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.

Comportamiento biológico de mallas para la reparación de paredes abdominales. Resultados preliminares en un modelo animal / Biological behavior of meshes used for abdominal wall surgery. Preliminary results in an animal model

Resumen La falla en la reparación de los defectos de la pared abdominal se relaciona con una alteración en la integración del material protésico. El objetivo de este trabajo fue evaluar el comportamiento biológico de mallas utilizadas en cirugía de paredes abdominales en un modelo animal. Luego de la confección de un defecto parietal se colocó una malla intraperitoneal, utilizando 4 grupos de 10 ratas; 1) Prolene (polipropileno microporo de alto peso), 2) Ultrapro (polipropileno + poliglecaprone, macroporo de bajo peso), 3) Proceed (polipropileno + polidoxanona + celulosa oxidada regenerada, macroporo de peso intermedio), y 4) Physiomesh (polipropileno + poliglecaprone, macroporo de bajo peso). Se realizó análisis macroscópico y microscópico a los 30 días y los resultados fueron evaluados por dos observadores independientes. Al examen macroscópico, la integración de la prótesis fue > 75% en todos los grupos. El análisis microscópico mostró mayor inflamación global y número de células gigantes multinucleadas en Prolene (p < 0.01) y menor cantidad de células inflamatorias en la interface músculo-malla en Physiomesh < Ultrapro (p < 0.05). La organización de las fibras de colágeno fue similar para todas las mallas, aunque hubo mayor depósito de colágeno en los espacios inter-filamento para las mallas macroporosas (p < 0.01). Concluimos que las mallas de polipropileno microporo y alto peso producen mayor reacción inflamatoria y de cuerpo extraño. Por lo tanto, las mallas compuestas tendrían una mejor biocompatibilidad y serían mejor toleradas por el huésped.

Abstract An adequate integration of the prosthetic materials used to repair abdominal wall defects is necessary for satisfactory outcomes. We aimed to evaluate, in an animal model, the biological behavior of meshes used for abdominal wall surgery. Four groups of 10 rats were separated. After laparotomy, intraperitoneal prostheses were placed: 1) Prolene (polypropylene microporous, heavy-weight), 2) Ultrapro (polypropylene + poliglecaprone, macroporous low-weight), 3) Proceed (polypropylene + polidoxanone + regenerated oxidized cellulose, microporous medium-weight), 4) Physiomesh (polypropylene + poliglecaprone, macroporous lowweight). Macroscopic and microscopic analyses were performed at 30 days. The results were evaluated by two independent observers and expressed in means with standard deviation. For statistical analysis p < 0.05 was considered significant. On macroscopic examination, mesh integration was greater than 75% in all cases. Microscopic analysis showed greater global inflammation and more multinucleated giant cells in Prolene (p < 0.01). Less inflammatory cells were observed at the muscle-mesh interface in Physiomesh vs. Ultrapro (p < 0.05). Collagen fibers disposition was similar in all meshes, although, microporous meshes had higher collagen deposit in the interfilamentous spaces (p < 0.01). In conclusion, in our animal model, microporous and heavy-weight polypropylene meshes produce greater inflammatory and foreign body reaction. Thus, composite meshes would have greater biocompatibility and better tolerance by the host.

Experimental comparative study of arterial implants made of silicone reinforced with polyester fabric and expanded polytetrafluoroethylene (PTFE) in rabbits aorta / Estudo experimental comparativo de implantes arteriais em silicone com reforço de poliéster e politetrafluoretileno expandido (PTFE) em aorta de coelhos

ABSTRACT Objectives: the aim of this study was to compare the outcomes of a new silicone vascular prostheses with PTFE vascular prostheses, on a rabbit experimental model. Methods: forty rabbits underwent infra-renal aorta replacement with 4 mm diameter prostheses, twenty animals with PDMS and twenty animals with PTFE (control group). Retrograde aortic angiography was performed to assess patency. Histological graft samples were examined by electron microscopy to evaluate prostheses endothelialization. Results: patency rates were 100% for both grafts after 30 days; after 60 days, patency rate for PDMS was 92.3% (±7.4), and 73,8% (±13.1) at 90 days. PTFE grafts had patency rates of 87.5% (±11.7) at 60 and 90 days. No statistically significant difference was found in between groups for patency rates (p=0.62). Postoperative complications (death, paraplegia) rates (p=0.526) and aortic clamping times (p=0.299) were comparable in both groups. No statistically significant difference for stenosis was found on angiographical analysis between groups (p=0.650). Electron microscopy revealed limited anastomotic endothelial ingrowth in both prostheses. Conclusion: in this experimental model, PDMS and PTFE vascular prostheses had comparable outcomes and PDMS prosthesis could be used as a vascular graft.

