Glycosaminoglycan-based hydrogels for the cytokine management in wound healing

Impaired wound healing and the resulting chronic wounds may cause significant morbidity and mortality. In these pathogenic wound environments, the ratio of inflammatory and anti-inflammatory cytokines is highly biased to the pro-inflammatory side. While the inflammatory process is an essential step in healthy wound healing, chronic wounds remain in a constant self-sustaining state of inflammation. Thus, decreased cell proliferation, reduced matrix deposition and delayed wound closure are the results. Although various cytokine-based therapies have shown promising results on skin regeneration in preliminary studies, their overall clinical use has been considerably limited by the short half-life time of the signaling molecules due to rapid dilution and degradation in the protease-rich chronic wound environment. In this work, we explored the ability of starPoly(ethylene glycol)-GAG hydrogels to modulate the hallmarks of chronic wound development, such as the prolonged inflammation, increased cell influx and delayed proliferative phase. Therefore, different strategies were developed to shape the cytokine levels in the wound towards a more pro-regenerative direction, finally promoting the natural repair process in chronic skin wounds. By biomimetically utilizing the interactions between cytokines and the tissue ECM in a GAG-based biohybrid hydrogel, we could engineer the concentrations of various signaling factors involved in the regulation of the repair process. More in detail, we utilized customized functionalized starPEG-GAG hydrogels to (1) reduce the extensive levels of inflammatory chemokines by scavenging them via GAG component of the hydrogel and thus diminish immune cell influx in a mouse wound model; (2) locally deliver the immunomodulatory IL-4 and IL-10 to shift the signaling balance into the pro-regenerative direction and thus resolve inflammation and (3) administer pre-conjugated TGF-β to enhance myofibroblast differentiation and extracellular matrix deposition. We believe that the presented hydrogel platform may become a promising tool in the management of cytokines in regenerative applications, which can be translated towards the clinical use for the treatment of chronic wounds and other diseases characterized by uncontrolled inflammation.:1 introduction 1.1 Motivation 1.2 Current state of biomaterial-based concepts in dermal wound healing 1.3 Objective 2 fundamentals 2.1 The physiological process of wound healing 2.1.1 The role of macrophages in wound healing 2.1.2 The role of fibroblasts in wound healing 2.1.3 The role of cytokines and their interaction with the ECM 2.2 The pathophysiology of chronic wounds 2.3 Strategies for treatment of chronic wounds 2.4 Biomaterials in medicine 2.4.1 Polymers in medicine 2.4.2 Mechanical properties 2.4.3 Cellular adhesion 2.4.4 Interaction with cytokines 2.4.5 Scaffold degradability 2.4.6 StarPEG-GAG hydrogels as potential material in wound healing 3 materials & methods 3.1 Preparation of hydrogels 3.1.1 Functionalization of glass surfaces 3.1.2 Hydrogel formation with EDC - NHS chemistry 3.1.3 Hydrogel formation with thiol - maleimide chemistry 3.1.4 Rheometric measurement of hydrogel discs 3.1.5 Characterization of cytokine uptake and release 3.2 Culture of human & murine cells 3.2.1 Isolation and differentiation of murine dermal fibroblasts 3.2.2 Isolation & differentiation of murine macrophages 3.2.3 Culture of human & murine cell lines 3.3 In vitro methods 3.3.1 Enzyme-linked immunosorbent assay (ELISA) 3.3.2 Bead-based multiplex immunoassay 3.3.3 Live/Dead Staining 3.3.4 Crystal violet staining 3.3.5 Cell proliferation assay 3.3.6 RNA extraction & analysis 3.3.7 cDNA synthesis 3.3.8 Quantitative real time rt-PCR 3.4 Statistical analysis 3.5 Software use 4 scavenging inflammatory chemokines to control immune cell influx in the wound 4.1 Results 4.1.1 Engineering heparin-based hydrogels to scavenge chemokines 4.1.2 Heparin-based hydrogels reduce migration of immune cells 4.1.3 Heparin-based hydrogels decrease wound immune cell influx and inflammatory signaling 4.2 Discussion 5 promotion of regenerative macrophage polarizationin inflammatory environments 5.1 Results 5.1.1 Reversible complexation of IL-4 & IL-10 to starPEG-heparin gels 5.1.2 Stabilizing effects of starPEG-heparin gels on IL-4 5.1.3 IL-4 & IL-10-laden starPEG-heparin hydrogels modulate macrophage polarization 5.1.4 IL-4-laden starPEG-heparin induce collagen deposition in dermal fibroblasts 5.2 Discussion 6 modulation of human dermal fibroblast proliferation and differentiation 6.1 Results 6.1.1 Reversible complexation of TGF-b to starPEG heparin gels 6.1.2 Cell attachment, spreading and proliferation 6.1.3 Matrix deposition by fibroblasts grown on starPEG-heparin hydrogels 6.1.4 Degradation of starPEG-heparin hydrogels 6.1.5 TGF-b-laden starPEG-heparin that efficiently induces myofibroblast differentiation 6.2 Discussion 7 general discussion 7.1 Summary and conclusion 7.2 Future perspective Appendix 8 supplementary materials & methods 9 declaration of authorship 10 publications and conference contributions bibliography list of figures list of tables nomenclature selbstständigkeitserklärun

