Biomineralization of lithium nanoparticles by Li-resistant Pseudomonas rodhesiae isolated from the Atacama salt flat

BACKGROUND: The Atacama salt flat is located in northern Chile, at 2300 m above sea level, and has a high concentration of lithium, being one of the main extraction sites in the world. The effect of lithium on microorganism communities inhabiting environments with high concentrations of this metal has been scarcely studied. A few works have studied the microorganisms present in lithium-rich salt flats (Uyuni and Hombre Muerto in Bolivia and Argentina, respectively). Nanocrystals formation through biological mineralization has been described as an alternative for microorganisms living in metal-rich environments to cope with metal ions. However, bacterial lithium biomineralization of lithium nanostructures has not been published to date. In the present work, we studied lithium-rich soils of the Atacama salt flat and reported for the first time the biological synthesis of Li nanoparticles. RESULTS: Bacterial communities were evaluated and a high abundance of Cellulomonas, Arcticibacter, Mucilaginibacter, and Pseudomonas were determined. Three lithium resistant strains corresponding to Pseudomonas rodhesiae, Planomicrobium koreense, and Pseudomonas sp. were isolated (MIC > 700 mM). High levels of S2− were detected in the headspace of P. rodhesiae and Pseudomonas sp. cultures exposed to cysteine. Accordingly, biomineralization of lithium sulfide-containing nanomaterials was determined in P. rodhesiae exposed to lithium salts and cysteine. Transmission electron microscopy (TEM) analysis of ultrathin sections of P. rodhesiae cells biomineralizing lithium revealed the presence of nanometric materials. Lithium sulfide-containing nanomaterials were purified, and their size and shape determined by dynamic light scattering and TEM. Spherical nanoparticles with an average size < 40 nm and a hydro-dynamic size ~ 44.62 nm were determined. CONCLUSIONS: We characterized the bacterial communities inhabiting Li-rich extreme environments and reported for the first time the biomineralization of Li-containing nanomaterials by Li-resistant bacteria. The biosynthesis method described in this report could be used to recover lithium from waste batteries and thus provide a solution to the accumulation of batteries.

Ex Vivo Analysis of MTA FLOW® Biomineralization and Push-Out Strength: A Pilot Study / Análisis ex vivo del proceso de biomineralización y resistencia al desplazamiento del MTA FLOW®: Estudio piloto

Abstract: This study evaluated the biomineralization processes and push-out strength of MTA Flow® with radicular dentine in three different consistencies. The push-out test was performed on an ex vivo model, using 2mm thick dentin discs from the middle third of the root with standardized cavities of 1.5 mm. Samples were filled with MTA-Angelus (Angelus Dental, Brazil), Biodentine (Septodont, France), MTA Flow® Putty (Ultradent, USA), MTA Flow® Thick or MTA Flow® Thin. The samples were divided into 3 groups: subgroup 1 (n=5), analysis of the biomineralization process; 2 (n=20), evaluation of the bonding strength and push-out resistance; and 3 (n=5), evaluation of the cement/ dentin interface. The samples filled with Biodentine had a higher precipitation of carbonate apatite. However, there was no significant difference between MTA-Angelus, MTA Flow® Putty, or Thick (p=0.0536), but there was a significant difference in the Thin group (P<0.05). The samples with Biodentine displayed the greatest release of calcium ions. The formation of a partially carbonated intermediate apatite layer was observed in all groups. Zones of biomineralization were observed at the interface but were not continuous. After 72 hours, a significant difference was found between the Biodentine and MTA Flow® Thin groups (p=0.0090) in the push-out test. The samples submerged in phosphate-buffered saline (PBS) for 15 days showed a significant difference between all groups and MTA Flow® Thin (p=0.0147). Putty or Thick consistencies presented a similar bonding strength to MTA-Angelus and Biodentine. MTA Flow® Putty and Thick consistencies show a good adaptation to dentin, similar to MTA-Angelus. However, the thickness of the interface was lower compared to that of Biodentine. MTA Flow® Thin, despite their tubular infiltration, results in gaps and a defective peripheral seal.Therefore, MTA Flow®, in Putty or Thick consistencies, presents a biomineralization process and push-out strength similar to MTA Angelus and Biodentine, however, both characteristics decreases considerably in Thin consistency.

Resumen: El objetivo de este estudio fue evaluar, en un modelo ex vivo, el proceso de biomineralización y fuerza de adhesión del MTA Flow® en sus tres diferentes consistencias por medio de la prueba de resistencia al desplazamiento (Push-out). Se utilizaron discos de dentina de 2mm de espesor del tercio medio radicular con cavidades estandarizadas de 1.5mm de diámetro, las cuales se obturaron con diferentes materiales entre ellos: MTA Angelus (Angelus Dental, Brasil), Biodentine (Septodont, Francia), MTA Flow® Consistencia Putty (Ultradent, E.E.U.U), MTA Flow® Consistencia Thick (Ultradent, E.E.U.U) y MTA Flow® Consistencia Thin (Ultradent, E.E.U.U). Las muestras se sometieron al proceso de biomineralización y a pruebas de Push-out. Las muestras obturadas con Biodentine promovieron una mayor precipitación de apatita carbonatada, sin embargo, no se presentó diferencia estadística significativa con respecto al MTA Angelus, MTA Flow® Putty ni Thick (p=0.0536). No obstante, si presentó una diferencia significativa con respecto al grupo de MTA Flow® consistencia Thin (P<0.05). Las muestras con Biodentine presentaron la mayor liberación de iones calcio. De acuerdo a las pruebas de resistencia al desplazamiento, a las 72 horas post-obturación, solamente se encontró diferencia significativa entre las muestras obturadas con Biodentine y las correspondientes al MTA Flow consistencia Thin (p=0.0090), sin embargo las muestras sumergidas 15 días en PBS presentaron diferencia significativa entre todos los grupos con respecto al MTA Flow Thin (p=0.0147). En general se observaron zonas de biomineralización en la interface, sin embargo, no fueron continuas. Se concluye que el MTA Flow en consistencia Putty o Thick presenta un proceso de biomineralización y una resistencia al desplazamiento similar al MTA Angelus y al Biodentine, sin embargo, esta última disminuye considerablemente en presentación Thin.