RESUMO Objetivo: analisar novo tubo de silicone como material para prótese vascular e compará-lo a prótese de PTFE, em modelo experimental com coelhos. Métodos: quarenta coelhos foram submetidos a interposição, na aorta infrarrenal, de próteses de 4mm de diâmetro, sendo 20 animais com PDMS e 20 com PTFE (grupo controle). Foi realizada arteriografia retrógrada da aorta para avaliar a patência das próteses. Para avaliar a endotelização das próteses foi realizada microscopia eletrônica de maneira amostral pareada. Resultados: a patência em 30 dias foi de 100% para as duas próteses. Aos 60 dias, a taxa de patência do PDMS foi de 92,3% (± 7,4), e de 73,8% (±13,1) em 90 dias; as próteses de PTFE tiveram taxas de patência de 87,5% (± 11,7) aos 60 e 90 dias. Não foi observada diferença significativa entre as taxas de patência dos grupos (p=0,62). Não foi observada diferença entre os grupos quanto à evolução com complicações pós-operatórias (p=0,526) e quanto ao tempo de clampeamento da aorta (p=0,299). Não houve diferença significativa entre os grupos quanto ao grau de estenose das próteses (p=0,650) à avaliação angiográfica. A microscopia eletrônica mostrou crescimento endotelial limitado às regiões próximas às anastomoses nos dois tipos de próteses. Conclusões: o PDMS mostrou-se passível de utilização como prótese vascular, com resultados comparáveis aos do PTFE no modelo utilizado.

Histological evaluation of capsules formed by texturized silicone implants with and without polyester mesh coverage (Parietex®). A study on female rats

ABSTRACT Purpose To evaluate capsules formed by microtextured silicone implants with and without Parietex® mesh coverage histologically. Methods Sixty Wistar rats were divided in two groups (meshed and unmeshed). Each group was, then, divided into two subgroups for evaluation at 30 and 90 days. Capsules were analyzed based on hematoxylin and eosin (HE) and picrosirius staining. Results The number of fibroblasts, neutrophils and macrophages was similar among all subgroups. There was a higher lymphocyte reaction in the 30-day meshed group (p = 0.003). Giant cell reaction, granulation tissue and neoangiogenesis were similar among the subgroups. Synovial metaplasia was milder at 90-day in the unmeshed (p = 0.002) and meshed group (p < 0.001). Capsular thickness was significantly greater in the meshed samples (30-day p < 0.001 and 90-day p < 0.001). There was a similar amount of collagen types I and III in both groups. Conclusions The mesh-covered implants produced capsules similar to the microtextured ones when analyzing inflammatory variables. Synovial metaplasia was milder at 90 than at 30 days, and the capsular thickness was significantly greater in the meshed group. A similar amount of collagen types I and III was observed. Due to these characteristics, the mesh coverage did not seem to significantly affect the local inflammatory activity.

Thirty years of metabolic engineering for biosynthesis of polyhydroxyalkanoates / 生物工程学报

Polyhydroxyalkanoate (PHA) is a family of biodegradable polyesters synthesized by microorganisms. It has various monomer structures and physical properties with broad application prospects. However, its large-scale production is still hindered by the high cost. In the past 30 years, metabolic engineering approach has been used to tune the metabolic flux, engineer and introduce pathways. The efficiency of PHA synthesis by microorganisms has been significantly improved, and the diversity of PHA monomer, structure and substrate have also been enriched. Meanwhile, by changing cell morphology and PHA particle size, more efficient downstream production process has achieved and PHA production costs have been reduced. In recent years, "Next generation industrial biotechnology" (NGIB) based on extremophiles, especially halophilic Halomonas spp., has been rapidly developed. NGIB has achieved the opening and continuous production of PHA, which simplifies the production process and saves energy and fresh water. Combined with metabolic engineering, Halomonas spp. can be transformed into low-cost production platform of numerous PHA. It is expected to improve the market competitiveness and promote the commercialization of PHA.

Effect of short-chain thioesterase deficiency on P(3HB-co-LA) biosynthesis in Escherichia coli / 生物工程学报

Polyhydroxyalkanoates (PHAs) have obtained much attention in biomaterial fields due to their similar physicochemical properties to those of the petroleum-derived plastics. Poly(3-hydroxybutyrate-co-lactate) [P(3HB-co-LA)] is one member of the PHAs family, and has better toughness and transparency compared to existing polylactic acid (PLA) and poly[(R)-3-hydroxybutyrate] [P(3HB)]. First, we confirmed the one-step biosynthesis of P(LA-co-3HB) with the lactate fraction of 23.8 mol% by introducing P(3HB-co-LA) production module into Escherichia coli MG1655. Then, the lactate fraction was increased to 37.2 mol% in the dld deficient strain WXJ01-03. The genes encoding the thioesterases, ydiI and yciA, were further knocked out, and the lactate fraction in the P(3HB-co-LA) was improved to 42.3 mol% and 41.1 mol% respectively. Strain WXJ03-03 with dld, ydiI and yciA deficient was used for the production of the LA-enriched polymer, and the lactate fraction was improved to 46.1 mol%. Notably, the lactate fraction in P(3HB-co-LA) from xylose was remarkably higher than from glucose, indicating xylose as a potent carbon source for P(3HB-co-LA) production. Therefore, the deficiency of thioesterase may be considered as an effective strategy to improve the lactate fraction in P(3HB-co-LA) in xylose fermentation.

