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The aim of this study was to evaluate the clinical outcome at 5-year follow-up of a one-step procedure combining anterior cruciate ligament (ACL) reconstruction and partial meniscus replacement using a polyurethane scaffold for the treatment of symptomatic patients with previously failed ACL reconstruction and partial medial meniscectomy. Moreover, the implanted scaffolds have been evaluated by MRI protocol in terms of morphology, volume, and signal intensity.Twenty patients with symptomatic knee laxity after failed ACL reconstruction and partial medial meniscectomy underwent ACL revision combined with polyurethane-based meniscal scaffold implant. Clinical assessment at 2- and 5-year follow-ups included VAS, Tegner Activity Score, International Knee Documentation Committee (IKDC), Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC), and the Lysholm Score. MRI evaluation of the scaffold was performed according to the Genovese scale with quantification of the scaffold's volume at 1- and 5-year follow-ups.All scores revealed clinical improvement as compared with the preoperative values at the 2- and 5-year follow-ups. However, a slight, but significant reduction of scores was observed between 2 and 5 years. Concerning the MRI assessment, a significant reduction of the scaffold's volume was observed between 1 and 5 years. Genovese Morphology classification at 5 years included two complete resorptions (Type 3) and all the remaining patients had irregular morphology (Type 2). With regard to the Genovese Signal at the 5-year follow-up, three were classified as markedly hyperintense (Type 1), 15 as slightly hyperintense (Type 2), and two as isointense (Type 1).Simultaneous ACL reconstruction and partial meniscus replacement using a polyurethane scaffold provides favourable clinical outcomes in the treatment of symptomatic patients with previously failed ACL reconstruction and partial medial meniscectomy at 5 years. However, MRI evaluation suggests that integration of the scaffold is not consistent.Level IV.

Stimulating brain tissue regeneration is a major challenge after central nervous system (CNS) injury, such as those observed from trauma or cerebrovascular accidents. Full regeneration is difficult even when a neurogenesis-associated repair response may occur. Currently, there are no effective treatments to stimulate brain tissue regeneration. However, biomaterial scaffolds are showing promising results, where hydrogels are the materials of choice to develop these supportive scaffolds for cell carriers. Their combination with growth factors, such as brain-derived neurotrophic factor (BDNF), basic fibroblast growth factor (bFGF), or vascular endothelial growth factor (VEGF), together with other cell therapy strategies allows the prevention of further neuronal death and can potentially lead to the direct stimulation of neurogenesis and vascularisation at the injured site. Imaging of the injured site is particularly critical to study the reestablishment of neural cell functionality after brain tissue injury. This review outlines the latest key advances associated with different strategies aiming to promote the neuroregeneration, imaging, and functional recovery of brain tissue. Graphical abstract

Intervertebral disc (IVD) herniation often causes severe pain and is frequently associated with the degeneration of the IVD. As the IVD degenerates, more fissures with increasing size appear within the outer region of the IVD, the annulus fibrosus (AF), favoring the initiation and progression of IVD herniation. For this reason, we propose an AF repair approach based on methacrylated gellan gum (GG-MA) and silk fibroin. Therefore, coccygeal bovine IVDs were injured using a biopsy puncher (â 2 mm) and then repaired with 2% GG-MA as a filler material and sealed with an embroidered silk yarn fabric. Then, the IVDs were cultured for 14 days either without any load, static loading, or complex dynamic loading. After 14 days of culture, no significant differences were found between the damaged and repaired IVDs, except for a significant decrease in the IVDsâ relative height under dynamic loading. Based on our findings combined with the current literature that focuses on ex vivo AF repair approaches, we conclude that it is likely that the repair approach did not fail but rather insufficient harm was done to the IVD., DFG -Deutsche Forschungsgemeinschaft(437213841)

Breast cancer is still the leading cause of women's death due to relapse and metastasis. In vitro tumor models are considered reliable tools for drug screening and understanding cancer-driving mechanisms due to the possibility of mimicking tumor heterogeneity. Herein, a 3D breast cancer model (3D-BCM) is developed based on enzymatically-crosslinked silk fibroin (eSF) hydrogels. Human MCF7 breast cancer cells are encapsulated into eSF hydrogels, with and without human mammary fibroblasts. The spontaneously occurring conformational change from random coil to β-sheet is correlated with increased eSF hydrogels’ stiffness over time. Moreover, mechanical properties analysis confirms that the cells can modify the stiffness of the hydrogels, mimicking the microenvironment stiffening occurring in vivo. Fibroblasts support cancer cells growth and assembly in the eSF hydrogels up to 14 days of culture. Co-cultured 3D-BCM exhibits an upregulated expression of genes related to extracellular matrix remodeling and fibroblast activation. The 3D-BCM is subjected to doxorubicin and paclitaxel treatments, showing differential drug response. Overall, these results suggest that the co-culture of breast cancer cells and fibroblasts in eSF hydrogels allow the development of a mimetic in vitro platform to study cancer progression. This opens up new research avenues to investigate novel molecular targets for anti-cancer therapy.

