Culture of rat mesenchymal stem cells on PHBV-PCL scaffolds: analysis of conditioned culture medium by FT-Raman spectroscopy

Abstract Mesenchymal stem cells (MSCs) have great potential for application in cell therapy and tissue engineering procedures because of their plasticity and capacity to differentiate into different cell types. Given the widespread use of MSCs, it is necessary to better understand some properties related to osteogenic differentiation, particularly those linked to biomaterials used in tissue engineering. The aim of this study was to develop an analysis method using FT-Raman spectroscopy for the identification and quantification of biochemical components present in conditioned culture media derived from MSCs with or without induction of osteogenic differentiation. All experiments were performed between passages 3 and 5. For this analysis, MSCs were cultured on scaffolds composed of bioresorbable poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) and poly(-caprolactone) (PCL) polymers. MSCs (GIBCO®) were inoculated onto the pure polymers and 75:25 PHBV/PCL blend (dense and porous samples). The plate itself was used as control. The cells were maintained in DMEM (with low glucose) containing GlutaMAX® and 10% FBS at 37oC with 5% CO2 for 21 days. The conditioned culture media were collected and analyzed to probe for functional groups, as well as possible molecular variations associated with cell differentiation and metabolism. The method permitted to identify functional groups of specific molecules in the conditioned medium such as cholesterol, phosphatidylinositol, triglycerides, beta-subunit polypeptides, amide regions and hydrogen bonds of proteins, in addition to DNA expression. In the present study, FT-Raman spectroscopy exhibited limited resolution since different molecules can express similar or even the same stretching vibrations, a fact that makes analysis difficult. There were no variations in the readings between the samples studied. In conclusion, FT-Raman spectroscopy did not meet expectations under the conditions studied.

Resumo As células-tronco mesenquimais (MSCs) possuem grande potencial para aplicação em procedimentos terapêuticos ligados a terapia celular e engenharia de tecidos, considerando-se a plasticidade e capacidade de formação em diferentes tipos celulares por elas. Dada a abrangência no emprego das MSCs, há necessidade de se compreender melhor algumas propriedades relacionadas à diferenciação osteogênica, particularmente liga à biomateriais usados em engenharia de tecidos. Este projeto objetiva o desenvolvimento de uma metodologia de análise empregando-se a FT-Raman para identificação e quantificação de componentes bioquímicos presentes em meios de cultura condicionados por MSCs, com ou sem indução à diferenciação osteogênica. Todos os experimentos foram realizados entre as passagens 3 e 5. Para essas análises, as MSCs foram cultivadas sobre arcabouços de polímeros biorreabsorvíveis de poli (hidroxibutirato-co-hidroxivalerato) (PHBV) e o poli (-caprolactona) (PCL). As MSCs (GIBCO®) foram inoculadas nos polímeros puros e na mistura 75:25 de PHBV / PCL (amostras densas e porosas). As células foram mantidas em DMEM (com baixa glicose) contendo GlutaMAX® e 10% de SFB a 37oC com 5% de CO2 por 21 dias. A própria placa foi usada como controle. Os meios de cultura condicionados foram coletados e analisadas em FT-Raman para sondagem de grupos funcionais, bem como possíveis variações moleculares associadas com a diferenciação e metabolismo celular. Foi possível discernir grupos funcionais de moléculas específicas no meio condicionado, como colesterol, fosfatidilinositol, triglicerídeos, forma Beta de polipeptídeos, regiões de amida e ligações de hidrogênio de proteínas, além da expressão de DNA. Na presente avaliação, a FT-Raman apresentou como uma técnica de resolução limitada, uma vez que modos vibracionais de estiramento próximos ou mesmo iguais podem ser expressos por moléculas diferente, dificultando a análise. Não houve variações nas leituras entre as amostras estudadas, concluindo-se que a FT-Raman não atendeu às expectativas nas condições estudadas.

Hancornia speciosa serum latex fraction: a non-allergenic biomaterial / Fração soro do látex de Hancornia speciosa: um biomaterial não alergênico

Abstract Plant-derived products can assist in the healing process of dermal wounds. It has been demonstrated that Hancornia speciosa latex present angiogenic, osteogenic, anti-inflammatory, and antioxidant activities. Then, it could contribute to the wound healing process. However, natural products in contact with skin may cause dermatitis. The objective of this work was to evaluate the allergic and irritant potential of H. speciosa serum fraction latex using in vitro assays. The obtained results showed that the H. speciosa serum fraction latex has a slightly irritant potential and is not cytotoxic neither allergenic for human cells. Moreover, we identified a remarkable low amount of proteins in this material in comparison to Hevea brasiliensis latex. This result could explain the non-allergenic potential of H. speciosa serum fraction latex because proteins present in latex are the main responsible for allergy. This biomaterial could be used as a non-allergenic source for development of new medicines.

Resumo Produtos derivados de plantas podem auxiliar no processo de cicatrização de feridas cutâneas. Foi demonstrado que o látex de Hancornia speciosa apresenta atividades angiogênicas, osteogênicas, antiinflamatórias e antioxidantes. Então, este biomaterial pode contribuir para o processo de cicatrização de feridas. No entanto, produtos naturais em contato com a pele podem causar dermatites. O objetivo deste trabalho foi avaliar o potencial alérgico e irritante do látex da fração soro de H. speciosa por meio de ensaios in vitro. Os resultados obtidos mostraram que o látex da fração do soro de H. speciosa possui um potencial pouco irritante e não é citotóxico nem alergênico para células humanas. Além disso, foi identificado uma notável baixa quantidade de proteínas neste material em comparação ao látex de Hevea brasiliensis. Esse resultado poderia explicar o potencial não alergênico do látex da fração soro de H. speciosa, pois as proteínas presentes no látex são as principais responsáveis ​​pela alergia. Este biomaterial pode ser utilizado como fonte não alergênica para desenvolvimento de novos medicamentos.

