Biomineralization potential and biological properties of a new tantalum oxide (Ta2O5)-containing calcium silicate cement.

OBJECTIVE: The present study evaluated the biological effects and biomineralization potential of a new tantalum oxide (Ta2O5)-containing material designed for vital pulp therapy or perforation repair (NeoMTA 2), compared to NeoMTA Plus and Bio-C Repair. MATERIAL AND METHODS: Human dental pulp stem cells (hDPSCs) were exposed to different eluates from NeoMTA Plus, NeoMTA 2, and Bio-C Repair. Ion release from each material was determined using inductively coupled plasma-optical emission spectrometry (ICP-MS). The biological experiments performed were MTT assays, apoptosis/necrosis assays, adhesion assays, migration assays, morphology evaluation, and reactive oxygen species (ROS) production analysis. Biomineralization was assessed by Alizarin red S staining. Finally, osteo/odontogenic gene expression was determined by real-time quantitative reverse-transcriptase polymerase chain reaction (RT-qPCR). Data were analyzed using one-way ANOVA followed by Tukey's multiple comparison test. RESULTS: NeoMTA 2 displayed a significantly higher calcium release compared to the other materials (p < 0.05). When hDPSCs were cultured in presence of the different material eluates, all groups exhibited similar hDPSC viability and migration rates when compared to untreated cells. Substantial cell attachment and spreading were observed in all materials' surfaces, without significant differences. hDPSCs treated with NeoMTA 2 displayed an upregulation of ALP, Col1A1, RUNX2 (p < 0.001), ON, and DSPP genes (p < 0.05), and showed the highest mineralization potential compared to other groups (p < 0.001). Finally, the more concentrated eluates from these materials, specially NeoMTA Plus and NeoMTA 2, promoted higher ROS production in hDPSCs compared to Bio-C Repair and control cells (p < 0.001), although these ROS levels did not result in increased cell death. CONCLUSIONS: The new tantalum oxide (Ta2O5)-containing material shows an adequate cytocompatibility and the ability to promote biomineralization without using chemical osteogenic inducers, showing great potential as a new material for vital pulp therapy. CLINICAL RELEVANCE: NeoMTA 2 seems to be a promising material for vital pulp therapy. Further studies considering its biocompatibility and biomineralization potential are necessary.

Cytocompatibility and bioactive properties of the new dual-curing resin-modified calcium silicate-based material for vital pulp therapy.

OBJECTIVE: The aim of the present study was to evaluate the in vitro biocompatibility of Theracal PT, Theracal LC, and MTA Angelus, considered as bioactive materials used for vital pulp treatment, on human dental pulp stem cells (hDPSCs). MATERIALS AND METHODS: Human dental pulp stem cells (hDPSCs) were isolated from third molars, and material eluates were prepared (undiluted, 1:2, and 1:4 ratios). The hDPSC cytotoxicity, adhesion, morphology, viability, and cell migration were assessed. The mineralization nodule formation was determined by Alizarin red S staining (ARS). The odonto/osteogenic differentiation potential was assessed by osteo/odontogenic marker expression real-time qPCR. The chemical composition and ion release of the vital pulp materials were determined by energy dispersive X-ray (EDX) and inductively coupled plasma-mass spectrometry (ICP-MS), respectively. Statistical differences were assessed by ANOVA and Tukey's test (p < 0.05). RESULTS: The three vital pulp materials showed variable levels of calcium, tungsten, silicon, and zirconium release and in their chemical composition. Cytocompatibility assays revealed higher hDPSC viability and migration rates when treated with Theracal PT than with Theracal LC. The lowest cell adhesion and spreading were observed in all Theracal LC-treated groups, whereas the highest were observed when treated with MTA. Theracal PT and MTA promoted the upregulation of DSPP and RUNX2 gene expression (p < 0.05). After 21 days, both MTA Angelus and Theracal PT-treated cells exhibited a significantly higher mineralized nodule formation than the negative control (p < 0.05). CONCLUSIONS: This study demonstrates the favorable in vitro cytocompatibility and bioactive properties of the recently introduced Theracal PT and the well-established MTA Angelus on hDPSCs, as opposed to Theracal LC. More studies, including in vivo animal testing are suggested before these new formulations might be used in the clinical setting. CLINICAL RELEVANCE: Theracal PT is a new material that could be clinically suitable for vital pulp therapy. Further studies considering its biocompatibility and bioactivity are necessary.

Chemical composition and bioactivity potential of the new Endosequence BC Sealer formulation HiFlow.