Influence of the Vehicle on the Tissue Reaction and Biomineralization of Fast Endodontic Cement

ABSTRACT Objective: To investigate the tissue response and the biomineralization ability of CER prepared with epoxy resin or water compared to Mineral Trioxide Aggregate (MTA). Material and Methods: Polyethylene tubes containing materials or empty tubes for control were inserted into the subcutaneous tissues of 30 rats. After 7, 15, 30, 60, and 90 days, the rats were killed and the tubes were removed for analysis using hematoxylin-eosin staining, von Kossa staining, and under polarized light. Inflammation was graded through a score system; the thickness of the fibrous capsule was classified as thin or thick; the biomineralization ability was recorded as present or absent. The results were statistically analyzed using the Kruskal-Wallis test (p<0.05). Results: Histologic analysis performed after 7 and 15 days for CER prepared with epoxy resin or water and for MTA showed moderate inflammation and a thick fibrous capsule (p>0.05). After 30, 60, and 90 days, mild inflammation, and a thin fibrous capsule were observed in all groups (p>0.05). Conclusion: All materials had structures positive for von Kossa and birefringent to polarized light. CER epoxy resin showed biocompatibility and biomineralization similar to CER water and MTA.

Advances in biomineralization-inspired materials for hard tissue repair / 国际口腔科学杂志·英文版

Biomineralization is the process by which organisms form mineralized tissues with hierarchical structures and excellent properties, including the bones and teeth in vertebrates. The underlying mechanisms and pathways of biomineralization provide inspiration for designing and constructing materials to repair hard tissues. In particular, the formation processes of minerals can be partly replicated by utilizing bioinspired artificial materials to mimic the functions of biomolecules or stabilize intermediate mineral phases involved in biomineralization. Here, we review recent advances in biomineralization-inspired materials developed for hard tissue repair. Biomineralization-inspired materials are categorized into different types based on their specific applications, which include bone repair, dentin remineralization, and enamel remineralization. Finally, the advantages and limitations of these materials are summarized, and several perspectives on future directions are discussed.

Biocompatibility, biomineralization, and maturation of collagen by rtr®, bioglass and dm bone® materials

Abstract This study evaluated the biocompatibility, biomineralization, and collagen fiber maturation induced by Resorbable Tissue Replacement (RTR®; β-tricalcium phosphate [TCP]), Bioglass (BIOG; bioactive glass), and DM Bone® (DMB; hydroxyapatite and β-TCP) in vivo. Sixty-four polyethylene tubes with or without (control group; CG) materials (n=8/group/period) were randomly implanted in the subcutaneous tissue of 16 male Wistar rats (four per rat), weighting 250 to 280 g. The rats were killed after 7 and 30 days (n=8), and the specimens were removed for analysis of inflammation using hematoxylin-eosin; biomineralization assay using von Kossa (VK) staining and polarized light (PL); and collagen fiber maturation using picrosirius red (PSR). Nonparametric data were statistically analyzed by Kruskal-Wallis and Dunn tests, and parametric data by one-way ANOVA test (p<0.05). At 7 days, all groups induced moderate inflammation (p>0.05). At 30 days, there was mild inflammation in the BIOG and CG, and moderate inflammation in the RTR and DMB groups, with a significant difference between the CG and RTR (p<0.05). The fibrous capsule was thick at 7 days and predominantly thin at 30 days in all groups. All materials exhibited structures that stained positively for VK and PL. Immature collagen fibers were predominant at 7 and 30 days in all groups (p>0.05), although DMB exhibited more mature fibers than BIOG at 30 days (p<0.05). RTR, BIOG, and DMB were biocompatible, inducing inflammation that reduced over time and biomineralization in the subcutaneous tissue of rats. DMB exhibited more mature collagen fibers than BIOG over a longer period.