Does low-intensity pulsed ultrasound accelerate phasic calcium phosphate ceramic-induced bone formation?

Purpose: Low-intensity pulsed ultrasound (LIPUS) has been used to stimulate the healing of the fresh fracture, delayed union, and non-union in both animal and clinical studies. Besides, biphasic calcium phosphate ceramic (BCP) is a promising biomaterial for bone repair as it shows favorable biocompatibility, osteoinduction, and osteoconduction. However, scarcity is known about the combined effect of LIPUS and BCP on bone formation. Methods: The combined effect of LIPUS and BCP was studied in a beagle model. Twelve dogs were used. BCP granules without any additions were implanted into bilateral erector spinae muscles. One side is the BCP group, while the counterlateral side is LIPUS + BCP group. Histological and histomorphometric analyses, and quantitative real-time polymerase chain reaction were evaluated. Results: Compared with BCP alone, the LIPUS + BCP showed no advantages in early bone formation. Furthermore, the Notch signaling pathway-related mRNA has no significant difference between the two groups. Conclusions: The preliminary results showed that the BCP, which has intrinsic osteoinduction nature, was an effective and promising material. However, LIPUS has no enhanced effect in BCP induced ectopic bone formation. Furthermore, LIPUS has no effect on the Notch signaling pathway. Whether costly LIPUS could be used in combination with BCP should be a rethink.

Culture of rat mesenchymal stem cells on PHBV-PCL scaffolds: analysis of conditioned culture medium by FT Raman spectroscopy / Cultivo de células-tronco mesenquimais de rato sobre arcabouços de PHBV-PCL: análise do meio de cultura condicionado por espectroscopia de FT-Raman

Mesenchymal stem cells (MSCs) have great potential for application in cell therapy and tissue engineering procedures because of their plasticity and capacity to differentiate into different cell types. Given the widespread use of MSCs, it is necessary to better understand some properties related to osteogenic differentiation, particularly those linked to biomaterials used in tissue engineering. The aim of this study was to develop an analysis method using FT-Raman spectroscopy for the identification and quantification of biochemical components present in conditioned culture media derived from MSCs with or without induction of osteogenic differentiation. All experiments were performed between passages 3 and 5. For this analysis, MSCs were cultured on scaffolds composed of bioresorbable poly(hydroxybutyrate co-hydroxyvalerate) (PHBV) and poly(ε-caprolactone) (PCL) polymers. MSCs (GIBCO®) were inoculated onto the pure polymers and 75:25 PHBV/PCL blend (dense and porous samples). The plate itself was used as control. The cells were maintained in DMEM (with low glucose) containing GlutaMAX® and 10% FBS at 37ºC with 5% CO2 for 21 days. The conditioned culture media were collected and analyzed to probe for functional groups, as well as possible molecular variations associated with cell differentiation and metabolism. The method permitted to identify functional groups of specific molecules in the conditioned medium such as cholesterol, phosphatidylinositol, triglycerides, beta-subunit polypeptides, amide regions and hydrogen bonds of proteins, in addition to DNA expression. In the present study, FT-Raman spectroscopy exhibited limited resolution since different molecules can express similar or even the same stretching vibrations, a fact that makes analysis difficult. There were no variations in the readings between the samples studied. In conclusion, FT-Raman spectroscopy did not meet expectations under the conditions studied.

As células-tronco mesenquimais (MSCs) possuem grande potencial para aplicação em procedimentos terapêuticos ligados a terapia celular e engenharia de tecidos, considerando-se a plasticidade e capacidade de formação em diferentes tipos celulares por elas. Dada a abrangência no emprego das MSCs, há necessidade de se compreender melhor algumas propriedades relacionadas à diferenciação osteogênica, particularmente liga à biomateriais usados em engenharia de tecidos. Este projeto objetiva o desenvolvimento de uma metodologia de análise empregando-se a FT-Raman para identificação e quantificação de componentes bioquímicos presentes em meios de cultura condicionados por MSCs, com ou sem indução à diferenciação osteogênica. Todos os experimentos foram realizados entre as passagens 3 e 5. Para essas análises, as MSCs foram cultivadas sobre arcabouços de polímeros biorreabsorvíveis de poli (hidroxibutirato-co-hidroxivalerato) (PHBV) e o poli (ε-caprolactona) (PCL). As MSCs (GIBCO®) foram inoculadas nos polímeros puros e na mistura 75:25 de PHBV / PCL (amostras densas e porosas). As células foram mantidas em DMEM (com baixa glicose) contendo GlutaMAX® e 10% de SFB a 37oC com 5% de CO2 por 21 dias. A própria placa foi usada como controle. Os meios de cultura condicionados foram coletados e analisadas em FT-Raman para sondagem de grupos funcionais, bem como possíveis variações moleculares associadas com a diferenciação e metabolismo celular. Foi possível discernir grupos funcionais de moléculas específicas no meio condicionado, como colesterol, fosfatidilinositol, triglicerídeos, forma Beta de polipeptídeos, regiões de amida e ligações de hidrogênio de proteínas, além da expressão de DNA. Na presente avaliação, a FT-Raman apresentou como uma técnica de resolução limitada, uma vez que modos vibracionais de estiramento próximos ou mesmo iguais podem ser expressos por moléculas diferente, dificultando a [...].