AIM: To evaluate in a laboratory setting the effects of Endosequence BC Sealer HiFlow (Brasseler USA, Savannah, GA, USA), a novel calcium silicate-based sealer developed for use in warm canal filling techniques, on human periodontal ligament stem cells (hPDLSCs). METHODOLOGY: Eluates of EndoSequence BC Sealer HiFlow (BCHiF) (Brasseler USA), EndoSequence BC Sealer (BCS) (Brasseler USA) and AH Plus (AHP) (Dentsply DeTrey GmbH, Konstanz, Germany) were placed in contact with hPDLSCs. The characterization of the chemical elements of the root canal sealers was assessed using scanning electron microscopy and energy-dispersive X-ray analysis (SEM-EDX). Inductively coupled plasma-mass spectrometry (ICP-MS) was used to determine the ion release of the sealers. MTT assay and wound healing techniques were used to determine cell viability and migration, respectively. Cell morphology and cell attachment were assessed using a direct contact technique of hPDLSCs onto the surface of the sealers and analysed by SEM. The bioactivity potential was carried out with the Alizarin Red and qPCR testing methods. The statistical differences were evaluated using one-way anova and Tukey's test (P < 0.05). RESULTS: ICP-MS and EDX revealed significantly more zirconium in BCHiF than BCS (P < 0.05), whereas BCS had slightly higher levels of Ca2+ than BCHiF (P < 0.05). The cell viability assay revealed no relevant differences between BCS and BCHiF when compared with the control group (P > 0.05). Both BCS and BCHiF had similar rates of cell migration to the control group at 24 and 48 h. Cell morphology and adhesion capacity were also similar for BCS and BCHiF groups, whilst the AHP group was associated with reduced adhesion capacity. The Alizarin Red assay revealed a significant difference between the BCS and the control group (P < 0.001), as well as for the BCHiF group (P < 0.001). Finally, BCS and BCHiF promoted overexpression of osteo/cementogenic genes. CONCLUSIONS: In general, EndoSequence BC Sealer HiFlow possesses suitable biological properties to be safely used as a root canal filling material and promote increased expression of oste/cementogenic genes by hPDLSCs.

Cytocompatibility, bioactivity potential, and ion release of three premixed calcium silicate-based sealers.

OBJECTIVE: Compositional modifications may alter the biological and physicochemical characteristics of calcium silicate-based sealers (CSBS) and, ultimately, their bioactivity. The main objective of this study was to evaluate the biological properties of three CSBS: EndoSequence BC Sealer, Ceraseal, and Endoseal mineral trioxide aggregate. MATERIALS AND METHODS: Human periodontal ligament stem cells (hPDLSCs) were exposed to several eluates of CSBS. The ion release profile and pH were determined, and metabolic activity and cell migration were assessed using the MTT and wound healing assays. hPDLSCs were cultured in direct contact with the surface of each material, and cell morphology and attachment were analyzed by scanning electron microscopy (SEM). Bioactivity potential was assessed by RT-qPCR and mineralization assays. Statistical differences between biomaterials were assessed using one- or two-way ANOVA (α < 0.05). RESULTS: All materials showed an alkaline pH, although Endoseal exhibited a significantly higher pH compared with the other CSBS (p < 0.05). Ceraseal released significantly more Ca2+ (p < 0.05) than EndoSequence BC Sealer and Endoseal. Interestingly, Endoseal induced a significant reduction in cell viability and cell migration compared with the control (p < 0.001). Moreover, SEM showed abundant cells adhering to EndoSequence BC Sealer and Ceraseal surfaces, whereas very few round cells were detected on the surface of Endoseal. Finally, Ceraseal and EndoSequence induced ALP, CAP, and CEMP-1 expression and a significantly higher mineralization capacity than Endoseal (***p < 0.001). CONCLUSIONS: The eluates from EndoSequence BC Sealer and Ceraseal displayed higher cell viability, cell attachment, cell migration rates, and ion release rates than Endoseal. Ceraseal and EndoSequence BC Sealer exhibited significantly more gene expression and mineralization capacity than Endoseal. CLINICAL RELEVANCE: The results obtained in the present work suggest that EndoSequence BC Sealer and Ceraseal possess biological properties that make them suitable materials for root canal treatment.

Evaluation of changes in ion release and biological properties of NeoMTA-Plus and Endocem-MTA exposed to an acidic environment.