Resumo Este estudo avaliou a biocompatibilidade, biomineralização e maturação das fibras de colágeno induzidas por Resorbable Tissue Replacement (RTR®; fosfato β-tricálcico [TCP]), Bioglass (BIOG; vidro bioativo) e DM Bone® (DMB; hidroxiapatita e β-TCP) in vivo. Sessenta e quatro tubos de polietileno com ou sem (grupo controle; GC) os materiais (n=8/grupo/período) foram implantados aleatoriamente em tecido subcutâneo de 16 ratos machos Wistar (quatro por rato), pesando entre 250 a 280g. Os ratos foram mortos após 7 e 30 dias (n=8), e as amostras foram removidas para análise da inflamação utilizando hematoxilina-eosina; avaliação da biomineralização utilizando coloração de von Kossa (VK) e luz polarizada (LP); e maturação das fibras colágenas, utilizando picrosirius red (PSR). Os dados não-paramétricos foram analisados pelos testes de Kruskal-Wallis e Dunn, e os paramétricos pelo teste de one-way ANOVA (p<0.05). Aos 7 dias, todos os grupos induziram inflamação moderada (p>0,05). Aos 30 dias, houve inflamação leve nos grupos BIOG e GC, e inflamação moderada nos grupos RTR e DMB, com diferença significativa entre os GC e RTR (p<0,05). A cápsula fibrosa foi espessa aos 7 dias, e predominantemente fina aos 30 dias em todos os grupos. Todos os materiais exibiram estruturas positivas para VK e LP. Fibras colágenas imaturas foram predominantes aos 7 e 30 dias em todos os grupos (p>0,05), embora o DMB exibiu fibras mais maduras do que o BIOG aos 30 dias (p<0,05). RTR, BIOG e DMB foram biocompatíveis, induzindo inflamação que reduziu com o tempo, e biomineralização no tecido subcutâneo de ratos. O DMB exibiu mais fibras colágenas maduras do que o BIOG em período mais longo.

Ex Vivo Evaluation of a Restoration Protocol for Teeth with Simulated Incomplete Rhizogenesis / Evaluación ex vivo de un protocolo de restauración para dientes con rizogénesis incompleta simulada

Abstract: The use of phosphate buffered saline (PBS) as an intracanal medication triggers a biomineralization process within mineral trioxide aggregate (MTA) apical plugs during the apexification process in teeth with incomplete rhizogenesis. However, no consensus is available in the literature regarding a restorative protocol for this type of treatment. Thus, the objective of this study was to use scanning electron microscopy (SEM) to evaluate the processes of biomineralization and adhesion in a restorative protocol for teeth with simulated incomplete rhizogenesis. Methodology: Root sections with a thickness of 2mm and cavities with a diameter of 2mm were used. The sections were randomly prepared and filled with the following materials: Group 1 (n=12), ProRoot MTA; and Group 2 (n=12): MTA Exp. Subsequently, the samples were immersed in PBS for 35 days. Every 5 days, the PBS was replaced, and the precipitates were collected, dried, and weighed. Two samples from each group were analyzed by SEM. Moreover, 24 single-rooted teeth were standardized, incomplete rhizogenesis was simulated, and 5-mm-long apical plugs were created with Pro Root MTA. As an intracanal medication, PBS was used for different periods of time: Group 1:48 h; Group 2:7 days; and Group 3:15 days. Then, fiberglass posts were cemented with the REBILDA® Post System. The samples were prepared and analyzed by SEM. Results: ProRoot MTA and MTA Exp effectively promoted the formation of carbonated apatite precipitates and biomineralization with dentin. ProRoot MTA yielded more carbonated apatite precipitates compared to MTA Exp (p=0.0536). The use of PBS as an intracanal medication for 7 and 15 days promoted intratubular mineralization (MIT), and treatment for 15 days was more effective (p < 0.05). The REBILDA® Post System effectively promoted the microimbrication of the adhesive system and the formation of resinous tags with lateral adhesive branches. Conclusion: Apexification with MTA associated with the use of PBS as an intracanal medication for 15 days, in addition to the use of the REBILDA® Post System, seems to be a feasible restorative protocol.

Resumen: El uso de solución salina tamponada con fosfato (PBS) como medicamento intracanal desencadena un proceso de biomineralización en los plugs apicales con agregado de trióxido mineral (MTA) durante el proceso de apexificación en dientes con rizogénesis incompleta. Sin embargo, no hay consenso disponible en la literatura sobre un protocolo restaurador para este tipo de tratamiento. Por lo tanto, el objetivo de este estudio fue utilizar microscopía electrónica de barrido (MEB) para evaluar los procesos de biomineralización y adhesión en un protocolo restaurador para dientes con rizogénesis incompleta simulada. Metodología: Se utilizaron secciones de raíz con un espesor de 2mm y se realizaron cavidades con un diámetro de 2 mm. Las cavidades en las secciones se obturaron con: Grupo 1 (n=12), ProRoot MTA; y Grupo 2 (n=12): MTA Exp. Posteriormente, las muestras se sumergieron en PBS durante 35 días. Cada 5 días, se reemplazó el PBS y se recogieron los precipitados, se secaron y pesaron. Dos muestras de cada grupo fueron analizadas por MEB. Además, se estandarizaron 24 dientes de raíz única, se simuló la rizogénesis incompleta y se crearon tapones apicales de 5mm de longitud con Pro Root MTA. Como medicamento intracanal, se utilizó PBS durante diferentes períodos de tiempo: Grupo 1:48 h; Grupo 2:7 días; y Grupo 3:15 días. Posteriormente, los postes de fibra de vidrio se cementaron con el sistema de postes REBILDA®. Las muestras fueron preparadas y analizadas por MEB. Resultados: ProRoot MTA y MTA Exp promovieron efectivamente la formación de precipitados de apatita carbonatada y la biomineralización con dentina. ProRoot MTA produjo más precipitados de apatita carbonatada en comparación con MTA Exp (p=0.0536). El uso de PBS como medicamento intracanal durante 7 y 15 días promovió la mineralización intratubular (MIT), siendo el tratamiento durante 15 días más efectivo (p <0.05). El sistema de postes REBILDA® promovió efectivamente la microimbricación del sistema adhesivo y la formación de tags resinosos. Conclusión: La apexificación con MTA asociada con el uso de PBS como medicación intracanal durante 15 días, además del uso del sistema de postes REBILDA®, parece ser un protocolo factible y eficaz en este tipo de tratamientos.

