL RELEVANCE: Biodentine has a higher remineralization ability than that of GIC for carious dentin, and its interfacial properties make it a promising bioactive dentin restorative material.

Laponite/lactoferrin hydrogel loaded with eugenol for methicillin-resistant Staphylococcus aureus-infected chronic skin wound healing.

Severe bacterial infections can give rise to protracted wound healing processes, thereby posing a significant risk to a patient's well-being. Consequently, the development of a versatile hydrogel dressing possessing robust bioactivity becomes imperative, as it holds the potential to expedite wound healing and yield enhanced clinical therapeutic outcomes. In this context, the present study involves the formulation of an injectable multifunctional hydrogel utilizing laponite (LAP) and lactoferrin (LF) as foundational components and loaded with eugenol (EG). This hydrogel is fabricated employing a straightforward one-pot mixing approach that leverages the principle of electrostatic interaction. The resulting LAP/LF/EG2% composite hydrogel can be conveniently injected to address irregular wound geometries effectively. Once administered, the hydrogel continually releases lactoferrin and eugenol, mitigating unwarranted oxidative stress and eradicating bacterial infections. This orchestrated action culminates in the acceleration of wound healing specifically in the context of MRSA-infected wounds. Importantly, the LAP/LF/EG2% hydrogel exhibits commendable qualities including exceptional injectability, potent antioxidant attributes, and proficient hemostatic functionality. Furthermore, the hydrogel composition notably encourages cellular migration while maintaining favorable cytocompatibility. Additionally, the hydrogel manifests noteworthy bactericidal efficacy against the formidable multidrug-resistant MRSA bacterium. Most significantly, this hydrogel formulation distinctly expedites the healing of MRSA-infected wounds by promptly inducing hemostasis, curbing bacterial proliferation, and fostering angiogenesis, collagen deposition, and re-epithelialization processes. As such, the innovative hydrogel material introduced in this investigation emerges as a promising dressing for the facilitation of bacterial-infected wound healing and consequent tissue regeneration.

A Composite Hydrogel Functionalized by Borosilicate Bioactive Glasses and VEGF for Critical-Size Bone Regeneration.

Critical-size bone defects pose a formidable challenge in clinical treatment, prompting extensive research efforts to address this problem. In this study, an inorganic-organic multifunctional composite hydrogel denoted as PLG-g-TA/VEGF/Sr-BGNPs is developed, engineered for the synergistic management of bone defects. The composite hydrogel demonstrated the capacity for mineralization, hydroxyapatite formation, and gradual release of essential functional ions and vascular endothelial growth factor (VEGF) and also maintained an alkaline microenvironment. The composite hydrogel promoted the proliferation and osteogenic differentiation of rat bone marrow mesenchymal stem cells (rBMSCs), as indicated by increased expression of osteogenesis-related genes and proteins in vitro. Moreover, the composite hydrogel significantly enhanced the tube-forming capability of human umbilical vein endothelial cells (HUVECs) and effectively inhibited the process of osteoblastic differentiation of nuclear factor kappa-B ligand (RANKL)-induced Raw264.7 cells and osteoclast bone resorption. After the implantation of the composite hydrogel into rat cranial bone defects, the expression of osteogenic and angiogenic biomarkers increased, substantiating its efficacy in promoting bone defect repair in vivo. The commendable attributes of the multifunctional composite hydrogel underscore its pivotal role in expediting hydrogel-associated bone growth and repairing critical bone defects, positioning it as a promising adjuvant therapy candidate for large-segment bone defects.

A three-dimensional cell culture approach to investigate cytotoxic effects and production of inflammatory mediators by epoxy resin-based and calcium silicate-based endodontic sealer.

