Numerical analysis of a mechanotransduction dynamical model reveals homoclinic bifurcations of extracellular matrix mediated oscillations of the mesenchymal stem cell fate

We perform one and two-parameter numerical bifurcation analysis of a mechanotransduction model approximating the dynamics of mesenchymal stem cell differentiation into neurons, adipocytes, myocytes and osteoblasts. For our analysis, we use as bifurcation parameters the stiffness of the extracellular matrix and parameters linked with the positive feedback mechanisms that up-regulate the production of the YAP/TAZ transcriptional regulators (TRs) and the cell adhesion area. Our analysis reveals a rich nonlinear behaviour of the cell differentiation including regimes of hysteresis and multistability, stable oscillations of the effective adhesion area, the YAP/TAZ TRs and the PPAR$\gamma$ receptors associated with the adipogenic fate, as well as homoclinic bifurcations that interrupt relatively high-amplitude oscillations abruptly. The two-parameter bifurcation analysis of the Andronov-Hopf points that give birth to the oscillating patterns predicts their existence for soft extracellular substrates ($<1kPa$), a regime that favours the neurogenic and the adipogenic cell fate. Furthermore, in these regimes, the analysis reveals the presence of homoclinic bifurcations that result in the sudden loss of the stable oscillations of the cell-substrate adhesion towards weaker adhesion and high expression levels of the gene encoding Tubulin beta-3 chain, thus favouring the phase transition from the adipogenic to the neurogenic fate

Development of flow focusing device for the visualization of leukocyte rolling adhesion

La microfluídica es un área de la microtecnología basada en chips de PDMS que está siendo utilizada cada vez más en multitud de aplicaciones. Una de estas aplicaciones es la investigación biomédica. La microfluídica o “Lab on a Chip” se ha convertido en una manera de realizar experimentos biomédicos y diagnósticos de una manera barata, rápida y eficaz. Cuando se realizan estudios sobre la extravasación leucocitaria utilizando chips microfluídicos, podemos observar la inconsistencia en la trayectoria de rodadura de los leucocitos debido a un flujo laminar. En este trabajo de fin de grado presentamos un método para centrar la interfaz de células en el centro de canal microfluídico. Cuando las células circulan por los sistemas microfluídicos, las células tienden a circular de manera aleatoria por los canales. Por tanto, con el sistema propuesto en este trabajo, dichas células serán redirigidas a la porción central del canal con el fin de recrear el fenómeno de rodadura presente en nuestro sistema circulatorio y así obtener información más detallada. Los resultados de este trabajo muestran la utilidad y la versatilidad de este dispositivo para experimentos relacionados

NASA space biology accomplishments, 1983-84

Approximately 42 project summaries from NASA's Space Biology Program are presented. Emphasis is placed on gravitational effects on plant and animal life. The identification of gravity perception; the effects of weightlessness on genetic integrity, cellular differentiation, reproduction, development, growth, maturation, and senescence; and how gravity affects and controls physiology, morphology, and behavior of organisms are studied

Three dimensional oral mucosa models: development and applications

Animal experimentation has been extensively and for a long time applied in several research fields, but since 2011 it has been substantially limited by the Commission of the European Parliament to ensure people/animals safety and reduce research costs. To respond to these directives, many attempts have been focused on the development and validation of new in vitro 3D systems, bypassing the traditional 2D cell cultures. In this regard, diverse approaches to tissue-engineered bone and oral mucosa have been developed. Despite the promising premises and the cutting-edge results, the used 3D in vitro bone-oral mucosal models still lack interaction between the mucosal and the bone components. Therefore, this project aimed to create 3D models, entirely made with primary human cells (keratinocytes, fibroblasts, and osteoblasts), able to mimic the natural structure and interaction of bone and oral mucosa. In the present work, the regulatory role of the mesenchymal tissue onto epithelia was evaluated. The main results showed that that during the differentiation hMSC produce and secrete factors that induce the keratinization and the expression of the marker of differentiation CK10; in particular in the middle stage of differentiation (OB14). The proteomic analysis revealed that this effect can be ascribable to KGF secretion. This finding may impact the design of new implantable devices able to induce, alone, the epithelial growth and keratinization to improve implant graft avoiding epithelial graft linked to the morbidity of another zone. Moreover, we also showed that OM might have a pro-innervation effect, at least during the last stages of keratinocytes stratification. Finally, we obtained and characterized an innervated mucoperiosteal model that could open new in vitro frontiers for oral biomaterials validation as well as improve knowledge regarding the mesenchymal stem cells roles onto oral mucosa development

Establishment of 3d oral mucosa model using differentiated stem cells from human exfoliated deciduous teeth