Chemokine‐Capturing Wound Contact Layer Rescues Dermal Healing

Excessive inflammation often impedes the healing of chronic wounds. Scavenging of chemokines by multiarmed poly(ethylene glycol)-glycosaminoglycan (starPEG-GAG) hydrogels has recently been shown to support regeneration in a diabetic mouse chronic skin wound model. Herein, a textile-starPEG-GAG composite wound contact layer (WCL) capable of selectively sequestering pro-inflammatory chemokines is reported. Systematic variation of the local and integral charge densities of the starPEG-GAG hydrogel component allows for tailoring its affinity profile for biomolecular signals of the wound milieu. The composite WCL is subsequently tested in a large animal (porcine) model of human wound healing disorders. Dampening excessive inflammatory signals without affecting the levels of pro-regenerative growth factors, the starPEG-GAG hydrogel-based WCL treatment induced healing with increased granulation tissue formation, angiogenesis, and deposition of connective tissue (collagen fibers). Thus, this biomaterials technology expands the scope of a new anti-inflammatory therapy toward clinical use

Integrative review: indicators of result process of organ donation and transplants

OBJECTIVE: Identify by integrated review the main result indicators in the process of organ donation and transplants used in Brazil and worldwide. The integrated review of this literature was performed on bibliographic database MedLine/PubMed and LILACS and governmental and nongovernmental sites between 1995 and 2011. The describers/keywords used were organ donations, organ transplant and results in health, being selected 26 articles and nine sites. The evidence level classification in the articles changed from one to six. RESULTS: The evidence level in the articles in its totality was 66.6% was four (12:18 PubMed) all the articles of LILACS database (8). The indicators showed in the articles intends to evaluate, assess, measure and control data related to the profile of the donor, clinical and hospital conditions, ischemia timing, organ size, surgical procedure and the complications that came from transplants.OBJETIVO: Identificar por meio da revisão integrativa os principais indicadores de resultado do processo de doação de órgãos e transplantes utilizados e transplantes no Brasil e no mundo. A revisão integrativa da literatura foi realizada nas bases de dados bibliográficas MedLine/PubMed e LILACS e sites governamentais e não governamentais no período de 1995 e 2011. Os descritores/palavras chaves utilizados foram doação de órgãos, transplante de órgãos e resultados em saúde, sendo selecionados 26 artigos e nove sites. A classificação do nível de evidência dos artigos variou de um a seis. RESULTADOS: Nível de evidência dos artigos na sua maioria 66,6% foi quatro (12:18 PubMed) todos os artigos da base LILACS (8). Os indicadores apresentados nos artigos se propõem a avaliar, mensurar e controlar dados relacionados ao perfil do doador, condições clínicas e hospitalares, tempo de isquemia e tamanho do órgão, procedimento cirúrgico e as complicações advindas do transplante.Secretaria de Saúde do Estado de São Paulo Central de Transplantes de São PauloUniversidade Federal de São Paulo (UNIFESP) Escola Paulista de MedicinaUniversidade Federal de São Paulo (UNIFESP) Escola Paulista de EnfermagemCentro Universitário para o Desenvolvimento do Alto Vale do ItajaíUNIFESP, EPMUNIFESP, EPESciEL

