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Bio-based polymers are attracting great interest due to their potential for several applications in place of conventional polymers. In the field of electrochemical devices, the electrolyte is a fundamental element that determines their performance, and polymers represent good candidates for developing solid-state and gel-based electrolytes toward the development of full-solid-state devices. In this context, the fabrication and characterization of uncrosslinked and physically cross-linked collagen membranes are reported to test their potential as a polymeric matrix for the development of a gel electrolyte. The evaluation of the membrane’s stability in water and aqueous electrolyte and the mechanical characterization demonstrated that cross-linked samples showed a good compromise in terms of water absorption capability and resistance. The optical characteristics and the ionic conductivity of the cross-linked membrane, after overnight dipping in sulfuric acid solution, demonstrated the potential of the reported membrane as an electrolyte for electrochromic devices. As proof of concept, an electrochromic device was fabricated by sandwiching the membrane (after sulfuric acid dipping) between a glass/ITO/PEDOT:PSS substrate and a glass/ITO/SnO2 substrate. The results in terms of optical modulation and kinetic performance of such a device demonstrated that the reported cross-linked collagen membrane could represent a valid candidate as a water-based gel and bio-based electrolyte for full-solid-state electrochromic devices.

Type I collagen is the most abundant protein of the human body. Due to its favourable properties, collagen extracted from animal tissues is adopted to manufacture a wide range of devices for biomedical applications. Compared to bovine and porcine collagens, which are the most largely used, equine collagen is free from the risk of zoonosis, has no reported immune reactions, and has not religious constraints. In this work, a recently available type I collagen extracted from horse tendon was evaluated and compared with a commercially available collagen isoform derived from the same species and tissue. Detailed physical, chemical and biological investigations were performed, in agreement with the requirements of the current standard for the characterization of type I collagen to be used for the manufacture of Tissue Engineering Medical Products. To the best of our knowledge, this is the first report on the complete primary structure of the investigated collagen.

Soft tissues diseases significantly affect patients quality of life and usually require targeted, costly and sometimes constant interventions. With the average lifetime increase, a proportional increase of age-related soft tissues diseases has been witnessed. Due to this, the last two decades have seen a tremendous demand for minimally invasive one-step resolutive procedures. Intensive scientific and industrial research has led to the recognition of injectable formulations as a new advantageous approach in the management of complex diseases that are challenging to treat with conventional strategies. Among them, collagen-based products are revealed to be one of the most promising among bioactive biomaterials-based formulations. Collagen is the most abundant structural protein of vertebrate connective tissues and, because of its structural and non-structural role, is one of the most widely used multifunctional biomaterials in the health-related sectors, including medical care and cosmetics. Indeed, collagen-based formulations are historically considered as the “gold standard” and from 1981 have been paving the way for the development of a new generation of fillers. A huge number of collagen-based injectable products have been approved worldwide for clinical use and have routinely been introduced in many clinical settings for both aesthetic and regenerative surgery. In this context, this review article aims to be an update on the clinical outcomes of approved collagen-based injectables for both aesthetic and regenerative medicine of the last 20 years with an in-depth focus on their safety and effectiveness for the treatment of diseases of the integumental, gastrointestinal, musculoskeletal, and urogenital apparatus.

Vascularization is a highly conserved and considerably complex and precise process that is finely driven by endogenous regulatory processes at the tissue and systemic levels. However, it can reveal itself to be slow and inadequate for tissue repair and regeneration consequent to severe lesions/damages. Several biomaterial-based strategies were developed to support and enhance vasculogenesis by supplying pro-angiogenic agents. Several approaches were adopted to develop effective drug delivery systems for the controlled release of a huge variety of compounds. In this work, a microparticulate system was chosen to be loaded with the essential amino acid L-lysine, a molecule that has recently gained interest due to its involvement in pro-angiogenic, pro-regenerative, and anti-inflammatory mechanisms. Poly (lactic-co-glycolic acid), the most widely used FDA-approved biodegradable synthetic polymer for the development of drug delivery systems, was chosen due to its versatility and ability to promote neovascularization and wound healing. This study dealt with the development and the effectiveness evaluation of a PLGA-based microparticulate system for the controlled release of L-lysine. Therefore, in order to maximize L-lysine encapsulation efficiency and tune its release kinetics, the microparticle synthesis protocol was optimized by varying some processing parameters. All developed formulations were characterized from a morphological and physicochemical point of view. The optimized formulation was further characterized via the evaluation of its preliminary biological efficacy in vitro. The cellular and molecular studies revealed that the L-lysine-loaded PLGA microparticles were non-toxic, biocompatible, and supported cell proliferation and angiogenesis well by stimulating the expression of pro-angiogenic genes such as metalloproteinase-9, focal adhesion kinases, and different growth factors. Thus, this work showed the potential of delivering L-lysine encapsulated in PLGA microparticles as a cost-effective promoter system for angiogenesis enhancement and rapid healing.