Hydrogels are three-dimensional platforms that serve as substitutes for native extracellular matrix. These materials are starting to play important roles in regenerative medicine because of their similarities to native matrix in water content and flexibility. It would be very advantagoues for researchers to be able to regulate cell behavior and fate with specific hydrogels that have tunable mechanical properties as biophysical cues. Recent developments in dynamic chemistry have yielded designs of adaptable hydrogels that mimic dynamic nature of extracellular matrix. The current review provides a comprehensive overview for adaptable hydrogel in regenerative medicine as follows. First, we outline strategies to design adaptable hydrogel network with reversible linkages according to previous findings in supramolecular chemistry and dynamic covalent chemistry. Next, we describe the mechanism of dynamic mechanical microenvironment influence cell behaviors and fate, including how stress relaxation influences on cell behavior and how mechanosignals regulate matrix remodeling. Finally, we highlight techniques such as bioprinting which utilize adaptable hydrogel in regenerative medicine. We conclude by discussing the limitations and challenges for adaptable hydrogel, and we present perspectives for future studies., Graphical abstract Image 1, Highlights • Introduction of adaptable hydrogels with dynamic mechanical properties as 3D extracellular matrix. • Summary of reversible linkages based on supramolecular interactions and dynamic covalent bonds. • Discussion of how adaptable hydrogels provide dynamic mechanical microenvironment and influence cell behaviors and fate. • Overview of applications of adaptable hydrogel in regenerative medicine.

Basic pre-clinical research based on 2D cultures have been very valuable in colorectal cancer (CRC) research but still have failed to improve patient prognostic outcomes. This is because they simply do not replicate what happens in vivo, i.e. 2D cultured cells system cannot replicate the diffusion constraints usually found in the body. Importantly, they also do not mimic the dimensionality of the human body and of a CRC tumour (3D). Moreover, 2D cultures lack the cellular heterogeneity and the tumour microenvironment (TME) such as stromal components, blood vessels, fibroblasts, and cells of the immune system. Cells behave differently whether in 2D and 3D, in particular their different genetic and protein expression panels are very different and therefore we cannot fully rely on drug tests done in 2D. A growing field of research based on microphysiological systems involving organoids/spheroids or patient-derived tumour cells has become a solid base for a better understanding of the TME and as a result is a step towards personalized medicine. Furthermore, microfluidic approaches have also started to open possibilities of research, with tumour-on-chips and body-on-chips being used in order to decipher complex inter-organ signalling and the prevalence of metastasis, as well as CRC early-diagnosis through liquid biopsies. Herein, we focus on the state-of-the-art of CRC research with emphasis on 3D microfluidic in vitro cultures—organoids, spheroids—drug resistance, circulating tumour cells and microbiome-on-a-chip technology.

Advanced strategies to bioengineer a fibrocartilaginous tissue to restore the function of the meniscus are necessary. Currently, 3D bioprinting technologies have been employed to fabricate clinically relevant patient-specific complex constructs to address unmet clinical needs. In this study, a highly elastic hybrid construct for fibrocartilaginous regeneration is produced by coprinting a cell-laden gellan gum/fibrinogen (GG/FB) composite bioink together with a silk fibroin methacrylate (Sil-MA) bioink in an interleaved crosshatch pattern. We characterize each bioink formulation by measuring the rheological properties, swelling ratio, and compressive mechanical behavior. For in vitro biological evaluations, porcine primary meniscus cells (pMCs) are isolated and suspended in the GG/FB bioink for the printing process. The results show that the GG/FB bioink provides a proper cellular microenvironment for maintaining the cell viability and proliferation capacity, as well as the maturation of the pMCs in the bioprinted constructs, while the Sil-MA bioink offers excellent biomechanical behavior and structural integrity. More importantly, this bioprinted hybrid system shows the fibrocartilaginous tissue formation without a dimensional change in a mouse subcutaneous implantation model during the 10-week postimplantation. Especially, the alignment of collagen fibers is achieved in the bioprinted hybrid constructs. The results demonstrate this bioprinted mechanically reinforced hybrid construct offers a versatile and promising alternative for the production of advanced fibrocartilaginous tissue., United States National Institutes of Health (1P41EB023833-346 01) and the Portuguese Foundation for Science and Technology (PTDC/BBB-ECT/2690/2014 and PTDC/EMD-EMD/ 31367/2017). FCT/MCTES is acknowledged for the Ph.D. scholarship attributed to J.B.C. (PD/BD/113803/2015) and the financial support provided to J.S.-C. (IF/00115/2015) and J.M.O. (IF/01285/2015) under the program “Investigador FCT”

This study proposes a green and innovative ionic liquid (IL) methodology for processing sucrose acetate isobutyrate (SAIB) porous structures for tissue engineering. The solubilization of SAIB in an...

The increasing rate of musculoskeletal pathologies has compelled the development of improved and novel treatment strategies in order to address unmet clinical needs. Tissue engineering approaches comprising the use of scaffolds for bone regeneration have been showing to be a promising alternative to conventional bone repair/substitution approaches. In particular, hierarchical scaffolds as methods of structural support and osteogenic differentiation promoters are among the most used tools in bone tissue engineering (BTE). In this reasoning, hierarchical scaffolds have sparked the field, striving toward mimicking the natural bone tissue in both, its complex 3D structure and composition. A recent and promising trend has been the merging of nanotechnology and tissue engineering concepts. As such the incorporation of nanoparticles and nanocomposites into micro- or macroscaffold systems can result in an improvement of scaffoldsâ biofunctionality at different levels. These tools are versatile in nature and can be used for multiple purposes such as drug delivery, thermal conductors, and mechanical reinforcement. Taking into consideration multidisciplinary approaches, several strategies have been pursued. The recent reports dealing with the approaches pursued in the hierarchical scaffolds production and enhancement, ranging from the nanoscale to the macroscale, are overviewed herein., The authors thank the funds provided by the project FROnTHERA (NORTE-01-0145-FEDER-000023), supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF), the project 3BioMeD (FCT/4773/ 4/5/2017/S) supported by Fundação para a Ciência e a Tecnologia (FCT) and the R&D Project KOAT PTDC/BTMMAT/29760/2017 (POCI 01-0145-FEDER-029760) financed by FCT and cofinanced by FEDER and POCI. F.R.M. acknowledges FCT for her contract under the Transitional Rule DL 57/2016 (CTTI-57/18-I3BS (5)).