AIM: To analyse in vitro changes in ion release and biological properties of Endocem-MTA (Maruchi, Wonju, Korea) and NeoMTA-Plus (Avalon Biomed Inc, Bradenton, FL, USA) exposed to acidic or neutral environment on human dental periodontal ligament stem cells (hPDLSCs). METHODOLOGY: Cell viability and wound healing assays were performed using eluates of each material. Cell death and changes in phenotype induced by the set endodontic sealer eluates were evaluated through flow cytometry. To evaluate cell attachment to the different materials, hPDLSCs were directly seeded onto the material surfaces and analysed by scanning electron microscopy. The chemical composition of the materials was determined by energy-dispersive X-ray (EDX), and ion release was evaluated by inductively coupled plasma-mass spectrometry. Statistical analysis was performed with analysis of variance and a Bonferroni or Tukey post-test (α < 0.05). RESULTS: The MTT assay revealed non-cytotoxic effects of NeoMTA-Plus and Endocem-MTA at pH 5.2 and 7.4. However, there were minor differences compared with the control, especially at pH 5.2, where both materials were associated with significantly greater cell viability (P < 0.05). In both environments, the materials stimulated hPDLSCs to migrate. hPDLSCs were attached to the bioactive cements, with multiple prolongations proliferated on the surface of the samples. Moreover, there were no changes to cell phenotype or apoptosis/necrosis rates, indicating that the acidic environment did not induce cell death. Prismatic crystalline structures were seen on the surface of the cements exposed to butyric acid and EDX analysis identified a marked peak of Ca2+ from NeoMTA-Plus and Endocem-MTA in acidic and physiological environments. CONCLUSIONS: An acidic environment favoured the release of Ca2+ ions from both bioactive cements, and the cytotoxicity of these bioactive cements was low in both environments studied.

Biological effects of silk fibroin 3D scaffolds on stem cells from human exfoliated deciduous teeth (SHEDs).

The aim is to investigate in vitro biological effects of silk fibroin 3D scaffolds on stem cells from human exfoliated deciduous teeth (SHEDs) in terms of proliferation, morphological appearance, cell viability, and expression of mesenchymal stem cell markers. Silk fibroin 3D scaffolding materials may represent promising suitable scaffolds for their application in regenerative endodontic therapy approaches. SHEDs were cultured in silk fibroin 3D scaffolds. Then, cell numbers were counted and the Alamar blue colorimetric assay was used to analyse cell proliferation after 24, 48, 72, and 168 h of culture. The morphological features of SHEDs cultured on silk fibroin scaffolds were evaluated by scanning electron microscopy (SEM). Finally, cell viability and the expression of mesenchymal stem cell markers were analysed by flow cytometry. One-way analysis of variance (ANOVA) followed by a Bonferroni post-test was performed (P < 0.05). At 24 and 48 h of culture, SHED proliferation on scaffolds was modest compared to the control although still significant (p < 0.05). However, cell proliferation progressively increased from 72 to 168 h compared with the control (p < 0.001; p < 0.01). In addition, flow cytometry analysis showed that the culture of SHEDs on silk fibroin scaffolds did not significantly alter the level of expression of the mesenchymal markers CD73, CD90, or CD105 up to 168 h; in addition, cell viability in silk fibroin was similar to than obtained in plastic. Moreover, SEM studies revealed a suitable degree of proliferation, cell spreading, and attachment, especially after 168 h of culture. The findings from the current study suggest that silk fibroin 3D scaffolds had a favourable effect on the biological responses of SHEDs. Further in vivo investigations are required to confirm these results.

On the presence and immunoregulatory functions of extracellular microRNAs in the trematode Fasciola hepatica.

Liver flukes represent a paraphyletic group of endoparasitic flatworms that significantly affect man either indirectly due to economic damage on livestock or directly as pathogens. A range of studies have focussed on how these macroscopic organisms can evade the immune system and live inside a hostile environment such as the mammalian liver and bile ducts. Recently, microRNAs, a class of short noncoding gene regulators, have been proposed as likely candidates to play roles in this scenario. MicroRNAs (miRNAs) are key players in development and pathogenicity and are highly conserved between metazoans: identical miRNAs can be found in flatworms and mammalians. Interestingly, miRNAs are enriched in extracellular vesicles (EVs) which are secreted by most cells. EVs constitute an important mode of parasite/host interaction, and recent data illustrate that miRNAs play a vital part. We have demonstrated the presence of miRNAs in the EVs of the trematode species Dicrocoelium dendriticum and Fasciola hepatica (Fhe) and identified potential immune-regulatory miRNAs with targets in the host. After our initial identification of miRNAs expressed by F. hepatica, an assembled genome and additional miRNA data became available. This has enabled us to update the known complement of miRNAs in EVs and speculate on potential immune-regulatory functions that we review here.