Fish collagen garnered significant academic and commercial focus in the last decades featuring prospective applications in a variety of health-related industries, including food, medicine, pharmaceutics, and cosmetics. Due to its distinct advantages over mammalian-based collagen, including the reduced zoonosis transmission risk, the absence of cultural-religious limitations, the cost-effectiveness of manufacturing process, and its superior bioavailability, the use of collagen derived from fish wastes (i.e., skin, scales) quickly expanded. Moreover, by-products are low cost and the need to minimize fish industry waste’s environmental impact paved the way for the use of discards in the development of collagen-based products with remarkable added value. This review summarizes the recent advances in the valorization of fish industry wastes for the extraction of collagen used in several applications. Issues related to processing and characterization of collagen were presented. Moreover, an overview of the most relevant applications in food industry, nutraceutical, cosmetics, tissue engineering, and food packaging of the last three years was introduced. Lastly, the fish-collagen market and the open technological challenges to a reliable recovery and exploitation of this biopolymer were discussed.

In the last two decades, fisheries and fish industries by-products have started to be recovered for the extraction of type I collagen because of issues related to the extraction of traditional mammalian tissues. In this work, special attention has been paid to by-products from fish bred in aquaponic plants. The valorization of aquaponic fish wastes as sources of biopolymers would make the derived materials eco-friendlier and attractive in terms of profitability and cost effectiveness. Among fish species, Nile Tilapia is the second-most farmed species in the world and its skin is commonly chosen as a collagen extraction source. However, to the best of our knowledge, no studies have been carried out to investigate, in depth, the age-related differences in fish skin with the final aim of selecting the most advantageous fish size for collagen extraction. In this work, the impact of age on the structural and compositional properties of Tilapia skin was evaluated with the aim of selecting the condition that best lends itself to the extraction of type I collagen for biomedical applications, based on the known fact that the properties of the original tissue have a significant impact on those of the final product. Performed analysis showed statistically significant age-related differences. In particular, an increase in skin thickness (+110 µm) and of wavy-like collagen fiber bundle diameter (+3 µm) besides their organization variation was observed with age. Additionally, a preferred collagen molecule orientation along two specific directions was revealed, with a higher fiber orientation degree according to age. Thermal analysis registered a shift of the endothermic peak (+1.7 °C) and an increase in the enthalpy (+3.3 J/g), while mechanical properties were found to be anisotropic, with an age-dependent brittle behavior. Water (+13%) and ash (+0.6%) contents were found to be directly proportional with age, as opposed to protein (−8%) and lipid (−10%) contents. The amino acid composition revealed a decrease in the valine, leucine, isoleucine, and threonine content and an increase in proline and hydroxyproline. Lastly, fatty acids C14:0, C15:0, C16:1, C18:2n6c, C18:3n6, C18:0, C20:3n3, and C23:0 were revealed to be upregulated, while C18:1n9c was downregulated with age.

Three-dimensional (3D) cell culture systems mimic the structural complexity of the tissue microenvironment that includes the extracellular matrix (ECM) in addition to the cellular components Thus, 3D culture systems are increasingly important as they resemble the ECM-cell and cell-cell physical interactions occurring in vivo. So far, several scaffold-based culture systems and techniques have been proposed as valuable approaches for large-scale production of spheroids, but often suffering of poor reproducible conditions or high costs of production. In this work we present a reliable 3D culture system based on collagen I-blended agarose hydrogels and show how the variation of the agarose weight percentage affects the physical and mechanical properties of the resulting hydrogel, being that with a lower amount of agarose more permeable, softer and more prone to degradation compared to hydrogels with higher agarose concentrations. We have also evaluated the effect of the different physical and mechanical properties of the agarose hydrogels on the growth, size, morphology and cell motility of spheroids obtained by culturing three different breast cancer cell lines (MCF-7, MDA-MB-361and MDA-MB-231). As proof of concept, the cisplatin penetration and its cytotoxic effect on the tumor spheroids was evaluated as function of the hydrogel stiffness. Noteworthy, the possibility to recover the spheroids from the hydrogels for further processing and other biological studies has been also considered.

In the present work, we investigated the potential of novel semi-interpenetrating polymer network (semi-IPN) cryogels, obtained through ultraviolet exposure of aqueous mixtures of poly(ethylene glycol) diacrylate and type I collagen, as tunable off-the-shelf platforms for 3D cancer cell research. We synthesized semi-IPN cryogels with variable collagen amounts (0.1% and 1% w/v) and assessed the effect of collagen on key cryogel properties for cell culture, for example, porosity, degradation rate and mechanical stiffness. Then, we investigated the ability of the cryogels to sustain the long-term growth of two pancreatic ductal adenocarcinoma (PDAC) cell populations, the parenchymal Panc1 cells and their derived cancer stem cells. Results revealed that both cell lines efficiently infiltrated, attached and expanded in the cryogels over a period of 14 days. However, only when grown in the cryogels with the highest collagen concentration, both cell lines reproduced their characteristic growth pattern previously observed in collagen-enriched organotypic cultures, biomimetic of the highly fibrotic PDAC stroma. Cellular preembedding in Matrigel, that is, the classical approach to develop/grow organoids, interfered with an efficient intra-scaffold migration and growth. Although preliminary, these findings highlight the potential of the proposed cryogels as reproducible and tunable cancer cell research platforms.

