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    Celulose bacteriana como penso curativo

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    Tese de doutoramento em Biomedical EngineeringWounds, in particular traumatic (e.g. burns) and chronic ones, are a major cause of morbidity, impaired life quality and high health care costs. They often result in long hospitalization stays, taking up substantial health resources in developed countries. Conventional treatments are often painful, expensive and may increase the infection risk, compromising the treatments‚Äô time and success. In recent years, there have been efforts to develop new advanced methodologies to heal chronic wounds, including the topic use of growth factors or cell-based therapies. However, in many cases, the therapeutic efficacy is low, the therapies are expensive and require application in a clinical facility. Therefore, development of new therapeutics is absolutely necessary and important to satisfy these unmet clinical needs. So, this work comprised the development of a safe, easy-to-use and non expensive novel dressing, aimed at efficiently addressing these issues, by attaining faster and proper wound healing. The use of bacterial nanocellulose (BNC) has already demonstrated positive results in the treatment of different kinds of wounds. Additionally, BNC is considered a promising drug delivery system. In this work, BNC was conceived as a protective barrier against exogenous agents (particles, microorganisms) that can impair wound healing, and as a drug carrier for the controlled release of hydrophobic drugs, namely of vitamin D3 (Vit D3 ), an inducer of the endogenous expression of antimicrobial peptide (AMP) LL37, known for accelerating the wound healing process. In a first part of this project, the optimization of the static BNC production was performed, aiming at making it viable and economic at large scale. First, an experimental design, based on response surface methodology (RSM) - central composite design (CCD) - was used to optimize the culture medium for BNC production by Komagataeibacter xylinus BPR 2001, using a simple culture medium composition based on byproducts from the food industry. The optimal conditions for BNC production were (% (m/v)): molasses 5.38; CSL 1.91; ammonium sulphate 0.63; disodium phosphate 0.270; citric acid 0.115 and ethanol 1.38 % (v/v). The experimental and predicted maximum BNC production yields were 7.5 ¬Ī0.54 g/L and 6.64 ¬Ī0.079 g/L, respectively, after 9 days at 30 ¬ļC. Furthermore, the effect of the surface area and culture medium depth on the BNC production yield and productivity were evaluated. BNC dry mass production increased with the surface area and with the medium volume (depth) and fermentation time. Also, as long as nutrients were still available in the culture media, the BNC mass productivity was maintained overtime. The pre-inoculum preparation (PIP) step was also optimized with regards to the (a) identification of an inexpensive culture medium for pre-inoculum leading to a high cell density; (b) analysis of the effect of the initial cellular concentration on the static production of BNC and (c) kinetics of cell growth throughout the different steps of pre-inoculum preparation, including static and stirred - laboratorial and pilot-scale ‚Äď fermentations. The best composition for PIP medium was (% (m/v)): Glucose and Fructose syrup 1.5- 2.0; Corn Step Liquor (protein basis) 0.7; citric acid 0.115; Na2HPO4 0.27. The analysis of the cell growth kinetics in the different steps of PIP showed that a careful control on the culture time in each stage is advisable. The time required to reach the exponential phase was very different in each stage of PIP, reducing significantly from the static culture to the stirred culture and for large scale stirred culture, in a 75 L Bioreactor. In a second part of this work, the use of BNC as a drug carrier was addressed. Since Vit D3 is poorly water soluble, and thus not easily incorporated in the highly hydrophilic environment of the BNC membrane, Vit D3 was encapsulated in a self-assembled hyaluronic acid (HA)-based amphiphilic nanogel and then incorporated in the BNC membrane. The carrier was obtained by grafting hexadecylamine (Hexa) into the HA backbone (HA-Hexa). Vit D3 was successfully loaded into the nanogel (HA-Vit D3 ) with an encapsulation efficiency between 60-91 %. The loaded system- HA-Vit D3 - was embedded into BNC, conceived as a transdermal delivery system. The release of Vit D3 was monitored over time using a Franz cell device. Around 70 % of the initial Vit D3 available was released from BNC membranes in the first 48 h. Most importantly, we observed that the released Vit D3 still remained within the HA-Hexa nanogel carrier. Vit D3 is known to stimulate the endogenous production of human cathelicidin (LL37), which is known to accelerate wound healing. Thus, formulations of HA-Vit D3 and HA-LLKKK18 (an analogue of LL37) were tested in vivo, using excision and chronic wound in dexamethasone treated C57BL/6 and db+/db+ mice models, as to evaluate and compare their efficiency in wound repairing. However, the results did not confirm any wound healing improvement.As feridas cr√≥nicas e traum√°ticas (e.g. queimaduras) apresentam uma elevada morbilidade, afetando severamente a qualidade de vida dos pacientes. Os tratamentos convencionais implicam longos per√≠odos de interna√ß√£o hospitalar, com