The simultaneous generation of multiple tissues and their functional assembly into complex tissues remains a critical challenge for regenerative medicine. The tissue-to-tissue interface connecting two adjacent tissues is vital in effective tissue function. The presented worked hypothesize that differential functional property can be engineered by modulating the macromolecular composition of a 3D hydrogel construct and distinctively endow stem cell fate. Hence, it was possible to successfully generate macromolecular constructs by using the extracellular matrix (ECM)-based materials; type I collagen (Col I) and hyaluronic acid (HA); and natural-derived biomaterials as methacrylated gellan-gum (GGMA). The 3D hydrogel constructs consisted of two dissimilar layers: 1) Col I: HA hydrogel and 2) GGMA hydrogel. The tissue-to-tissue interface was created by seeding human mesenchymal stem cells (MSCs) between the two layers. Differential functional rheological and mechanical properties characterized the acellular 3D gradient hydrogel constructs. The cell-based 3D hydrogel constructs were assessed for MSCs viability by live/dead staining. Assessing apoptosis by flow cytometry, data showed the feasibility of the 3D hydrogel constructs in maintaining cell viability with no apoptosis induction onto MSCs. A homogeneous distribution was achieved in a successful cellular tissue-to-tissue interface. Human MSCs low proliferative rate and low ECM deposition were seen for all constructs; however, lower proliferative rate within the ECM microenvironment highlights controlled self-renewal of MSCs. The 3D hydrogel constructs maintained the human MSCs phenotype, yet the macromolecular modulation allowed tuning the human MSCs morphology from round to spindle-shaped phenotype. The intrinsic properties of the 3D cell-based hydrogel construct induced differential inflammatory and angiogenic paracrine secretory profiles owing to the dissimilar engineered biophysical milieu. Human MSCs sense the nearby macromolecular environment adjusting the cell-ECM interactions, which influence cell behaviour and fate. Beyond multi-tissue regeneration, the engineered cellular 3D hydrogel constructs may simultaneously address immune regeneration., This material is based on works supported by the European Union funding under the 7th Framework Programme under Grant Agreement Number 317304 and supported in part by a research grant from Science Foundation Ireland (SFI) and co-funded under the European Regional De-velopment Fund under Grant Number 13/RC/2073_P2. D.R. Pereira ac-knowledges the Foundation for Science and Technology (FCT) , Portugal, for an individual grant (SFRH/BD/81356/2011) . J. Silva-Correia and J.M. Oliveira also acknowledges the FCT for the funds provided under the prestigious program Investigator FCT 2015 (IF/00115/2015 and IF/01285/2015, respectively).The authors acknowledge the scientific and technical assistance of the Electron Microscopy unit within the Centre for Microscopy & Imaging, the flow cytometry core facility and histology core facility at the National University of Ireland (NUI)Galway. All facilities are funded by NUI Galway and the Irish Government's Programme for Research in Third Level Institutions (PRTLI), Cycles 4 and 5, National Development Plan 2007-2013. The authors would like to also acknowledge the Centre for Microscopy & Imaging funded by the National University of Ireland Galway and MSSI in the University of Limerick for the rheometer facilities, both funded by the Programme for Research in Third-Level Institutions (PRTLI), Cycles 4 and 5, National Development Plan 2007-2013.

Supplementary Materials: The following are available online at https://www.mdpi.com/article/10 .3390/pharmaceutics13111925/s1. Figure S1 (A) Intraperitoneal injection of STZ. (B) Mice are kept in cages with ad libitum fed regimen. Figure S2 MIN-6 cell transplantation. A) Abdominal viscera of female mice. (B) Intramuscular administration of anaesthetics. (C) Topical application of Vaseline to protect eyes from drying during surgery, lateral hair removal for incision. (D) Graft injection inside the kidney capsule. (E–F) Suture and stiches. (G) Post-operation recovery., Female mice (Black 6 strain) (C57BL/6) aged 6 weeks were subject to low dose streptozotocin (STZ) treatment for five consecutive days to mimic type 1 diabetes mellitus (T1DM) with insulitis. At two weeks after STZ injections, evaluation of the elevated glucose levels was used to confirm diabetes. The diabetic mice were then subject to the transplantation of pancreatic β-cells (MIN-6 line). Four groups of mice were studied. The first group was injected with saline-only acting as the placebo surgery control, also known as SHAM group, the second and third groups were injected with MIN-6 single cells and polyethylene glycol-modified dipalmitoyl-glycerol-phosphatidyl ethanolamine (PEG-DPPE) modified MIN-6 single cells (500 µg per 1.106 cells), respectively, while the fourth group was injected with hyaluronic acid (HA)-coated MIN-6 single cells (5 bilayers). At seven- and fourteen-days following transplantation, the mice were euthanised. The renal and pancreatic tissues were then collected and histologically analysed. The induction of diabetes in female mice, through five-consecutive daily STZ injections resulted in inconsistent glycaemic levels. Interestingly, this shows an incomplete diabetes induction in female mice, of which we attribute to sex dimorphism and hormonal interferences. Transplantation failure of free-floating encapsulated cells was unable to decrease blood glucose hyperglycaemia to physiological ranges. The result is attributed to deprived cell–cell interactions, leading to decreased β-cells functionality. Overall, we highlight the necessity of refining T1DM disease models in female subjects when using multiple low-dose STZ injections together with transplantation protocols. Considerations need to be made regarding the different developmental stages of female mice and oestrogen load interfering with pancreatic β-cells susceptibility to STZ. The use of pseudo islets, cell aggregates and spheroids are sought to improve transplantation outcome in comparison to free-floating single cells., Irish Research Council Postgraduate Scholarship (GOIPG/2015/3577) and ERASMUS+ grant student mobility placement (University of Limerick/Erasmus+ 1920) for F.Z. The authors also thank the financial support under the FRONTHERA (NORTE-01-0145-FEDER-000023) project (J.M.O. and I.F.C.), and the project 2IQBIONEURO (ref. 0624_2IQBIONEURO_6_E) (J.M.O. and I.F.C.), and TERM RES-Hub, Tissue Engineering and Regenerative Medicine Infrastructure project, funded by the Portuguese Foundation for Science and Technology (FCT) (I.F.C.).