Determinação da linha de base e adicionalidade de um projeto de MDL a partir da emissão de poluentes utilizando diferentes combustíveis

In many urban centers, bus and minibus usually are the main means of transport of the population, making public transport a potential source of emission of polluting and greenhouse gases. As it is well known that it is possible to estimate and compare the amount of pollutants emitted by such vehicles using alternative fuels, this study aims to estimate the less polluter fuel, taking as a parameter the main gases from their burning, considering diesel, biodiesel (100%) and natural gas, as well to determine the potential of Cascavel (assessed city) to elaborate a CDM project. To estimate the emission, it was used the methods "bottom-up" and mass balance, in order to compare the gaseous emissions found in the two methodologies. Among the fuel measured, which showed higher emissions it was the diesel, concerned to particulate matter, carbon dioxide and hydrocarbons. Regarding the CDM project, it was found a potential reduction of 8,000 t/CO2.year when one replaces the diesel by biodiesel. This work only represents an estimate of emissions, however already demonstrates the need to seek alternatives to the diesel use. Key words: biodiesel, diesel, natural gas, clear development mechanism, collective transport.Em muitos centros urbanos, ônibus e microônibus estão entre os principais meios de transporte da população, tornando o transporte público uma fonte potencial da emissão de poluentes e de gases do efeito estufa. Sabendo-se que é possível estimar e comparar a quantidade de poluentes emitidos por esses veículos utilizando combustíveis alternativos, este trabalho objetiva estimar o combustível menos poluidor, tomando-se como parâmetro os principais gases provenientes de sua combustão, considerando diesel, biodiesel (100%) e gás natural, bem como determinar o potencial de Cascavel (cidade avaliada) em gerar um projeto de MDL. No cálculo das emissões foram utilizados os métodos bottom-up e balanço de massa, de modo a se comparar as emissões gasosas encontradas nas duas metodologias. Dentre os combustíveis avaliados, o que apresentou maiores emissões foi o diesel, em termos de material particulado, dióxido de carbono e hidrocarbonetos. No que se refere ao projeto de MDL, foi encontrado um potencial de redução de 8.000 t/CO2.ano ao se substituir o diesel pelo biodiesel. Este estudo representa apenas uma estimativa da emissão de poluentes, todavia já demonstra a necessidade de se buscar alternativas à utilização do diesel. Palavras-chave: biodiesel, diesel, gás natural, mecanismos de desenvolvimento limpo, transporte coletivo

Rosácea cutânea e ocular - uma revisão abrangente sobre etiologia, fisiopatologia, manifestações clínicas, fatores de agravamento, diagnóstico, classificação e tratamento

A rosácea é uma condição crônica que afeta a pele e, em alguns casos, os olhos, caracterizada por inflamação e vermelhidão. Sua etiologia não é totalmente compreendida, mas fatores genéticos, ambientais e vasculares estão implicados. Na fisiopatologia, acredita-se que a disfunção do sistema imunológico e a resposta inflamatória desempenhem papéis importantes. As manifestações clínicas da rosácea cutânea incluem rubor facial persistente, pápulas, pústulas e telangiectasias. Além disso, a rosácea ocular pode causar sintomas como olhos secos, sensação de corpo estranho, queimação e fotofobia. Os fatores de agravamento da rosácea são diversos e incluem exposição ao sol, temperaturas extremas, alimentos picantes, álcool, estresse e certos medicamentos. O diagnóstico da rosácea é geralmente clínico, baseado na apresentação dos sintomas e na exclusão de outras condições similares. No entanto, exames adicionais, como cultura bacteriana, podem ser realizados para descartar outras doenças. Quanto à classificação, a rosácea é comumente dividida em subtipos: eritemato telangiectásica, pápulo-pustulosa, fimatosa e ocular. A classificação auxilia no direcionamento do tratamento mais adequado. O tratamento da rosácea envolve uma abordagem multifacetada, que pode incluir medidas tópicas, como cremes e géis contendo agentes anti-inflamatórios, antibióticos ou agentes vasoconstritores. Antibióticos orais e isotretinoína também podem ser prescritos em casos mais graves. Além disso, medidas não farmacológicas, como evitar gatilhos conhecidos e proteger a pele dos raios solares, são fundamentais para controlar os sintomas e prevenir recorrências. No caso da rosácea ocular, tratamentos específicos, como lágrimas artificiais, antibióticos tópicos e corticosteroides, podem ser necessários para aliviar os sintomas e proteger a saúde ocular. O manejo adequado da rosácea requer uma abordagem individualizada, adaptada às necessidades e gravidade de cada paciente

Single-cell genomics identifies cell type-specific molecular changes in autism.