significativo consumo de recursos dos sistemas de sa√ļde nos pa√≠ses desenvolvidos. Al√©m disso, s√£o dolorosos, caros e podem aumentar o risco de infe√ß√£o, comprometendo a dura√ß√£o e o sucesso dos tratamentos. Recentemente, t√™m sido desenvolvidos esfor√ßos para o desenvolvimento de novas metodologias avan√ßadas para o tratamento de feridas cr√≥nicas, incluindo a aplica√ß√£o t√≥pica de fatores de crescimento ou terapias baseadas em c√©lulas. Em muitos casos, estas novas abordagens s√£o caras, devendo ser realizadas numa unidade hospitalar, e a sua efic√°cia terap√™utica √© baixa. Assim, o desenvolvimento de novas solu√ß√Ķes para satisfazer esta necessidade cl√≠nica ainda n√£o satisfeita √© absolutamente necess√°rio. Com este trabalho pretende se desenvolver um penso curativo eficiente, inovador, f√°cil de usar e n√£o dispendioso, atrav√©s de uma abordagem segura, visando uma cicatriza√ß√£o mais r√°pida e adequada da ferida. A nanocelulose bacteriana (BNC) demonstrou j√° resultados positivos no tratamento de diferentes tipos de feridas, assim como foi j√° demonstrado tamb√©m o seu potencial como sistema de entrega de f√°rmacos. Neste trabalho, a BNC foi utilizada como ve√≠culo para a liberta√ß√£o controlada de mol√©culas hidrof√≥bicas, nomeadamente a vitamina D3 (Vit D3 ), que √© um indutor da express√£o end√≥gena do pept√≠do antimicrobiano LL37, conhecido por acelerar o processo de cicatriza√ß√£o de feridas. Al√©m disso, a BNC funciona como uma barreira protetora contra agentes ex√≥genos (poeiras, microorganismos) que podem prejudicar a cicatriza√ß√£o de feridas. Numa primeira parte, foram desenvolvidos trabalhos visando tornar a produ√ß√£o em grande escala de BNC em cultura est√°tica econ√≥mica e vi√°vel. Nesse sentido, foi usado um desenho experimental, baseado na metodologia de superf√≠cie de resposta (RSM) - planeamento composto central (CCD) - para otimizar o meio de cultura, usando subprodutos da ind√ļstria alimentar. Foi utilizada a estirpe Komagataeibacter xylinus BPR 2001, a 30 ¬ļC. Foram identificadas as seguintes condi√ß√Ķes √≥timas para a produ√ß√£o de BNC (% (m/v)): mela√ßo 5,38, xarope de milho (CSL) 1,91; sulfato de am√≥nio 0,63; fosfato diss√≥dico 0,270; √°cido c√≠trico 0,115 e etanol 1,38 % (v/v). Os rendimentos m√°ximos experimentais e previstos de produ√ß√£o de BNC foram 7,5 ¬Ī0,54 g/L e 6,64 ¬Ī0,079 g/L, respetivamente, ap√≥s 9 dias. Adicionalmente, foram avaliados o efeito da √°rea superficial e da profundidade/altura do meio de cultura no rendimento e produtividade em BNC. Verificou-se que a produ√ß√£o de BNC aumenta com a √°rea superficial, com o volume de meio de cultura (profundidade) e com o tempo de fermenta√ß√£o. Al√©m disso, observou-se que a produtividade de BNC se mant√©m constante at√© se esgotarem os nutrientes no meio de cultura. Para a etapa de prepara√ß√£o pr√©-in√≥culo (PIP), a otimiza√ß√£o consistiu em diferentes estudos, especificamente: (a) otimiza√ß√£o dum meio de cultura de custos reduzidos, que permita a obten√ß√£o de uma elevada densidade celular; (b) avalia√ß√£o do efeito da concentra√ß√£o celular inicial na produ√ß√£o est√°tica de BNC e (c) estudo da cin√©tica de crescimento celular ao longo das diferentes etapas de PIP. A melhor composi√ß√£o para o PIP foi (% (m/v)): xarope de glucose e frutose 1,5- 2,0; CSL 0,7; √°cido c√≠trico 0,115 e Na2HPO4 0,27. Os estudos de cin√©tica de crescimento celular para as diferentes etapas do PIP evidenciam a necessidade dum controle cuidadoso do tempo de cultura em cada etapa do PIP. O tempo necess√°rio para atingir a fase exponencial foi muito diferente em cada fase do PIP, reduzindo significativamente da cultura est√°tica, para a cultura agitada, e para cultura agitada em larga escala num bioreator de 75 L. A segunda parte do trabalho relaciona-se com o desenvolvimento da BNC como sistema de entrega de f√°rmacos. A Vit D3 √© pouco sol√ļvel em √°gua e, portanto, n√£o √© facilmente incorporada no ambiente altamente hidrof√≠lico como o da membrana de BNC. Para esse efeito foi usado um nanogel anfif√≠lico auto-organizado obtido pela liga√ß√£o de hexadecilamina (Hexa) na cadeia do √°cido hialur√≥nico (HA). A Vit D3 foi ent√£o encapsulada no nanogel de (HA-Hexa) e em seguida impregnada na membrana de BNC, com uma efici√™ncia de encapsula√ß√£o entre 60-91 %. A liberta√ß√£o da Vit D3 foi monitorizada ao longo do tempo, usando uma c√©lula de Franz e realizando estudos de permea√ß√£o. Observou-se a liberta√ß√£o de cerca de 70 % da Vit D3 , ainda dentro do nanogel de HA-Hexa, das membranas de BNC em 48h. Finalmente, foi testada a utiliza√ß√£o de HA-Vit D3 e de HA-LLKKK18 (um p√©ptido an√°logo √† LL37) em modelos de feridas de excis√£o e cr√≥nicas em ratinhos tratados com dexametasona e diab√©ticos tipo II (db + / db +) C57BL/6. No entanto, os resultados n√£o revelaram uma maior efici√™ncia na cicatriza√ß√£o de feridas na presen√ßa das referidas formula√ß√Ķes.Ao Projeto BioTecNorte (NORTE-01-0145-FEDER-000004), n¬ļ 003435: ‚ÄúBUILD ‚Äď Bacterial cellulose Leather‚ÄĚ, financiado pelo Fundo Europeu de Desenvolvimento Regional (FEDER) atrav√©s do Programa Operacional do Regional do Norte (NORTE 2020) e ao projeto SkinChip: Disruptive cellulose-based microfluidic device for 3D skin modelling, PTDC/BBB-BIO/1889/2014 e ainda √† Funda√ß√£o para a Ci√™ncia e Tecnologia no √Ęmbito do financiamento estrat√©gico da unidade UID / BIO / 04469/2019 e pela atribui√ß√£o da bolsa de doutoramento SFRH/BD/89547/2012