Three-dimensional printed poly (L-lactide) and hydroxyapatite composite for reconstruction of critical bone defect in rabbits

ABSTRACT Purpose To use a 3D printed poly (L-lactide) acid (PLLA) and hydroxyapatite (HA) composite as a bone substitute for reconstruction of a critical bone defect in the radius of rabbits. Methods A 1.5 cm ostectomy was performed in the radial diaphysis of 60 New Zealand white rabbits. The rabbits were divided into three groups according to surgical treatment of the bone defect (group I - control, group II - bone graft, group III - 3D PLLA). Each group was divided into four subgroups with different radiographic and histopathologic evaluation times (T1 - 15 days, T2 - 30 days, T3 - 60 days, T4 - 90 days). Results The implant group had greater clinically lameness (p = 0.02), edema (p = 0.007), pain (p = 0.04) and more complications at the surgical site (p = 0.03). Histologically, this group showed greater congestion (p = 0.04), hemorrhage (p = 0.04) and inflammation. Osteogenesis was microscopically similar between days (p = 0.54) and treatments (p = 0.17), even though radiographically, more effective bone healing occurred in the graft group (II), with more callus and bone bridge formation. Conclusions The customization of a 3D PLLA/HA scaffold was successful. However, in animals receiving the polymer-ceramic composite less bone callus and bone bridge was formed compared to the graft group.

Respiration in Azotobacter vinelandii and its relationship with the synthesis of biopolymers

Azotobacter vinelandii is a gram-negative soil bacterium that produces two biopolymers of biotechnological interest, alginate and poly(3-hydroxybutyrate), and it has been widely studied because of its capability to fix nitrogen even in the presence of oxygen. This bacterium is characterized by its high respiration rates, which are almost 10-fold higher than those of Escherichia coli and are a disadvantage for fermentation processes. On the other hand, several works have demonstrated that adequate control of the oxygen supply in A. vinelandii cultivations determines the yields and physicochemical characteristics of alginate and poly(3-hydroxybutyrate). Here, we summarize a review of the characteristics of A. vinelandii related to its respiration systems, as well as some of the most important findings on the oxygen consumption rates as a function of the cultivation parameters and biopolymer production.

Magnesium versus poly-L-lactic acid bioresorbable scaffolds: in vivo optical coherence tomography comparison of mechanical performance / Andamiajes bioresorbibles de magnesio versus poliméricos: comparación in vivo de su comportamiento mecánico por OCT

Abstract Background: Different mechanical properties have been suggested for metallic bioresorbable vascular scaffolds (BVS) in comparison to polymeric BVS. We aim to evaluate the acute mechanical performance of Magmaris® scaffold in comparison to Absorb®. Materials and Methods: Two groups of 10 coronary lesions treated with Magmaris® and Absorb® 1.1 (20584 vs. 21016 struts) were compared. In all cases, optical coherence tomographic (OCT) images were acquired after scaffold deployment. Baseline clinical, angiographic, and procedural characteristics were compared, including OCT evaluations. Results: No baseline clinical or angiographic significant differences were found between groups. The most common indication for revascularization was effort angina (60% vs. 70% p = 0.45) with no ST-elevation myocardial infarction (MI) cases. Main target artery was left anterior descending, with a mean vessel diameter of 3.46 ± 0.23 in Absorb® and 3.52 ± 0.19mm in Magmaris® groups (p = 0.56). All cases underwent pre- and post-dilatation with a procedural success rate of 100%. OCT analyses showed larger scaffold and vessel diameters in Magmaris® group: 3.11 ± 0.38 mm versus 3.07 ± 0.36 mm, p = 0.03 and 4.12 ± 0.51 mm versus 4.04 ± 0.46 mm, p = 0.04. Despite the application of slightly higher postdilatation pressures to Magmaris® devices (18.01 ± 2.15 vs. 17.20 ± 3.80 atm, p = 0.05), significantly lower percentages of disrupted and malapposed struts were identified within Magmaris® scaffolds (0.15% vs. 0.27%, p = 0.03 and 1.06% vs. 1.46% p = 0.01). No cardiac death, target vessel-related MI, or clinically driven target lesion revascularization was reported in a 30-day follow-up. Conclusion: Mechanical properties of Magmaris® scaffold allow achieving larger vessel and scaffold diameters in a safe manner, with lower rates of malapposition and scaffold disruption.