Influência das suplementações de ômega-3 e melatonina sobre a resposta tecidual de diferentes cimentos endodônticos / The influence of the supplementation of Omega 3 and melatonin on tissue response of different endodontic sealers

O objetivo deste estudo foi avaliar a influência das suplementações de ômega-3 e melatonina sobre a resposta tecidual de cimentos endodônticos. Foram utilizados 72 ratos Wistar alocados em 3 grupos principais: ratos controle tratados com água (TA), ratos tratados com ômega-3 (TO) e ratos tratados com melatonina (TM). Em cada rato foram implantados 4 tubos de polietileno, sendo 1 tubo vazio para controle, e os demais contendo um dos cimentos endodônticos: Sealapex®, AH Plus® e Endofil®. Os períodos de avaliação foram de 5, 15 e 30 dias (n=8/tempo). Após eutanásia, os tubos foram removidos e processados para análise microscópica. A inflamação foi avaliada em coloração de H.E. e técnica imunohistoquímica para IL-6 e TNF-α, o colágeno pela coloração de Picrosírios red e a deposição de cálcio, pela técnica de von Kossa e luz polarizada. A estatística foi realizada de acordo com cada tipo de análise, com nível de significância de 5% (p <0,05) e considerando cada tipo de suplementação comparado ao controle. Podese observar infiltrado inflamatório reduzido com o tempo e de menor intensidade nos grupos TO ou TM comparado a TA, assim como a imunomarcação para IL-6 e TNF-α. O cimento Sealapex induziu menor inflamação aos 30 dias, tanto em TO quanto em TM. Em relação ao colágeno, foi observado maior expressão de fibras imaturas no grupo TO aos 15 e 30 dias, entretanto, no grupo TM, foi observado maior quantidade de fibras maduras aos 5 dias, ambos comparados a TA. O cimento Sealapex foi único a apresentar marcação positiva para cálcio e cristais de calcita birrefringentes à luz polarizada, independente das suplementações. Pode-se concluir que a suplementação com ômega-3 ou melatonina modularam a inflamação, diminuindo o infiltrado inflamatório e a imunomarcação de IL-6 e TNF- α, estimularam o reparo tecidual e não influenciaram no processo de deposição de cálcio(AU)

The aim of this study was to evaluate the influence of omega-3 and melatonin supplements on the tissue response of endodontic sealers. 72 Wistar rats were allocated in 3 main groups: control rats treated with water (TW), rats treated with omega-3 (TO) and rats treated with melatonin (TM). In each rat, 4 polyethylene tubes were implanted, 1 empty tube for control, and the others containing one of the endodontic sealers: Sealapex®, AH Plus® and Endofil®. The evaluation periods were of 5, 15 and 30 days (n = 8 / time). After euthanasia, the tubes were removed and processed for microscopic analysis. Inflammation was evaluated in H.E. staining and immunohistochemical technique for IL-6 and TNF-α, collagen by Picrosírios red staining and calcium deposition by von Kossa and polarized light technique. Statistics were performed according to each type of analysis, with a significance level of 5% (p <0.05) and considering each type of supplementation compared to the control. It is possible to observe a reduced inflammatory infiltrate over time and less intensity in the TO or TM groups compared to TW, as well as the immunolabeling for IL-6 and TNF-α. Sealapex sealer induced less inflammation at 30 days, both in TO and TM. Regarding collagen, a greater expression of immature fibers was observed in the TO group at 15 and 30 days, however, in the TM group, a greater amount of mature fibers was observed at 5 days, both compared to TW. Sealapex sealer was the only one to present positive marking for calcium and birefringent calcite crystals under polarized light, regardless of supplementation. It can be concluded that supplementation with omega-3 or melatonin modulated the inflammation, reducing the inflammatory infiltrate and the immunolabeling of IL-6 and TNF-α, they stimulated tissue repair and did not influence the calcium deposition process(AU)

Fracture resistance of teeth obturated with two different types of mineral trioxide aggregate cements / Resistência à fratura de dentes obturados com dois tipos diferentes de cimentos agregados de trióxido minera

Objective: Endodontically obturated teeth have lower fracture resistance depending on the obturating material and technique. The purpose of this study was therefore to evaluate the influence of ProRoot MTA (Dentsply Sirona, Tulsa Division) and OrthoMTA III (BioMTA, Daejeon, Korea) as an obturating material on the fracture resistance of endodontically treated teeth. Material and Methods: Thirty extracted human maxillary central incisors were decoronated and instrumented using Protaper instruments (size F5). Irrigation was performed with 2.5% sodium hypochlorite between each instrument change followed by 7% maleic acid for one minute. Finally, canals were flushed with 5 ml of PBS solution for one minute. Samples were then divided into three groups. Group I- positive control (no root canal filling); Group II- obturation with ProRoot MTA; Group III- obturation with OrthoMTA III. Ten teeth were randomly selected as a negative control in which no treatment was performed. All the specimens were then subjected to fracture strength testing using universal testing machine. For evaluation of biomineralization, six maxillary central incisors were divided into two groups. Group I obturated with ProRoot MTA and group II obturated with OrthoMTA III. These samples were subjected to SEM analysis. Results: Positive control group demonstrated the least fracture resistance, while OrthoMTA III group showed the highest fracture resistance. There was no significant difference between negative control group and ProRoot MTA groups (p=0.821). OrthoMTA III group showed better tubular biomineralization when compared to ProRoot MTA. Conclusions: Root canals obturated with OrthoMTA III had better fracture resistance and increased tubular biomineralization compared to ProRoot MTA. Since root canals obturated with OrthoMTA III had better fracture resistance, it can be used as a promising obturating material.(AU)