    Clinical evaluation of antimicrobial hidrophobic dressing for managing infection of chronic wounds

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    RESUMEN: Introducci√≥n: Las √ļlceras en extremidades inferiores de etiolog√≠a venosa son muy prevalentes en la actualidad, generando un problema tanto a nivel individual como para la Sociedad y para el Sistema Sanitario. La cicatrizaci√≥n es un proceso muy complejo en el cual hay que tener en cuenta muchos aspectos, como la infecci√≥n de la herida, entre otros. Existen muchas alternativas para el tratamiento de la infecci√≥n de las heridas cr√≥nicas y se trata de antibi√≥ticos y antis√©pticos, cuya desventaja principal es que pueden llegar a crear resistencias y/o reacciones al√©rgicas. La alternativa, a mencionados anteriormente, son los ap√≥sitos con propiedades hidr√≥fobas, cuyo objetivo es atraer y atrapar los microorganismos presentes en el lecho de las heridas en sus fibras, mediante la interacci√≥n hidr√≥foba sin contener ning√ļn principio activo desde el punto de vista qu√≠mico o farmacol√≥gico, bas√°ndose en un mecanismo de acci√≥n f√≠sica gracias al revestimiento hidr√≥fobo con cloruro de diaquilcarbamilo (DACC). Objetivo principal: Determinar la efectividad de los ap√≥sitos antimicrobianos con propiedades hidr√≥fobas en el tratamiento de heridas cr√≥nicas de las extremidades inferiores de etiolog√≠a venosa con sospecha de infecci√≥n o infecci√≥n local. Metodolog√≠a: Se plantea un estudio pre ‚Äď experimental de tipo pre ‚Äď post sin grupo control en los 5 Centros de Atenci√≥n Primaria de la ciudad de Girona. Se incluir√°n a los pacientes que cumplan los criterios de selecci√≥n y ser√°n elegidos de manera aleatoria. Para poder calcular el tama√Īo muestral se realizar√° un estudio piloto para calcular el Riesgo Relativo. El per√≠odo de seguimiento de los pacientes tendr√° una duraci√≥n de 4 semanas, con una valoraci√≥n inicial y posteriormente semanal mediante una hoja de recogida de datos elaborada al efecto del estudio. Se realizar√° un an√°lisis secuencial de tipo univariante y, posteriormente, de tipo bivariante.ABSTRACT: Background: Ulcers in lower extremities of venous etiology are very prevalent at present, generating a problem individually, for the Society and for the Health System. Healing is a very complex process in which many aspects have to be taken into account, such as wound infection, among others. There are many alternatives for the treatment of infection of chronic wounds and it is antibiotics and antiseptics, whose main disadvantage is that they can end up creating resistance and/or allergic reactions. The alternative, aforementioned, are the dressings with hydrophobic properties, whose objective is to attract and trap the microorganisms present in the wound bed in their fibers, through hydrophobic interaction without containing any active substance from the chemical or pharmacological point of view, based on a mechanism of physical action due to the hydrophobic coating with diaquilcarbamyl chloride (DACC). Objective: To determine the effectiveness of antimicrobial dressings with hydrophobic properties in the treatment of chronic wounds of the lower extremities of venous etiology with suspicion of infection or local infection. Method: A pre ‚Äď experimental, pre ‚Äď post study without a control group will be proposed in the 5 Primary Care Centers of the city of Girona. Patients who complete the selection criteria will be included and will be chosen randomly. In order to calculate the sample size, a pilot study will be carried out to calculate the Relative Risk. The period of follow-up of the patients will last for 4 weeks, with an initial and later weekly assessment through a data collection sheet prepared for the purpose of the study. A sequential analysis of univariate type and, later, of bivariate type will be carried out.M√°ster en Gesti√≥n Integral e Investigaci√≥n de las Heridas Cr√≥nica