Supplementary Information: The online version contains supplementary material available at https://doi.org/10.1038/s41598-021-99390-x., The collective dynamics of cells on surfaces and interfaces poses technological and theoretical challenges in the study of morphogenesis, tissue engineering, and cancer. Different mechanisms are at play, including, cellâ cell adhesion, cell motility, and proliferation. However, the relative importance of each one is elusive. Here, experiments with a culture of glioblastoma multiforme cells on a substrate are combined with in silico modeling to infer the rate of each mechanism. By parametrizing these rates, the time-dependence of the spatial correlation observed experimentally is reproduced. The obtained results suggest a reduction in cellâ cell adhesion with the density of cells. The reason for such reduction and possible implications for the collective dynamics of cancer cells are discussed., The authors acknowledge fnancial support from the Portuguese Foundation for Science and Technology (FCT) under Contracts no. PTDC/FIS-MAC/28146/2017 (LISBOA-01-0145-FEDER-028146), UIDB/00618/2020, and UIDP/00618/2020. F.R.M. also acknowledges FCT for her contract under the Transitional Rule DL 57/2016 (CTTI-57/18-I3BS(5)).

Digestive system cancers account for nearly half of all cancers around the world and have a high mortality rate. Cell culture and animal models represent cornerstones of digestive cancer research. However, their ability to en- able cancer precision medicine is limited. Cell culture models cannot retain the genetic and phenotypic heteroge- neity of tumors and lack tumor microenvironment (TME). Patient-derived xenograft mouse models are not suitable for immune-oncology research. While humanized mouse models are time- and cost-consuming. Suitable preclinical models, which can facilitate the understanding of mechanisms of tumor progression and develop new therapeutic strategies, are in high demand. This review article summarizes the recent progress on the establish- ment of TME by using tumor organoid models and microfluidic systems. The main challenges regarding the translation of organoid models from bench to bedside are discussed. The integration of organoids and a microflu- idic platform is the emerging trend in drug screening and precision medicine. A future prospective on this field is also provided., This study was supported by the National Natural Science Foundation of China (Grant No.82073148), the Guangdong Provincial Key Laboratory of Digestive Cancer Research (No. 2021B1212040006), the Sanming Project of Medicine in Shenzhen (SZSM201911010), the Shenzhen Key Medical Discipline Construction Fund (SZXK016), the Shenzhen Sustainable Project (KCXFZ202002011010593), and the Shenzhen-Hong Kong-Macau Technology Research Programme (Type C) (Grant No. SGDX2020110309260100).

Oxaliplatin is the first-line regime for advanced gastric cancer treatment, while its resistance is a major problem that leads to the failure of clinical treatments. Tumor cell heterogeneity has been considered as one of the main causes for drug resistance in cancer. In this study, the mechanism of oxaliplatin resistance was investigated through in vitro human gastric cancer organoids and gastric cancer oxaliplatin-resistant cell lines and in vivo subcutaneous tumorigenicity experiments. The in vitro and in vivo results indicated that CD133+â stem cell-like cells are the main subpopulation and PARP1 is the central gene mediating oxaliplatin resistance in gastric cancer. It was found that PARP1 can effectively repair DNA damage caused by oxaliplatin by means of mediating the opening of base excision repair pathway, leading to the occurrence of drug resistance. The CD133+â stem cells also exhibited upregulated expression of N6-methyladenosine (m6A) mRNA and its writer METTL3 as showed by immunoprecipitation followed by sequencing and transcriptome analysis. METTTL3 enhances the stability of PARP1 by recruiting YTHDF1 to target the 3â ²-untranslated Region (3â ²-UTR) of PARP1 mRNA. The CD133+â tumor stem cells can regulate the stability and expression of m6A to PARP1 through METTL3, and thus exerting the PARP1-mediated DNA damage repair ability. Therefore, our study demonstrated that m6A Methyltransferase METTL3 facilitates oxaliplatin resistance in CD133+â gastric cancer stem cells by Promoting PARP1 mRNA stability which increases base excision repair pathway activity., NSFC -National Natural Science Foundation of China(2021B1212040006)

Cell therapy is a promising tool for treating central nervous system (CNS) disorders; though, the translational efforts are plagued by ineffective delivery methods. Due to the large contact surface with CNS and relatively easy access, the intrathecal route of administration is attractive in extensive or global diseases such as stroke or amyotrophic lateral sclerosis (ALS). However, the precision and efficacy of this approach are still a challenge. Hydrogels were introduced to minimize cell sedimentation and improve cell viability. At the same time, contrast agents were integrated to allow image-guided injection. Here, we report using manganese ions (Mn2+) as a dual agent for cross-linking alginate-based hydrogels and magnetic resonance imaging (MRI). We performed in vitro studies to test the Mn2+ alginate hydrogel formulations for biocompatibility, injectability, MRI signal retention time, and effect on cell viability. The selected formulation was injected intrathecally into pigs under MRI control. The biocompatibility test showed a lack of immune response, and cells suspended in the hydrogel showed greater viability than monolayer culture. Moreover, Mn2+-labeled hydrogel produced a strong T1 MRI signal, which enabled MRI-guided procedure. We confirmed the utility of Mn2+ alginate hydrogel as a carrier for cells in large animals and a contrast agent at the same time., This research was funded by The National Centre for Research and Development, grant number 12/EuroNanoMed/2016.