Evaluation of cytocompatibility of calcium silicate-based endodontic sealers and their effects on the biological responses of mesenchymal dental stem cells.

AIM: To investigate in vitro the cytocompatibility of the calcium silicate-containing endodontic sealers MTA Fillapex and TotalFill BC Sealer on human periodontal ligament stem cells (hPDLSCs) by assaying their biological responses and compare them with that observed when using an epoxy resin-based sealer (AH Plus). METHODOLOGY: Specimens from the three different endodontic sealers were eluated with culture medium for 24 h. The cytotoxicity of these eluates was evaluated using the MTT assay. In addition, an in vitro scratch wound healing model was used to determine their effects on cell migration. Cell adhesion to collagen type I after treatment with the different sealer eluates was also measured, whereas cytotoxicity was determined using the DNA-specific fluorochrome Hoechst 33342. Finally, to assess cell morphology and attachment to the different sealers, hPDLSCs were directly seeded onto the material surfaces and analysed by scanning electron microscopy (SEM). One-way analysis of variance (anova) followed by a Bonferroni post-test were performed (P < 0.05). RESULTS: hPDLSCs exposed to different dilutions of TotalFill BC Sealer eluates had significantly higher cell proliferation compared with that observed when cells were treated with AH Plus and MTA Fillapex eluates (P < 0.001). In addition, TotalFill eluates were associated with significantly increased cell adhesion to collagen type I and migration of hPDLSCs in a concentration-dependent manner than displayed after treatment with MTA Fillapex or AH Plus eluates (P < 0.001). Moreover, TotalFill BC Sealer-induced cytotoxicity was significantly lower than observed using AH Plus and MTA Fillapex eluates (P < 0.001). Finally, SEM studies revealed suitable proliferation, cell spreading and attachment, especially when using TotalFill BC Sealer discs. CONCLUSION: TotalFill BC Sealer exhibited a higher cytocompatibility than AH Plus and MTA Fillapex. Further investigations using in vivo animal models are required to validate the potential biological responses of TotalFill BC Sealer on hPDLSCs.

Biocompatibility of three new calcium silicate-based endodontic sealers on human periodontal ligament stem cells.

AIM: To evaluate the biocompatibility of three calcium silicate-based endodontic sealers, Bioroot BC Sealer (Septodont, Saint-Maur-des-Fosses, France), Endoseal MTA (EndoSeal, Maruchi, Seoul, Korea) and Nano-ceramic Sealer (B&L Biotech, Fairfax, VA, USA) (NCS), on human periodontal ligament stem cells (hPDLSCs). METHODOLOGY: Human periodontal ligament stem cells were cultured in the presence of various endodontic sealer eluates for 24 h. Cell viability was determined using the MTT assay. Cell death and changes in phenotype induced by the set endodontic sealer eluates were evaluated through flow cytometry. Also, an in vitro scratch wound-healing model was used to determine their effects in cell migration. Finally, to assess cell morphology and attachment to the different sealers, hPDLSCs were directly seeded onto the material surfaces and analysed by scanning electron microscopy (SEM). One-way analysis of variance (anova) followed by a Bonferroni post-test was performed (P < 0.05). RESULTS: At 24 h, cell spreading was evident in the presence of Bioroot BC Sealer (BR) and Nano-ceramic Sealer (NCS), but not Endoseal MTA (ES). At 72 h, BR and NCS exhibited high and moderate cell proliferation, respectively, whereas ES revealed low rates of cell proliferation (P < 0.05). Similar results were obtained in a cell death assay. In addition, hPDLSCs maintained their mesenchymal phenotype in all conditions although their capacity to migrate was higher in the presence of BR. Finally, SEM studies revealed a good degree of proliferation, cell spreading and attachment, especially when using BR and NCS discs. CONCLUSIONS: BR and NCS were associated with better cytocompatibility than ES. Further in vitro and in vivo investigations are required to confirm the suitability of these calcium silicate-based endodontic sealers for clinical application.

Cytotoxicity and bioactivity of various pulpotomy materials on stem cells from human exfoliated primary teeth.