    Efficacy of a nurse nutritional intervention on venous leg ulcer’s healing

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    RESUMEN: Introducci√≥n Se ha comprobado la alta prevalencia de las √ļlceras de extremidad inferior, encontr√°ndose en primer lugar las de etiolog√≠a venosa. Este dato unido a la creciente esperanza de vida en nuestro pa√≠s, han hecho que sea un problema de salud importante que hay que abordar de manera hol√≠stica. Diferentes aspectos influyen en la cicatrizaci√≥n de las √ļlceras de etiolog√≠a venosa, uno de los m√°s importantes es el aspecto nutricional de la persona que la sufre. La nutrici√≥n est√° directamente relacionada con la cicatrizaci√≥n de las heridas y el pron√≥stico de las mismas. Realizar una adecuada valoraci√≥n nutricional y una intervenci√≥n enfocada en este aspecto puede reducir el tiempo de cicatrizaci√≥n de las √ļlceras y mejorar la calidad de vida de las personas diagnosticadas con esta dolencia. Objetivos - Determinar la eficacia de una intervenci√≥n enfermera nutricional, que incluya suplementos de arginina, vitamina C y zinc, frente a la intervenci√≥n nutricional habitual (dieta hiperproteica) en la cicatrizaci√≥n de las UV. - Evaluar la fiabilidad y concordancia de dos instrumentos de cribado nutricional (MUST y MNA) en pacientes con UV. M√©todo Ensayo cl√≠nico aleatorizado realizado en los Centros de Atenci√≥n Primaria seleccionados del √Ārea de Salud de Burgos. Se incluir√°n pacientes mayores de 65 a√Īos con diagnostico de √ļlcera de etiolog√≠a venosa con una puntuaci√≥n en la escala RESVECH 2.0. de 17 puntos o superior. Se excluir√°n del mismo los pacientes con deterioro cognitivo o que no puedan realizar el seguimiento de la √ļlcera en el Centro de Salud. Se compara una intervenci√≥n nutricional de pr√°ctica habitual con una intervenci√≥n nutricional basada en la evidencia cient√≠fica que contempla un aumento en la densidad de nutrientes esenciales en el proceso de cicatrizaci√≥n. Se ha calculado un tama√Īo muestral de 216 participantes ajustado a las p√©rdidas. Se va a realizar un an√°lisis de los datos por intenci√≥n de tratar. Para ello se llevar√° a cabo estad√≠stica descriptiva e inferencial. Se utilizar√°n frecuencias y porcentajes para las variables cualitativas. En el caso de las cuantitativas, dependiendo de su normalidad, las medidas de tendencia central correspondientes. En el caso de la estad√≠stica inferencial, se utilizar√°n las pruebas param√©tricas o no param√©tricas m√°s adecuadas a las variables de inter√©s, as√≠ como se calcular√° el riesgo relativo y diferencia de riesgos para la incidencia de cicatrizaci√≥n entre el grupo experimental y control. Adem√°s, se llevar√° a cabo an√°lisis multivariante mediante curvas de supervivencia de Kaplan-Meier y la prueba de Log-Rank.ABSTRACT: Introduction The high prevalence of lower limb ulcers has been confirmed, finding in the first place the venous ulcers. This fact, joined with the growing life expectancy in our country, has performed it as a relevant health problem that needs to be approached holistically. Various aspects influence on the wound healing of venous etiology ulcers; one of the most important is the nutritional aspect of the person who suffers it. Nutrition is directly related to wound healing and their prediction. Performing an adequate nutritional assessment and an intervention focused on this aspect can reduce the healing time of ulcers and improve the quality of life of people diagnosed with this disease. Objectives - Determine the efficacy of a nutritional nursing intervention, including supplements of arginine, vitamin C and zinc, against to the usual nutritional intervention (hyperprotein diet) in the healing of UV. - Evaluate the reliability and concordance of two nutritional screening instruments (MUST and MNA) in UV patients. Method Randomized clinical study conducted at the Primary Health-Care Centres selected from the Health Area of Burgos. Patients older than 65 years with diagnosis of venous etiology ulcer with a score of more than 17 points on the RESVECH 2.0 scale will be included. Patients with cognitive impairment or which are unable to follow up the ulcer at the Health- Care Centre will be excluded. A nutritional intervention of an habitual practice is compared with a nutritional intervention based on scientific evidence that contemplates an increase in the density of essential nutrients during the healing process. A sample size of 216 participants has been calculated in accordance with the sample losses. An analysis of the data will be performed, which will be done by descriptive and inferential statistics. Frequencies and percentages will be used for qualitative variables. In the case of quantitative variables, depending on their normality, the corresponding central trend measures. For inferential statistics, parametric or nonparametric tests, which are more adequate to the variables of the present study, shall be used; as well as calculating the relative risk and risk difference for the incidence of healing between the experimental group and control. In addition, multivariate analysis will be performed through Kaplan-Meier and Log-Rank test.M√°ster en Gesti√≥n Integral e Investigaci√≥n de las Heridas Cr√≥nica

    Abstracts Book : XXVI - SILAE 2017 Societ√† Italo-Latinoamericana di etnomedicina y IX Congreso colombiano de cromatograf√≠a ‚Äď COCOCRO 2017