Objetivo: Dentes obturados endodonticamente apresentam menor resistência à fratura, dependendo do material e da técnica de obturação. Portanto, o objetivo deste estudo foi avaliar a influência do ProRoot MTA (Dentsply Sirona, Tulsa Division) e OrthoMTA III (BioMTA, Daejeon, Coréia) como material obturador na resistência à fratura de dentes tratados endodonticamente. Material e Métodos: Trinta incisivos centrais superiores humanos extraídos foram decoronados e instrumentados com instrumentos Protaper (tamanho F5). A irrigação foi realizada com hipoclorito de sódio a 2,5% entre cada troca de instrumento, seguida por ácido maleico a 7% por um minuto. Finalmente, os canais foram lavados com 5 ml de solução de PBS por um minuto. As amostras foram então divididas em três grupos. Grupo I - controle positivo(sem preenchimento do canal radicular); Grupo II - obturação com ProRoot MTA; Grupo III -obturação com OrthoMTA III. Dez dentes foram selecionados aleatoriamente como controle negativo, no qual nenhum tratamento foi realizado. Todas as amostras foram então submetidas atestes de resistência à fratura usando uma máquina de teste universal. Para avaliação da biomineralização, seis incisivos centrais superiores foram divididos em dois grupos: grupo Iobturado com ProRoot MTA e grupo II obturado com OrthoMTA III. Essas amostras foram submetidas à análise SEM. Resultados: O grupo controle positivo demonstrou a menor resistência à fratura, enquanto o grupo OrthoMTA III apresentou a maior resistência à fratura. Não houve diferença significativa entre o grupo controle negativo e os grupos ProRoot MTA (p= 0,821). O grupo OrthoMTA III apresentou melhor biomineralização tubular quando comparado ao ProRoot MTA. Conclusões: Os canais radiculares obturados com OrthoMTA III apresentaram melhor resistência à fratura e maior biomineralização tubular em comparação como ProRoot MTA. Como os canais radiculares obturados com OrthoMTA III apresentaram melhor resistência à fratura, podendo ser utilizado como um material obturador promissor.(AU)

Cytotoxicity, inflammation, biomineralization, and immunoexpression of IL-1β and TNF-α promoted by a new bioceramic cement

Abstract Aim To evaluate the cytotoxicity, biocompatibility and mineralization capacity of BIO-C PULPO, and MTA. Methodology L929 fibroblasts were cultured and MTT assay was used to determine the material cytotoxicity on 6, 24, and 48 h. A total of 30 male rats (Wistar) aged between 4 and 6 months, weighing between 250 and 300 g were used. Polyethylene tubes containing BIO-C PULPO, MTA, and empty tubes were implanted into dorsal connective tissue. After the experimental periods (7, 15, 30, 60, and 90 days) the tubes were histologically analyzed using hematoxylin-eosin (H&E), immunolabeling of IL-1β and TNF-α, and von Kossa staining, or without staining for polarized light analysis. The average number of inflammatory cells was quantified; the mineralization assessment was determined by the area marked in μm2 and semiquantitative immunolabeling analyses of IL-1β and TNF-α were performed. Then, data underwent statistical analysis with a 5% significance level. Results It was observed that BIO-C PULPO and MTA presented cytocompatibility at 6, 24, and 48 similar or higher than control for all evaluated period. On periods 7 and 15 days, BIO-C PULPO was the material with the highest number of inflammatory cells (p<0.05). On periods 30, 60, and 90 days, BIO-C PULPO and MTA presented similar inflammatory reactions (p>0.05). No statistical differences were found between Control, BIO-C PULPO, and MTA for immunolabeling of IL-1β and TNF-α in the different periods of analysis (p<0.05). Positive von Kossa staining and birefringent structures under polarized light were observed in all analyzed periods in contact with both materials, but larger mineralization area was found with BIO-C PULPO on day 90 (p<0.05). Conclusion BIO-C PULPO was biocompatible and induced mineralization similar to MTA.

Cytotoxicity, biocompatibility and biomineralization of a new ready-for-use bioceramic repair material

Abstract New mineral trioxide aggregate (MTA) formulations are constantly introduced in the market, usually in a powder-and-liquid form. Bioceramic (Bio-C) Repair is a ready-for-use material suggested as substitute for MTA, but its properties need to be studied. This study evaluated the cytotoxicity, biocompatibility and biomineralization of Bio-C Repair compared to MTA Repair High-Plasticity (MTA-HP) and white MTA-Angelus (MTA-Ang). L929 fibroblasts were exposed to material-extracted (undiluted, ½ and » dilutions; 6, 24 and 48h). Polyethylene tubes with material or empty (control) were implanted in the subcutaneous tissue of rats. After 7 and 30 days (n=8), the specimens were removed for analysis (hematoxylin-eosin, von Kossa and polarized light). Cytotoxicity data were statistically analyzed by two-way ANOVA, and biocompatibility data by Kruskal-Wallis and Dunn tests (p<0.05). The cells exposed to the materials had greater viability at most of the periods compared with control (p<0.05). The undiluted and ½ dilutions of MTA-HP extract showed higher cytocompatibility than Bio-C Repair at 6 h and with the » dilution at 24 h (p<0.05); the white MTA-Ang showed higher cytocompatibility than Bio-C Repair at most of periods (p<0.05). The undiluted white MTA-Ang extract had higher cytocompatibility at 6 and 24h than MTA-HP, and with ½ dilution at 24h (p<0.05). The materials' cytocompatibility was similar at 48h for most dilutions (p>0.05). At 7 and 30 days, the groups had moderate and mild inflammation, respectively (p>0.05). All materials showed positive structures for von Kossa and polarized light. In conclusion, Bio-C Repair had similar cytocompatibility to MTA-based materials is biocompatible and induces biomineralization.