AIMS: To investigate the cytotoxicity and bioactivity of several pulpotomy materials: Biodentine (Septodont, Saint-Maur-des-Fosses, France) MTA (Angelus, Londrina, PR, Brazil), Theracal LC (Bisco Inc., Schamburg, IL, USA) and IRM (Dentsply DeTrey GmbH, Konstanz, Germany), after contact with stem cells isolated from human exfoliated primary teeth (SHEDs). METHODOLOGY: SHEDs were cultured in the presence of the eluates of various pulpotomy materials for 24, 48 and 72 h. Cell viability was determined by mitochondrial dehydrogenase enzymatic (MTT) assay. Apoptosis and changes in cell phenotype were evaluated by flow cytometry. Also, an in vitro scratch wound-healing assay was used to determine their effects on cell migration. To assess cell morphology and attachment to the different pulpotomy materials, SHEDs were directly seeded onto the material surfaces and analysed by scanning electron microscopy (SEM). Finally, the deposition of a calcified matrix in presence of these materials was verified by Alizarin Red staining. Statistical analysis was performed with analysis of variance and Bonferroni or Tukey post-test (α = 0.05). RESULTS: Cell viability in the presence of Biodentine eluates was significantly higher to that obtained using complete medium alone (control; P < 0.01) and was also significantly higher than using MTA Angelus from 48 h of incubation (P < 0.01). However, Theracal LC and IRM were associated with low rates of cell viability (P < 0.001). Similar results were obtained in an apoptosis assay. In addition, SHEDs maintained their mesenchymal phenotype in all conditions although their capacity to migrate was higher in the presence of Biodentine. SEM studies revealed a suitable proliferation rate, cell spreading and attachment, especially when using Biodentine and MTA Angelus discs. Finally, Biodentine eluates significantly induced calcified matrix deposition from 7 days of culture (P < 0.01). CONCLUSIONS: Biodentine exhibited better cytocompatibility and bioactivity than MTA Angelus, Theracal LC and IRM.

Comparative analysis of the biological effects of the endodontic bioactive cements MTA-Angelus, MTA Repair HP and NeoMTA Plus on human dental pulp stem cells.

AIM: To evaluate the biological effects in vitro of MTA-Angelus (MTA-Ang; Angelus, Londrina, PR, Brazil), MTA Repair HP (MTA-HP; Angelus) and NeoMTA Plus (NeoMTA-P; Avalon Biomed Inc, Bradenton, FL, USA) on human dental pulp stem cells (hDPSCs). METHODOLOGY: Cell viability and cell migration assays were performed using eluates of each material. To evaluate cell morphology and cell attachment to the different materials, hDPSCs were directly seeded onto the material surfaces and analysed by immunocytofluorescence and scanning electron microscopy, respectively. The chemical composition of the materials was determined by energy-dispersive X-ray (EDX), and eluates were analysed by inductively coupled plasma-mass spectrometry (ICP-MS). Statistical analysis was performed with the analysis of variance and Bonferroni or Tukey post-test (α < 0.05). RESULTS: Undiluted MTA-Ang, MTA-HP and NeoMTA-P displayed a significant increase in cell viability greater than that obtained using complete medium alone (control) (*P < 0.05; **P < 0.01; ***P < 0.001). Moreover, a cell migration assay revealed cell migration rates after incubation with extracts of MTA-Ang, MTA-HP and NeoMTA-P that were similar to levels obtained in the control group. In addition, stretched cytoskeletal F-actin fibres were detected in the cells treated with the three material extracts. SEM studies revealed a high degree of cell proliferation and attachment on all three materials. EDX analysis demonstrated similar weight percentages of C, O and Ca in all three materials, whilst other elements such as Al, Si and S were also found. CONCLUSIONS: MTA-Ang, MTA-HP and NeoMTA-P were associated with biological effects on hDPSCs in terms of cell proliferation, morphology, migration and attachment.

Angiotensin II induces Fat1 expression/activation and vascular smooth muscle cell migration via Nox1-dependent reactive oxygen species generation.

Fat1 is an atypical cadherin that controls vascular smooth muscle cell (VSMC) proliferation and migration. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 1 (Nox1) is an important source of reactive oxygen species (ROS) in VSMCs. Angiotensin II (Ang II) induces the expression and/or activation of both Fat1 and Nox1 proteins. This study tested the hypothesis that Ang II-induced Fat1 activation and VSMC migration are mediated by Nox1-dependent ROS generation and redox signaling. Studies were performed in cultured VSMCs from Sprague­Dawley rats. Cells were treated with Ang II (1 µmol/L) for short (5 to 30 min) or long term stimulations (3 to 12 h) in the absence or presence of the antioxidant apocynin (10 µmol/L), extracellular-signal-regulated kinases 1/2 (Erk1/2) inhibitor PD98059 (1 µmol/L), or Ang II type 1 receptor (AT1R) valsartan (1 µmol/L). siRNA was used to knockdown Nox1 or Fat1. Cell migration was determined by Boyden chamber assay. Ang II increased Fat1 mRNA and protein levels and promoted Fat1 translocation to the cell membrane, responses that were inhibited by AT1R antagonist and antioxidant treatment. Downregulation of Nox1 inhibited the effects of Ang II on Fat1 protein expression. Nox1 protein induction, ROS generation, and p44/p42 MAPK phosphorylation in response to Ang II were prevented by valsartan and apocynin, and Nox1 siRNA inhibited Ang II-induced ROS generation. Knockdown of Fat1 did not affect Ang II-mediated increases in Nox1 expression or ROS. Inhibition of p44/p42 MAPK phosphorylation by PD98059 abrogated the Ang II-induced increase in Fat1 expression and membrane translocation. Knockdown of Fat1 inhibited Ang II-induced VSMC migration, which was also prevented by valsartan, apocynin, PD98059, and Nox1 siRNA. Our findings indicate that Ang II regulates Fat1 expression and activity and induces Fat1-dependent VSMC migration via activation of AT1R, ERK1/2, and Nox1-derived ROS, suggesting a role for Fat1 downstream of Ang II signaling that leads to vascular remodeling.

