Combinatory approach for developing silk fibroin-based scaffolds with hierarchical porosity and enhanced performance for cartilage tissue engineering applications

Introduction: The combination of several processing technologies can open the possibility for producing scaffolds with superior performance for tissue engineering (TE) applications. Hydrogels are structurally similar to the natural extracellular matrix microenvironment presenting high elasticity and resistance to compression forces. They have been extensively used in biomedical devices fabrication and for TE applications, including for cartilage defects repair[1]. Recently, it was found that proteins like silk fibroin (SF), presenting tyrosine groups can be used to prepare fast formed hydrogels with controlled gelation properties, via an enzyme-mediated cross-linking reaction using horseradish peroxidase (HRP) and hydrogen peroxide (H2O2)[2],[3]. Moreover, the high versatility, processability and tailored mechanical properties of SF, make this natural polymer attractive for the development of innovative scaffolding strategies for cartilage TE applications[4],[5]. Materials and Methods: The present work proposes a novel route for developing SF-based scaffolds derived from high- concentrated SF (16wt%) enzymatically cross-linked by a HRP/H2O2 complex. The combination of salt-leaching and freeze-drying methodologies was used to prepare macro/microporous SF scaffolds with an interconnected structure and specific features regarding biodegradation and mechanical properties (Fig. 1a). The scaffolds morphology and porosity were analyzed by SEM and micro-CT. The mechanical properties (Instron) and protein conformation (FTIR, XRD) were also assessed. In order to evaluate the scaffolds structural integrity, swelling ratio and degradation profile studies were performed for a period of 30 day. This work also aims to evaluate the in vitro chondrogenic differentiation response by culturing human adipose derived stem cells (hASCs) over 21 days in basal and chondrogenic conditions. Cell behaviour in the presence of the macro/microporous structures will be evaluated through different quantitative (Live/Dead, DNA, GAGs, RT PCR) and qualitative (SEM, histology, immunocytochemistry) assays. Results and Discussion: The macro/microporous SF scaffolds showed high porosity and interconnectivity with the trabecular structures evenly distributed (Fig. 1b,c). A dramatic decrease of compressive modulus was observed for samples in hydrated state. Chemical analysis revealed that SF scaffolds displayed the characteristic peaks for β-sheet conformation. Swelling ratio data demonstrated a large swelling capacity, maintaining their structural integrity for 30 days. As expected, when immersed in protease XIV the degradation rate of SF scaffolds increased. Based on the promising morphology and physicochemical properties of the developed SF scaffolds, in vitro chondrogenic differentiation studies with hASCs are envisioned in order to validate their performance for cartilage regeneration applications. Conclusion: This study proposes an innovative approach to produce fast-formed porous SF scaffolds using enzymatically cross- linked SF hydrogels structured by the combination of salt-leaching and freeze-drying methodologies. The obtained results can provide a valuable reference of SF as a tunable and versatile biomaterial with great potential for applications in cartilage TE scaffolding. Portuguese Foundation for Science and Technology (FCT) project PEst-C/SAU/LA0026/201; ERDP funding through POCTEP Project 0687_NOVOMAR_1_P; Investigator FCT program IF/00423/2012 and IF/00411/2013 References: [1] Xia, L.-W., R. Xie, X.-J. Ju, W. Wang, Q. Chen, and L.-Y. Chu, Nano-structured smart hydrogels with rapid response and high elasticity. Nature communications, 2013. 4. [2] Sofia, S.J., A. Singh, and D.L. Kaplan, Peroxidase-catalyzed crosslinking of functionalized polyaspartic acid polymers. Journal of Macromolecular Science, Part A, 2002. 39(10): p. 1151-1181. [3] Reis, R.L., L.-P. Yan, A.L. Oliveira, J.M. Oliveira, D.R. Pereira, C. Correia, and R.A. Sousa, Hydrogels derived from silk fibroin: Methods and uses thereof. 2014. 107426. [4] Chen, C.-H., J.M.-J. Liu, C.-K. Chua, S.-M. Chou, V.B.-H. Shyu, and J.-P. Chen, Cartilage tissue engineering with silk fibroin scaffolds fabricated by indirect additive manufacturing technology. Materials, 2014. 7(3): p. 2104-2119. [5] Yan, L.-P., J.M. Oliveira, A.L. Oliveira, S.G. Caridade, J.F. Mano, and R.L. Reis, Macro/microporous silk fibroin scaffolds with potential for articular cartilage and meniscus tissue engineering applications. Acta biomaterialia, 2012. 8(1): p. 289-301.Â