Resumo Novas formulações de agregado de trióxido mineral (MTA) são constantemente introduzidas no mercado, geralmente em forma de pó e líquido. O Biocerâmico (Bio-C) Reparador (Repair) é um material pronto para uso sugerido como substituto do MTA, mas suas propriedades precisam ser estudadas. Este estudo avaliou a citotoxicidade, biocompatibilidade e biomineralização do Bio-C Repair comparado ao MTA-High Plasticity (MTA-HP) e MTA branco da Angelus (MTA-Ang). Fibroblastos L929 foram expostos a extratos dos materiais (não diluído, ½ e » diluições; 6, 24 e 48 h). Tubos de polietileno contendo os materiais ou vazios (controle) foram implantados no tecido subcutâneo de ratos. Após 7 e 30 dias (n=8), os espécimes foram removidos para análises (hematoxilina-eosina, von Kossa e luz polarizada). Os dados da citotoxicidade foram analisados estatisticamente pelo teste de two-way ANOVA, e os dados da biocompatibilidade pelos testes de Kruskal-Wallis e Dunn (p<0,05). As células expostas aos materiais apresentaram maior viabilidade celular na maior parte dos períodos, comparados com o controle (p<0,05). O extrato não diluído e ½ diluição do MTA-HP apresentaram maior citocompatibilidade do que Bio-C Repair às 6h, e com » diluição às 24h (p<0,05); o MTA-Ang branco apresentou maior citocompatibilidade do que o Bio-C Repair na maior parte dos períodos (p<0,05). O extrato não diluído do MTA-Ang branco apresentou maior citocompatibilidade às 6 e 24 h comparado ao MTA-HP, e com ½ diluição às 24h (p<0,05). A citocompatibilidade dos materiais foi semelhante às 48 h para a maior parte das diluições (p>0,05). Aos 7 e 30 dias, os grupos apresentaram inflamação moderada e leve, respectivamente (p>0,05). Todos os materiais mostraram estruturas positivas para von Kossa e luz polarizada. Em conclusão, o Bio-C Repair teve citocompatibilidade semelhante aos materiais à base de MTA, é biocompatível e induz à biomineralização.

Estudo de novos dispositivos, instrumentos e materiais empregados no tratamento endodôntico / Evaluation of new devices, instruments and materials used in endodontic treatment

Introdução: O tratamento endodôntico visa manter ou restabelecer a saúde do periodonto através do preparo biomecânico, utilizando instrumentos de níqueltitânio automatizados e associados com insertos ultrassônicos para potencializar a ação dos irrigantes. Os materiais empregados na endodontia que permanecerão em contato com tecidos devem apresentar biocompatibilidade e preferencialmente ser biomineralizador, a fim de selar comunicações (fisiológicas/iatrogênicas), isolando o sistema de canais radiculares. Objetivos: 1- Avaliar a resistência à fadiga cíclica das limas reciprocantes Genius e EdgeFile X1 Small, comparados à lima WaveOne Gold Primary; 2- Avaliar a capacidade de limpeza de um inserto ultrassônico de níquel titânio em ativação da irrigação de forma contínua e passiva; 3- Avaliar a biocompatiblidade e biomineralização do cimento reparador MTA Flow. Material e Métodos: Para avaliar a resistência à fadiga cíclica dos sistemas Genius, EdgeFile e WaveOne Gold, 80 instrumentos foram utilizados (n=20), divididos em 4 grupos experimentais: Genius 25.04, Genius 30.04; EdgeFile X1 Small e Wave One Gold Primary, acionados em um canal artificial de aço inoxidável com ângulo de curvatura de 60o e raio de curvatura de 5 mm. O tempo decorrido da ativação do motor foi gravado em um cronômetro digital e parado assim que a fratura foi detectada. O número de ciclos até a falha (NCF) e o tempo para fratura (TF) foram calculados e os comprimentos dos segmentos fraturados medidos. Para avaliar a capacidade de limpeza do inserto ultrassônico, 45 prémolares inferiores, padronizados em 16 mm foram utilizados. A instrumentação foi realizada até lima 50.04 sob irrigação com hipoclorito de sódio e foram divididos em 3 grupos (n = 15) de acordo com a técnica de ativação final da solução irrigadora: irrigação convencional (IC) (passiva com pressão positiva/sucção concomitante) como controle; ativação ultrassônica passiva (PUI) e ativação ultrassônica contínua (CUI) do irrigante. Os três grupos tiveram os protocolos de ativação/irrigação final em soluções de hipoclorito de sódio e EDTA. As amostras foram clivadas, as imagens obtidas com microscopia eletrônica de varredura (MEV) e avaliadas quanto à capacidade de remoção da camada de smear layer no terço cervical, médio e apical, por um sistema de escores. Para análise biológica do material reparador MTA Flow, os materiais MTA Angelus e ProRoot MTA foram utilizados para comparação. Quarenta ratos receberam implantes subcutâneo de tubos de polietileno contendo os 3 materiais e tubo vazio como controle (n=10). Após 7, 15, 30 e 60 dias, os animais foram eutanasiados e os tubos removidos com o tecido conjuntivo circundantes. Infiltrado inflamatório e espessura da cápsula fibrosa foram avaliados histologicamente e indução da mineralização analisada por Von Kossa e sob luz polarizada. Os dados foram analisados com testes estatísticos específicos (p<5%). Resultados: Quanto à fadiga cíclica, os instrumentos EdgeFile X1 Small apresentaram maior resistência (NCF: 6175.74 ± 1608.99; TF: 1058.7 ± 275.82), seguido por ambos instrumentos Genius (p<0.05), de forma significativa comparados ao Waveone Gold Primary. O inserto de NiTi aumentou a limpeza, comparado com IC, principalmente no terço apical e CUI mostrou os melhores resultados (p<0.05). Nos resultados biológicos, MTA Angelus induziu a reação mais leve após 15 dias (p<0.05), seguido do MTA Flow. ProRoot MTA induziu uma inflamação severa no dia 7, reduzindo após 15 dias. Nenhuma diferença foi observada entre os materiais após 30 ou 60 dias (p>0.05). Estruturas coradas por Von Kossa e birrefringentes foram positivas para todos os materiais. Conclusões: 1- Os instrumentos EdgeFile X1 Small demonstraram maior resistência à fadiga cíclica que Genius e WaveOne Gold Primary. Ambos instrumentos Genius apresentaram resistência superior à WaveOne Gold Primary. 2- A ativação final do irrigante com o inserto NiTi melhorou a remoção de smear layer, com protocolo CUI superior à PUI ou IC. 3- O MTA Flow mostrou biocompatibilidade e induziu a biomineralização em todos os períodos observados(AU)