OBJECTIVES: The aim of the present study was to assess the cytocompatibility of epoxy resin-based AH Plus Jet (Dentsply De Trey, Konstanz, Germany), Sealer Plus (MK Life, Porto Alegre, Brazil), calcium silicate-based Bio-C Sealer (Angelus, Londrina, PR, Brazil), Sealer Plus BC (MK Life) and AH Plus BC (Dentsply) through a tridimensional (3D) culture model of human osteoblast-like cells. METHODS: Spheroids of MG-63 cells were produced and exposed to fresh root canal sealers extracts by 24 h, and the cytotoxicity was assessed by the Lactate Dehydrogenase assay (LDH). The distribution of dead cells within the microtissue was assessed by fluorescence microscopy, and morphological effects were investigated by histological analysis. The secreted inflammatory mediators were detected in cell supernatants through flow luminometry (XMap Luminex). RESULTS: Cells incubated with AH Plus Jet, AH Plus BC, Sealer Plus BC and Bio-C Sealer extracts showed high rates of cell viability, while the Sealer Plus induced a significant reduction of cell viability, causing reduction on the spheroid structure. Sealer Plus and Seaker Plus BC caused alterations on 3D microtissue morphology. The AH Plus BC extract was associated with the downregulation of secretion of pro-inflammatory cytokines IL-5, IL-7, IP-10 and RANTES. CONCLUSIONS: The new AH Plus BC calcium silicate-based endodontic sealer did not reduce cell viability in vitro, while led to the downregulation of pro-inflammatory cytokines. CLINICAL SIGNIFICANCE: Choosing the appropriate endodontic sealer is a crucial step. AH Plus BC demonstrated high cell viability and downregulation of pro-inflammatory cytokines, appearing reliable for clinical use, while Sealer Plus presented lower cytocompatibility.

Hierarchical Chiral Calcium Silicate Hydrate Films Promote Vascularization for Tendon-to-Bone Healing.

Revascularization after rotator cuff repair is crucial for tendon-to-bone healing. The chirality of materials has been reported to influence their performance in tissue repair. However, data on the use of chiral structures to optimize biomaterials as a revascularization strategy remain scarce. Here, calcium silicate hydrate (CSO) films with hierarchical chirality on the atomic to micrometer scale are developed. Interestingly, levorotatory CSO (L-CSO) films promote the migration and angiogenesis of endothelial cells, whereas dextral and racemic CSO films do not induce the same effects. Molecular analysis demonstrates that L-chirality can be recognized by integrin receptors and leads to the formation of focal adhesion, which activates mechanosensitive ion channel transient receptor potential vanilloid 4 to conduct Ca2+ influx. Consequently, the phosphorylation of serum response factor is biased by Ca2+ influx to promote the vascular endothelial growth factor receptor 2 signaling pathway, resulting in enhanced angiogenesis. After implanted in a rat rotator cuff tear model, L-CSO films strongly enhance vascularization at the enthesis, promoting collagen maturation, increasing bone and fibrocartilage formation, and eventually improving the biomechanical strength. This study reveals the mechanism through which chirality influences angiogenesis in endothelial cells and provides a critical theoretical foundation for the clinical application of chiral biomaterials.

A bioactive calcium silicate nanowire-containing hydrogel for organoid formation and functionalization.

Organoids, which are 3D multicellular constructs, have garnered significant attention in recent years. Existing organoid culture methods predominantly utilize natural and synthetic polymeric hydrogels. This study explored the potential of a composite hydrogel mainly consisting of calcium silicate (CS) nanowires and methacrylated gelatin (GelMA) as a substrate for organoid formation and functionalization, specifically for intestinal and liver organoids. Furthermore, the research delved into the mechanisms by which CS nanowires promote the structure formation and development of organoids. It was discovered that CS nanowires can influence the stiffness of the hydrogel, thereby regulating the expression of the mechanosensory factor yes-associated protein (YAP). Additionally, the bioactive ions released by CS nanowires in the culture medium could accelerate Wnt/ß-catenin signaling, further stimulating organoid development. Moreover, bioactive ions were found to enhance the nutrient absorption and ATP metabolic activity of intestinal organoids. Overall, the CS/GelMA composite hydrogel proves to be a promising substrate for organoid formation and development. This research suggested that inorganic biomaterials hold significant potential in organoid research, offering bioactivities, biosafety, and cost-effectiveness.

Continuous valorization of food waste and oily food waste using bacteria-pumice and bacteria-smectite nanocomposites: Alternative cell immobilization and zooplankton lifespan impact.

Recycling waste into commercial products is a profitable strategy but the lifetime of immobilized cells for long-term waste treatment remains a problem. This study presents alternative cell immobilization methods for valorizing food waste (FW) and oily food waste (OFW) to microbial carotenoids and proteins. Carriers (pumice or smectite), magnetite nanoparticles, and isolated photosynthetic bacteria were integrated to obtain magnetically recoverable bacteria-pumice and bacteria-smectite nanocomposites. After recycling five batches (50 d), chemical oxygen demand removal from FW reached 76% and 78% with the bacteria-pumice and bacteria-smectite nanocomposite treatments, respectively, and oil degradation in OFW reached 71% and 62%, respectively. Destructive changes did not occur, suggesting the durability of nanocomposites. The used nanocomposites had no impact on the lifespan of Moina macrocopa or water quality as assessed by toxicity analysis. Bacteria-pumice and bacteria-smectite nanocomposites are efficient for food waste recycling and do not require secondary treatment before being discharged into the environment.