Resumen Introducción: Se ha sugerido la presencia de un distinto comportamiento mecánico entre los dos grupos principales de dispositivos bioresorbibles: metálicos y poliméricos. En este estudio evaluamos el comportamiento mecánico agudo del andamiaje bioresorbible metálico Magmaris® frente al del polimérico Absorb®. Métodos: Se compararon dos grupos de 10 lesiones coronarias tratadas con Magmaris® y Absorb® 1.1 (20584 vs. 21016 struts). En todos los casos se realizó estudio postimplante del dispositivo mediante tomografia de coherencia óptica (OCT). Se compararon las características basales clínicas y angiográficas, así como aspectos del procedimiento (incluídos los estudios de OCT) entre ambos grupos. Resultados: No se encontraron diferencias clínicas o angiográficas estadísticamente significativas entre ambos grupos. La indicación más frecuente de revascularización coronaria fué la presencia de angina de esfuerzo (60% vs. 70% p = 0.45), sin incluirse casos de IAMCEST. La arteria descendente anterior fué el principal vaso diana, con un diámetro medio de 3.46 ± 0.23 mm en el grupo de Absorb® y de 3.52 ± 0.19mm en el grupo de Magmaris® (p = 0.56). En todos los casos se realizó pre y postdilatación, con una tasa de éxito del procedimiento del 100%. Los estudios mediante OCT demostraron un mayor diámetro de stent y del vaso en el grupo de Magmaris®: 3.11 ± 0.38mm versus 3.07 ± 0.36 mm, p = 0.03 y 4.12 ± 0.51mm versus 4.04 ± 0.46mm, p = 0.04. A pesar de someter a los dispositivos Magmaris® a presiones de postdilatación ligeramente superiores (18.01 ± 2.15 vs. 17.20 ± 3.80 atm, p = 0.05), se identificó un menor porcentaje estadísticamente significativo de struts rotos o malapuestos en dicho grupo (0.15% vs. 0.27 %, p = 0.03 y 1.06 % vs. 1.46% p = 0.01). En un seguimiento a 30 días no se registraron eventos mayores: muerte cardíaca, IM relacionado con vaso diana o TLR. Conclusión: Las propiedades mecánicas del scaffold metálico bioresorbible Magmaris® permiten alcanzar mayores diámetros de stent y vaso de forma segura tras su implante, con una baja tasa de malaposición y disrupción.

Statistical analysis of shrinkage levels of human brain slices preserved by sheet plastination technique with polyester resin / Análisis estadístico de los niveles de retracción de cortes de cerebro humano conservados mediante la técnica de plastinación de cortes con resina poliéster

Plastination is currently the most important anatomical preservation technique due to the possibility of preserving bodies and organs for an indefinite period, in a dry and biosecure form, while preserving the morphological characteristics of the tissues. However, the shrinkage of the samples is also part of the plastination, perhaps becoming one of its few disadvantages. This paper presents the shrinkage caused by the classic technique of sheet plastination with polyester resin (Biodur® P40) in human brain slices, with the aim of statistically establishing the percentages of tissue shrinkage caused by this plastination protocol.

La plastinación es actualmente la técnica de preservación anatómica más importante debido a la posibilidad de preservar los cuerpos y órganos por un período indefinido, en forma seca y biosegura, al tiempo que preserva las características morfológicas de los tejidos. Sin embargo, la retracción de las muestras también es parte de la plastinación, quizás convirtiéndose en una de sus pocas desventajas. Este artículo presenta la retracción causada por la técnica clásica de plastinación de cortes con resina poliéster (Biodur® P40) en cortes de cerebro humano, con el objetivo de establecer estadísticamente los porcentajes de retracción de tejidos causados por este protocolo de plastinación.

Jalecos em têxteis de poliéster agem como barreira contra fluidos e bactérias? / ¿Batas médicas de telas de poliéster actúan como barrera contra fluidos y bacterias? / Do white coats on polyester fabrics act as a barrier against fluids and bacteria?

Resumo Objetivo Avaliar tecidos de poliéster quanto à função de barreira física contra fluidos e bactérias. Métodos Trata-se de uma pesquisa experimental laboratorial in vitro realizada em três etapas: avaliação do tempo de passagem de fluido através dos tecidos, cronometrado desde o início do escoamento do fluido até a formação e queda da última gota; determinação microbiológica da carga bacteriana presente no fluido, após a sua passagem através dos tecidos; e análise das características estruturais dos tecidos por meio de microscopia eletrônica de varredura. Os dados foram submetidos aos testes de normalidade e ao teste de U de Mann-Whitney, com nível de significância de a=5%. Resultados as comparações dos tempos obtidos na primeira etapa entre os dois tipos de tecidos utilizados demonstraram diferença estatística ( p <0,001). Com relação à avaliação microbiológica, não foi observada diferença entre as cargas bacterianas após a passagem do fluido através dos tecidos, tanto para Staphylococcus aureus ( p =0,056) quanto para Pseudomonas aeruginosa ( p= 0,320). A análise por microscopia eletrônica de varredura evidenciou diferenças estruturais entre os tecidos, no entanto não foi constatada a presença bacteriana na superfície dos tecidos. Conclusão Ambos os tecidos de poliéster empregados para confecção de jalecos não apresentaram função de barreira física contra fluidos e bactérias. Assim, os resultados nos permitem especular que o jaleco de poliéster ao entrar em contato com fluidos corporais pode possibilitar a contaminação do profissional.