Introduction: The endodontic treatment aims to maintain or restore periodontal health through biomechanical instrumentation with nickel-titanium automatized instruments, associated with an ultrasonic tip to activate the irrigant, enhancing cleanness. Materials used in endodontics that will remain in contact with tissues, should demonstrate properties as biocompatibility and biomineralization ability, in order to seal communications (physiological/iatrogenic) isolating the root canal system. Objectives: 1- Evaluate the cyclic fatigue resistance of reciprocating Genius and EdgeFile X1 Small instruments compared to Waveone Gold Primary instruments; 2- Evaluate the cleaning effectiveness of a nickel-titanium ultrasonic tip in continuous and passive irrigant activation; 3-Evaluate the biocompatibility and biomineralization of MTA Flow repair cement. Materials and Methods: To evaluate the cyclic fatigue resistance of Genius, EdgeFile and WaveOne Gold, 80 instruments were used (n = 20), divided into 4 experimental groups: Genius 25.04, Genius 30.04; EdgeFile X1 Small and Wave One Gold Primary, reciprocating in a stainless steel artificial canal with a 60° angle of curvature and 5mm radius of curvature. The elapsed time of the motor activation was recorded in a digital timer and stopped as soon as the fracture was detected. The number of cycles to fracture (NCF) and time to fracture (TF) were calculated and the fractured segments lengths measured. To evaluate the ultrasonic tip cleanness capacity, forty-five mandibular premolars, standardized at 16 mm were used. The instrumentation was performed until file 50.04 under irrigation with sodium hypochlorite. The specimens were divided into 3 groups (n = 15) according to the final irrigant activation protocol: conventional irrigation (CI) (passive, with positive pressure/simultaneous aspiration) as control; passive ultrasonic irrigant activation (PUI) and continuous ultrasonic irrigant activation (CUI). All groups had the final activation / irrigation protocols with solutions of sodium hypochlorite and EDTA, standardized with same volume. Samples were sodium hypochlorite and EDTA, standardized with same volume. Samples were cleaved and images obtained through scanning electron microscopy (SEM) to assess smear layer removal in the cervical, middle and apical thirds, via a score system. To biologically analyze MTA Flow repair cement, MTA Angelus and ProRoot MTA cements were used as comparison. Forty rats received subcutaneous implants of polyethylene tubes containing the 3 cements and empty tube as control (n = 10). After days 7, 15, 30 and 60, the animals were euthanized and the tubes removed with the surrounding tissues. Inflammatory infiltrate and fibrous capsule thickness were histologically evaluated and mineralization induction assessed by Von Kossa staining and under polarized light. The data were analyzed with specific statistical tests (p<5%). Results: The cyclic fatigue test showed EdgeFile X1 Small with the highest resistance (NCF: 6175.74 ± 1608.99; TF: 1058.7 ± 275.82), followed by both Genius instruments (p<0.05). The nickel-titanium tip enhanced cleanness, compared to CI, mainly in the apical area, whereas CUI showed the best results (p<0.05). In the biological results, MTA Angelus induced the mildest reaction after 7 (p>0.05) and 15 days (p<0.05), followed by MTA Flow. ProRoot MTA induced severe inflammation on day 7, reducing after day 15 (p>0.05). No difference was observed after 30 or 60 days (p>0.05). Von Kossa staining and birefringents structures were positive for all materials. Conclusion: 1- EdgeFile X1 Small instruments demonstrated superior cyclic fatigue resistance than Genius and WaveOne Gold Primary. In addition, both Genius instruments showed superior resistance than WaveOne Gold Primary. 2- The final irrigant activation with the NiTi tip improved smear layer removal, with CUI protocol superior to PUI or CI. 3 - MTA Flow showed biocompatibility and induced biomineralization in all observed periods(AU)