Injectable, self-healing hydrogels based on gelatin, quaternized chitosan, and laponite as localized celecoxib delivery system for nucleus pulpous repair.

Utilization of injectable hydrogels stands as a paradigm of minimally invasive intervention in the context of intervertebral disc degeneration treatment. Restoration of nucleus pulposus (NP) function exerts a profound influence in alleviating back pain. This study introduces an innovative class of injectable shear-thinning hydrogels, founded on quaternized chitosan (QCS), gelatin (GEL), and laponite (LAP) with the capacity for sustained release of the anti-inflammatory drug, celecoxib (CLX). First, synthesis of Magnesium-Aluminum-Layered double hydroxide (LDH) was achieved through a co-precipitation methodology, as a carrier for celecoxib and a source of Mg ions. Intercalation of celecoxib within LDH layers (LDH-CLX) was verified through a battery of analytical techniques, including FTIR, XRD, SEM, EDAX, TGA and UV-visible spectroscopy confirmed a drug loading efficiency of 39.22 ± 0.09 % within LDH. Then, LDH-CLX was loaded in the optimal GEL-QCS-LAP hydrogel under physiological conditions. Release behavior (15 days profile), mechanical properties, swelling ratio, and degradation rate of the resulting composite were evaluated. A G* of 15-47 kPa was recorded for the hydrogel at 22-40 °C, indicating gel stability in this temperature range. Self-healing properties and injectability of the composite were proved by rheological measurements. Also, ex vivo injection into intervertebral disc of sheep, evidenced in situ forming and NP cavity filling behavior of the hydrogel. Support of GEL-QCS-LAP/LDH-CLX (containing mg2+ ions) for viability and proliferation (from ~94 % on day 1 to ~134 % on day 7) of NP cells proved using MTT assay, DAPI and Live/Dead assays. The hydrogel could significantly upregulate secretion of glycosaminoglycan (GAG, from 4.68 ± 0.1 to 27.54 ± 1.0 µg/ml), when LHD-CLX3% was loaded. We conclude that presence of mg2+ ion and celecoxib in the hydrogel can lead to creation of a suitable environment that encourages GAG secretion. In conclusion, the formulated hydrogel holds promise as a minimally invasive candidate for degenerative disc repair.

An Acid-Responsive Iron-Based Nanocomposite for OSCC Treatment.

Oral squamous cell carcinoma (OSCC) is the most common type of oral cancer, characterized by invasiveness, local lymph node metastasis, and poor prognosis. Traditional treatment and medications have limitations, making the specific inhibition of OSCC growth, invasion, and metastasis a challenge. The tumor microenvironment exhibits mildly acidity and high concentrations of H2O2, and its exploitation for cancer treatment has been widely researched across various cancers, but research in the oral cancer field is relatively limited. In this study, by loading ultra-small Prussian blue nanoparticles (USPBNPs) into mesoporous calcium-silicate nanoparticles (MCSNs), we developed an acid-responsive iron-based nanocomposite, USPBNPs@MCSNs (UPM), for the OSCC treatment. UPM demonstrated excellent dual enzyme activities, generating toxic ·OH in a mildly acidic environment, effectively killing OSCC cells and producing O2 in a neutral environment to alleviate tissue hypoxia. The results showed that UPM could effectively inhibit the proliferation, migration, and invasion of OSCC cells, as well as the growth of mice solid tumors, without obvious systemic toxicity. The mechanisms may involve UPM inducing ferroptosis of OSCC cells by downregulating the xCT/GPX4/glutathione (GSH) axis, characterized by intracellular iron accumulation, reactive oxygen species accumulation, GSH depletion, lipid peroxidation, and abnormal changes in mitochondrial morphology. Therefore, this study provides empirical support for ferroptosis as an emerging therapeutic target for OSCC and offers a valuable insight for future OSCC treatment.

Comparison of push-out bond strength and apical microleakage of different calcium silicate-based cements after using EDTA, chitosan and phytic acid irrigations.