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    El presente documento recoge 2 investigaciones realizadas en modo ‚ÄúOral communications‚ÄĚ y 11 en modo ‚ÄúPoster‚ÄĚ presentadas para el congreso XXVI‚Äď SILAE 2017 Societ√† Italo-Latinoamericana di etnomedicina y IX Congreso colombiano de cromatograf√≠a ‚Äď COCOCRO 2017; realizado en Cartagena. Estos res√ļmenes recogen la investigaci√≥n sobre la leishmaniasis y el desarrollo de fitomedicinas basadas en plantas, que permitan tratar la esta condici√≥n y sus manifestaciones como medicinas alternativas, que sean efectivas y a bajo costo. Los trabajos presentados fueron: ‚ÄúOral communications‚ÄĚ: (1) Cytotoxicity and leishmanicidal activity of essential oil from plants present in Colombia and commonly used in traditional medicine; (2) Studies in vitro and in vivo of antileishmanial activity and differential cytotoxicity of Cannabis spp. En modo ‚ÄúPoster‚ÄĚ (1) Phytochemistry and ethnopharmacological research on isla grande, Cartagena; (2) Ethnobotanical study of medicinal plants in Isla Grande, Cartagena Colombia; (3) In vitro evaluation of interaction of anti-inflammatory, wound healing and anti-leishmanial activities. An useful strategy to identify hit compounds for drug development to treat cutaneous leishmaniasis; (4) Chemical composition and evaluation of the biological potential of three vegetable species of the caribbean region; (5) Characterization by means of gas chromatography / mass spectrometry (gc / ms) and evaluation of the antibacterial activity of the essential oil of the bark from Astronium graveolens; (6) Synthesis and trypanocide activity of chloro and bromo l-tyrosine derivatives; (7) Leishmanicidal and citotoxic activity of benzothiopyrans. Structure-activity relationship; (8) Chemical composition, antioxidant activity, antimicrobial and antiprotozoal properties of ethanol propolis extracts of Bolivia; (9) Leishmanicidal activity and cytotoxicity in vitro of styrylquinolines; (10) Antileishmanial activity of rich fractions in alkaloid and flavonoid to design and develop a new low cost topical phytomedicine product from medicinal plants in the northern coast of Colombia; (11) Synthesis, leishmanicidal activity and chiral resolution of 4-oxothiochromane-2-carboxylic acid and its esters and amides derivatives.Departamento Administrativo de Ciencia, Tecnolog√≠a e Innovaci√≥n [CO] Colciencias1107-569-33684Aislamiento y purificaci√≥n de compuestos con potencial actividad antileishmanial a partir de Cordia dentada Poir. y Heliotropium indicum L.n