Resumen Objetivo Evaluar telas de poliéster con relación a la función de barrera física contra fluidos y bacterias. Métodos Se trata de un estudio experimental de laboratorio in vitro realizado en tres etapas: evaluación del tiempo de pasaje del fluido a través de las telas, cronometrado desde el inicio del derrame del fluido hasta la formación y caída de la última gota; determinación microbiológica de la carga bacteriana presente en el fluido después del pasaje a través de las telas; y análisis de las características estructurales de las telas mediante microscopio electrónico de barrido. Con los datos obtenidos se realizaron las pruebas de normalidad y la prueba U de Mann-Whitney, con nivel de significación de a=5%. Resultados La comparación de los tiempos obtenidos en la primera etapa entre los dos tipos de telas utilizados demostró diferencia estadística ( p <0,001). Respecto a la evaluación microbiológica, no se observó diferencia entre las cargas bacterianas después del pasaje del fluido a través de las telas, tanto de Staphylococcus aureus ( p =0,056) como de Pseudomonas aeruginosa ( p= 0,320). El análisis mediante microscopio electrónico de barrido constató diferencias estructurales entre las telas; sin embargo, no se observó la presencia bacteriana en la superficie de las telas. Conclusión Las dos telas de poliéster empleadas para la confección de batas médicas no presentan función de barrera física contra fluidos y bacterias. De esta forma, los resultados nos permiten suponer que la bata médica de poliéster, al entrar en contacto con fluidos corporales, puede posibilitar la contaminación del profesional.

Abstract Objective To evaluate polyester fabrics as physical barrier function against fluids and bacteria. Methods This is an in vitro experimental laboratory research carried out in three stages: evaluation of the length of time for the fluid to pass through the fabrics, timed from the beginning of the fluid flow until the formation and fall of the last drop; microbiological determination of the bacterial load in the fluid, after its passage through the fabrics; and analysis of the structural characteristics of the fabrics by scanning in electron microscopy. The data were submitted to normality tests and the Mann-Whitney U test, with a significance level of a=5%. Results Comparisons of length of time in the first stage between the two types of fabrics used showed a statistical difference ( p <0.001). Regarding the microbiological evaluation, there was no difference among bacterial loads after the fluid passed through the fabrics, both for Staphylococcus aureus ( p =0.056) and Pseudomonas aeruginosa ( p =0.320). The analysis by scanning electron microscopy showed structural differences between the fabrics, however, there were no bacteria on the fabric surface. Conclusion Both polyester fabrics used to make white coats did not work as a physical barrier against fluids and bacteria. Thus, the results allowed us to speculate that the polyester coat when in contact with body fluids may allow contamination of the professional.

Characterization and assessment of the genotoxicity and biocompatibility of poly (hydroxybutyrate) and norbixin membranes

Abstract Purpose To synthesize and characterize poly(hydroxybutyrate) (PHB) and norbixin membranes to evaluate them for genotoxicity in rats and wound healing in mice by histological staining. Methods For the evaluation of genotoxicity, male rats ( Rattus novegicus ) were divided into three groups (n= 5): 5% PHB/Norbixin membrane introduced into the peritoneum by laparotomy; B - negative control; C - positive control (intraperitoneal dose of cyclophosphamide 50 mg/kg). For the evaluation of biocompatibilty, a cutaneous wound was induced on the back of males mice ( Mus musculus ) divided into two experimental treatment groups: control and membrane that underwent euthanasia after 7 and 14 days treatment. Statistical analysis ware made by One Way Anova post hoc Tukey Test (p<0.05). Results Regarding the incidence of polychromatic erythrocytes, there was no difference between negative control and 5% PHB/Norbixin membrane; however, when compared to the positive control represented by cyclophosphamide, there was a significant difference (p <0.001). As for DNA damage, the changes induced in the first 4h were repaired in 24h. In addition, the membrane was effective in abbreviating the inflammatory process and served as a scaffold due to the stimulus to reepithelialization mainly on the 7 days of treatment. Conclusion The non-genotoxic PHB/Norbixin 5% membrane presented promising results that suggest its effectiveness as a guide for tissue regeneration given its biocompatibility.