This research was aimed to evaluate push-out bond strength and apical-microleakage after application of three different calcium silicate-based cements with irrigation solutions on simulated immature teeth. 40 maxillary permanent canine teeth were used for push-out bond strength test, and 120 maxillary permanent incisors were used for microleakage evaluation. 120 root slices were divided into four main groups (EDTA, Chitosan, Phytic acid, and Saline) and immersed these solutiouns according to irrigation procedures. Each irrigation group was divided into 3 subgroups (Biodentine, MTA Repair HP, and NeoPUTTY). The prepared teeth were divided into four groups according to irrigation procedure for microleakage test. EDTA irrigation with Biodentine group showed highest push-out bond strength value and saline group with Neoputty showed the lowest push-out bond strength value. The highest microleakage value was seen in saline group with MTA Repair HP, while the lowest microleakage value was observed chitosan with Biodentine group. Chitosan and phytic acid solutions can be recommended as an alternative irrigation solution to 17% EDTA in single-session apexification treatment, since they are non-toxic, naturally occurring materials, effectively remove the smear layer, and have a positive effect on bond strength and apical leakage. RESEARCH HIGHLIGHTS: One of the factors affecting the long-term success of root canal treatment is a hermetic seal. Non-hermetic or inadequate filling triggers a chronic inflammatory reaction in periapical tissues, causing fluids to enter the spaces and negatively affecting the success of the treatment. Therefore, this study will help clinicians choose the right biomaterial and irrigation solution that will affect the success of root canal treatment.

A comparative in vitro and in vivo analysis of the impact of copper substitution on the cytocompatibility, osteogenic, and angiogenic properties of a borosilicate bioactive glass.

The 0106-B1-bioactive glass (BG) composition (in wt %: 37.5 SiO2, 22.6 CaO, 5.9 Na2O, 4.0 P2O5, 12.0 K2O, 5.5 MgO, and 12.5 B2O3) has demonstrated favorable processing properties and promising bone regeneration potential. The present study aimed to evaluate the biological effects of the incorporation of highly pro-angiogenic copper (Cu) in 0106-B1-BG in vitro using human bone marrow-derived mesenchymal stromal cells (BMSCs) as well as its in vivo potential for bone regeneration. CuO was added to 0106-B1-BG in exchange for CaO, resulting in Cu-doped BG compositions containing 1.0, 2.5 and 5.0 wt % CuO (composition in wt %: 37.5 SiO2, 21.6/ 20.1/17.6 CaO, 5.9 Na2O, 4.0 P2O5, 12.0 K2O, 5.5 MgO, 12.5 B2O3, and 1.0/ 2.5/ 5.0 CuO). In vitro, the BGs' impact on the viability, proliferation, and growth patterns of BMSCs was evaluated. Analyses of protein secretion, matrix formation, and gene expression were used for the assessment of the BGs' influence on BMSCs regarding osteogenic differentiation and angiogenic stimulation. The presence of Cu improved cytocompatibility, osteogenic differentiation, and angiogenic response when compared with unmodified 0106-B1-BG in vitro. In vivo, a critical-size femoral defect in rats was filled with scaffolds made from BGs. Bone regeneration was evaluated by micro-computed tomography. Histological analysis was performed to assess bone maturation and angiogenesis. In vivo effects regarding defect closure, presence of osteoclastic cells or vascular structures in the defect were not significantly changed by the addition of Cu compared with undoped 0106-B1-BG scaffolds. Hence, while the in vitro properties of the 0106-B1-BG were significantly improved by the incorporation of Cu, further evaluation of the BG composition is necessary to transfer these effects to an in vivo setting.

Development of a tannic acid- and silicate ion-functionalized PVA-starch composite hydrogel for in situ skeletal muscle repairing.

The repair capacity of skeletal muscle is severely diminished in massive skeletal muscle injuries accompanied by inflammation, resulting in muscle function loss and scar tissue formation. In the current work, we developed a tannic acid (TA)- and silicate ion-functionalized tissue adhesive poly(vinyl alcohol) (PVA)-starch composite hydrogel, referred to as PSTS (PVA-starch-TA-SiO32-). It was formed based on the hydrogen bonding of TA to organic polymers, as well as silicate-TA ligand interaction. PSTS could be gelatinized in minutes at room temperature with crosslinked network formation, making it applicable for injection. Further investigations revealed that PSTS had skeletal muscle-comparable conductivity and modulus to act as a temporary platform for muscle repairing. Moreover, PSTS could release TA and silicate ions in situ to inhibit bacterial growth, induce vascularization, and reduce oxidation, paving the way to the possibility of creating a favorable microenvironment for skeletal muscle regeneration and tissue fibrosis control. The in vivo model confirmed that PSTS could enhance muscle fiber regeneration and myotube formation, as well as reduce infection and inflammation risk. These findings thereby implied the great potential of PSTS in the treatment of formidable skeletal muscle injuries.