Organoid culture is a recently developed in-vitro three-dimensional (3D) cell culture technology. It has wide applications in tissue engineering studies. However, histological analysis of organoid is quite complex and tedious for researchers. This study proposes a user-friendly, affordable and efficient method for making formalin-fixed paraffin embedded (FFPE) organoid blocks and Optimal Cutting Temperature compound (OCT) embedded frozen organoid blocks. This method implements a key pre-embedding step for preparing paraffin embedded organoid blocks, which could concentrate organoid togethor without damaging or loss of samples. This method could be used to process even a small number of organoids with high efficiency. In addition, with minor modifications, the method is readily applied for OCT embedded organoid blocks. The slides generated were ready for H&E staining, immunohistochemistry staining and immunofluorescent staining. The method described in this study can be easily used for routine histological analysis of organoid, and could be performed in general pathology labs and requires no dedicated equipment and reagent. This article is protected by copyright. All rights reserved.

The application of nanoparticles in magnetic resonance imaging (MRI) has been greatly increasing, due to their advantageous properties such as nanoscale dimension and tuneability. In this context, manganese (Mn2+)-based nanoparticles have been greatly investigated, due to their valuable use as a contrast agent, improving signal intensity and specificity in MRI (manganese-enhanced MRI, MEMRI). Additionally, Mn2+ can act as scavengers of reactive oxygen species (ROS), commonly present in the inflammatory processes of neurodegenerative diseases. The aim of the present study was to develop nanoreactors, which can be used as contrast-agent in MEMRI. Several blends of methacrylated gellan gum (GG-MA) and hyaluronic acid (HA) were embedded with different types of manganese dioxide (MnO2) nanoparticles and further physico-chemically characterized. Dynamic light scattering, scanning electron microscopy, water uptake and degradation studies were performed. In vitro cytotoxicity of the different formulations was also evaluated using an immortalized rat fibroblast cell line L929, up to 72 h of culturing. Synthesized nanoparticles were obtained with an average size of 70 nm and round-shaped morphology. The stability of the different formulations of hydrogels was not affected by nanoparticles' concentration or HA ratio. The presence of synthesized MnO2 (MnO2_S) nanoparticles reduced hydrogels' cytocompatibility, whereas the commercially available type 1 (MnO2_C1) nanoparticles were less toxic to cells. Additionally, cell proliferation and viability were enhanced when a lower content of HA was present. Higher concentrations (75 and 100 ng/mL) of MnO2_S and MnO2_C1 nanoparticles did not negatively affected cell viability, whereas the opposite effect was observed for the commercial type 2 (MnO2_C2) nanoparticles. Further studies are required to evaluate the potential application of the most promising nanoreactors' formulations for combined application in MEMRI and as ROS scavengers., The authors would like to acknowledge the financial support pro vided through the EC Funded project NanoTech4ALS (ENMed/0008/ 2015) and the funding provided under the projects FROnTHERA (No. NORTE-01-0145-FEDER-000023) and JUSThera (No. NORTE-01-0145- FEDER-000055), supported by Norte Portugal Regional Operational Programme (No. NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). The authors also thank the funding through the project 2IQBIONEURO (ref. 0624_2IQBIONEURO_6_E). S.V. Lopes thanks to the Portuguese Foundation for Science and Technology (FCT) for the PhD grant (SFRH/ BD/143954/2019). The FCT distinction to J. Silva-Correia (IF/00115/ 2015) under the Investigator FCT program is also greatly acknowledged.

uncorrected proof version, The generation of invasive fluctuating protrusions is a distinctive feature of tumor dissemination. During the invasion, individual cancer cells modulate the morphodynamics of protrusions to optimize their migration efficiency. However, it remains unclear how protrusion fluctuations govern the invasion of more complex multi-cellular structures, such as tumors, and their correlation with the tumor metastatic potential. Herein, a reductionist approach based on 3D tumor cell micro-spheroids with different invasion capabilities is used as a model to decipher the role of tumor-associated fluctuating protrusions in cancer progression. To quantify fluctuations, a set of key biophysical parameters that precisely correlate with the invasive potential of tumors is defined. It is shown that different pharmacological drugs and cytokines are capable of modulating protrusion activity, significantly altering protrusion fluctuations, and tumor invasiveness. This correlation is used to define a novel quantitative invasion index encoding the key biophysical parameters of fluctuations and the relative levels of cell-cell/matrix interactions, which is capable of assessing the tumor's metastatic capability solely based on its magnitude. Overall, this study provides new insights into how protrusion fluctuations regulate tumor cell invasion, suggesting that they may be employed as a novel early indicator, or biophysical signature, of the metastatic potential of tumors., Portuguese Foundation for Science and Technology (FCT) under the program CEEC Individual 2017 (CEECIND/00352/2017). D.C., V.B., A.C.L., C.M.A., and S.C.K. also thank the support from the FCT under the scope of the projects 2MATCH (PTDC/BTM-ORG/28070/2017) and BREAST-IT (PTDC/ BTM-ORG/28168/2017) funded by the Programa Operacional Regional do Norte supported by European Regional Development Funds (ERDF). The authors also thank the financial support from the European Union Framework Program for Research and Innovation Horizon 2020 on FoReCaST project under (Grant Number: 668983), and the Institution of Engineering and Technology (IET) for the funding provided under the ENG ThE CANCER project (Grant Number: IET Harvey Engineering Research Award 2018). J.M.O. is grateful for the FCT distinctions (IF/00423/2012 and IF/01285/2015)