Culture of rat mesenchymal stem cells on PHBV-PCL scaffolds: analysis of conditioned culture medium by FT-Raman spectroscopy / Cultivo de células-tronco mesenquimais de rato sobre arcabouços de PHBV-PCL: análise do meio de cultura condicionado por espectroscopia de FT-Raman

Abstract Mesenchymal stem cells (MSCs) have great potential for application in cell therapy and tissue engineering procedures because of their plasticity and capacity to differentiate into different cell types. Given the widespread use of MSCs, it is necessary to better understand some properties related to osteogenic differentiation, particularly those linked to biomaterials used in tissue engineering. The aim of this study was to develop an analysis method using FT-Raman spectroscopy for the identification and quantification of biochemical components present in conditioned culture media derived from MSCs with or without induction of osteogenic differentiation. All experiments were performed between passages 3 and 5. For this analysis, MSCs were cultured on scaffolds composed of bioresorbable poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) and poly(ε-caprolactone) (PCL) polymers. MSCs (GIBCO®) were inoculated onto the pure polymers and 75:25 PHBV/PCL blend (dense and porous samples). The plate itself was used as control. The cells were maintained in DMEM (with low glucose) containing GlutaMAX® and 10% FBS at 37oC with 5% CO2 for 21 days. The conditioned culture media were collected and analyzed to probe for functional groups, as well as possible molecular variations associated with cell differentiation and metabolism. The method permitted to identify functional groups of specific molecules in the conditioned medium such as cholesterol, phosphatidylinositol, triglycerides, beta-subunit polypeptides, amide regions and hydrogen bonds of proteins, in addition to DNA expression. In the present study, FT-Raman spectroscopy exhibited limited resolution since different molecules can express similar or even the same stretching vibrations, a fact that makes analysis difficult. There were no variations in the readings between the samples studied. In conclusion, FT-Raman spectroscopy did not meet expectations under the conditions studied.

Resumo As células-tronco mesenquimais (MSCs) possuem grande potencial para aplicação em procedimentos terapêuticos ligados a terapia celular e engenharia de tecidos, considerando-se a plasticidade e capacidade de formação em diferentes tipos celulares por elas. Dada a abrangência no emprego das MSCs, há necessidade de se compreender melhor algumas propriedades relacionadas à diferenciação osteogênica, particularmente liga à biomateriais usados em engenharia de tecidos. Este projeto objetiva o desenvolvimento de uma metodologia de análise empregando-se a FT-Raman para identificação e quantificação de componentes bioquímicos presentes em meios de cultura condicionados por MSCs, com ou sem indução à diferenciação osteogênica. Todos os experimentos foram realizados entre as passagens 3 e 5. Para essas análises, as MSCs foram cultivadas sobre arcabouços de polímeros biorreabsorvíveis de poli (hidroxibutirato-co-hidroxivalerato) (PHBV) e o poli (ε-caprolactona) (PCL). As MSCs (GIBCO®) foram inoculadas nos polímeros puros e na mistura 75:25 de PHBV / PCL (amostras densas e porosas). As células foram mantidas em DMEM (com baixa glicose) contendo GlutaMAX® e 10% de SFB a 37oC com 5% de CO2 por 21 dias. A própria placa foi usada como controle. Os meios de cultura condicionados foram coletados e analisadas em FT-Raman para sondagem de grupos funcionais, bem como possíveis variações moleculares associadas com a diferenciação e metabolismo celular. Foi possível discernir grupos funcionais de moléculas específicas no meio condicionado, como colesterol, fosfatidilinositol, triglicerídeos, forma Beta de polipeptídeos, regiões de amida e ligações de hidrogênio de proteínas, além da expressão de DNA. Na presente avaliação, a FT-Raman apresentou como uma técnica de resolução limitada, uma vez que modos vibracionais de estiramento próximos ou mesmo iguais podem ser expressos por moléculas diferente, dificultando a análise. Não houve variações nas leituras entre as amostras estudadas, concluindo-se que a FT-Raman não atendeu às expectativas nas condições estudadas.

Uso da termografia e ultrassonografia power dopplerpara avaliação da resposta tecidual em falhas ósseas preenchidas com biomaterial em ovinos / Termography and power doppler sonography assessment for tissue response evaluation in sheep bone defects filled with biomaterial

Biomateriaistem diversas indicações como auxiliares no processo de reparação óssea, além de terem função de substituto ósseo em perdas extensas. Diversas são as vantagens de sua utilização, como por exemplo, auxílio na osteocondutividade, estímulo de neovascularização, potencial antimicrobiano, entre outros. Falhas ósseas foram realizadas nas tíbias de seis ovinos da raça Santa Inês e preenchidas com biomaterial à base de quitosana, colágeno e hidroxiapatita. Um membro foi considerado controle e outro membro tratado segundo estudo randomizado. Foram realizadas avaliações termográficas e por ultrassonografia Power Dopplerem todos os animais do estudo, semanalmente, nos dias D0, D7, D14, D21, D28, D35, D42 e D56. Não houve diferenças significativas com relação à temperatura mínima, máxima e média entre os grupos com biomaterial e controle nas imagens termográficas. Houveram variações com relação ao tempo dentro de ambos os grupos. Com relação à presença de vasos na ultrassonografia Power Dopplernão houve diferenças estatísticas entre os grupos, exceto no dia 21 (P=0,031). Dentro das possibilidades de avaliação que os exames de imagem fornecem, a termografia e a ultrassonografia Power Dopplermostraram-se ferramentas não invasivas de avaliação pós-operatória de processo inflamatório e neovascularização, sendo realizadas semanalmente, permitindo acompanhamento fidedigno e detalhado ao longo do experimento, sem gerar desconforto ou estresse aos animais. Não houve indícios de complicações relacionadas ao biomaterial.(AU)

Biomaterials have several indications supporting the bone repair process, besides having bone substitute function in extensive losses. There are several advantages of its use, such as contribution in osteoconductivity, stimulation of neovascularization, antimicrobial potential, among others.Tibial bone defects was performed in six Santa Inês breed ewes and implanted with chitosan, collagen and hydroxyapatite biomaterial. One limb was considered the control limb and the other one the biomateriallimb, chosenrandomly. Thermographic and Power Dopplersonography was performed in all animals, weekly for 56 days. The thermography showed no significant differences related to low, high and mean temperature between the control and the biomaterial groups. Statisticalvariations were found between time in both groups. The visualization of neovascularization with the Power Dopplersonography did not present statistical differences, except for day 21 (P=0,031). Within the possibilities provided by imaging exams, thermography and Power Doppler sonography demonstrate to be non-invasive methods for post-operative evaluation of inflammatory process and neovascularization. The weekly management allowed reliable and detailed monitoring throughout the experiment, without causingdiscomfort or stress to the animals. There was no evidence of biomaterial complications.(AU)