Effects of composite films of silk fibroin and graphene oxide on the proliferation, cell viability and mesenchymal phenotype of periodontal ligament stem cells.

In regenerative dentistry, stem cell-based therapy often requires a scaffold to deliver cells and/or growth factors to the injured site. Graphene oxide (GO) and silk fibroin (SF) are promising biomaterials for tissue engineering as they are both non toxic and promote cell proliferation. On the other hand, periodontal ligament stem cells (PDLSCs) are mesenchymal stem cells readily accessible with a promising use in cell therapy. The purpose of this study was to investigate the effects of composite films of GO, SF and GO combined with fibroin in the mesenchymal phenotype, viability, adhesion and proliferation rate of PDLSCs. PDLSCs obtained from healthy extracted teeth were cultured on GO, SF or combination of GO and SF films up to 10 days. Adhesion level of PDSCs on the different biomaterials were evaluated after 12 h of culture, whereas proliferation rate of cells was assessed using the MTT assay. Level of apoptosis was determined using Annexin-V and 7-AAD and mesenchymal markers expression of PDLSCs were analyzed by flow cytometry. At day 7 of culture, MTT experiments showed a high rate of proliferation of PDLSCs growing on GO films compared to the other tested biomaterials, although it was slightly lower than in plastic (control). However PDLSCs growing in fibroin or GO plus fibroin films showed a discrete proliferation. Importantly, at day 10 of culture it was observed a significant increase in PDLSCs proliferation rate in GO films compared to plastic (P < 0.05), as well as in GO plus fibroin compared to fibroin alone (P < 0.001). Flow cytometry analysis showed that culture of PDLSCs in fibroin, GO or GO plus fibroin films did not significantly alter the level of expression of the mesenchymal markers CD73, CD90 or CD105 up to 168 h, being the cell viability in GO even better than obtained in plastic. Our findings suggest that the combination of human dental stem cells/fibroin/GO based-bioengineered constructs have strong potential for their therapeutic use in regenerative dentistry.

Therapeutic effect of adipose-derived mesenchymal stem cells in a porcine model of abdominal sepsis.

BACKGROUND: The term sepsis refers to a complex and heterogeneous syndrome. Although great progress has been made in improving the diagnosis and treatment of this condition, it continues to have a huge impact on morbidity and mortality worldwide. Mesenchymal stem cells are a population of multipotent cells that have immunomodulatory properties, anti-apoptotic effects, and antimicrobial activity. We studied these capacities in a porcine model of peritoneal sepsis. METHODS: We infused human adipose-derived mesenchymal stem cells (ADSCs) into a porcine model of peritoneal sepsis. Twenty piglets were treated with antibiotics alone (control group) or antibiotics plus peritoneal infusion of ADSCs at a concentration of 2 × 106 cells/kg or 4 × 106 cells/kg (low- and high-dose experimental groups, respectively). The animals were evaluated at different time points to determine their clinical status, biochemical and hematologic parameters, presence of inflammatory cytokines and chemokines in blood and peritoneal fluid, and finally by histologic analysis of the organs of the peritoneal cavity. RESULTS: One day after sepsis induction, all animals presented peritonitis with bacterial infection as well as elevated C-reactive protein, haptoglobin, IL-1Ra, IL-6, and IL-1b. Xenogeneic ADSC infusion did not elicit an immune response, and peritoneal administration of the treatment was safe and feasible. One day after infusion, the two experimental groups showed a superior physical condition (e.g., mobility, feeding) and a significant increase of IL-10 and TGF-ß in blood and a decrease of IL-1Ra, IL-1b, and IL-6. After 7 days, all animals treated with ADSCs had better results concerning blood biomarkers, and histopathological analysis revealed a lower degree of inflammatory cell infiltration of the organs of the peritoneal cavity. CONCLUSIONS: Intraperitoneal administration of ADSCs as an adjuvant therapy for sepsis improves the outcome and diminishes the effects of peritonitis and associated organ damage by regulating the immune system and reducing intra-abdominal adhesions in a clinically relevant porcine model of abdominal sepsis.

