Biofabrication approaches and regulatory framework of metastatic tumor-on-a-chip models for precision oncology.

The complexity of the tumor microenvironment (TME) together with the development of the metastatic process are the main reasons for the failure of conventional anticancer treatment. In recent years, there is an increasing need to advance toward advanced in vitro models of cancer mimicking TME and simulating metastasis to understand the associated mechanisms that are still unknown, and to be able to develop personalized therapy. In this review, the commonly used alternatives and latest advances in biofabrication of tumor-on-chips, which allow the generation of the most sophisticated and optimized models for recapitulating the tumor process, are presented. In addition, the advances that have allowed these new models in the area of metastasis, cancer stem cells, and angiogenesis are summarized, as well as the recent integration of multiorgan-on-a-chip systems to recapitulate natural metastasis and pharmacological screening against it. We also analyze, for the first time in the literature, the normative and regulatory framework in which these models could potentially be found, as well as the requirements and processes that must be fulfilled to be commercially implemented as in vitro study model. Moreover, we are focused on the possible regulatory pathways for their clinical application in precision medicine and decision making through the generation of personalized models with patient samples. In conclusion, this review highlights the synergistic combination of three-dimensional bioprinting systems with the novel tumor/metastasis/multiorgan-on-a-chip systems to generate models for both basic research and clinical applications to have devices useful for personalized oncology.

Unifying Different Cancer Theories in a Unique Tumour Model: Chronic Inflammation and Deaminases as Meeting Points.

The increase in cancer incidences shows that there is a need to better understand tumour heterogeneity to achieve efficient treatments. Interestingly, there are several common features among almost all types of cancers, with chronic inflammation induction and deaminase dysfunctions singled out. Deaminases are a family of enzymes with nucleotide-editing capacity, which are classified into two main groups: DNA-based and RNA-based. Remarkably, a close relationship between inflammation and the dysregulation of these molecules has been widely documented, which may explain the characteristic intratumor heterogeneity, both at DNA and transcriptional levels. Indeed, heterogeneity in cancer makes it difficult to establish a unique tumour progression model. Currently, there are three main cancer models-stochastic, hierarchic, and dynamic-although there is no consensus on which one better resembles cancer biology because they are usually overly simplified. Here, to accurately explain tumour progression, we propose interactions among chronic inflammation, deaminases dysregulation, intratumor genetic heterogeneity, cancer phenotypic plasticity, and even the previously proposed appearance of cancer stem-like cell populations in the edges of advanced solid tumour masses (instead of being the cells of origin of primary malignancies). The new tumour development model proposed in this study does not contradict previously accepted models and it may open up a window to interesting therapeutic approaches.

Current Status of the Diagnosis and Management of Osteoporosis.

Osteoporosis has been defined as the silent disease of the 21st century, becoming a public health risk due to its severity, chronicity and progression and affecting mainly postmenopausal women and older adults. Osteoporosis is characterized by an imbalance between bone resorption and bone production. It is diagnosed through different methods such as bone densitometry and dual X-rays. The treatment of this pathology focuses on different aspects. On the one hand, pharmacological treatments are characterized by the use of anti-resorptive drugs, as well as emerging regenerative medicine treatments such as cell therapies and the use of bioactive hydrogels. On the other hand, non-pharmacological treatments are associated with lifestyle habits that should be incorporated, such as physical activity, diet and the cessation of harmful habits such as a high consumption of alcohol or smoking. This review seeks to provide an overview of the theoretical basis in relation to bone biology, the existing methods for diagnosis and the treatments of osteoporosis, including the development of new strategies.

Osteoarthritis: Trauma vs Disease.

Osteoarthritis (OA) is the most prevalent joint disease characterized by pain and degenerative lesions of the cartilage, subchondral bone, and other joint tissues. The causes of OA remain incompletely understood. Over the years, it has become recognized that OA is a multifactorial disease. In particular, aging and trauma are the main risk factors identified for the development of OA; however, other factors such as genetic predisposition, obesity, inflammation, gender and hormones, or metabolic syndrome contribute to OA development and lead to a more severe outcome. While this disease mainly affects people older than 60 years, OA developed after joint trauma affects all range ages and has a particular impact on young individuals and people who have highest levels of physical activity such as athletes. Traumatic injury to the joint often results in joint instability or intra-articular fractures which lead to posttraumatic osteoarthritis (PTOA). In response to injury, several molecular mechanisms are activated, increasing the production and activation of different factors that contribute to the progression of OA.In this chapter, we have focused on the interactions and contribution of the multiple factors involved in joint destruction and progression of OA. In addition, we overview the main changes and molecular mechanisms related to OA pathogenesis.