Culture of rat mesenchymal stem cells on PHBV-PCL scaffolds: analysis of conditioned culture medium by FT Raman spectroscopy / Cultivo de células-tronco mesenquimais de rato sobre arcabouços de PHBV-PCL: análise do meio de cultura condicionado por espectroscopia de FT-Raman

Mesenchymal stem cells (MSCs) have great potential for application in cell therapy and tissue engineering procedures because of their plasticity and capacity to differentiate into different cell types. Given the widespread use of MSCs, it is necessary to better understand some properties related to osteogenic differentiation, particularly those linked to biomaterials used in tissue engineering. The aim of this study was to develop an analysis method using FT-Raman spectroscopy for the identification and quantification of biochemical components present in conditioned culture media derived from MSCs with or without induction of osteogenic differentiation. All experiments were performed between passages 3 and 5. For this analysis, MSCs were cultured on scaffolds composed of bioresorbable poly(hydroxybutyrate co-hydroxyvalerate) (PHBV) and poly(ε-caprolactone) (PCL) polymers. MSCs (GIBCO®) were inoculated onto the pure polymers and 75:25 PHBV/PCL blend (dense and porous samples). The plate itself was used as control. The cells were maintained in DMEM (with low glucose) containing GlutaMAX® and 10% FBS at 37ºC with 5% CO2 for 21 days. The conditioned culture media were collected and analyzed to probe for functional groups, as well as possible molecular variations associated with cell differentiation and metabolism. The method permitted to identify functional groups of specific molecules in the conditioned medium such as cholesterol, phosphatidylinositol, triglycerides, beta-subunit polypeptides, amide regions and hydrogen bonds of proteins, in addition to DNA expression. In the present study, FT-Raman spectroscopy exhibited limited resolution since different molecules can express similar or even the same stretching vibrations, a fact that makes analysis difficult. There were no variations in the readings between the samples studied. In conclusion, FT-Raman spectroscopy did not meet expectations under the conditions studied.(AU)

As células-tronco mesenquimais (MSCs) possuem grande potencial para aplicação em procedimentos terapêuticos ligados a terapia celular e engenharia de tecidos, considerando-se a plasticidade e capacidade de formação em diferentes tipos celulares por elas. Dada a abrangência no emprego das MSCs, há necessidade de se compreender melhor algumas propriedades relacionadas à diferenciação osteogênica, particularmente liga à biomateriais usados em engenharia de tecidos. Este projeto objetiva o desenvolvimento de uma metodologia de análise empregando-se a FT-Raman para identificação e quantificação de componentes bioquímicos presentes em meios de cultura condicionados por MSCs, com ou sem indução à diferenciação osteogênica. Todos os experimentos foram realizados entre as passagens 3 e 5. Para essas análises, as MSCs foram cultivadas sobre arcabouços de polímeros biorreabsorvíveis de poli (hidroxibutirato-co-hidroxivalerato) (PHBV) e o poli (ε-caprolactona) (PCL). As MSCs (GIBCO®) foram inoculadas nos polímeros puros e na mistura 75:25 de PHBV / PCL (amostras densas e porosas). As células foram mantidas em DMEM (com baixa glicose) contendo GlutaMAX® e 10% de SFB a 37oC com 5% de CO2 por 21 dias. A própria placa foi usada como controle. Os meios de cultura condicionados foram coletados e analisadas em FT-Raman para sondagem de grupos funcionais, bem como possíveis variações moleculares associadas com a diferenciação e metabolismo celular. Foi possível discernir grupos funcionais de moléculas específicas no meio condicionado, como colesterol, fosfatidilinositol, triglicerídeos, forma Beta de polipeptídeos, regiões de amida e ligações de hidrogênio de proteínas, além da expressão de DNA. Na presente avaliação, a FT-Raman apresentou como uma técnica de resolução limitada, uma vez que modos vibracionais de estiramento próximos ou mesmo iguais podem ser expressos por moléculas diferente, dificultando a [...].(AU)

Efficiency analysis of commercial polymeric membranes for bone regeneration in rat cranial defects

Purpose: To evaluate the in vivo efficiency of commercial polymeric membranes for guided bone regeneration. Methods: Rat calvarial critical size defects was treated with LuminaCoat (LC), Surgitime PTFE (SP), GenDerm (GD), Pratix (PR), Techgraft (TG) or control (C-) and histomorphometric analysis determined the percentage of new bone, connective tissue and biomaterial at 1 or 3 months. Statistical analysis used ANOVA with Tukey's post-test for means at same experimental time and the paired Student's t test between the two periods, considering p < 0.05. Results: New bone at 1 month was higher for SP, TG and C-, at 3 months there were no differences, and between 1 and 3 months PR had greater increase growthing. Connective tissue at 1 month was higher for C-, at 3 months for PR, TG and C-, and between 1 and 3 months C- had sharp decline. Biomaterial at 1 month was higher for LC, in 3 months for SP and TG, and between 1 and 3 months, LC, GD and TG had more decreasing mean. Conclusion: SP had greater osteopromotive capacity and limitation of connective ingrowth, but did not exhibit degradation. PR and TG had favorable osteopromotion, LC less connective tissue and GD more accelerated biodegradation.