Premixed calcium silicate-based ceramic sealers promote osteogenic/cementogenic differentiation of human periodontal ligament stem cells: A microscopy study.

To evaluate the effects of premixed calcium silicate based ceramic sealers on the viability and osteogenic/cementogenic differentiation of human periodontal ligament stem cells (hPDLSCs). The materials evaluated were TotalFill BC Sealer (TFbc), AH Plus Bioceramic Sealer (AHPbc), and Neosealer Flo (Neo). Standardized discs and 1:1, 1:2, and 1:4 eluates of the tested materials were prepared. The following in vitro experiments were carried out: ion release, cell metabolic activity 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, cell migration, immunofluorescence experiment, cell attachment, gene expression, and mineralization assay. Statistical analyses were performed using one-way ANOVA followed by Tukey's post hoc test (p < .05). Increased Ca2+ release was detected in TFbc compared to AHPbc and Neo (*p < .05). Biological assays showed a discrete cell metabolic activity and cell migration in Neo-treated cell, whereas scanning electronic microscopy assay exhibited that TFbc group had a better cell adhesion process of substrate attachment, spreading, and cytoskeleton development on the niche-like structures of the cement than AHPbc and Neo. The sealers tested were able to induce overexpression of the CEMP-1, ALP, and COL1A1 genes in the first days of exposure, particularly in the case of TFbc (***p < .001). All materials tested significantly increased the mineralization of hPDLSCs when compared to the negative control, although more pronounced calcium deposition was observed in the TFbc-treated cells (***p < .001). Our results suggested that TFbc promotes cell differentiation, both by increasing the expression of key osteo/odontogenic genes and by promoting mineralization of the extracellular matrix, whereas this phenomenon was less evident in Neo and AHPbc. RESEARCH HIGHLIGHTS: TFbc group had a better cell adhesion process of substrate attachment, spreading, and cytoskeleton development on the niche-like structures of the cement than AHPbc and Neo. The sealers tested were able to induce overexpression of the CEMP-1, ALP, and COL1A1 genes in the first days of exposure, particularly in the case of TFbc. All materials tested significantly increased the mineralization of hPDLSCs when compared to the negative control, although more pronounced calcium deposition was observed in the TFbc-treated cells.

Biocompatibility and bioactive potential of NeoPUTTY calcium silicate-based cement: An in vivo study in rats.

AIM: To evaluate the inflammatory reaction and the ability to induce mineralization activity of a new repair material, NeoPUTTY (NPutty; NuSmile, USA), in comparison with Bio-C Repair (BC; Angelus, Brazil) and MTA Repair HP (MTA HP; Angelus, Brazil). METHODOLOGY: Polyethylene tubes were filled with materials or kept empty (control group, CG) and implanted in subcutaneous tissue of rats for 7, 15, 30, and 60 days (n = 6/group). Capsule thickness, number of inflammatory cells (ICs), fibroblasts, collagen content, and von Kossa analysis were performed. Unstained sections were evaluated under polarized light and by immunohistochemistry for osteocalcin (OCN). Data were submitted to two-way anova followed by Tukey's test (p ≤ .05), except for OCN. OCN data were submitted to Kruskal-Wallis and Dunn and Friedman post hoc tests followed by the Nemenyi test at a significance level of 5%. RESULTS: At 7, 15, and 30 days, thick capsules containing numerous ICs were seen around the materials. At 60 days, a moderate inflammatory reaction was observed for NPutty, BC while MTA HP presented thin capsules with moderate inflammatory cells. In all periods, NPutty specimens contained the highest values of ICs (p < .05). From 7 to 60 days, the number of ICs reduced significantly while an increase in the number of fibroblasts and birefringent collagen content was observed. At 7 and 15 days, no significant difference was observed in the immunoexpression of OCN (p > .05). At 30 and 60 days, NPutty showed the lowest values of OCN (p < .05). At 60 days, a similar immunoexpression was observed for BC and MTA HP (p > .05). In all time intervals, capsules around NPutty, BC, and MTA HP showed von Kossa-positive and birefringent structures. CONCLUSIONS: Despite the greater inflammatory reaction promoted by NeoPutty than BC and MTA HP, the reduction in the thickness of capsules, the increase in the number of fibroblasts, and the reduction in the number of ICs indicate that this bioceramic material is biocompatible Furthermore, NeoPutty presents the ability to induce mineralization activity.