Poly (Lactic Acid) membrane and Sedum dendroideum extract favors the repair of burns in rats

Abstract Purpose: To evaluate the healing potential of the electrospinning membranes of Poly (Lactic Acid) (PLA) associated with Sedum dendroideum extract in burn injuries in rats. Methods: Seventy-five rats were submitted to burn injury on their back skin: (C) untreated; (F) with daily topical application of S. dendroideum extract; (M) with electrospinning membranes of PLA; (MF10) with electrospinning membranes of PLA with 10% S. dendroideum extract; (MF25) with electrospinning membranes of PLA with 25% S. dendroideum extract. Tissue samples were taken after 2, 6 and 14 days of the burn injury and were subjected to histomorfometric analysis of quantification of fibroblasts, collagen fibers, blood vessels, and inflammatory infiltrate Results: The histomorphometric analysis showed an increase in the number of fibroblasts, collagen fibers and blood vessels in the burns treated with membranes of PLA, associated or not with the 10% and 25% extract. The extract of S. dendroideum promoted the increase of collagen fibers. Conclusion: The electrospinning PLA membrane, isolated or associated with the S. dendrodeum extract, favored the healing of burn injuries in this experimental model, with an increase of fibroblasts, collagen fibers, and blood vessels. S. dendroideum isolated only stimulated the collagenesis.

Desarrollo de un protocolo de plastinación de cortes con resina poliéster aplicado a secciones de cerebro humano / Development of a sheet plastination protocol with polyester resin applied to human brain slices

La plastinación es una técnica anatómica de conservación cadavérica creada en 1977 por Gunther von Hagens, en Heidelberg, Alemania, y que sustituye los líquidos biológicos y/o de fijación por acetona, para luego impregar las muestras con distintas resinas, dependiendo de la técnica de plastinación desarrollada, para finalmente llevar a cabo la polimerización de los componentes incorporados a las muestras, para obtener muestras biológicas secas y totalmente duraderas. El objetivo de este trabajo consistió en desarrollar un protocolo de plastinación de cortes con resina poliéster (Biodur® P40) en secciones de 3 mm de espesor de cerebro humano. La muestras fueron fijadas y conservadas con formalina al 10 %. Los cerebros luego fueron seccionados con una maquina cortadora de tejidos, obteniéndose láminas delgadas de 3 mm de espesor. Inmediatamente los cortes de cerebro fueron colocados en deshidratación en acetona al 100 %, a -25 ºC, durante 7 días el primer baño de acetona, y durante otros 3 días más, para el segundo baño de acetona. Una vez deshidratados los cortes, estos fueron colocados en resina poliéster Biodur® P40 y se llevó a cabo la impregnación forzada de los cortes, en cámara de vacío a temperatura ambiente (20 ºC). Una vez finalizada la impregnación forzada, se procedió a la etapa de curado, la cual en primer lugar consiste en el armado de las cámaras de curado dentro de las cuales se colocaran los cortes con resina poliéster. Las cámaras de curado fueron colocadas bajo luz UV para acelerar la polimerización del poliéster y finalizar el proceso de plastinación. Se logró desarrollar satisfactoriamente en el Laboratorio de Plastinación y Técnicas Anatómicas de la Universidad de La Frontera un protocolo de plastinación de cortes con resina poliéster, obteniendo una excelente conservación de cortes de cerebro, con diferenciación de sustancias gris y blanca, y conservación de todas las características morfológicas.

Plastination is an anatomical technique of cadaveric conservation created in 1977 by Gunther von Hagens, in Heidelberg, Germany, and that substitutes biological and / or fixation fluids with acetone, to then impregnate the samples with different resins, depending on the developed plastination technique, to finally carry out the polymerization of the components incorporated into the samples, to obtain dry and totally durable biological samples. The aim of this work was to develop a sheet plastination protocol with polyester resin (Biodur® P40) in 3 mm thick slices of human brain. The samples were fixed and preserved with 10 % formalin. The brains were sectioned with a slice cut machine, obtaining thin sheets of 3 mm thick. Immediately the slices of brain were placed in dehydration in 100 % acetone, at -25 °C, for 7 days the first acetone bath, and for another 3 more days, for the second acetone bath. Once the cuts were dehydrated, they were placed in Biodur® P40 polyester resin and the forced impregnation was carried out in a vacuum chamber at room temperature (20 °C). Once the forced impregnation was finished, the curing stage was carried out, which first consists in the assembly of the curing chambers within which the slices with polyester resin were placed. The curing chambers were placed under UV light to accelerate the polymerization of the polyester and finished the plastination process. A sheet plastination protocol with polyester resin was successfully developed in the Laboratory of Plastination and Anatomical Techniques of Universidad de La Frontera, obtaining excellent conservation of brain slices, with differentiation of gray and white substances, and conservation of all morphological characteristics.

Effect of norbixin-based poly(hydroxybutyrate) membranes on the tendon repair process after tenotomy in rats

Abstract Purpose: To determine the efficacy of norbixin-based poly(hydroxybutyrate) (PHB) membranes for Achilles tendon repair. Methods: Thirty rats were submitted to total tenotomy surgery of the right Achilles tendon and divided into two groups (control and membrane; n = 15 each), which were further subdivided into three subgroups (days 7, 14, and 21; n = 5 each). Samples were analyzed histologically. Results: Histological analysis showed a significant reduction in inflammatory infiltrates on days 7, 14 (p < 0.0001 for both), and 21 (p = 0.0004) in the membrane group compared to that in the control group. There was also a significant decrease in the number of fibroblasts in the control group on days 7, 14 (p < 0.0001), and 21 (p = 0.0032). Further, an increase in type I collagen deposition was observed in the membrane group compared to that in the control group on days 7 (p = 0.0133) and 14 (p = 0.0107). Conclusion: Treatment with norbixin-based PHB membranes reduces the inflammatory response, increases fibroblast proliferation, and improves collagen production in the tendon repair region, especially between days 7 and 14.