Comparative study of bullfrog (Rana catebeiana) skin preservation for use as a biomaterial / Estudo comparativo de meios de conservação de pele de rã-touro (Rana catesbeiana) para utilização como biomaterial

The use of biological membranes in wound dressings has increasingly become a reality. Accordingly, an ideal means of preservation is sought that can provide tissue maintenance for long periods without interfering with its quality or clinical applicability. Therefore, the objective of the present study was to evaluate and histologically and microbiologically compare frog skins subjected to two different preservation methods. Sixteen frog skins were evaluated and, depending on the preservation method, subdivided into two groups with eight skins each, namely, the Freezing Group, in which the skins were frozen at -4º in a 20% glycerin solution; and the Glycerin Group, whose skins were kept in 98% glycerin at room temperature (average of 28 ºC and average humidity of 78%). The skins were analyzed fresh (T0) and at 30 (T1), 60 (T2), 90 (T3), and 120 (T4) days of preservation. Data were analyzed comparatively. There was no bacterial or fungal growth, and the skin structure and collagen arrangement remained intact at all time points in both treatments. In conclusion, both preservation methods are efficient and capable of maintaining the tissue morphological structure and preventing the growth and proliferation of contaminants for up to 120 days.

A utilização de membranas biológicas em curativos tem se tornado cada vez mais uma realidade. Concomitante, busca-se um meio de conservação ideal que possa proporcionar a manutenção do tecido por longos períodos de tempo sem interferir em sua qualidade e aplicabilidade clínica. Desta forma, o objetivo deste trabalho foi o de avaliar e comparar histologicamente e microbiologicamente peles de rã submetidas a dois diferentes métodos de conservação. Foram avaliadas 16 peles de rã-touro as quais foram, em função do método de conservação, subdivididas em dois grupos com 08 peles cada: O Grupo Congelamento (GC) no qual as peles foram submetidas ao congelamento a -4º, em solução de glicerina a 20%; e o grupo Glicerina (GG), no qual as peles foram conservadas em temperatura ambiente (média de 28ºC e umidade média de 78%) em glicerina a 98%. As peles foram analisadas a fresco (T0) e com 30 (T1), 60 (T2), 90 (T3) e 120 (T4) dias de conservação. Os dados foram analisados de forma comparativa. Em todos os tempos analisados e em ambos os tratamentos, não houve crescimento bacteriano ou fúngico e a estrutura da pele e o arranjo de colágeno mantiveram-se íntegros. Conclui-se que, ambos os métodos de conservação são eficientes e capazes de manter a estrutura morfológica tecidual, e impedem o crescimento e a proliferação de contaminantes por até 120 dias.

Effect of ascorbic acid and epidermal growth factor in a rat tibia defect

Purpose: Bone repair aims to restore the anatomical, biomechanical, and functional integrity of the affected structure. Here we study the effects of ascorbic acid (AA) and epidermal growth factor (EGF) applied in a single dose and in combination on the repair of a noncritical bone defect model. Methods: Twenty-four rats were divided into four groups: an intact G-1 control group, and three groups that underwent a noncritical bone defect in the right tibia: G-2 treated with AA, G-3 treated with EGF, and G-4 treated with AA in combination with EGF. After 21 days of treatment, rats were sacrificed, the tibias were dissected and a destructive biomechanical analysis of three-point flexion test was performed in a universal testing machine; the values of stiffness, resistance, maximum energy, and energy at maximum load were statistically compared. Results: G-3 and G-4 recovered the biomechanical properties of strength and stiffness of an intact tibia 3 weeks after their application. Not so the energy and energy at maximum load. For G-2, only the stiffness of an intact tibia was recovered. Conclusion: EGF and AA-EGF applied to a noncritical bone defect in the rat tibia favors the recovery of bone resistance and stiffness.

Development and characterization of poultry collagen-based hybrid hydrogels for bone regeneration

Purpose: Poultry by-products can contribute as an innovative natural source for the development of composites based on polymers and minerals aiming at bone regeneration. The objective of this study was the physicochemical and biological characterization of collagen-based hydrogels crosslinked with ultraviolet (UV)-riboflavin. Methods: Pure hydrogels of 100% collagen (G1) or hybrid hydrogels, 90% collagen:10% apatite (G2), 90% collagen:10% nanokeratin (G3), and 90% collagen:5% apatite:5% nanokeratin (G4) were characterized by scanning electron microscope, Fourier-transform infrared spectroscopy, differential scanning calorimetry, swelling degree and quali-quantitative histological analysis. Ectopic implantation in subcutaneous tissue in mice at one, three and nine weeks allowed to assess the inflammation (neutrophils, lymphocytes, macrophages, and giant cells) and repair (neovascularization, and connective tissue) to determine biocompatibility and the integrity of biomaterials to score their biodegradability. Histomorphometry on critical size defects in rat calvaria at one and three months evaluated the percentage of bone, connective tissue, and biomaterials in all groups. Results: The hydrogels presented porous microstructure, water absorption and physicochemical characteristics compatible with their polymeric and/or mineral composition. All materials exhibited biocompatibility, biodegradability, and low osteoconductivity. G2 showed greater density of new bone and biomaterial than the G1, G3 and G4. Conclusions: The collagen-apatite group formulation suggests potential for development as osteopromoting membrane.