The recent progress in biomaterials science and development of tubular conduits (TCs) still fails in solving the current challenges in the treatment of peripheral nerve injuries (PNIs), in particular when disease-related and long-gap defects need to be addressed. Nanotechnology-based therapies that seemed unreachable in the past are now being considered for the repair and reconstruction of PNIs, having the power to deliver bioactive molecules in a controlled manner, to tune cellular behavior, and ultimately guide tissue regeneration in an effective manner. It also offers opportunities in the imaging field, with a degree of precision never achieved before, which is useful for diagnosis, surgery and in the patientâ s follow-up. Nanotechnology approaches applied in PNI regeneration and theranostics, emphasizing the ones that are moving from the lab bench to the clinics, are herein overviewed., The authors acknowledge the Portuguese Foundation for Science and Technology (FCT) for the financial support provided to Joaquim M. Oliveira (IF/01285/2015) and Joana Silva-Correia (IF/00115/2015) under the program “Investigador FCT”., info:eu-repo/semantics/publishedVersion

 Purpose of Review Overview the outcomes of the latest use of platelet-rich plasma (PRP) for the treatment of knee lesions in the clinics and discuss the challenges and limitations. Recent Findings Recent clinical studies mainly indicate there may be benefit of PRP usage for the treatment of knee lesions. As an autologous source of bioactive components, PRP has been shown to be typically safe, free of major adverse outcomes. The use of PRP has been continuously increasing, and some well-designed, double-blinded, placebo-controlled clinical trials have been published. Summary Clinical outcomes relating to PRP usage are multifactorial and depend on the severity of the lesion and patient characteristics. Although PRP is safe to use and it can be easily applied in the clinics, case-specific considerations are needed to determine whether PRP could be beneficial or not. If the use of PRP is favored, then, the configuration/optimization of the preparation and administration/delivery strategy with or without a concomitant treatment may further enhance the clinical outcomes and patients experience., The authors would like to thank the Portuguese Foundation for Science and Technology (FCT) project M-ERA-NET/0001/2014 project. This work is also a result of the project FROnTHERA(NORTE-01-0145-FEDER-000023), supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). IFC thanks the FCT for the grant SFRH/BD/99555/2014. JMO also thanks the FCT for the funds provided under the program Investigador FCT 2015 (IF/01285/2015).

Cartilage degeneration or damage treatment is still a challenge, but, tissue engineering strategies, which combine cell therapy strategies, which combine cell therapy and scaffolds, and have emerged as a promising new approach. In this regard, polyurethanes and polyacrylates polymers have been shown to have clinical potential to treat osteochondral injuries. Here, we have used polymer microarrays technology to screen 380 different polyurethanes and polyacrylates polymers. The top polymers with potential to maintain chondrocyte viability were selected, with scale-up studies performed to evaluate their ability to support chondrocyte proliferation during long-term culture, while maintaining their characteristic phenotype. Among the selected polymers, poly (methylmethacrylate-co-methacrylic acid), showed the highest level of chondrogenic potential and was used to create a 3D hydrogel. Ultrastructural morphology, microstructure and mechanical testing of this novel hydrogel revealed robust characteristics to support chondrocyte growth. Furthermore, in vitro and in vivo biological assays demonstrated that chondrocytes cultured on the hydrogel had the capacity to produce extracellular matrix similar to hyaline cartilage, as shown by increased expression of collagen type II, aggrecan and Sox9, and the reduced expression of the fibrotic marker's collagen type I. In conclusion, hydrogels generated from poly (methylmethacrylate-co-methacrylic acid) created the appropriate niche for chondrocyte growth and phenotype maintenance and might be an optimal candidate for cartilage tissue-engineering applications. SIGNIFICANCE STATEMENT: Articular cartilage has limited self-repair ability due to its avascular nature, therefore tissue engineering strategies have emerged as a promising new approach. Synthetic polymers displaygreat potential and are widely used in the clinical setting. In our study, using the polymer microarray technique a novel type of synthetic polyacrylate was identified, that was converted into hydrogels for articular cartilage regeneration studies. The hydrogel based on poly (methylmethacrylate-co-methacrylic acid-co-PEG-diacrylate) had a controlable ultrastructural morphology, microstructure (porosity) and mechanical properties (stiffness) appropriate for cartilage engineering. Our hydrogel created the optimal niche for chondrocyte growth and phenotype maintenance for long-term culture, producing a hyaline-like cartilage extracellular matrix. We propose that this novel polyacrylate hydrogel could be an appropriate support to help in the treatment efficient cartilage regeneration.

Guided tissue regeneration (GTR) is a surgical procedure applied in the reconstruction of periodontal defects, where an occlusive membrane is used to prevent the fast-growing connective tissue from migrating into the defect. In this work, silk fibroin (SF) membranes were developed for periodontal guided tissue regeneration. Solutions of SF with glycerol (GLY) or polyvinyl alcohol (PVA) where prepared at several weight ratios up to 30%, followed by solvent casting and thermal annealing at 85 °C for periods of 6 and 12 h to produce high flexible and stable membranes. These were characterized in terms of their morphology, physical integrity, chemical structure, mechanical and thermal properties, swelling capability and in vitro degradation behavior. The developed blended membranes exhibited high ductility, which is particular relevant considering the need for physical handling and adaptability to the defect. Moreover, the membranes were cultured with human periodontal ligament fibroblast cells (hPDLs) up to 7 days. Also, the higher hydrophilicity and consequent in vitro proteolytic degradability of these blends was superior to pure silk fibroin membranes. In particular SF/GLY blends demonstrated to support high cell adhesion and viability with an adequate hPDLs’ morphology, make them excellent candidates for applications in periodontal regeneration.