Proliferation and differentiation of mouse spermatogonial stem cells on a three-dimensional surface composed of PCL/gel nanofibers / Proliferación y diferenciación de células madre espermatogónicas de ratón en una superficie tridimensional compuesta de nanofibras PCL / gel

Spermatogonial stem cells (SSCs) have self-renewal and differentiation capacity essential for sperm production throughout the male reproductive life. The electrospun polycaprolactone/gelatin (PCL/Gel) nanofibrous scaffold mimics important features of the extracellular matrix (ECM), which can provide a promising technique for the proliferation and differentiation of SSCs in vitro. The goal of the present study was to investigate the effects of PCL/Gel nanofibrous scaffold on the propagation and differentiation of neonate mouse SSCs (mSSCs). mSSCs were enzymatically isolated, and the cells were purified by differential plating method and seeded on scaffold. After 2 weeks, viability, colony number and diameter, and expression of specific spermatogonial cell genes were investigated. After mSSCs propagation, the cells were cultivated in a differentiation medium on the scaffold for another 2 weeks, and differentiating cells were analyzed by real-time PCR. The number of mSSC colony (P<0.01) and expression levels of specific spermatogonial genes Plzf and Inga6 (P<0.01) and also differentiation genes c-Kit, Tp1 and Ptm1 (P<0.05) were higher in scaffold group compared with control during the culture period. We conclude that mSSCs can be expanded and can differentiate toward spermatid cells on PCL/Gel nanofibrous scaffold with improved developmental parameters.

Las células madre espermatogónicas (CME) tienen capacidad de auto renovación y diferenciación esenciales para la producción de esperma a lo largo de la vida reproductiva masculina. El «scaffold¼ nanofibroso de policaprolactona / gelatina (PCL / Gel) electrohilado imita características importantes de la matriz extracelular (MEC), que puede proporcionar una técnica prometedora para la proliferación y diferenciación de CME in vitro. El objetivo del presente estudio fue investigar los efectos del «scaffold¼ nanofibroso PCL / Gel en la propagación y diferenciación de CME de ratones neonatos (mSSC). Los mSSC se aislaron enzimáticamente y las células se purificaron mediante un método de siembra diferencial y se sembraron en un «scaffold¼. Después de 2 semanas, se investigaron la viabilidad, el número y el diámetro de las colonias y la expresión de genes específicos de células espermatogónicas. Después de la propagación de mSSC, las células se cultivaron en un medio de diferenciación en el «scaffold¼ durante otras 2 semanas, y las células se analizaron mediante PCR en tiempo real. El número de colonias mSSC (P <0,01) y los niveles de expresión de los genes espermatogónicos específicos Plzf e Inga6 (P <0,01) y también los genes de diferenciación c-Kit, Tp1 y Ptm1 (P <0,05) fueron mayores en el grupo de «scaffold¼ en comparación con el control durante el período de cultivo. Concluimos que los mSSC pueden expandirse y diferenciarse en células espermátidas en un «scaffold¼ de nanofibras PCL / Gel con parámetros de desarrollo mejorados.

Development of biodegradation process for Poly(DL-lactic acid) degradation by crude enzyme produced by Actinomadura keratinilytica strain T16-1

Background: Plastic waste is a serious problem because it is difficult to degrade, thereby leading to global environment problems. Poly(lactic acid) (PLA) is a biodegradable aliphatic polyester derived from renewable resources, and it can be degraded by various enzymes produced by microorganisms. This study focused on the scale-up and evaluated the bioprocess of PLA degradation by a crude microbial enzyme produced by Actinomadura keratinilytica strain T16-1 in a 5 L stirred tank bioreactor. Results: PLA degradation after 72 h in a 5 L bioreactor by using the enzyme of the strain T16-1 under controlled pH conditions resulted in lactic acid titers (mg/L) of 16,651 mg/L and a conversion efficiency of 89% at a controlled pH of 8.0. However, the PLA degradation process inadvertently produced lactic acid as a potential inhibitor, as shown in our experiments at various concentrations of lactic acid. Therefore, the dialysis method was performed to reduce the concentration of lactic acid. The experiment with a dialysis bag achieved PLA degradation by weight loss of 99.93%, whereas the one without dialysis achieved a degradation of less than approximately 14.75%. Therefore, the dialysis method was applied to degrade a commercial PLA material (tray) with a conversion efficiency of 32%, which was 6-fold more than that without dialysis. Conclusions: This is the first report demonstrating the scale-up of PLA degradation in a 5 L bioreactor and evaluating a potential method for enhancing PLA degradation efficiency.

