Microbial infections from post-surgery or other medical-related procedure is a serious health problem. Nowadays, the research is focused on the development of new drug-free materials with antibacterial properties to prevent or minimize the risk of infections. Spider silk is known for its unique biomechanical properties allied with biocompatibility. Recombinant DNA technology allows to bioengineering spider silk with antimicrobial peptides (AMP). Thus, our goal was to bioengineered spider silk proteins with AMP (6mer-HNP1) as an antibacterial drug-free coating for commercial silk sutures (Perma-Hand Ò ) for decreasing bacterial infections. Perma-Hand Ò sutures were coated with 6mer-HNP1 by dip coating. In vitro tests, using human fetal lung fibroblasts (MRC5), showed that coated sutures sustained cell viability, and also, the contact with red blood cells (RBCs) demonstrate blood compatibility. Also, the coatings inhibited significantly the adherence and formation of biofilm, where sutures coated with 6mer-HNP1 produced a 1.5 log reduction of Methicillin-Resistant Staphylococcus aureus (MRSA) and a 2 log reduction of Escherichia coli (E. coli) compared to the uncoated Perma-Hand Ò suture. The mechanical properties of Perma-Hand Ò sutures were not affected by the presence of bioengineered spider silk proteins. Thus, the present work demonstrated that using spider silk drug-free coatings it is possible to improve the antibacterial properties of the commercial sutures. Furthermore, a new class of drug-free sutures for reducing post-implantation infections can be developed. Statement of SignificanceMicrobial infections from post-surgery or other medical-related procedure is a serious health problem. Developing new drug-free materials with antibacterial properties is an approach to prevent or minimize the risk of infections. Spider silk is known for its unique biomechanical properties allied with biocompatibility. Recombinant DNA technology allow to bioengineering spider silk with antimicrobial peptides (AMP). Our goal is bioengineered spider silk proteins with AMP as an antibacterial coating for silk sutures. The coatings showed exceptional antibacterial properties and maintained intrinsic mechanical features. In vitro studies showed a positive effect of the coated sutures on the cell behavior. With this new drugfree bioengineered spider silk coating is possible to develop a new class of drug-free sutures for reducing post-implantation infections.

Tendinopathies represent half of all musculoskeletal injuries worldwide. Inflammatory events contribute to both tendon healing and to tendinopathy conditions but the cellular triggers leading to one or the other are unknown. In previous studies, we showed that magnetic field actuation modulates human tendon cells (hTDCs) behavior in pro-inflammatory environments, and that magnetic responsive membranes could positively influence inflammation responses in a rat ectopic model. Herein, we propose to investigate the potential synergistic action of the magnetic responsive membranes, made of a polymer blend of starch with polycaprolactone incorporating magnetic nanoparticles (magSPCL), and the actuation of pulsed electromagnetic field (PEMF): 5 Hz, 4mT of intensity and 50% of duty cycle, in IL-1 β-treated-hTDCs, and in the immunomodulatory response of macrophages. It was found that the expression of pro-inflammatory (TNF α, IL-6, IL-8, COX-2) and ECM remodeling (MMP-1,-2,-3) markers tend to decrease in cells cultured onto magSPCL membranes under PEMF, while the expression of TIMP-1 and anti-inflammatory genes (IL-4, IL-10) increases. Also, CD16 ++ and CD206 + macrophages were only found on magSPCL membranes with PEMF application. Magnetic responsive membranes show a modulatory effect on the inflammatory profile of hTDCs favoring anti-inflammatory cues which is also supported by the anti-inflammatory/repair markers expressed in macrophages. These results suggest that magnetic responsive magSPCL membranes can contribute for inflammation resolution acting on both resident cell populations and inflammatory cells, and thus significantly contribute to tendon regenerative strategies. Statement of significance Magnetically-assisted strategies have received great attention in recent years to remotely trigger and guide cell responses. Inflammation plays a key role in tendon healing but persistent pro-inflammatory molecules can contribute to tendon disorders, and therefore provide a therapeutic target for advanced treatments. We have previously reported that magnetic fields modulate the response of human tendon cells (hT-DCs) conditioned to pro-inflammatory environments (IL-1 β-treated-hTDCs), and that magnetic responsive membranes positively influence immune responses. In the present work, we combined pulsed electromagnetic field (PEMF) and magnetic responsive membranes to guide the inflammatory profile of IL-1 βtreated-hTDCs and of macrophages. The results showed that the synergistic action of PEMF and magnetic membranes supports the applicability of magnetically actuated systems to regulate inflammatory events and stimulate tendon regeneration.

Bone marrow cells are a potential source to induce different lineage cells which can be used to rebuild or replace damaged tissues using a Tissue Engineering (TE) approach. However, TE strategies usually require the use of a material to support the development of a biological tissue. Beta-polyvinylidene fluoride (β-PVDF) is a biocompatible, thermoplastic with piezo-electrical properties that has been shown to provide a good cellular attachment and therefore might present advantageous properties as a scaffold material for cell seeding/culturing. The present study describes the characterization of β-PVDF membranes as a support material for growth and differentiation of goat marrow cells (GMCs) into osteoblasts, leading to the formation of substitutes for tissue regeneration. The obtained results suggest that β-PVDF piezoelectric properties influence cellular behavior. β- PVDF membranes not only enhance GMCs adherence and proliferation but also improve differentiation towards the osteogenic phenotype both in static and dynamic culture conditions. Furthermore, β-PVDF membranes exhibit very promising properties, suggesting that this material provides adequate support for the seeding and the development of undifferentiated cells towards a desired phenotype.

Human tendon-derived cell sheets created by magnetic force-based tissue engineering hold tenogenic and immunomodulatory potential,

Identification of a suitable cell source and bioactive agents guiding cell differentiation towards tenogenic phenotype represents a prerequisite for advancement of cellbased therapies for tendon repair. Human adipose-derived stem cells (hASCs) are a promising, yet intrinsically heterogenous population with diversified differentiation capacities. In this work, we investigated antigenically-defined subsets of hASCs expressing markers related to tendon phenotype or associated with pluripotency that might be more prone to tenogenic differentiation, when compared to unsorted hASCs. Subpopulations positive for tenomodulin (TNMD+ hASCs) and stage specific early antigen 4 (SSEA-4+ hASCs), as well as unsorted ASCs were cultured up to 21 days in basic medium or media supplemented with TGF-β3 (10 ng/ml), or GDF-5 (50 ng/ml). Cell response was evaluated by analysis of expression of tendon-related markers at gene level and protein level by real time RT-PCR, western blot, and immunocytochemistry. A significant upregulation of scleraxis was observed for both subpopulations and unsorted hASCs in the presence of TGF-β3. More prominent alterations in gene expression profile in response to TGF-β3 were observed for TNMD+ hASCs. Subpopulations evidenced an increased collagen III and TNC deposition in basal medium conditions in comparison with unsorted hASCs. In the particular case of TNMD+ hASCs, GDF-5 seems to influence more the deposition of TNC.Within hASCs populations, discrete subsets could be distinguished offering varied sensitivity to specific biochemical stimulation leading to differential expression of tenogenic components suggesting that cell subsets may have distinctive roles in the complex biological responses leading to tenogenic commitment to be further explored in cell based strategies for tendon tissues. KEYWORDS adipose derived stromal/stem cells, subpopulation, tenogenic differentiation, tenomodulin, transforming growth factor beta 3 Ana I. Gonçalves and Dominika Berdecka contributed equally.