    Aplica√ß√Ķes biom√©dicas de materiais fotoativos √† base de amido

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    The centrality that light presents in the human being‚Äôs life since antiquity to its use in industrial and biomedical applications makes it a topic of intense research. The knowledge of its properties and the way it affects the environment and living beings allows us to find new applications, namely biomedical. Photodynamic therapy is known since the end of the 1970‚Äôs as a promising approach in the treatment of neoplasia. This therapeutic approach takes advantage of light‚Äôs interaction with photosensitive compounds, named photosensitizers, which in the presence of molecular oxygen produce reactive oxygen species capable of leading tumoral cells to cellular death. From tumoral cells to microbial cells was only a step, and this photodynamic approach has proved very successful, more specifically using porphyrins as photosensitizers, in inactivating microorganisms, including microbial strains multi-resistant to conventional antimicrobial agents (e.g., antibiotics, antifungals). With the rise of microbial resistance to conventional antimicrobials and increasing emergence of chronical conditions such as diabetes and its complications, it becomes imperative to find new therapies which allow to treat and help meliorate patients‚Äô quality of life. The goal of this study was to evaluate the possibility of using cost-friendly materials, such as starch, to support porphyrinic derivatives and in this way to produce photoactive materials with the ability to act as photosensitizers and inactivate common skin infections such as the ones incited by methicillin resistant Staphylococcus aureus and Pseudomonas aeruginosa, and which arise in diabetic ulcers, and furthermore to evaluate its ability to promote tissue regeneration. The porphyrins selected to incorporate in starch were cationic porphyrin 5,10,15,20-tetrakis(1-methylpyridinium-4-yl)porphyrin (TPP5F) and neutral porphyrin 5,10,15,20-tetrakis(pentafluorophenyl)porphyrin (TMPyP) which were prepared and later immobilized in the starch-based materials. The photoactive films prepared by incorporation of porphyrin in starch were then tested and their antimicrobial properties evaluated in in vitro and ex vivo studies. The TMPyP incorporated starch-based film revealed promising antimicrobial activity as it was able to inactivate S.aureus (MRSA), both in vitro and ex vivo, after white light exposure with an irradiance of 50 mW cm-2 for 60 min and 24h, respectively. Using porcine skin as skin infection model, this TMPyP film revealed potential in preventing the onset of bacterial infections. To evaluate the biocompatibility and possible stimulating effects in tissue regeneration of the starch/ porphyrin films, in vitro studies were conducted in cell lines of fibroblasts (HDF) and endothelial cells (HMEC), two important components of tissue regeneration and wound healing. Four different parameters were tested: cellular viability, cellular migration, ROS formation and cellular adhesion. For that, the starch/porphyrin-based materials were applied and irradiated with a red light from a LED system with an irradiance of 5 mW cm-2 for 15 min. The films of 7.06 mm2 revealed the best results in viability and at these conditions the cells present high levels of cellular ROS. The application of light seems to have influenced all parameters except ROS formation. Low dose red-light seems to positively influence viability and wound healing in endothelial cells, revealing itself as a potential promoter of vascularization. The results obtained with the starch/porphyrin films allow for a perspective of their potential application in the treatment of diabetics‚Äô wounds/ulcers.A centralidade que a luz apresenta na vida do ser Humano desde a antiguidade at√© √† sua utiliza√ß√£o em aplica√ß√Ķes industriais e biom√©dicas faz com que esta continue a ser objeto de intensa investiga√ß√£o. O conhecimento das suas propriedades e da forma como afeta o ambiente e os seres vivos permite encontrar novas aplica√ß√Ķes nomeadamente biom√©dicas. A terapia fotodin√Ęmica √© reconhecida desde finais da d√©cada de 70 como uma abordagem promissora no tratamento de neoplasias. Esta abordagem terap√™utica tira partido da intera√ß√£o da luz com compostos fotossens√≠veis designados de fotosensibilizadores que na presen√ßa de oxig√©nio molecular produzem esp√©cies reativas de oxig√©nio capaz de levar √† morte celular de c√©lulas tumorais. Das c√©lulas tumorais √†s c√©lulas microbianas foi um passo, e esta abordagem fotodin√Ęmica tem sido muito bem sucedida nomeadamente utilizando porfirinas como fotosensibilizadores, na inativa√ß√£o de microorganismos incluindo estirpes microbianas multi-resistentes aos agentes antimicrobianos convencionais (eg. antibi√≥ticos, antif√ļngicos, etc). Com o aumento da resist√™ncia microbiana aos antimicrobianos convencionais e o surgimento crescente de condi√ß√Ķes cr√≥nicas como diabetes e as suas complica√ß√Ķes, torna-se imperativo encontrar novas terapias que permitam tratar e melhorar a qualidade de vida dos pacientes. Este estudo teve como objetivo avaliar a possibilidade de utilizar materiais baratos, como seja o amido, para suportar derivados porfir√≠nicos e desta forma produzir materiais fotoactivos com capacidade de atuar como fotosensibilizadores e inactivarem comuns infe√ß√Ķes de pele como sejam as provocadas por Staphylococcus aureus resistente √† meticilina (MRSA) e Pseudomonas aeruginosa e que surgem em √ļlceras de diab√©ticos, e mais do que isso, avaliar a sua capacidade de promoverem a regenera√ß√£o de tecidos. As porfirinas selecionadas para incorporar no amido foram a porfirina cati√≥nica 5,10,15,20-tetraquis(1- metilpiridinium-4-il)porfirina (TMPyP) e porfirina neutra 5,10,15,20- tetraquis(pentafluorofenil)porfirina (TPP5F) que foram preparadas e posteriormente imobilizadas nos materiais √† base de amido. Os filmes fotoactivos preparados por incorpora√ß√£o de porfirina em amido, foram ent√£o testados e as suas propriedades antimicrobianas em estudos in vitro e ex vivo avaliadas. O filme √† base de amido com a TMPyP incorporada revelou promissora atividade antimicrobiana sendo que conseguiu inativar a S. aureus (MRSA), tanto in vitro como ex vivo, ap√≥s exposi√ß√£o a luz branca com uma irradi√Ęncia de 50 mW cm-2 por 60 min e 24h, respetivamente. Utilizando pele de porco como um modelo de infe√ß√£o de pele, este filme contendo TMPyP revelou potencial em prevenir a instala√ß√£o de infe√ß√Ķes bacterianas. Para avaliar a biocompatibilidade dos filmes de amido-porfirina preparados e poss√≠veis efeitos estimulantes na regenera√ß√£o de tecidos, foram realizados estudos in vitro em linhas celulares de fibroblastos (HDF) e c√©lulas endoteliais (HMEC), dois componentes importantes da regenera√ß√£o de tecido e cura de feridas. Quatro par√Ęmetros diferentes foram testados: viabilidade celular, migra√ß√£o celular, forma√ß√£o de ROS e ades√£o celular. Para tal os filmes √† base de amido-porfirinas foram aplicados e irradiados com luz vermelha proveniente de um sistema LED com uma irradi√Ęncia de 5 mW cm-2 por 15 min. Os filmes de dimens√Ķes 7.06 mm2 revelaram os melhores resultados nos ensaios de viabilidade celular, e as c√©lulas nestas condi√ß√Ķes apresentam elevada quantidade de ROS celular. A aplica√ß√£o de luz parece ter influenciado todos os par√Ęmetros exceto a forma√ß√£o de ROS. A luz vermelha em baixa dose parece influenciar positivamente a viabilidade e regenera√ß√£o em c√©lulas endoteliais, revelando-se um potencial promotor de vasculariza√ß√£o. Os resultados obtidos com os filmes amido-porfirina permitem perspetivar a sua potencial aplica√ß√£o no tratamento de feridas/√ļlceras de diab√©ticos.Mestrado em Biomedicina Molecula