Poly(trimethylene carbonate-co-L-lactide) electrospun scaffolds for use as vascular grafts

Currently, there is great clinical need for suitable synthetic grafts that can be used in vascular diseases. Synthetic grafts have been successfully used in medium and large arteries, however, their use in small diameter vessels is limited and presents a high failure rate. In this context, the aim of this study was to develop tissue engineering scaffolds, using poly(trimethylene carbonate-co-L-lactide) (PTMCLLA), for application as small diameter vascular grafts. For this, copolymers with varying trimethylene carbonate/lactide ratios - 20/80, 30/70, and 40/60 - were submitted to electrospinning and the resulting scaffolds were evaluated in terms of their physicochemical and biological properties. The scaffolds produced with PTMCLLA 20/80, 30/70, and 40/60 showed smooth fibers with an average diameter of 771±273, 606±242, and 697±232 nm, respectively. When the degradation ratio was evaluated, the three scaffold groups had a similar molecular weight (Mw) on the final day of analysis. PTMCLLA 30/70 and 40/60 scaffolds exhibited greater flexibility than the PTMCLLA 20/80. However, the PTMCLLA 40/60 scaffolds showed a large wrinkling and their biological properties were not evaluated. The PTMCLLA 30/70 scaffolds supported the adhesion and growth of mesenchymal stem cells (MSCs), endothelial progenitor cells, and smooth muscle cells (SMCs). In addition, they provided a spreading of MSCs and SMCs. Given the results, the electrospun scaffolds produced with PTMCLLA 30/70 copolymer can be considered promising candidates for future applications in vascular tissue engineering.

Comparative study of peritoneal adhesions after intraperitoneal implantation in rats of meshes of polypropylene versus polypropylene/polyglecaprone versus polyester/porcine collagen

Abstract Purpose To Compare the extent and intensity of adhesions formed between the intra-abdominal organs and the intraperitoneal implants of polypropylene mesh versus polypropylene/polyglecaprone versus polyester/porcine collagen used for correction of abdominal wall defect in rats. Methods After the defect in the abdominal wall, thirty Wistar rats were placed in three groups (ten animals each) for intraperitoneal mesh implant: polypropylene group, polypropylene/polyglecaprone group, and polyester/porcine collagen group. The macroscopic evaluation of the extent and intensity of adhesions was performed 21 days after the implant. Results The polypropylene group had a higher statistically significant impairment due to visceral adhesions (p value = 0.002) and a higher degree of intense adherence in relation to polypropylene/polyglecaprone and polyester/porcine collagen groups (p value<0.001). The polyester/porcine collagen group showed more intense adhesions than the polypropylene/polyglecaprone group (p value=0.035). Conclusions The intraperitoneal implantation of polypropylene meshes to correct defects of the abdominal wall caused the appearance of extensive and firm adhesions to intra-abdominal structures. The use of polypropylene/polyglecaprone or polyester/porcine collagen tissue-separating meshes reduces the number and degree of adhesions formed.

In vitro evaluation, cellular uptake and anti-acute myocardial ischemia effect of puerarin PEG-PLGA micelles / 中国中药杂志

PUE@PEG-PLGA micelles has excellent characteristics such as small particle size, high drug loading and slow drug release. The results of TEM electron microscopy showed that PUE@PEG-PLGA micelles had obvious core-shell structure. The critical micelle concentration(CMC) of PEG-PLGA micelles determined by pyrene assay was about 4.8 mg·L~(-1). Laser confocal experiments showed that PEG-PLGA micelles can enhance the cellular uptake of coumarin-6 and aggregate around the mitochondria; quantitative results of extracellular drug residues also indirectly confirmed that PEG-PLGA micelles can promote cellular uptake of the drug. Acute ischemic myocardial model rats were prepared by coronary artery ligation, and then the model rats were randomly divided into six groups: Sham operation group, model group, puerarin(PUE) group, as well as low-, mid-, and high-dose PUE@PEG-PLGA micelles groups. Drugs were given by iv administration 5 min after the ligation. The ST segment changes in the electrocardiogram were monitored; serum creatine kinase(CK), lactate dehydrogenase(LDH), aspartate aminotransferase(AST), and malondialdehyde(MDA) levels were detected and myocardial infarct size was also measured. Both PUE and PUE@PEG-PLGA micelles can reduce the elevated ST segment, reduce serum CK, LDH, AST and MDA levels, and reduce myocardial infarct size. The efficacy of PUE@PEG-PLGA medium and high dose groups was significantly better than that in the PUE group, and the efficacy in PUE@PEG-PLGA low dose group was basically equivalent to that in the PUE group. PUE@PEG-PLGA micelles can greatly improve the cardiomyocytes uptake of PUE, enhance the anti-acute myocardial ischemia effect of drugs, and reduce its dosage.