Despite the clinical and genetic heterogeneity of autism, bulk gene expression studies show that changes in the neocortex of autism patients converge on common genes and pathways. However, direct assessment of specific cell types in the brain affected by autism has not been feasible until recently. We used single-nucleus RNA sequencing of cortical tissue from patients with autism to identify autism-associated transcriptomic changes in specific cell types. We found that synaptic signaling of upper-layer excitatory neurons and the molecular state of microglia are preferentially affected in autism. Moreover, our results show that dysregulation of specific groups of genes in cortico-cortical projection neurons correlates with clinical severity of autism. These findings suggest that molecular changes in upper-layer cortical circuits are linked to behavioral manifestations of autism

Tumor gástrico de colisão: adenocarcinoma associado a tumor carcinoide

Relato de caso de paciente masculino, 62 anos, com quadro de dor epigástrica e perda ponderal. A tomografia abdominal subsequente revelou lesão vegetante de 3,0 x 1,5 cm no antro gástrico. O paciente foi submetido à gastrectomia parcial com reconstrução em Y de Roux. Após análise anatomopatológica da peça cirúrgica, evidenciou-se tumor carcinoide maligno de 2,0 cm, associado a adenocarcinoma gástrico do tipo intestinal de Laurén e ausência de metástases linfonodais. A Imunohistoquímica confirmou estes achados. Esse caso é de grande importância para a literatura médica, visto que, ao nosso conhecimento, representa o nono relato de um tumor gástrico dotado dessas características

Gastric collision tumor : adenocarcinoma associated with carcinoid tumor

Relato de caso de paciente masculino, com 62 anos, com quadro de dor epigástrica e perda ponderal. A tomografia abdominal subsequente revelou lesão vegetante de 3,0 x 1,5 cm no antro gástrico. O paciente foi submetido à gastrectomia parcial com reconstrução em Y de Roux. Após análise anatomopatológica da peça cirúrgica, evidenciou-se tumor carcinoide maligno de 2,0 cm associado ao adenocarcinoma gástrico do tipo intestinal de Lauren e à ausência de metástases linfonodais. A imuno-histoquímica confirmou esses achados. Esse caso é de grande importância para a literatura médica, visto que, ao nosso conhecimento, representa o nono relato de um tumor gástrico dotado dessas características.A case report of a 62-year-old male patient with a history of epigastric pain and weight loss. A subsequent abdominal computed tomography (CT) scan revealed a vegetative lesion of 3.0 x 1.5 cm in the gastric antrum. The patient underwent partial gastrectomy with Rouxen- Y reconstruction. After anatomic pathological analysis of the surgical specimen, a 2-cm malignant carcinoid tumor, associated with a Lauren intestinal-type gastric adenocarcinoma and no lymph node metastasis, was detected. Immunohistochemistry confirmed these findings. This case is of great importance to the medical literature, because this report represents, to our knowledge, the ninth case of a gastric tumor with these characteristics reported in the literature

Heparin-based, injectable microcarriers for controlled delivery of interleukin-13 to the brain

Interleukin-13 (IL-13) drives cells of myeloid origin towards a more anti-inflammatory phenotype, but delivery to the brain remains problematic. Herein, we show that heparin-based cryogel microcarriers load high amounts of IL-13, releasing it slowly. Intra-striatal injection of loaded microcarriers caused local up-regulation of ARG1 in myeloid cells for pro-regenerative immunomodulation in the brain. © 2020 The Royal Society of Chemistry

Heparin-based, injectable microcarriers for controlled delivery of interleukin-13 to the brain

Interleukin-13 (IL-13) drives cells of myeloid origin towards a more anti-inflammatory phenotype, but delivery to the brain remains problematic. Herein, we show that heparin-based cryogel microcarriers load high amounts of IL-13, releasing it slowly. Intra-striatal injection of loaded microcarriers caused local up-regulation of ARG1 in myeloid cells for pro-regenerative immunomodulation in the brain