In Vitro Biocompatibility of CPP-ACP and Fluoride-containing Desensitizers on Human Gingival Cells.

OBJECTIVES: To analyze the biocompatibility of different desensitizers containing casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) and fluoride in their composition: MI Varnish (MV), Clinpro White Varnish (3M Oral Care), Profluorid Varnish (VOCO), Duraphat (Colgate) and Embrace Varnish (Pulpdent) on human gingival fibroblast cells (hGF). METHODS AND MATERIALS: Human gingival fibroblast (hGF) cells were exposed to several desensitizer extracts at different concentrations (0.1%, 1%, and 4% eluates). Then, in vitro biocompatibility was studied by analyzing the IC50 value, cell proliferation (MTT assay and cell cycle), cell migration (wound healing assay), cell morphology and F-actin content (immunocytofluorescence), and induction of apoptosis/necrosis (flow cytometry). Data were analyzed by one-way analysis of variance (ANOVA) followed by Tukey test. RESULTS: The lowest cell viability and IC50 were observed in all concentrations of Embrace Varnish-treated hGFs (p<0.001), whereas the highest were exhibited by those treated with Clinpro White Varnish. Similar effects were evidenced when induction of apoptosis/necrosis and cell migration assays were assessed. Finally, MI Varnish, Profluorid Varnish, Duraphat, and Embrace Varnish extracts showed lower numbers of attached cells, some of them with an unusual fibroblastic morphology when cultured with 4% concentration of the varnishes, while Clinpro White Varnish exhibited a similar number of cells with an evident actin cytoskeleton compared to the control group. CONCLUSIONS: The results obtained in this study indicate that hGFs show better in vitro biocompatibility after exposure to Clinpro White Varnish, even at the highest concentration employed, making it the most eligible for topical applications. In contrast, Embrace Varnish exhibited a high cytotoxicity towards hGFs that could potentially delay the healing process and regeneration of the oral mucosa, although more studies are needed to confirm this hypothesis.

Proteomic analysis of Strongyloides stercoralis L3 larvae.

Strongyloidiasis can be perpetuated by autoinfection with the filariform larvae L3, causing asymptomatic chronic infections and creating a population of carriers, affecting not only developing countries. So far, very little is known about the proteins that interact with the human host, and few proteins from the infective Strongyloides stercoralis L3 have been characterized. Here, we report results obtained from a proteomic analysis of the proteins from S. stercoralis L3 larvae obtained from patients. Since the genome of S. stercoralis is not yet available, we used proteomic analysis to identify 26 different proteins, 13 of them released by short digestion with trypsin, which could represent surface-associated proteins. The present work extends our knowledge of host-parasite interactions by identifying proteins that could be of interest in the development of diagnostic tools, vaccines, or treatments for a neglected disease like strongyloidiasis.

The functional role of the caudal fin in the feeding ecology of the common thresher shark Alopias vulpinus.

This study tests the hypothesis that the common thresher shark Alopias vulpinus uses its elongate caudal fin to both produce thrust and immobilize prey during feeding. Underwater video recorded in southern California from 2007 to 2009 revealed 34 feeding events, all of which were initiated with the upper lobe of the caudal fin.

Assessment of the effectiveness and safety of two radiofrequency techniques for the treatment of knee pain secondary to gonarthrosis. Prospective randomized double blind study. / Valoración de la efectividad y seguridad de dos técnicas de radiofrecuencia para el tratamiento de la gonalgia secundaria a gonartrosis. Estudio prospectivo randomizado doble ciego.