A biocomplex to repair experimental critical size defects associated with photobiomodulation therapy

Abstract Background: The association of scaffolds to repair extensive bone defects can contribute to their evolution and morphophysiological recomposition. The incorporation of particulate biomaterials into three-dimensional fibrin bioproducts together with photobiomodulation therapy (PBM) has potential and can improve regenerative medicine procedures. The objective of this experiment was to evaluate the effects of PBM therapy on critical size defects filled with xenogenic bone substitute associated with fibrin biopolymer. Methods: A critical defect of 8 mm was performed in 36 Wistar male adult rats that were divided into four groups. Groups BC and BC-PBM were defined as controls with defects filled by a clot (without or with PBM, respectively) and groups XS and XS-PBM that comprised those filled with biocomplex Bio-OssTM in association with fibrin biopolymer. PBM was applied immediately after the surgery and three times a week every other day, with the parameters: wavelength of 830 nm, energy density 6.2 J/cm2, output power 30 mW, beam area of 0.116 cm2, irradiance 0.258,62 W/cm2, energy/point 0.72 J, total energy 2.88 J. Fourteen and 42 days after the surgery, animals were euthanatized and subjected to microtomography, qualitative and quantitative histological analysis. Results: The BC-PBM and XS-PBM groups had a similar evolution in the tissue repair process, with a higher density of the volume of new formed bone in relation to the groups without PBM (p = 0.04086; p = 0.07093, respectively). Intense vascular proliferation and bone deposition around the biomaterial particles were observed in the animals of the groups in which biocomplex was applied (XS and XS-PBM). Conclusion: PBM therapy allowed an improvement in the formation of new bone, with a more organized deposition of collagen fibers in the defect area. Biocomplex favored the insertion and permanence of the particulate material in bone defects, creating a favorable microenvironment for accelerate repair process.

A biocomplex to repair experimental critical size defects associated with photobiomodulation therapy

The association of scaffolds to repair extensive bone defects can contribute to their evolution and morphophysiological recomposition. The incorporation of particulate biomaterials into three-dimensional fibrin bioproducts together with photobiomodulation therapy (PBM) has potential and can improve regenerative medicine procedures. The objective of this experiment was to evaluate the effects of PBM therapy on critical size defects filled with xenogenic bone substitute associated with fibrin biopolymer. Methods: A critical defect of 8 mm was performed in 36 Wistar male adult rats that were divided into four groups. Groups BC and BC-PBM were defined as controls with defects filled by a clot (without or with PBM, respectively) and groups XS and XS-PBM that comprised those filled with biocomplex Bio-OssTM in association with fibrin biopolymer. PBM was applied immediately after the surgery and three times a week every other day, with the parameters: wavelength of 830 nm, energy density 6.2 J/cm2, output power 30 mW, beam area of 0.116 cm2, irradiance 0.258,62 W/cm2, energy/point 0.72 J, total energy 2.88 J. Fourteen and 42 days after the surgery, animals were euthanatized and subjected to microtomography, qualitative and quantitative histological analysis. Results: The BC-PBM and XS-PBM groups had a similar evolution in the tissue repair process, with a higher density of the volume of new formed bone in relation to the groups without PBM (p = 0.04086; p = 0.07093, respectively). Intense vascular proliferation and bone deposition around the biomaterial particles were observed in the animals of the groups in which biocomplex was applied (XS and XS-PBM). Conclusion: PBM therapy allowed an improvement in the formation of new bone, with a more organized deposition of collagen fibers in the defect area. Biocomplex favored the insertion and permanence of the particulate material in bone defects, creating a favorable microenvironment for accelerate repair process.(AU)

Biobancos para a conservação da vida silvestre: desafios e perspectivas / Biobanks for wildlife conservation: challenges and prospects

Um repositório de amostras biológicas, ou biobanco, pode ser definido como uma coleção-base de amostras orgânicas de origem humana, animal, vegetal ou microbiana, destinados para a conservação da ampla variabilidade dos recursos genéticos, apresentando-se como uma alternativa para a manutenção da biodiversidade. Esses repositórios são instrumentos indispensáveis no proposito de salvaguardar e coordenar um conjunto de informações da fauna silvestre, dando origem a uma rede de dados para o desenvolvimento de estratégias para a conservação de espécimes com genótipos raros ou ameaçados. Portanto, esta revisão tem como objetivo ressaltar a importância e aplicabilidade dos biobancos para a conservação da vida silvestre, destacando o estado da arte, e enfatizando os aspectos técnicos, procedimentos necessários para sua implementação e quais os principais desafios e pespectivas para o aprimoramento e utilização desse recurso na biotecnologia reprodutiva.(AU)

A repository of biological samples or biobanks can be defined as a base collection of organic samples of human, animal, plant or microbial origin, intended for the conservation of the wide variability of genetic resources, presenting itself as an alternative for the maintenance of the biodiversity. These repositories are indispensable instruments for the purpose of safeguarding and coordinating a set of information on wildlife, giving rise to a data network for the development of strategies for the conservation of specimens with rare or endangered genotypes. Therefore, this review aims to highlight the importance and applicability of biobanks for the conservation of wildlife, highlighting the state of the art, and emphasizing the technical aspects, as well as the necessary procedures for its implementation and what are the main challenges and perspectives for the improvement and use of this resource in reproductive biotechnology.(AU)

Biocompatibility in vivo of elastic cartilage treated in alkaline solutions / [Biocompatibilidade in vivo de cartilagem elástica tratada em soluções alcalinas]

This study verified the in vivo biocompatibility of bovine elastic cartilage decellularized with alkaline solution in relation to the non-decellularized cartilage implanted in rats. Fifty Wistar rats were divided into two groups, with the experimental group (EG) receiving subcutaneous implants of cartilage treated in alkaline solution and the control group (CG) receiving subcutaneous implants of untreated cartilage. In both groups, the implants were removed on days 3, 7, 14, 21 and 42 with the preparation of histological slides stained with Hematoxylin and Eosin for the quantification of inflammatory cells, fibroblasts, angiogenesis, and cartilage degradation. The results showed that EG presented a less intense inflammatory infiltrate and better organization of collagen fibers compared to CG. It was concluded that the alkaline treatment provided better biocompatibility for elastic cartilage when implanted subcutaneously in rats.