Finely tuned fiber-based porous structures for bone tissue engineering applications

[Excerpt] Scaffolds developed for bone tissue engineering (TE) must possess specific structural properties to allow neo-tissue formation and integration within the material[1]. Several polymeric systems and processing methodologies have been proposed to develop bone TE scaffolds. Nevertheless, the so far proposed strategies do not fulfil all the requirements for effective bone regeneration. Textile technologies have recently emerged as an industrial route for producing more complex fibre-based porous scaffolds[2]. Silk fibroin (SF) from Bombyx mori has already proved to be a good biomaterial for bone TE[3]. SF-based structures are known for the impressive mechanical properties and biocompatibility, which meet the basic requirements for developing bone TE scaffolds[4],[5]. [...]Portuguese Foundation for Science and Technology (FCT) for the project TISSUE2TISSUE (PTDC/CTM/105703/2008); Investigator FCT program IF/00423/2012 and IF/00411/2013

Importância do uso profilático de Ferro e Vitamina D em lactentes

Introdução: O aleitamento materno é a fonte primordial de nutrição para os recém-nascidos. Todavia, o leite materno possui um teor reduzido de vitaminas e sais minerais, sendo necessário a realização de suplementação. Objetivo: Compreender a importância do uso preventivo de ferro e vitamina D em lactentes externalizando os benefícios dessa prática. Método: Revisão bibliográfica da literatura realizada em janeiro de 2023 nas bases de dados BVS, MEDLINE, LILACS e Google Scholar através dos seguintes DeCS: “Suplementos Nutricionais”, “Fatores de Risco” e “Lactentes” combinados entre si pelo operador booleano AND. Foram encontrados 98 estudos e após aplicar os critérios de inclusão e exclusão, foram selecionados 8 estudos para compor a revisão. Utilizou-se como pergunta norteadora:“Qual a importância do uso preventivo de Ferro e Vitamina D em lactentes?” Resultados: É notório os benefícios da suplementação de ferro e de vitamina D não apenas para os lactentes, mas em diversas fases da vida, essa intervenção pontual pode trazer diversos benefícios para a saúde em geral. Visto que a falta de ferro durante a gravidez estabelece uma ameaça para o desenvolvimento saudável do feto, principalmente em relação ao cérebro. Além disso, compreende-se que a hipovitaminose é considerada uma problemática de saúde pública mundial. Conclusão: As deficiências ocasionadas pela falta de ferro e vitamina D ocorrem com frequência, principalmente nos lactentes que apresentam fatores de risco dietéticos e não necessariamente nutricionais, os quais geralmente ocasionam desfechos clínicos adversos que afetam em sua maioria o desenvolvimento neurológico, impactando consideravelmente a qualidade de vida

Síndrome de DiGeorge: um relato de caso: DiGeorge's Syndrome: a case report

O presente artigo objetivou apresentar o caso clínico de uma paciente pediátrica diagnosticada com a síndrome de DiGeorge e que foi admitida no setor de Cardiologia Pediátrica para correção de malformação cardíaca. Este trabalho se trata de um estudo descritivo, do tipo relato de caso, que visou analisar as principais manifestações desse distúrbio genético, bem como suas abordagens diagnósticas e terapêuticas. A criança foi submetida à correção de defeito cardíaco característico da anomalia e evoluiu com parada cardiorrespiratória, prontamente revertida, e com crise convulsiva no pós-operatório. A anomalia possui espectro clínico diverso, com repercussões que impactam sobremaneira no equilíbrio eletrolítico e nos sistemas imunológico e cardiovascular, exigindo reconhecimento em tempo hábil e a adoção de condutas assertivas para reduzir a morbidade do portador

Chagasic Thymic Atrophy Does Not Affect Negative Selection but Results in the Export of Activated CD4+CD8+ T Cells in Severe Forms of Human Disease