Mineral matrix deposition of MC3T3-E1 pre-osteoblastic cells exposed to silicocarnotite and nagelschmidtite bioceramics: In vitro comparison to hydroxyapatite.

This work presents the effect of the silicocarnotite (SC) and nagelschmidtite (Nagel) phases on in vitro osteogenesis. The known hydroxyapatite of biological origin (BHAp) was used as a standard of osteoconductive characteristics. The evaluation was carried out in conventional and osteogenic media for comparative purposes to assess the osteogenic ability of the bioceramics. First, the effect of the material on cell viability at 24 h, 7 and 14 days of incubation was evaluated. In addition, cell morphology and attachment on dense bioceramic surfaces were observed by fluorescence microscopy. Specifically, alkaline phosphatase (ALP) activity was evaluated as an osteogenic marker of the early stages of bone cell differentiation. Mineralized extracellular matrix was observed by calcium phosphate deposits and extracellular vesicle formation. Furthermore, cell phenotype determination was confirmed by scanning electron microscope. The results provided relevant information on the cell attachment, proliferation, and osteogenic differentiation processes after 7 and 14 days of incubation. Finally, it was demonstrated that SC and Nagel phases promote cell proliferation and differentiation, while the Nagel phase exhibited a superior osteoconductive behavior and could promote MC3T3-E1 cell differentiation to a higher extent than SC and BHAp, which was reflected in a higher number of deposits in a shorter period for both conventional and osteogenic media.

"Biological responses of two calcium-silicate-based cements on a tissue-engineered 3D organotypic deciduous pulp analogue".

OBJECTIVES: The biological responses of MTA and Biodentine™ has been assessed on a three-dimensional, tissue-engineered organotypic deciduous pulp analogue. METHODS: Human endothelial (HUVEC) and dental mesenchymal stem cells (SHED) at a ratio of 3:1, were incorporated into a collagen I/fibrin hydrogel; succeeding Biodentine™ and MTA cylindrical specimens were placed in direct contact with the pulp analogue 48 h later. Cell viability/proliferation and morphology were evaluated through live/dead staining, MTT assay and Scanning Electron Microscopy (SEM), and expression of angiogenic, odontogenic markers through real time PCR. RESULTS: Viable cells dominated at day 3 after treatment presenting typical morphology, firmly attached within the hydrogel structures, as shown by live/dead staining and SEM images. MTT assay at day 1 presented a significant increase of cell proliferation in Biodentine™ group. Real-time PCR showed significant upregulation of odontogenic markers DSPP, BMP-2 (day 3,6), RUNX2, ALP (day 3) in contact with Biodentine™ compared to MTA and the control, whereas MTA promoted significant upregulation of DSPP, BMP-2, RUNX2, Osterix (day 3) and ALP (day 6) compared to the control. MSX1 presented downregulation in both experimental groups. Expression of angiogenic markers VEGFa and ANGPT-1 at day 3 was significantly upregulated in contact with Biodentine™ and MTA respectively, while the receptors VEGFR1, VEGFR2 and Tie-2, as well as PECAM-1 were downregulated. SIGNIFICANCE: Both calcium silicate-based materials are biocompatible and exert positive angiogenic and odontogenic effects, although Biodentine™ during the first days of culture, seems to induce higher cell proliferation and provoke a more profound odontogenic and angiogenic response from SHED.

Effects of vehicles on the physical properties and biocompatibility of premixed calcium silicate cements.

Premixed calcium silicate cements (pCSCs) contain vehicles which endow fluidity and viscosity to CSCs. This study aimed to investigate the effects of three vehicles, namely, polyethylene glycol (PEG), propylene glycol (PG), and dimethyl sulfoxide (DMSO), on the physicochemical properties and biocompatibility of pCSCs. The setting time, solubility, expansion rate, and mechanical strength of the pCSCs were evaluated, and the formation of calcium phosphate precipitates was assessed in phosphate-buffered saline (PBS). The effects of pCSC extracts on the osteogenic differentiation of mesenchymal stem cells (MSCs) were investigated. Finally, the tissue compatibility of pCSCs in rat femurs was observed. CSC containing PEG (CSC-PEG) exhibited higher solubility and setting time, and CSC-DMSO showed the highest expansion rate and mechanical strength. All pCSCs generated calcium phosphate precipitates. The extract of CSC-PG induced the highest expressions of osteogenic markers along with the greatest calcium deposites. When implanted in rat femurs, CSC-PEG was absorbed considerably, whereas CSC-PG remained relatively unaltered inside the femur.