Este estudo verificou a biocompatibilidade in vivo de cartilagem elástica bovina descelularizada com solução alcalina em relação à cartilagem não descelularizada implantada em ratos. Cinquenta ratos Wistar foram divididos em dois grupos; o grupo experimental (GE) recebeu implantes subcutâneos de cartilagem tratada em solução alcalina, e o grupo controle (GC) recebeu implantes subcutâneos de cartilagem não tratada. Em ambos os grupos, os implantes foram retirados nos dias três, sete, 14, 21 e 42, com a preparação de lâminas histológicas coradas com hematoxilina e eosina para a quantificação de células inflamatórias, de fibroblastos, de angiogênese e para a degradação da cartilagem. Os resultados mostraram que o GE apresentou infiltrado inflamatório menos intenso e melhor organização das fibras de colágeno em relação ao GC. Concluiu-se que o tratamento alcalino proporcionou melhor biocompatibilidade para a cartilagem elástica quando implantado por via subcutânea em ratos.

Femoral orthopedic implants in dogs with titanium - mechanical evaluation

Background: Orthopedic implants are commonly used for different types of surgical procedures to gain optimal functionand to provide stability to both bones and tendon structures. When inserting these implants, the characteristics of the material are important for surgical success, and the ideal implant must be biocompatible and nonallergenic. However, whenmolding an implant to the bone structure, its resistance can change significantly. Implants can be temporary or permanentin the body, and metal possesses properties that make it acceptable for bone repair. In biomedical implants, 2 types aremost common, commercially pure (CP)-Ti and Ti-6A1-4V. They both provide stable fixation and low risk of loosening.Implants made with the same material and composition can perform differently if the material has been altered by processing techniques for different scenarios. Stress, strain and elastic modulus are the primary metrics used in the description ofimplant materials. They can be calculated based on mechanical tests of specimens with defined geometry, most commonlytensile, bending and torsional tests. In order to better evaluate those changes, we compared the mechanical characteristicsof titanium bone plates, before and after they were molded to the bone, to verify and quantify the loss of stiffness andresistance after molding the plate.Materials, Methods & Results: The study was prospective. Orthopedic implant made of commercially pure titanium (CPTi) were divided into 2 groups, one group without plate molding and the other with plate molding to a dog femora bone.Thirty-six plates of different sizes (5.0, 6.5, 8.0, 9.0, 10.0- and 11.0-mm diameter) were divided into 6 groups containing6 plates of each size and submitted to the 4-point flexion test of resistance, using a piece of dog femur (weights of 5, 10,15, 20 and 25 kg) as the bone in which the...

Femoral orthopedic implants in dogs with titanium - mechanical evaluation

Background: Orthopedic implants are commonly used for different types of surgical procedures to gain optimal functionand to provide stability to both bones and tendon structures. When inserting these implants, the characteristics of the material are important for surgical success, and the ideal implant must be biocompatible and nonallergenic. However, whenmolding an implant to the bone structure, its resistance can change significantly. Implants can be temporary or permanentin the body, and metal possesses properties that make it acceptable for bone repair. In biomedical implants, 2 types aremost common, commercially pure (CP)-Ti and Ti-6A1-4V. They both provide stable fixation and low risk of loosening.Implants made with the same material and composition can perform differently if the material has been altered by processing techniques for different scenarios. Stress, strain and elastic modulus are the primary metrics used in the description ofimplant materials. They can be calculated based on mechanical tests of specimens with defined geometry, most commonlytensile, bending and torsional tests. In order to better evaluate those changes, we compared the mechanical characteristicsof titanium bone plates, before and after they were molded to the bone, to verify and quantify the loss of stiffness andresistance after molding the plate.Materials, Methods & Results: The study was prospective. Orthopedic implant made of commercially pure titanium (CPTi) were divided into 2 groups, one group without plate molding and the other with plate molding to a dog femora bone.Thirty-six plates of different sizes (5.0, 6.5, 8.0, 9.0, 10.0- and 11.0-mm diameter) were divided into 6 groups containing6 plates of each size and submitted to the 4-point flexion test of resistance, using a piece of dog femur (weights of 5, 10,15, 20 and 25 kg) as the bone in which the...(AU)

Influence of silver nanoparticles on the tissue reaction of polyacrylic acid-based gel

Purpose: To study the influence of silver nanoparticles (AgNP) on tissue reaction when incorporated into a polymeric matrix of polyacrylic acid-based (Carbopol®) gel as a proposal for a new low-cost type of biomaterial that is simple to manufacture for use as an antimicrobial and antioxidative dressing. Methods: In-vivo tests of implantation in the subcutaneous tissue of the back of rats were performed using polyethylene tubes in three situations: empty, only the gel, and gel incorporated with AgNP. Then, the tissue reaction was studied by counting inflammatory cells. Additionally, in-vitro tests of the antioxidative and antimicrobial activity of AgNP were performed. The radical 2,2 diphenyl-1 picrylhydrazyl (DPPH) was used to test the antioxidative activity of AgNP using electron spin resonance. The antimicrobial activity of AgNP was determined by minimum inhibitory concentration against the microorganisms: Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli. Results: The results indicated that AgNP presents antioxidative activity and was able to inhibit the growth of the microorganisms tested. The addition of AgNP in Carbopol® did not alter the tissue inflammatory response (p>0.05, Kruskal-Wallis's test). Conclusions: The new biomaterial is promising for future use as a dressing for its beneficial properties for regenerative processes

Biological behavior of magnesium-substituted hydroxyapatite during bone repair / Comportamento biológico de hidroxiapatita substituída com magnésio durante o reparo ósseo