BACKGROUND AND OBJECTIVE: In patients with knee pain due to gonarthrosis, we have treatments that are not free of side effects. OBJECTIVE: to evaluate the analgesic efficacy of radiofrequency (pulsed and conventional) on the saphenous nerve at the subsartorial level and the genicular nerves of the knee, by ultrasonography. MATERIALS AND METHODS: Prospective, randomized, double-blind clinical trial. G1 (RDF1): subjects subjected to radiofrequency, G2 (PLCB): subjects subjected to placebo. A decrease ≥30% of the pain was considered clinically relevant, according to numerical rating scale and in the Western Ontario and McMaster Universities Osteoarthritis Index, global patient impression questionnaire (PGIC) and health status questionnaire (SF-12) in the evaluation at month, three months and six months after the completion of the technique. RESULTS: 28 patients (G1: 12, G2: 16), 72% women, age: 75.2±9.1 years, body mass index: 29.9±4.64. The analysis did not show a pain reduction, neither statistically significant, not clinically relevant, at one month, three, or six months with respect to the start of treatment, in the Western Ontario and McMaster Universities Osteoarthritis Index questionnaire and numerical rating scale (rest, movement). Regarding PGIC and the SF-12 questionnaire, there were no statistically significant differences between G1 and G2 either before or after treatment. CONCLUSIONS: The combination of two radiofrequency techniques, does not cause a reduction in the intensity of the knee pain, at month, three, or at six months after its completion. It is necessary to change the radiofrequency technique and include more variables to continue with the efficacy study.

[Consensus statement on the use of acetaminophen/tramadol in patients with moderate-severe pain]. / Documento de consenso sobre el uso de la combinación paracetamol/tramadol en pacientes con dolor moderado-intenso.

OBJECTIVES: To present recommendations on the use of the paracetamol/tramadol (P/T) combination in patients with moderate-intense pain based on best evidence and experience. METHODS: The method of nominal groups and Delphi was followed, and supported by a systematic literature review (SLR). A multidisciplinary panel of 12 experts in pain management was selected. In the first nominal group meeting, the aim, scope, users, and sections of the consensus document, were defined, along with the preliminary general recommendations. For the SLR, the inclusion and exclusion criteria, as well as the search strategies, were defined. Two reviewers selected and analysed the articles. This evidence was discussed in a second nominal group meeting, and definitive recommendations were developed. For each recommendation, the evidence levels and grade of recommendation grades were classified according to the Oxford model, and the grade according to the Delphi technique. It was defined as an agreement if at least 70% of the participants scored ≥7 for each recommendation (1=total disagreement to 10=total agreement). RESULTS: A total of 20 recommendations were produced, which covered general aspects, such as the assessment of pain, and those specific to P/T management. These latter included the indications of the P/T combination (patient profile, dosing, prescription, formulations), risk management (contraindications, precautions, interactions, concomitant use with other medications, follow-up, special situations), and patient education. CONCLUSIONS: These recommendations attempt to resolve any of the routine clinical questions, and help in the making of decisions on the use of the P/T combination in patients with moderate-intense pain.

GuttaFlow Bioseal promotes spontaneous differentiation of human periodontal ligament stem cells into cementoblast-like cells.

OBJECTIVES: To evaluate in vitro the cementogenic potential and the biological effects of GuttaFlow Bioseal, GuttaFlow 2, MTA Fillapex and AH Plus on human periodontal ligament stem cells (hPDLSCs). METHODS: Cell viability, cell migration and cell morphology assays were performed using eluates of each material. To evaluate cell attachment, hPDLSCs were directly seeded onto the material surfaces and analyzed by scanning electron microscopy (SEM). The effects of endodontic sealers on cementum protein 1 (CEMP1), cementum-derived attachment protein (CAP), bone sialoprotein (BSP), ameloblastin (AMBN), amelogenin (AMELX) and alkaline phosphatase (ALP) gene expression on hPDLSCs were investigated by qPCR and immunofluorescence (IF). Statistical analysis was performed with analysis of variance and Bonferroni or Tukey post-test (α<0.05). RESULTS: More than 90% of viable cells were obtained using extracts of GuttaFlow Bioseal and GuttaFlow2 after 72h of culture. By contrast, AH Plus and MTA Fillapex induced significantly lower levels of cell viability. GuttaFlow2 and GuttaFlow Bioseal promoted wound closure in a concentration-dependent manner, comparable to that observed with control extracts (*p<0.05). However, with AH Plus and MTA Fillapex, cell migration was significantly lower than in the control (***p<0.0001). SEM analysis pointed to an organized stress fiber assembly and high degree of cell adhesion on GuttaFlow Bioseal disks but low rates on GuttaFlow2, MTA Fillapex and AH Plus. When hPDLSCs were cultured with GuttaFlow Bioseal-conditioned media, qPCR assays and IF showed a higher level of AMELX, AMBN, CEMP1 and CAP expression than the control (*p<0.05)), whereas no such expression was observed in the other sealers. SIGNIFICANCE: Our results showed that GuttaFlow sealers were more cytocompatible than AH Plus and MTA Fillapex, while GuttaFlow Bioseal favored cementoblast differentiation of hPDLSCs in the absence of any growth factors.