Extrathymic CD4+CD8+ double-positive (DP) T cells are increased in some pathophysiological conditions, including infectious diseases. In the murine model of Chagas disease, it has been shown that the protozoan parasite Trypanosoma cruzi is able to target the thymus and induce alterations of the thymic microenvironment and the lymphoid compartment. In the acute phase, this results in a severe atrophy of the organ and early release of DP cells into the periphery. To date, the effect of the changes promoted by the parasite infection on thymic central tolerance has remained elusive. Herein we show that the intrathymic key elements that are necessary to promote the negative selection of thymocytes undergoing maturation during the thymopoiesis remains functional during the acute chagasic thymic atrophy. Intrathymic expression of the autoimmune regulator factor (Aire) and tissue-restricted antigen (TRA) genes is normal. In addition, the expression of the proapoptotic Bim protein in thymocytes was not changed, revealing that the parasite infection-induced thymus atrophy has no effect on these marker genes necessary to promote clonal deletion of T cells. In a chicken egg ovalbumin (OVA)-specific T-cell receptor (TCR) transgenic system, the administration of OVA peptide into infected mice with thymic atrophy promoted OVA-specific thymocyte apoptosis, further indicating normal negative selection process during the infection. Yet, although the intrathymic checkpoints necessary for thymic negative selection are present in the acute phase of Chagas disease, we found that the DP cells released into the periphery acquire an activated phenotype similar to what is described for activated effector or memory single-positive T cells. Most interestingly, we also demonstrate that increased percentages of peripheral blood subset of DP cells exhibiting an activated HLA-DR+ phenotype are associated with severe cardiac forms of human chronic Chagas disease. These cells may contribute to the immunopathological events seen in the Chagas disease

Síndrome de Chiari e Hidrossiringomielia com comprometimento neurológico: um relato de caso

A Malformação de Chiari (MC) pertence a um amplo grupo de raras deformidades estruturais da junção craniocerebelomedular. O tipo I da doença caracteriza-se pela herniação tonsilar ou amigdaliana cerebelar devido à anomalia da base do crânio e da parte superior da coluna cervical, além de a porção medial do lobo inferior do cerebelo pelo canal cervical também se protuberar através do forame magno, impedindo que o líquor flua normalmente através do canal. A real prevalência da doença é desconhecida, pois muitos pacientes com herniação cerebelar são assintomáticos e o problema agrava-se na fase adulta, com queixas de cefaleia intensa e, por vezes, parestesia. O objetivo deste estudo é relatar um caso de síndrome de Chiari (SC) em uma paciente de 53 anos, ao abordar sua apresentação clínica, diagnóstico e tratamento. Paciente do sexo feminino, 53 anos, foi admitida em um hospital da rede pública de referência se queixando de cefaleia occipital intensa e cervicalgia com irradiação da dor para os membros superiores, acompanhada de parestesia nos quatro segmentos. Relatou já sentir dor há 2 anos, mas apresentou piora do quadro clínico há 8 meses. Foi, também, observada incontinência urinária devido à dissinergia detrusora-esfincteriana por provável bexiga neurogênica. Foi, então, realizado exame de imagem de ressonância magnética (RNM) do crânio e da coluna cervical, com obtenção de sequências ponderadas em T1, T2 e STIR, nos planos sagital e transverso com contraste, o qual evidenciou leve alargamento medular, além de sinais de hidrossiringomielia difusa, com hipossinal na sequência T2 intramedular na altura de D1-D2 (coluna dorsal). Foi notada discreta herniação das tonsilas cerebelares junta ao forame magno, típica da SC, sendo, por fim, confirmado o diagnóstico. A paciente, no entanto, não apresentava hidrocefalia, mesmo com a interrupção do fluxo do líquido cefalorraquidiano (LCR) para o canal vertebral. Ela encaixou- se nos parâmetros de indicação cirúrgica, sendo realizada craniotomia occipital, com acesso ao plexo coroide do quarto ventrículo do tronco encefálico com o intuito de elevar as tonsilas cerebelares baixas, herniadas no canal espinhal cervical e bloqueando o fluxo do LCR. Após a descompressão craniocervical, o curso do líquor foi restaurado e a paciente foi, por fim, encaminhada à sala de recuperação pós-operatória. A SC é uma rara doença que apresenta quadro clínico e alterações radiológicas complexas e extensas e, por vezes, o diagnóstico é retardado devido à inespecificidade dos sintomas confundidos com cervicalgias e cefaleias comuns. A hipótese diagnóstica deve ser embasada nas queixas do paciente, na anamnese minuciosa, exame clínico e nos exames de imagens, sendo a prevalência desta patologia de difícil definição e com faixas etárias distintas

Mortality and pulmonary complications in patients undergoing surgery with perioperative SARS-CoV-2 infection: an international cohort study