Although surgical management of peripheral nerve injuries (PNIs) has improved over time, autographs are still the current â gold standardâ treatment for PNIs, which presents numerous limitations. In an attempt to improve natural biomaterial-based nerve guidance conduits (NGCs), chitosan (CHT), a derivative of the naturally occurring biopolymer chitin, has been explored for peripheral nerve regeneration (PNR). In addition to CHT, keratin has gained enormous attention as a biomaterial and tissue engineering scaffolding. In this study, biomimetic CHT/keratin membranes were produced using solvent casting technique. These membranes were broadly characterized in terms of surface topography and physicochemical properties, with techniques such as Fourier Transform Infrared Spectroscopy (FTIR), Differencial Scanning Calorimetry (DSC), contact angle measurments, weight loss and water uptake, as well as Scanning Electron Microscopy (SEM) and Atomic force microscopy (AFM). Biological in vitro assays were also performed, where a preliminary cytotoxicity screening with L929 fibroblast cell line revealed that the membranes and respective materials are suitable for cell culture. In addition, Schwann cells, fibroblasts and endothelial cells were directly seeded in the membranes. Quantitative and qualitative assays revealed that the addition of keratin enchanced cell viablity and adhesion. Based on the in vitro encouraging results, the in vivo angiogenic/antiangiogenic potential of CHT and CHT/keratin membranes was assessed, using an optimized chick embryo chorioallantoic membrane assay, where higher angiogenic responses were seen in keratin-enriched materials. Overall, the obtained results indicate the higher potential of CHT/keratin membranes for guided tissue regeneration applications in the field of PNR., This study was also supported by the European Community’s Seventh Framework Programme (FP7-HEALTH-2011) under grant agreement no. 278612 (BIOHYBRID). The authors acknowledge the Portuguese Foundation for Science and Technology (FCT) for the financial support provided to Joaquim M. Oliveira (IF/ 00423/2012 and IF/01285/2015) and Joana Silva-Correia (IF/ 00115/2015) under the program “Investigador FCT”.

Chemotherapeutic resistance is a major problem in effective cancer treatment. Cancer cells engage various cells or mechanisms to resist anti-cancer therapeutics, which results in metastasis and the recurrence of disease. Considering the cellular heterogeneity of cancer stroma, the involvement of stem cells is reported to affect the proliferation and metastasis of osteosarcoma. Hence, the duo (osteosarcoma: Saos 2 and human adipose-derived stem cells: ASCs) is co-cultured in present study to investigate the therapeutic response using a nonadherent, concave surface. Staining with a cell tracker allows real-time microscopic monitoring of the cell arrangement within the sphere. Cell–cell interaction is investigated by means of E-cadherin expression. Comparatively high expression of E-cadherin and compact organization is observed in heterotypic tumorspheres (Saos 2–ASCs) compared to homotypic ones (ASCs), limiting the infiltration of chemotherapeutic compound doxorubicin into the heterotypic tumorsphere, which in turn protects cells from the toxic effect of the chemotherapeutic. In addition, genes known to be associated with drug resistance, such as SOX2, OCT4, and CD44 are overexpressed in heterotypic tumorspheres post-chemotherapy, indicating that the duo collectively repels the effect of doxorubicin. The interaction between ASCs and Saos 2 in the present study points toward the growing oncological risk of using ASC-based regenerative therapy in cancer patients and warrants further investigation., This work is supported by the European Union Framework Programme for Research and Innovation Horizon 2020 (nº 668983 — FoReCaST; FROnTHERA—NORTE-01-0145-FEDER-000023), Investigator FCT program (IF/01214/2014—V.M.), FCT2015 (IF/01285/2015—J.M.O.) and PTDC/BTMORG/28168/2017 (V.B. and S.C.K.).

Rheumatoid arthritis (RA) is an autoimmune and chronic inflammatory disorder that mostly affects the synovial joints and can promote both cartilage and bone tissue destruction. Several conservative treatments are available to relieve pain and control the inflammation; however, traditional drugs administration are not fully effective and present severe undesired side effects. Hydrogels are a very attractive platform as a drug delivery system to guarantee these handicaps are reduced, and the therapeutic effect from the drugs is maximized. Furthermore, hydrogels can mimic the physiological microenvironment and have the mechanical behavior needed for use as cartilage in vitro model. The testing of these advanced delivery systems is still bound to animal disease models that have shown low predictability. Alternatively, hydrogel-based human dynamic in vitro systems can be used to model diseases, bypassing some of the animal testing problems. RA dynamic disease models are still in an embryonary stage since advances regarding healthy and inflamed cartilage models are currently giving the first steps regarding complexity increase. Herein, recent studies using hydrogels in the treatment of RA, featuring different hydrogel formulations are discussed. Besides, their use as artificial extracellular matrices in dynamic in vitro articular cartilage is also reviewed., Norte2020 project (NORTE-08-5369-FSE000044) and the Portuguese Foundation for Science and Technology (FCT) program (PD/BD/143081/2018). IFC thanks the TERM RES-Hub, Tissue Engineering and Regenerative Medicine Infrastructure project, funded by FCT. The FCT distinction attributed to JMO under the Investigator FCT program (number IF/01285/2015) is also greatly acknowledged