    Modelos 3D humanizados para o estudo de doenças pulmonares

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    Pulmonary fibrosis, characterized by progressive and irreversible lung tissue stiffening resulting in organ failure, is a growing health problem and belongs to the major causes of death worldwide. The pathological mechanisms of lung fibrosis are not fully understood; current pathogenic theories assume an impaired wound healing response to chronic lung injuries, in which the mechanical and chemical stimuli from the lung environment induces fibroblast activation. Currently, therapeutic options are severely limited, and lung transplantation remains the only effective treatment for patients in end-stage fibrotic diseases. Complex tridimensional (3D) lung platforms able to accurately recapitulate function, structure, and cell and matrix interactions found in fibrotic lung tissue, are therefore necessary to provide the means for understanding the pathological mechanisms and mediators involved in the fibrotic process. Of the vast array of biomaterials that have been used for Tissue Engineering (TE) applications, a major enthusiasm has been developed towards hydrogels: 3D water-swollen polymeric networks, that provide mechanical support to cells and allow for the diffusion of nutrients, waste, and oxygen. Hydrogels are particularly interesting to study lung diseases as they recapitulate the mechanical and viscoelastic properties found in load-bearing soft tissues like the lung. Natural-based hydrogels are appealing platforms as they are inherently biocompatible and bioactive. Platelet-rich plasma (PRP) and human platelet lysates (PL) provide interesting materials to create hydrogels as they are a source for human-derived growth factors (GF). However, they present poor mechanical properties and are easily degraded. Synthetic-derived hydrogels do not face these limitations, but they lack differentiative cues required for tissue development. Human methacryloyl platelet lysates (PLMA)- based hydrogels have been proposed as a biochemical and biomechanicalsuperior platform for cell culture purposes. These autologous, GF-rich, platforms are herein proposed as reliable 3D platforms to model the fibrotic lung. PLMA hydrogels recapitulated the pathological stiffness of the fibrotic lung and supported the viability of lung fibroblasts cells for at least 7 days in culture. Cells adopted different morphologies as matrix stiffness changed and were able to induce matrix deformations in PLMA hydrogels, suggesting the feasibility of this scaffold to induce a profibrotic phenotype in fibroblasts in 3D, therefore recapitulating the pathological remodeling of lung fibrosis.A fibrose pulmonar, que se caracteriza por cicatriza√ß√£o progressiva e irrevers√≠vel do tecido pulmonar, culminando em falha pulmonar, √© umas das principais causas de morte a n√≠vel mundial. Os mecanismos patol√≥gicos na g√©nese da fibrose pulmonar n√£o s√£o claros; as teorias atuais sugerem a ocorr√™ncia de uma cicatriza√ß√£o anormal em resposta a les√Ķes pulmonares cr√≥nicas, nas quais os est√≠mulos mec√Ęnicos e qu√≠micos do ambiente pulmonar induzem a ativa√ß√£o dos fibroblastos. As op√ß√Ķes de tratamento atuais s√£o muito limitadas, sendo o transplante pulmonar a √ļnica op√ß√£o vi√°vel para pacientes em est√°gio final de fibrose. Plataformas tridimensionais (3D) de pulm√£o capazes de simular a fun√ß√£o e estrutura pulmonares e as intera√ß√Ķes c√©lula-c√©lula e c√©lula-matriz s√£o, por isso, necess√°rias para compreender os mecanismos patol√≥gicos e os mediadores envolvidos no processo fibr√≥tico. Dos v√°rios biomateriais usados em engenharia de tecidos (ET), os hidrog√©is t√™m ganho destaque: s√£o redes polim√©ricas 3D que absorvem √°gua, capazes de fornecer suporte mec√Ęnico √†s c√©lulas e de permitir a difus√£o de nutrientes, metabolitos e oxig√©nio. S√£o particularmente interessantes para estudar doen√ßas pulmonares, uma vez que conseguem simular as propriedades mec√Ęnicas e viscoel√°sticas de tecidos moles como o pulm√£o. Os hidrog√©is naturais s√£o plataformas atrativas por serem biocompat√≠veis e bioativas. Plasma Rico em Plaquetas (PRP) e Lisados de Plaquetas (LP) humanos t√™m sido usados para criar hidrog√©is, por serem uma fonte humana de fatores de crescimento (FC). No entanto, t√™m fracas propriedades mec√Ęnicas e s√£o facilmente degrad√°veis. Os hidrog√©is sint√©ticos n√£o t√™m estas limita√ß√Ķes, mas faltam-lhes sinais indutores de diferencia√ß√£o, cruciais para o desenvolvimento dos tecidos. Hidrog√©is √† base de LP metacrilatados (LPM) foram recentemente propostos como plataformas bioquimicamente e biomecanicamente superiores para cultura de c√©lulas. Neste trabalho, propomos estes hidrog√©is aut√≥logos e ricos em FC como uma plataforma 3D capaz de mimetizar o pulm√£o fibr√≥tico. Os hidrog√©is de LPM recapitularam a rigidez do pulm√£o fibr√≥tico, e permitiram manter fibroblastos pulmonares vi√°veis durante pelo menos 7 dias em cultura. As c√©lulas adotaram diferentes morfologias consoante a rigidez da matriz e induziram marcadas deforma√ß√Ķes nos hidrog√©is de LPM, sugerindo que estas plataformas induziram um fen√≥tipo fibr√≥tico nos fibroblastos e que, por isso, mimetizam a remodela√ß√£o patol√≥gica da matriz extracelular que ocorre na fibrose pulmonar.Mestrado em Bioqu√≠mic

    PRISMA 2020 checklist.

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    MicroRNAs (miRNAs) are responsible for regulating gene expression post-transcriptionally. Are involved in several biological processes, such as wound healing. Understanding the miRNAs involved in this process is fundamental for the development of new therapies. So, due to the need to understand the role of these molecules, we aimed systematically review the literature in order to identify which miRNAs are involved in the wound healing and determine, through bioinformatics analysis, which signaling pathways are associated with these miRNAs. An electronic search was performed in the following databases: National Library of Medicine National Institutes of Health (PubMed), Science Direct, Scifinder, Scopus and Web of Science, using the descriptors: ‚Äú(microRNA [MeSH])‚ÄĚ and ‚Äú(skin [MeSH])‚ÄĚ and ‚Äú(wound healing [MeSH])‚ÄĚ. After the search, two independent and previously calibrated reviewers selected the articles that analyzed the expression pattern of miRNAs in wound healing in in vivo studies, using the software Zotero bibliography manager. Following, bioinformatic analysis was performed using the software DIANA Tools, mirPath v.3 and the data was interpreted. The bioinformatics analysis revealed that on the day 1 there were 13 union pathways, eight of which were statistically significant. Still on the day 1, among the miRNAs that had a decrease in their expression, 12 of 17 union pathways found were statistically significant. On the day 5, among the miRNAs with an increase in expression, 16 union pathways were found, 12 of which were statistically significant. Finally, among the miRNAs with decreased expression, 11 of 15 union pathways found were statistically significant. Although it has been found substantial heterogeneity in the studies, with this systematic review, it was possible to study the panorama of miRNAs that may be altered in the wound healing. The present review summarizes existing evidence of miRNAs associated to wound healing, and these findings can contribute to new therapeutic approaches.</div

