Gene therapy uses nucleic acids as functional molecules to activate biological treatment for a wide range of diseases, such as cancer 1,2 , cystic fibrosis 3 , heart disease 4 , diabetes 5 , haemophilia and HIV/AIDS 6 . Nucleic acids have been attracting increasing attention owing to the global effort in the human genome elucidation together with recent discoveries such as RNA interference (RNAi) and CRISPR-based genome editing [7][8][9] . Gene therapy uses genetic material to alter the expression of a target gene or to modify the biological properties of living cells for therapeutic needs. In recent years, multiple gene therapy products have been approved by the regulatory agencies for various applications 10 . Perhaps the most relevant example is the authorization of mRNA vaccines to fight the COVID-19 outbreak 11 .Gene therapy can be divided into three main avenues, as detailed in Fig. 1. First is editing mutated genes using CRISPR-Cas technology to cause gain or loss of function 12,13 . Second, upregulating gene expression can be achieved through the insertion of a functional gene copy to be expressed by using molecules such as DNA plasmid (pDNA), minicircle DNA (mcDNA), synthetic mRNA, circular RNA and self-amplifying RNA (saRNA) [14][15][16] . Last is downregulating gene expression using molecules such as small interfering RNA (siRNA), antisense oligonucleotides (ASOs), short hairpin RNA (shRNA) and microRNA (miRNA) 17,18 .Nucleic acids have promising advantages compared with conventional drugs 19 . Unlike the latter, the mechanism of action and high specificity of nucleic acids present a possible therapy route for viral infections, various cancers and undruggable genetic disorders with unmet clinical need. Moreover, theoretically, a single treatment of the genetic payload can achieve a durable and even curative effect 20 . However, delivering nucleic acids to reach their active site inside the cell is challenging owing to their low in vivo stability and rapid host clearance outside cells. Additionally, nucleic acids are poorly permeable through the cellular membrane owing to their negative charge, high molecular weight and hydrophilicity 21 . Nonetheless, few delivery challenges differ between DNA and RNA. For example, the payload and carrier toxicity are of greater concern when delivering RNA molecules usually associated with short-term activity and low retention inside the cell, hence requiring more frequent administration 22 . Alternatively, DNA activity inside the nucleus adds complexity related to low nuclear transport, thus leading to distinguishing design concepts regarding the delivery system compared with RNA molecules 23 . Together with specific challenges relevant to the delivered molecule, the fundamental challenge is to develop tailored systems that can facilitate nucleic acid uptake into target cells. The carrier itself needs to overcome extracellular and intracellular barriers, provide protection from nuclease activity in the bloodstream, enhance and assist with cellular uptake, and promote ...

Blood components play key roles in the modulation of the wound healing process and, together with the provisional fibrin matrix ability to selectively bind bioactive molecules and control its spatial-temporal presentation, define the complex microenvironment that characterize this biological process. As a biomimetic approach, the use of blood derivatives in regenerative strategies has awakened as a source of multiple therapeutic biomolecules. Nevertheless, and despite their clinical relevance, blood derivatives have been showing inconsistent therapeutic results due to several factors, including proper control over their delivery mechanisms. Herein, we highlight recent trends on the use biomaterials to protect, sequester and deliver these pools of biomolecules in tissue engineering and regenerative medicine approaches. Particular emphasis is given to strategies that enable to control their spatiotemporal delivery and improve the selectivity of presentation profiles of the biomolecules derived from blood derivatives rich in platelets. Finally, we discussed possible directions for biomaterials design to potentiate the aimed regenerative effects of blood derivatives and achieve efficient therapies.

Three-dimensional (3D) bioprinting holds the promise to fabricate tissue and organ substitutes for regenerative medicine. However, the lack of bioactive inks to fabricate and support functional living constructs is one of the main limitations hindering the progress of this technology. In this study, a biofunctional human-based nanocomposite bioink (HUink) composed of platelet lysate hydrogels reinforced by cellulose nanocrystals is reported. When combined with suspended bioprinting technologies, HUink allows the biofabrication of 3D freeform constructs with high resolution and integrity, mimicking the hierarchical nano-tomacro fibrillary composition of native tissues. Remarkably, HUink supports bioprinting of stem cells with high viability immediately after extrusion and over long-term cell culture without the need for additional biochemical or animal-derived media supplementation. As opposed to typical polymer-based bioinks, the pool of growth factors, cytokines and adhesion proteins in HUink boosts cell spreading and proliferation, stimulating the fast production of cell-secreted extracellular matrix. This innovative bioprinting platform with unpaired biofunctionality allows the fabrication of complex freeform cell-laden constructs that can ultimately be applied in the development of xeno-free 3D tissue models for in vitro research or to develop tissue and organ surrogates for clinical applications.

Injectable hydrogels are particularly interesting for applications in minimally invasive tissue engineering and regenerative medicine strategies. However, the typical isotropic microstructure of these biomaterials limits their potential for the regeneration of ordered tissues. In the present work, we decorated rod-shaped cellulose nanocrystals with magnetic nanoparticles and coated these with polydopamine and polyethylene glycol polymer brushes to obtain chemical and colloidal stable nanoparticles. Then, these nanoparticles (0.1-0.5 wt.%.) were incorporated within gelatin hydrogels, creating injectable and magnetically responsive materials with potential for various biomedical applications. Nanoparticles alignment within the hydrogel matrix was achieved under exposure to uniform low magnetic fields (108 mT), resulting in biomaterials with directional microstructure and anisotropic mechanical properties. The biological performance of these nanocomposite hydrogels was studied using adipose tissue derived human stem cells. Cells encapsulated in the nanocomposite hydrogels showed high rates of viability demonstrating that the nanocomposite biomaterials are not cytotoxic. Remarkably, the microstructural patterns stemming from nanoparticles alignment induced the directional growth of seeded and, to a lower extent, encapsulated cells in the hydrogels, suggesting that this injectable system might find application in both cellular and acellular strategies targeting the regeneration of anisotropic tissues.

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The extracellular matrix (ECM)-biomimetic fibrillar structure of platelet lysate (PL) gels along with their enriched milieu of biomolecules has drawn significant interest in regenerative medicine applications. However, PL-based gels have poor structural stability, which severely limits their performance as a bioinstructive biomaterial. Here, rod-shaped cellulose nanocrystals (CNC) are used as a novel approach to modulate the physical and biochemical microenvironment of PL gels enabling their effective use as injectable human-based cell scaffolds with a level of biomimicry that is difficult to recreate with synthetic biomaterials. The incorporation of CNC (0 to 0.61 wt%) into the PL fibrillar network during the coagulation cascade leads to decreased fiber branching, increased interfiber porosity (from 66 to 83%) and modulates fiber (from 1.4 ± 0.7 to 27 ± 12 kPa) and bulk hydrogel (from 18 ± 4 to 1256 ± 82 Pa) mechanical properties. As a result of these physicochemical alterations, nanocomposite PL hydrogels resist the typical extensive clot retraction (from 76 ± 1 to 24 ± 3 at day 7) and show favored retention of PL bioactive molecules. The feedback of these cues on the fate of human adipose-derived stem cells is evaluated, showing how it can be explored to modulate the commitment of encapsulated stem cells toward different genetic phenotypes without the need for additional external biological stimuli. These fibrillar nanocomposite hydrogels allow therefore the exploration of the outstanding biological properties of human-based PL as an efficient engineered ECM which can be tailored to trigger specific regenerative pathways in minimal invasive strategies.

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