Models of Disease.

Osteochondral (OC) lesions are a major cause of chronic musculoskeletal pain and functional disability, which reduces the quality of life of the patients and entails high costs to the society. Currently, there are no effective treatments, so in vitro and in vivo disease models are critically important to obtain knowledge about the causes and to develop effective treatments for OC injuries. In vitro models are essential to clarify the causes of the disease and the subsequent design of the first barrier to test potential therapeutics. On the other hand, in vivo models are anatomically more similar to humans allowing to reproduce the pattern and progression of the lesion in a controlled scene and offering the opportunity to study the symptoms and responses to new treatments. Moreover, in vivo models are the most suitable preclinical model, being a fundamental and a mandatory step to ensure the successful transfer to clinical trials. Both in vitro and in vitro models have a number of advantages and limitation, and the choice of the most appropriate model for each study depends on many factors, such as the purpose of the study, handling or the ease to obtain, and cost, among others. In this chapter, we present the main in vitro and in vivo OC disease models that have been used over the years in the study of origin, progress, and treatment approaches of OC defects.

Cardiomyogenic differentiation potential of human endothelial progenitor cells isolated from patients with myocardial infarction.

BACKGROUND AIMS: Endothelial progenitor cells (EPCs) are known to play a beneficial role by promoting postnatal vasculogenesis in pathological events, such as ischemic heart disease and peripheral artery disease. However, little is known about the potential of EPCs to restore heart damage tissue. We compared the cardiac differentiation capacity of EPCs isolated from peripheral blood of patients with acute myocardial infarction (AMI) with EPCs obtained from umbilical cord blood (UCB). METHODS: EPCs from both origins were isolated by density gradient centrifugation and characterized through the use of endothelial markers (UEA-1lectin, CD133 and KDR) and endothelial cell colony-forming unit assay. Cardiac differentiation capacity of EPCs was assessed by immunofluorescence and reverse transcriptase-polymerase chain reaction after 5-azacytidine (5-aza) induction. RESULTS: No significant differences were observed between the number of endothelial cell colony-forming units in peripheral blood of patients with AMI and samples from UCB. Moreover, 5-aza induced the appearance of myotube-like structures and the positive expression of sarcomeric α-actinin, cardiac troponin I and T and desmin in a similar pattern for both cell sources, which indicates a comparable acquisition of a cardiac-like phenotype. CONCLUSIONS: For the first time, we have compared, in vitro, the cardiomyogenic potential of EPCs derived from patients with AMI with UCB-derived EPCs. Our data indicate that EPCs obtained from both origins have similar plasticity and functions and suggest a potential therapeutic efficacy in cardiac cell therapy.

Design and evaluation of a bilayered dermal/hypodermal 3D model using a biomimetic hydrogel formulation.

Due to the limitations of the current skin wound treatments, it is highly valuable to have a wound healing formulation that mimics the extracellular matrix (ECM) and mechanical properties of natural skin tissue. Here, a novel biomimetic hydrogel formulation has been developed based on a mixture of Agarose-Collagen Type I (AC) combined with skin ECM-related components: Dermatan sulfate (DS), Hyaluronic acid (HA), and Elastin (EL) for its application in skin tissue engineering (TE). Different formulations were designed by combining AC hydrogels with DS, HA, and EL. Cell viability, hemocompatibility, physicochemical, mechanical, and wound healing properties were investigated. Finally, a bilayered hydrogel loaded with fibroblasts and mesenchymal stromal cells was developed using the Ag-Col I-DS-HA-EL (ACDHE) formulation. The ACDHE hydrogel displayed the best in vitro results and acceptable physicochemical properties. Also, it behaved mechanically close to human native skin and exhibited good cytocompatibility. Environmental scanning electron microscopy (ESEM) analysis revealed a porous microstructure that allows the maintenance of cell growth and ECM-like structure production. These findings demonstrate the potential of the ACDHE hydrogel formulation for applications such as an injectable hydrogel or a bioink to create cell-laden structures for skin TE.