    Synergistic Effect of <i>Sophora japonica</i> and <i>Glycyrrhiza glabra</i> Flavonoid-Rich Fractions on Wound Healing: <i>In Vivo</i> and Molecular Docking Studies

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    Glycyrrhiza glabra and Sophora japonica (Fabaceae) are well-known medicinal plants with valuable secondary metabolites and pharmacological properties. The flavonoid-rich fractions of G. glabra roots and S. japonica leaves were prepared using Diaion column chromatography, and the confirmation of flavonoid richness was confirmed using UPLC-ESI-MS profiling and total phenolics and flavonoids assays. UPLC-ESI-MS profiling of the flavonoid-rich fraction of G. glabra roots and S. japonica leaves resulted in the tentative identification of 32 and 23 compounds, respectively. Additionally, the wound healing potential of topical preparations of each fraction, individually and in combination (1:1) ointment and gel preparations, were investigated in vivo, supported by histopathological examinations and biomarker evaluations, as well as molecular docking studies for the major constituents. The topical application of G. glabra ointment and gel, S. japonica ointment and gel and combination preparations significantly increase the wound healing rate and the reduction of oxidative stress in the wound area via MDA reduction and the elevation of reduced GSH and SOD levels as compared to the wound and Nolaver¬ģ-treated groups. The molecular docking study revealed that that major compounds in G. glabra and S. japonica can efficiently bind to the active sites of three proteins related to wound healing: glycogen synthase kinase 3-ő≤ (GSK3-ő≤), matrix metalloproteinases-8 (MMP-8) and nitric oxide synthase (iNOS). Consequently, G. glabra roots and S. japonica leaves may be a rich source of bioactive metabolites with antioxidant, anti-inflammatory and wound healing properties

    Response to Article &ldquo;Evaluation of the Antioxidant and Wound Healing Properties of 80% Methanol Extract and Solvent Fractions of the Leaves of Vernonia auriculifera Hiern. (Asteraceae)&rdquo; [Letter]

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    Putri Reno Intan,1,&ast; Ariyani Noviantari,1,&ast; Sukmayati Alegantina2,&ast; 1Center for Biomedical Research, Research Organization for Health, National Research and Innovation Agency (BRIN), Cibinong Science Center, Cibinong - Bogor, West Java, Indonesia; 2Research Center for Pharmaceutical Ingredients and Traditional Medicine, Research Organization for Health, National Research and Innovation Agency (BRIN), Cibinong Science Center, Cibinong - Bogor, West Java, Indonesia&ast;These authors contributed equally to this workCorrespondence: Putri Reno Intan, Center for Biomedical Research, Research Organization for Health, National Research and Innovation Agency (BRIN), Genomic Building, Cibinong Science Center, Jalan Raya Bogor Km. 46, Cibinong - Bogor, West Java, 16911, Indonesia, Email [email protected]

    Phytochemical prospection and larvicidal bioactivity of the janaguba (Himatanthus drasticus) Mart. Plumel (Apocynaceae) latex against Aedes aegypti L. (Diptera: Culicidae)

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    Abstract The aim of this study was to carry out phytochemical prospecting and evaluate the larvicidal activity of Himatanthus drasticus latex extracts against Aedes aegypti. The extracts were obtained by maceration from 5 g of latex powder concentrated separately in 100 mL of methanol, ethyl acetate, and hexane solvents. The concentrations of 100, 200, 300, 400, and 500 ppm of each extract were tested in triplicate with a solution of pyriproxyfen as the positive control and distilled water and dimethylsulfoxide as the negative control. The phytochemical prospection of the methanolic extract showed the presence of phenolic compounds, such as anthocyanins, anthocyanidins, catechins, chalcones, aurones, leucoanthocyanidins, and condensed tannins. The insecticidal bioactivity was most significant for the methanolic extract. The methanolic extract lethal concentrations (LC) of 50 and 90% were 190.76 and 464.74 ppm, respectively. After 48 hours of exposure, the extracts using methanol, ethyl acetate, and hexane at their highest concentrations (500 ppm) caused larval mortality of 100, 73.33, and 66.67%, respectively. These extracts also promoted changes in the external morphology of the larvae, such as damage to the anal papillae, darkening of the body, and reduction in the number of bristles. The methanolic extract showed greater expressivity for morphological changes. The latex of H. drasticus has larvicidal activity against third-stade larvae of A. aegypti and it is more significant when obtained through maceration in methanol. The methanolic extract of H. drasticus latex contains phenolic compounds with insecticidal activity against A. aegypti larvae
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