Oral mucosa is a specialized type of tissue that lines the oral cavity. It consists of two main layers: stratified squamous epithelium and lamina propria. The epithelial layer is resided by the epithelial cells, while the lamina propria layer is majorly occupied by fibroblasts. As far as the in vitro oral mucosa is concerned, the construction of an oral mucosa model should be performed in full thickness architecture using both cells mentioned. Therefore, the present study aimed to differentiate stem cells from human exfoliated deciduous teeth (SHED) into fibroblastand epithelial-like cells to be subsequently used in the establishment of a 3D oral mucosa model. The differentiation of SHED was carried out by the involvement of growth factors, namely connective tissue growth factor (CTGF) for fibroblastic differentiation, whereas keratinocyte growth factor (KGF), epidermal growth factor (EGF), hepatocyte growth factor (HGF) and insulin-like growth factor-2 (IGF-II) were employed in epithelial differentiation, respectively. The characterisation of the induced cells was done by morphological observation, proliferation rate, gene and protein expression analyses using semi-quantitative reverse transcription-polymerase chain reaction (sqRT-PCR), immunofluorescence staining and flow cytometry. The collagen-glycosaminoglycan-chitosan (CGC) scaffold was constructed by combining collagen/chitosan/chondroitin sulphate/hyaluronic acid (100/12/5/1) thoroughly. The porous scaffold produced was characterized via their structural integrity, porosity, and density. The characterized differentiated cells were then co-cultured on CGC scaffold to generate a 3D oral mucosa model, which was later characterized via histological and immunofluorescence analyses. The results demonstrated the inductive effect of growth factors in both fibroblastic and epithelial differentiation of SHED. SHED derived-fibroblast-like cells are morphologically similar to SHED, while SHED derived-epithelial-like cells resembled native epithelial cells. Statistical analysis using one-way ANOVA of the proliferation assay showed a significant correlation (p<0.05) between the induced cells and growth factors involved. There were significant differences in gene and protein expressions between SHED and both differentiated cells. A white, porous lyophilized CGC scaffold produced was able to maintain its structural integrity and did not degrade throughout the whole experiments. The scaffold also exhibited good porosity and density. The co-culture system showed that the fibroblast- and epithelial-like cells derived from SHED were able to attach and proliferate when being seeded on CGC scaffold. The haematoxylin and eosin (H&E) staining of the established oral mucosa model also exhibited the infiltration and stratification of the fibroblast- and epithelial-like cells in some regions within CGC scaffolds. Also, the production of collagen could be observed via the Masson Trichrome staining. The immunofluorescence staining of the epithelial-like cells grown in the CGC scaffold also supported the presence of those cells. These findings hence provide a new understanding on the potential of SHED in the establishment of oral mucosa model for dental tissue regeneration

Análise Biomecânica de Calo Ósseo usando Método Sem Malha

O osso é um tecido fisiologicamente dinâmico e que quando lesionado tem a capacidade de se reparar com o próprio tecido, não envolvendo um tecido cicatrizante, ao contrário de outros tecidos. Esta característica torna-o particularmente interessante para investigar os processos inerentes de fraturas ósseas. A maior parte das fraturas cicatriza através de uma sequência de processos de diferenciação de tecidos, desde os processos iniciais de hematoma, aos tecidos conjuntivos, e através da cartilagem ao osso. No entanto, qualquer falha neste processo pode resultar em uniões tardias, más uniões ou não uniões. A compreensão na totalidade deste processo ainda constitui um desafio. Os mecanismos que envolvem os processos de estimulação mecânica não se encontram bem compreendidos, em consequência da complexidade dos testes experimentais in vivo, que se tornam dependentes de dados in vitro, tornando difícil validar os pressupostos biológicos. Consequentemente, os modelos computacionais têm demonstrado serem bastante úteis e eficazes na investigação sobre a cicatrização óssea. Desta forma, com o presente trabalho foi possível analisar as condições mecânicas de um calo ósseo resultante de uma fratura, assim como compreender as metodologias de análise numérica aplicadas. O modelo teve por base um estudo in vivo de forma a obter uma variação temporal progressiva da forma do calo e das propriedades mecânicas durante a cicatrização óssea. Com este modelo obtiveram-se os campos de tensão e deformação nas diferentes fases do processo de regeneração, obtendo-se resultados que se encontram em conformidade com a literatura. Adicionalmente, foi aplicado um algoritmo de remodelação óssea em combinação com o Radial Point Interpolation Method (RPIM) que foi capaz de reproduzir as condições apresentadas pela respetiva imagem histológica nesta fase. Por último, espera-se que os trabalhos desenvolvidos neste âmbito possibilitem a conceção de estratégias mais precisas e eficazes tanto para o tratamento como para aceleração da cura. De forma complementar, encontram-se em desenvolvimento modelos específicos dos pacientes e que incorporam variabilidade genética.Bone is a physiologically dynamic tissue that, when injured, has the ability to repair itself, not involving scar tissue, unlike other tissues. This characteristic makes it particularly interesting for investigating the inherent processes of bone fractures. Most fractures heal through a sequence of tissue differentiation processes, from the initial hematoma, to connective tissues and through cartilage to bone. However, any failure in this process can result in a delayed union, mal-union or non-union. A complete understanding of this process is still a challenge. The mechanisms surrounding the mechanical stimulation processes are relatively poorly understood as a result of the complexity of in vivo experimental tests, which become dependent on in vitro data, making it difficult to validate the biological assumptions. Consequently, computational models have proven to be very useful and effective in the investigation of bone healing. Therefore, in the present work, it was possible to analyse the mechanical conditions of a bone callus as a consequence of a fracture and to understand the methodologies of numerical analysis applied. The model was based on an in vivo experimental study in order to obtain a progressive temporal variation of the callus shape and mechanical properties during bone healing. With this model, the stress and strain fields in the different phases of the regeneration process were obtained, where the results are in agreement with the literature. Additionally, a bone remodelling algorithm was applied in combination with the Radial Point Interpolation Method (RPIM), which was able to reproduce the conditions presented by the respective histological image at this stage. Finally, it is expected that the work developed in this area will enable the design of more accurate and effective strategies for both treatment and accelerating healing. Complementarily, patient-specific models and the incorporation of genetic variability are being developed