Hypertension affects the biocompatibility and biomineralization of MTA, High-plasticity MTA, and Biodentine®

Abstract This study evaluated the effect of hypertension on tissue response and biomineralization capacity of white Mineral Trioxide Aggregate (MTA), High-plasticity MTA (MTA HP), and Biodentine® (BDT) in rats. Polyethylene tubes filled with MTA, MTA HP, BDT, and the control group (empty tubes) were placed into the dorsal subcutaneous tissue of 32 male rats (16 normotensive (NT) and 16 hypertensive rats - 8 per group). After 7 and 30 days, the polyethylene tubes surrounded by connective tissue were removed, fixed, and embedded in histological resin. The mean number of inflammatory cells was estimated in HE-stained sections, biomineralization was quantified as area (µm2) by Kossa (VK) staining, and examination by polarized light (LP) microscopy was performed. The differences amongst the groups were analyzed statistically by the Mann-Whitney or Student's t test, according to Shapiro-Wilk test of normality (p < 0.05). The inflammatory responses to all materials were greater in hypertensive rats than in NT rats (p < 0.05). Positive VK staining in MTA and BDT were more pronounced in NT rats at 7 and 30 days (p < 0.05). Birefringent structures in LP for MTA, MTA HP, and BDT were more pronounced in NT rats at 7 days (p<0.05). In rats, hypertension was able to increase inflammatory infiltrate and decrease biomineralization of the tested materials.

Molecular characterization of CHST11 and its potential role in nacre formation in pearl oyster Pinctada fucata martensii

Background: C4ST-1 catalyzes the transfer of sulfate groups in the sulfonation of chondroitin during chondroitin sulfate synthesis. Chondroitin sulfate consists of numerous copies of negatively charged sulfonic acid groups that participate in the nucleation process of biomineralization. In the present study, we obtained two CHST11 genes (PmCHST11a and PmCHST11b) which encoded the C4ST-1 and explored the functions of these genes in the synthesis of chondroitin sulfate and in the formation of the nacreous layer of shells. Results: Both PmCHST11a and PmCHST11b had a sulfotransferase-2 domain, a signal peptide and a transmembrane domain. These properties indicated that these genes localize in the Golgi apparatus. Real-time PCR revealed that both PmCHST11a and PmCHST11b were highly expressed in the central zone of the mantle tissue. Inhibiting PmCHST11a and PmCHST11b via RNA interference significantly decreased the expression levels of these genes in the central zone of the mantle tissue and the concentration of chondroitin sulfate in extrapallial fluid. Moreover, shell nacre crystallized irregularly with a rough surface after RNA interference. Conclusions: This study indicated that PmCHST11a and PmCHST11b are involved in the nacre formation of Pinctada fucata martensii through participating in the synthesis of chondroitin sulfate.

Biomineralization of a calcifying ureolytic bacterium Microbacterium sp. GM-1

Background: Biomineralization is a significant process performed by living organisms in which minerals are produced through the hardening of biological tissues. Herein, we focus on calcium carbonate precipitation, as part of biomineralization, to be used in applications for environmental protection, material technology, and other fields. A strain GM-1, Microbacterium sp. GM-1, isolated from active sludge, was investigated for its ability to produce urease and induce calcium carbonate precipitation in a metabolic process. Results: It was discovered that Microbacterium sp. GM-1 resisted high concentrations of urea up to 60 g/L. In order to optimize the calcification process of Microbacterium sp. GM-1, the concentrations of Ni2+ and urea, pH value, and culture time were analyzed through orthogonal tests. The favored calcite precipitation culture conditions were as follows: the concentration of Ni2+ and urea were 50 µM and 60 g/L, respectively, pH of 10, and culture time of 96 h. Using X-ray diffraction analysis, the calcium carbonate polymorphs produced by Microbacterium sp. GM-1 were proven to be mainly calcite. Conclusions: The results of this research provide evidence that Microbacterium sp. GM-1 can biologically induce calcification and suggest that strain GM-1 may play a potential role in the synthesis of new biominerals and in bioremediation or biorecovery.

Microbial-induced remediation of Zn2+ pollution based on the capture and utilization of carbon dioxide

Background: Microbial-induced remediation of Zn2+ pollution based on the capture and utilization of carbon dioxide was investigated. In this study, carbon dioxide was absorbed and transformed into carbonate ions under the enzymatic action of Paenibacillus mucilaginosus, which was being utilized to mineralize Zn2+. Results: The compositional and morphological properties of the precipitations were studied using Fourier transform infrared spectroscopy (FTIR), energy dispersive X-ray spectroscopy (EDS), X-ray diffraction (XRD), and scanning electron microscopy (SEM). The thermal properties of the precipitates were investigated by thermogravimetric-differential scanning calorimetry (TG-DSC). The FTIR results confirmed that the functional groups of the precipitates were CO3² − and OH−. The XRD and EDS patterns showed that basic zinc carbonate could be obtained successfully by Microbial-induced remediation. The SEM micrographs demonstrated that the precipitates were in the nanometer range with sizes of 100-200 nm and were sphere-like in shape. Conclusions: The TG-DSC results showed that weight loss of the precipitates occurred around 253°C. The FTIR and TG-DSC results were in accord with the XRD and EDS results and proved again that the precipitates were basic zinc carbonate. This work thus demonstrates a new method for processing Zn2+ pollution based on the utilization of carbon dioxide.