The aim of this study was to analyze the biological behavior and osteogenic potential of magnesium (Mg) substituted hydroxyapatite (HA) microspheres, implanted in a critical bone defect, considering that this ion is of great clinical interest, since it is closely associated with homeostasis and bone mineralization. For the purpose of this study, 30 rats were used to compose three experimental groups: GI - bone defect filled with HA microspheres; GII - bone defect filled with HA microspheres replaced with Mg; GIII - empty bone defect; evaluated at biological points of 15 and 45 days. The histological results, at 15 days, showed, in all the groups, a discrete chronic inflammatory infiltrate; biomaterials intact and surrounded by connective tissue; and bone neoformation restricted to the borders. At 45 days, in the GI and GII groups, an inflammatory response of discrete granulomatous chronic type was observed, and in the GIII there was a scarce presence of mononuclear inflammatory cells; in GI and GII, the microspheres were seen to be either intact or fragmented, surrounded by fibrous connective tissue rich in blood vessels; and discrete bone neoformation near the edges and surrounding some microspheres. In GIII, the mineralization was limited to the borders and the remaining area was filled by fibrous connective tissue. It was concluded that the biomaterials were biocompatible and osteoconductive, and the percentage of Mg used as replacement ion in the HA did not favor a greater bone neoformation in relation to the HA without the metal.(AU)

O objetivo deste estudo foi analisar o comportamento biológico de microesferas de hidroxiapatita (HA) substituída com magnésio (Mg) durante o reparo de defeito ósseo crítico, tendo em vista que este íon é de grande interesse clínico, pois está intimamente associado à homeostasia e à mineralização óssea. Para tanto, utilizou-se 30 ratos para compor três grupos experimentais: GI - defeito ósseo preenchido com microesferas de HA; GII - defeito ósseo preenchido com microesferas de HA substituída com Mg; GIII (controle) - defeito ósseo vazio; avaliados nos pontos biológicos de 15 e 45 dias. Os resultados histológicos evidenciaram, aos 15 dias, discreto infiltrado inflamatório crônico e neoformação óssea restrita às bordas, em todos os grupos. Nos grupos GI e GII, os biomateriais mantiveram-se íntegros e circundados por tecido conjuntivo frouxo. Aos 45 dias, notou-se resposta inflamatória do tipo crônica granulomatosa discreta nos grupos GI e GII, e no GIII presença escassa de células inflamatórias mononucleares. As microesferas implantadas no GI e GII mantiveram-se, em sua maioria, íntegras e envolvidas por tecido conjuntivo fibroso. Notou-se discreta neoformação óssea próxima às bordas e circunjacente a algumas microesferas. No GIII, a mineralização limitou-se às bordas e a área remanescente foi preenchida por tecido conjuntivo fibroso. Conclui-se que os biomateriais foram biocompatíveis, bioativos, osteocondutores e apresentaram biodegradação lenta, indicando seu grande potencial para em aplicações clínica como biomaterial de preenchimento.(AU)

Bovine tunica albuginea conserved in honey as xenograft for cystoplasty in rats / Túnica albugínea bovina conservada em mel como xenoenxerto na cistoplastia em ratos

ABSTRACT: Urinary bladder damages leading to few viable bladder tissue available might demand a challenging reconstructive surgery. In this context, biomaterials are valid alternatives for bladder reconstruction. This study evaluated the bovine tunica albuginea fragment as graft material for cystoplasty in rats and honey-preserved implant viability.Thirty Wistar rats were assigned to two groups: (1) a test group (T) with a circular 1.0-cm-diameterbovine tunica albuginea graft application in the apex region by a continuous absorbable (Polyglactin 910 5-0) suture with stitching of all bladder layers and (2) a simulation group (S) in which animals underwent only partial cystectomy. In addition, each of these groups was further divided into three subgroups according to euthanasia period on post-surgery day 7, 15 and 30.Two animals had self-limiting hematuria at postsurgical period. At necropsy, frequent crystals and adhesion to the peritoneum were observed. At the histopathological evaluation, animals from the T group euthanized by 15th postoperative day had layers disorganization and initial muscle development, while T group rats euthanized by 30th postoperative day showed complete urothelization. Urothelization pattern was similar in both groups. Moreover, the muscular layer formation was present in both groups, but more evident in S group animals. Nevertheless, inflammatory infiltrate and neovascularization were remarkably more intense in T group rats.It might be concluded that bovine tunica albuginea graft was successful in repairing rats' bladder, being a good biomaterial option in reconstructive urinary vesicle surgery.

RESUMO: Lesões na vesícula urinária que culminem em pouco tecido vesical viável podem demandar cirurgias reconstrutivas desafiadoras. Neste contexto, biomateriais são alternativas válidas para a reconstrução da bexiga. O objetivo deste trabalho foi avaliar a túnica albugínea bovina como material para enxerto na cistoplastia em ratos, assim como a viabilidade deste implante preservado em mel. Trinta ratos Wistar foram divididos em dois grupos: (1) um grupo teste (T), no qual os animais foram submetidos a cistectomia parcial seguida de cistoplastia com aplicação do enxerto circular com 1,0cm de diâmetro de túnica albugínea bovina na região do ápice com sutura absorvível (Poliglactina 910 5-0) em padrão contínuo englobando todas as camadas da bexiga; e (2) um grupo simulação (S), cujos ratos realizaram apenas cistectomia parcial. Além disso, cada grupo foi posteriormente dividido em três subgrupos de acordo com a data de eutanásia no dia 7, 15 ou 30 de pós-operatório.Dois animais apresentaram hematúria autolimitante no período de observação pós-operatória. Na necrópsia, notaram-se cristais frequentes e aderências entre bexiga e peritônio. Na avaliação histopatológica, animais do grupo T eutanasiados com 15 dias de pós-operatório apresentavam camadas vesicais desorganizadas e formação de musculatura inicial na bexiga, enquanto aqueles eutanasiados com 30 dias de cirurgia mostraram urotelização completa. O padrão de urotelização foi semelhante nos dois grupos. Além disso, a formação da camada muscular esteve presente em ambos os grupos, porém mais evidente nos animais do grupo S. No entanto, o infiltrado inflamatório bem como a neovascularização foram notavelmente mais intensos nos ratos do grupo T.Pode-se concluir que o enxerto de túnica albugínea bovina foi bem sucedido no reparo da vesícula urinária de ratos, demonstrando ser uma boa opção de biomaterial para cirurgias reconstrutivas da vesícula urinária.