Cellular extracts from post-mortem human cardiac tissue direct cardiomyogenic differentiation of human adipose tissue-derived stem cells.

BACKGROUND AIMS: Human adipose tissue-derived stem cells (hASCs) can be easily (and inexpensively) expanded in culture, and their high plasticity allows their conversion to different cell types. We study the potential capacity of postmortem cardiac tissue to direct cardiac differentiation of hASCs in vitro. METHODS: Cardiac tissue collected from autopsies was used to obtain cell extracts and conditioned medium, and both approaches were tested for cardiac induction. RESULTS: Gene expression analyses proved that post-mortem human cardiac tissue maintains genetic integrity. hASCs exposed to the cell extracts or conditioned medium for 2 weeks achieved the appearance of myotube-like structures and were positive for cardiac markers such as sarcomeric α-actinin, cardiac troponin I and T and desmin as proved by immunofluorescence. In addition, differentiated cells showed increased expression of cardiomyocyte-related genes analyzed by reverse transcriptase polymerase chain reaction (GATA-4, myocyte-enhancer factor-2c, α-cardiac actin and cardiac troponin I). CONCLUSIONS: For the first time, post-mortem human cardiac tissue was used to induce hASC differentiation into myocardial-like cells. The methodology described here would serve as a useful model to obtain cardiomyocyte-like cells in vitro.

Apatite-coated outer layer eggshell membrane: A novel osteoinductive biohybrid composite for guided bone/tissue regeneration.

Hybrid biomimetic materials aim to replicate the organic-inorganic constructs of mineralized tissues. During eggshell formation, the outer surface of the eggshell membrane (ESM) promotes calcium carbonate nucleation, while the inner one prevents mineralization toward the egg white and yolk. In the current study, the outer surface of the ESM acted as a heteronucleant in calcium phosphate precipitation by the vapor diffusion sitting drop method, while the inner one remained unmineralized. The aim was to fabricate a 2D biomaterial with dual functions, osteoinductive on one side and protective against cell invasion on the other side. The microstructural, physicochemical, morphological, and mechanical properties of the mineralized ESM were characterized by XRD, TGA, XPS, FTIR/Raman, HR-SEM, and mechanical testing techniques. The cytocompatibility and osteoinductive ability were assessed by biological assays of cell viability, proliferation, and osteogenic differentiation on human mesenchymal stromal cells (hMSCs). Results indicate that the outer surface of the ESM induces the heterogeneous precipitation of carbonate-apatite phase depicting biomimetic features. In addition, the apatite/ESM shows a much higher cytocompatibility than the pristine ESM and promotes the osteogenic differentiation of hMSCs more efficiently. Overall, the apatite/ESM composite exhibits compositional, crystalline, mechanical, and biological properties that resemble those of mineralized tissues, rendering it an approachable and novel material especially useful in guided tissue/bone regeneration.

Biofabrication of a tri-layered 3D-bioprinted CSC-based malignant melanoma model for personalized cancer treatment.

Conventionalin vitrocancer models do not accurately reproduce the tumor microenvironment (TME), so three-dimensional (3D)-bioprinting represents an excellent tool to overcome their limitations. Here, two multicellular tri-layered malignant melanoma (MM) models composed by cancer stem cells (CSCs) isolated from a MM established cell line or a primary-patient derived cell line, fibroblasts, mesenchymal stem cells, and endothelial cells, embedded within an agarose-collagen type I hydrogel were bioprinted. Embedded-cells showed high proliferation and metabolic activity, and actively remodeled their TME. MM hydrogels displayed similar rheological properties that skin and were able to support an early onset of vascularization. Besides, MM hydrogels displayed different response to vemurafenib compared with cell cultures, and supported tumorigenesis in murine xenotransplant achieving more mimeticin vivomodels. For the first time a tri-layered 3D-bioprinted CSC-based human MM model is developed recreating TMEin vitroandin vivoand response to treatment, being useful for precision treatment regimens against MM.