Hydrogel application feasibility is still limited mainly due to their low mechanical strength and fragile nature. Therefore, several physical and chemical cross-linking modifications are being used to improve their properties. In this research, methacrylated Kefiran was synthesized by reacting Kefiran with methacrylic anhydride (MA). The developed MA-Kefiran was physicochemically characterized, and its biological properties evaluated by different techniques. Chemical modification of MA-Kefiran was confirmed by 1H-NMR and FTIR and GPC-SEC showed an average Mw of 793 kDa (PDI 1.3). The mechanical data obtained revealed MA-Kefiran to be a pseudoplastic fluid with an extrusion force of 11.21 ± 2.87 N. Moreover, MA-Kefiran 3D cryogels were successfully developed and fully characterized. Through micro-CT and SEM, the scaffolds revealed high porosity (85.53 ± 0.15%) and pore size (33.67 ± 3.13 μm), thick pore walls (11.92 ± 0.44 μm) and a homogeneous structure. Finally, MA-Kefiran revealed to be biocompatible by presenting no hemolytic activity and an improved cellular function of L929 cells observed through the AlamarBlue® assay. By incorporating methacrylate groups in the Kefiran polysaccharide chain, a MA-Kefiran product was developed with remarkable physical, mechanical, and biological properties, resulting in a promising hydrogel to be used in tissue engineering and regenerative medicine applications., H. Radhouani and C. Goncalveswere supported by the Foundation for Science and Technology (FCT) fromPortugal, with references CEECIND/00111/2017 and SFRH/BPD/94277/2013, respectively. S. Correia and this work were funded by the R&D Project KOAT-Kefiran Exopolysaccharide: Promising Biopolymer for Use in Regenerative Medicine and Tissue Engineering, with reference PTDC/BTMMAT/29760/2017 (POCI-01-0145-FEDER-029760), financed by FCT and co-financed by FEDER and POCI. We also thank Duarte N. Carvalho for input on the schematic representation of the process.

This review focuses on vascularization and strategies involved in its evaluation and modulation. Clinical issues associated with engineered tissues of an atomically relevant size that require a vascular network to supply their cells with nutrients and oxygen are analyzed in terms of vascular network formation within scaffolds, which can be produced from varying biomaterials, with the capability of connecting to the vasculature of the patient. Developing angiogenesis techniques and monitoring of angiogenesis development as well as how these methods can be further utilized to tailor vascularization within large tissue engineered constructs are also discussed. Finally, we offer a glimpse toward the future by providing an outlook for vascularization and associated emerging bioprinting concepts in tissue engineering applications., The authors acknowledge the financial support provided through the projects B-FABULUS (PTDC/BBB-ECT/2690/2014) and Fun4TE (PTDC/EMD-EMD/31367/2017), financed by the Portuguese Foundation for Science and Technology (FCT) and cofinanced by European Regional Development Fund (FEDER) and Operational Programme for Competitiveness and Internationalisation (POCI). This work was also partly supported by H2020-MSCA-RISE BAMOS project, funded from the European Union's Horizon 2020 research and innovation program under Grant Agreement No. 734156. S.V.L. acknowledges FCT for the PhD grant (SFRH/BD/143954/2019). The FCT distinction attributed to J.S.-C. (IF/00115/2015) under the Investigator FCT program is also greatly acknowledged.

Bioengineering has been revolutionizing the production of biofunctional tissues for tackling unmet clinical needs. Bioengineers have been focusing their research in biofabrication, especially 3D bioprinting, providing cutting-edge approaches and biomimetic solutions with more reliability and cost–effectiveness. However, these emerging technologies are still far from the clinical setting and deep learning, as a subset of artificial intelligence, can be widely explored to close this gap. Thus, deep-learning technology is capable to autonomously deal with massive datasets and produce valuable outputs. The application of deep learning in bioengineering and how the synergy of this technology with biofabrication can help (more efficiently) bring 3D bioprinting to clinics, are overviewed herein., The authors acknowledge financial support provided through projects B-FABULUS (PTDC/BBB-ECT/2690/2014), Fun4TE (PTDC/EMD-EMD/31367/2017) and JUSThera (ref: NORTE-01-0145-FEDER-000055), financed by the Portuguese Foundation for Science and Technology (FCT) and co-financed by European Regional Development Fund (FEDER) and Operational Program for Competitiveness and Internationalization (POCI). JB Costa acknowledges the Junior Researcher contract (POCI-01-0145-FEDER 031367) attributed by FCT to Fun4TE. The FCT distinction attributed to J Silva-Correia (IF/00115/2015) under the Investigator FCT program is also greatly acknowledged.

"Publication Date (Web): February 20, 2015", In the last few years, great progress has been made to validate tissue engineering strategies in preclinical studies and clinical trials on the regeneration of osteochondral defects. In the preclinical studies, one of the dominant strategies comprises the development of biomimetic/bioactive scaffolds, which are used alone or incorporated with growth factors and/or stem cells. Many new trends are emerging for modulation of stem cell fate towards osteogenic and chondrogenic differentiations, but bone/cartilage interface regeneration and physical stimulus have been showing great promise. Besides the matrix-associated autologous chondrocyte implantation (MACI) procedure, the matrix-associated stem cells implantation (MASI) and layered scaffolds in acellular or cellular strategy are also applied in clinic. This review outlines the progresses at preclinical and clinical levels, and identifies the new challenges in osteochondral tissue engineering. Future perspectives are provided, e.g., the applications of extracellular matrix-like biomaterials, computer-aided design/manufacture of osteochondral implant and reprogrammed cells for osteochondral regeneration., The authors thank the Portuguese Foundation for Science and Technology (FCT) through the projects TISSUE2TISSUE (PTDC/CTM/105703/2008) and OsteoCart (PTDC/CTM-BPC/115977/2009). We also acknowledge European Union's Seventh Framework Programme (FP7/2007-2013) under grant agreement REGPOT-CT2012-316331-POLARIS. L-P.Y. acknowledges the PhD scholarship from FCT (SFRH/BD/64717/2009). The FCT distinction attributed to J.M.O. and A.L.O. under the Investigator FCT program (IF/00423/2012) and (IF/00411/2013) are also greatly acknowledged.