Background: The impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on postoperative recovery needs to be understood to inform clinical decision making during and after the COVID-19 pandemic. This study reports 30-day mortality and pulmonary complication rates in patients with perioperative SARS-CoV-2 infection. Methods: This international, multicentre, cohort study at 235 hospitals in 24 countries included all patients undergoing surgery who had SARS-CoV-2 infection confirmed within 7 days before or 30 days after surgery. The primary outcome measure was 30-day postoperative mortality and was assessed in all enrolled patients. The main secondary outcome measure was pulmonary complications, defined as pneumonia, acute respiratory distress syndrome, or unexpected postoperative ventilation. Findings: This analysis includes 1128 patients who had surgery between Jan 1 and March 31, 2020, of whom 835 (74·0%) had emergency surgery and 280 (24·8%) had elective surgery. SARS-CoV-2 infection was confirmed preoperatively in 294 (26·1%) patients. 30-day mortality was 23·8% (268 of 1128). Pulmonary complications occurred in 577 (51·2%) of 1128 patients; 30-day mortality in these patients was 38·0% (219 of 577), accounting for 81·7% (219 of 268) of all deaths. In adjusted analyses, 30-day mortality was associated with male sex (odds ratio 1·75 [95% CI 1·28–2·40], p\textless0·0001), age 70 years or older versus younger than 70 years (2·30 [1·65–3·22], p\textless0·0001), American Society of Anesthesiologists grades 3–5 versus grades 1–2 (2·35 [1·57–3·53], p\textless0·0001), malignant versus benign or obstetric diagnosis (1·55 [1·01–2·39], p=0·046), emergency versus elective surgery (1·67 [1·06–2·63], p=0·026), and major versus minor surgery (1·52 [1·01–2·31], p=0·047). Interpretation: Postoperative pulmonary complications occur in half of patients with perioperative SARS-CoV-2 infection and are associated with high mortality. Thresholds for surgery during the COVID-19 pandemic should be higher than during normal practice, particularly in men aged 70 years and older. Consideration should be given for postponing non-urgent procedures and promoting non-operative treatment to delay or avoid the need for surgery. Funding: National Institute for Health Research (NIHR), Association of Coloproctology of Great Britain and Ireland, Bowel and Cancer Research, Bowel Disease Research Foundation, Association of Upper Gastrointestinal Surgeons, British Association of Surgical Oncology, British Gynaecological Cancer Society, European Society of Coloproctology, NIHR Academy, Sarcoma UK, Vascular Society for Great Britain and Ireland, and Yorkshire Cancer Research

Rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART): Study protocol for a randomized controlled trial

Background: Acute respiratory distress syndrome (ARDS) is associated with high in-hospital mortality. Alveolar recruitment followed by ventilation at optimal titrated PEEP may reduce ventilator-induced lung injury and improve oxygenation in patients with ARDS, but the effects on mortality and other clinical outcomes remain unknown. This article reports the rationale, study design, and analysis plan of the Alveolar Recruitment for ARDS Trial (ART). Methods/Design: ART is a pragmatic, multicenter, randomized (concealed), controlled trial, which aims to determine if maximum stepwise alveolar recruitment associated with PEEP titration is able to increase 28-day survival in patients with ARDS compared to conventional treatment (ARDSNet strategy). We will enroll adult patients with ARDS of less than 72 h duration. The intervention group will receive an alveolar recruitment maneuver, with stepwise increases of PEEP achieving 45 cmH(2)O and peak pressure of 60 cmH2O, followed by ventilation with optimal PEEP titrated according to the static compliance of the respiratory system. In the control group, mechanical ventilation will follow a conventional protocol (ARDSNet). In both groups, we will use controlled volume mode with low tidal volumes (4 to 6 mL/kg of predicted body weight) and targeting plateau pressure <= 30 cmH2O. The primary outcome is 28-day survival, and the secondary outcomes are: length of ICU stay; length of hospital stay; pneumothorax requiring chest tube during first 7 days; barotrauma during first 7 days; mechanical ventilation-free days from days 1 to 28; ICU, in-hospital, and 6-month survival. ART is an event-guided trial planned to last until 520 events (deaths within 28 days) are observed. These events allow detection of a hazard ratio of 0.75, with 90% power and two-tailed type I error of 5%. All analysis will follow the intention-to-treat principle. Discussion: If the ART strategy with maximum recruitment and PEEP titration improves 28-day survival, this will represent a notable advance to the care of ARDS patients. Conversely, if the ART strategy is similar or inferior to the current evidence-based strategy (ARDSNet), this should also change current practice as many institutions routinely employ recruitment maneuvers and set PEEP levels according to some titration method.Hospital do Coracao (HCor) as part of the Program 'Hospitais de Excelencia a Servico do SUS (PROADI-SUS)'Brazilian Ministry of Healt