The aim of this paper is to model and to compare the results of the mechanical characterization, carried out on numerical models and real specimens, of uniform P-scaffolds with different porosity values. The analysis includes the morphological characterization of 3D printed specimens and the implementation of a FEM shell model to reproduce a compressive test suitable for mechanical properties evaluation of PLA scaffolds. Young modulus and yield strength were also obtained, in order to verify the numerical model accuracy, by experimental tests on 3D printed PLA scaffolds. Numerical results showed that the shell model was able to reproduce, more efficiently compared to a solid model proposed in a previous work, both elastic and plastic behavior of the scaffolds, providing elastic modulus values very close to the experimental ones. On the other hand, the not very high quality of the 3D printing, detected by MicroCT analysis, caused a significant dispersion in the yield strength numerical values respect to the real data. Anyway, an inverse correlation between mechanical properties and porosity was found as expected. The elastic modulus values were similar to the typical values of the trabecular bone for whose regeneration this kind of scaffolds is usually employed.

The optimization of collagen-based scaffolds for tissue engineering goes through the careful selection of the crosslinking method(s), which should impart the prerequisite mechanical and degradation properties without impairing the cell/tissue response. Here, we investigated the chemically effective (ρxch) and the elastically effective (ρxel) crosslink density of collagen-based scaffolds, induced by various crosslinking methods. The aim was to get a deeper insight into the influence of intramolecular and intermolecular crosslinks on several scaffold properties. Freeze-dried collagen matrices were crosslinked via a dehydrothermal treatment (DHT), and then treated with different chemical agents, including carbodiimide (EDC), glutaraldehyde (GTA), formaldehyde (FA), genipin (GP) and dimethyl suberimidate (DMS). Quantification of primary amines and stress-relaxation compressive tests were performed to evaluate ρxch and ρxel, respectively. Scaffolds were then assessed for their water uptake, thermal stability and in vitro resistance to enzymatic degradation. Interestingly, for the various crosslinking treatments ρxch was found to increase in the order DHT

Chitosan (Cs) mats obtained by electrospinning are potentially ideal scaffolds for tissue engineering. This technique allows obtaining nanometric fibrous structures with preferred orientation, which in turn enable cells to align themselves and produce extracellular matrix along desired orientations. In this study, we fabricated aligned Cs electrospun nanofiber mats and investigated the role of the amino acid l -Arginine (L-Arg) as stabilizing agent. Morphological, chemical, mechanical and biological characterizations were performed on untreated and L-Arg treated nanofibrous mats showing the role of L-Arg as biomimetic stabilizer. L-Arg acts as chemical stabilizer of nanofibrous mats, providing improved wettability behavior, mechanical properties and stability even after 60 days in aqueous medium in comparison to untreated mats. Moreover, preliminary biological tests demonstrated favorable cell-material interactions implying physiological responses in terms of viability and proliferation. The proposed L-Arg-treated Cs mats can be considered as potential scaffolds for highly oriented tissue patterning.

Collagen represents one of the most widely used biomaterial for scaffolds fabrication in tissue engineering as it represents the mechanical support of natural tissues. It also provides physical scaffolding for cells and it influences their attachment, growth, and tissue regeneration. Among all fibrillary collagens, type I is considered one of the gold standard for scaffolds fabrication, thanks to its high biocompatibility, biodegradability, and hemostatic properties. It can be extracted by chemical and enzymatic protocols from several collagen-rich tissues, such as tendon and skin, of different animal species. Both the extraction processes and the manufacturing protocols for scaffolds fabrication provide structural and mechanical changes that can be tuned in order to deeply impact the properties of the final biomaterial. The aim of this review is to discuss the role of X-rays to study structural changes of type I collagen from fresh collagen-rich tissues (bovine, equine, fish) to the final scaffolds, with the aim to screen across available collagen sources and scaffolds fabrication protocols to be used in tissue regeneration.

This work explored the use of chitosan (Cs) and poly(ethylene oxide) (PEO) blends for the fabrication of electrospun fiber-orientated meshes potentially suitable for engineering fiber-reinforced soft tissues such as tendons, ligaments, or meniscus. To mimic the fiber alignment present in native tissue, the CS/PEO blend solution was electrospun using a traditional static plate, a rotating drum collector, and a rotating disk collector to get, respectively, random, parallel, circumferential-oriented fibers. The effects of the different orientations (parallel or circumferential) and high-speed rotating collector influenced fiber morphology, leading to a reduction in nanofiber diameters and an improvement in mechanical properties.