The art of fin regeneration in zebrafish

The zebrafish fin provides a valuable model to study the epimorphic type of regeneration, by which the amputated part of the appendage is nearly perfectly replaced. To accomplish fin regeneration, two reciprocally interacting domains need to be established at the injury site, namely, a wound epithelium and a blastema. The wound epithelium provides a supporting niche for the blastema, which contains mesenchyme-derived progenitor cells for the regenerate. The fate of blastemal daughter cells depends on their relative position with respect to the fin margin. The apical compartment of the outgrowth maintains its undifferentiated character, whereas the proximal descendants of the blastema progressively switch from the proliferation program to the morphogenesis program. A delicate balance between self-renewal and differentiation has to be continuously adjusted during the course of regeneration. This review summarizes the current knowledge about the cellular and molecular mechanisms of blastema formation, and discusses several studies related to the regulation of growth and morphogenesis during fin regeneration. A wide range of canonical signaling pathways has been implicated during the establishment and maintenance of the blastema. Epigenetic mechanisms play a crucial role for the regulation of the cellular plasticity during the transition between differentiation states. Ion fluxes, gap-junctional communication and protein phosphatase activity have been shown to coordinate proliferation and tissue patterning in the caudal fin. The identification of the downstream targets of the fin regeneration signals and the discovery of mechanisms integrating the variety of input pathways represent exciting future aims in this fascinating field of research

Adipose Tissue in Health and Disease

Obesity, being an epidemy these days, is the trigger of metabolic disturbances such as cardiovascular disease, type 2 diabetes, and insulin resistance. Defined as an increase in fat storage, adipose tissue has been put under the spotlight as the culprit of these conditions, as it is composed not only by adipocytes but of any immune system cell and a singular extracellular matrix. Its behavior under acute and chronic hypercaloric states is quite different; persistent hypertrophy in the latter creates hypoxia, resulting in the release of reactive oxygen species and proinflammatory cytokines that impact on the immune response type of the resident leucocytes, mainly macrophages. Hypertrophy over hyperplasia, adipose tissue macrophages-M1 phenotype polarization, and the adipokines/myokines profile are thought to be regulated by foreign microRNAs, delivered from surrounding or distant cells by exosomes through the bloodstream. In this chapter, we focus on adipose tissue immunometabolism and how obesity causes the chronic inflammatory state, and, subsequently, this stablishes a pathologic adiposity, characterized by dyslipidemia and insulin resistance (IR)

Biomedical applications of three‐dimensional bioprinted craniofacial tissue engineering

Abstract Anatomical complications of the craniofacial regions often present considerable challenges to the surgical repair or replacement of the damaged tissues. Surgical repair has its own set of limitations, including scarcity of the donor tissues, immune rejection, use of immune suppressors followed by the surgery, and restriction in restoring the natural aesthetic appeal. Rapid advancement in the field of biomaterials, cell biology, and engineering has helped scientists to create cellularized skeletal muscle‐like structures. However, the existing method still has limitations in building large, highly vascular tissue with clinical application. With the advance in the three‐dimensional (3D) bioprinting technique, scientists and clinicians now can produce the functional implants of skeletal muscles and bones that are more patient‐specific with the perfect match to the architecture of their craniofacial defects. Craniofacial tissue regeneration using 3D bioprinting can manage and eliminate the restrictions of the surgical transplant from the donor site. The concept of creating the new functional tissue, exactly mimicking the anatomical and physiological function of the damaged tissue, looks highly attractive. This is crucial to reduce the donor site morbidity and retain the esthetics. 3D bioprinting can integrate all three essential components of tissue engineering, that is, rehabilitation, reconstruction, and regeneration of the lost craniofacial tissues. Such integration essentially helps to develop the patient‐specific treatment plans and damage site‐driven creation of the functional implants for the craniofacial defects. This article is the bird's eye view on the latest development and application of 3D bioprinting in the regeneration of the skeletal muscle tissues and their application in restoring the functional abilities of the damaged craniofacial tissue. We also discussed current challenges in craniofacial bone vascularization and gave our view on the future direction, including establishing the interactions between tissue‐engineered skeletal muscle and the peripheral nervous system