Full length articleGold nanoparticles for the in situ polymerization of near-infrared responsive hydrogels based on fibrin
Graphical abstract
Introduction
Fibrin, a water insoluble, degradable naturally-occurring biopolymer generated from fibrinogen, is involved in the last step of the blood coagulation cascade as major component of the clot [1]. Fibrin clot formation is catalyzed by thrombin, a member of the serine protease family that removes fibrinopeptides A and B from fibrinogen to produce fibrin monomer. Noncovalent interactions of fibrin monomers result in intermediate polymers which aggregate to form the fibrin clot [2]. Fibrin plays an important and ubiquitous role during tissue repair serving as provisional extracellular matrix for infiltrating cells and acting as a reservoir for growth factors [3]. As biomaterial for tissue repair and regeneration, fibrin shows attractive features over synthetic polymers, often limited by biocompatibility concerns, their inability to support cell attachment, undesirable degradation rate and toxic degradation products, potential immune response and acute inflammation [4,5]. Key advantages of fibrin include ease of purification of fibrinogen from donors, possibility of recombinant production, high tunability of the matrix architecture and mechanical properties by controlling the conditions of fibrin gelation, in situ curability, biodegradability, and minimal inflammation or foreign body reaction. These features support the clinical interest of products based on fibrin to be used as sealants or bioadhesives in surgical procedures [6], [7], [8], [9], cellular scaffolds for reconstituting full thickness wounds [4,10] or carriers for the delivery of drugs, genes and growth factors [11]. As scaffold, fibrin has been employed for supporting pre-vascularized tissues in the treatment of ischemic strokes [12]. Thus, fibrin patches containing endothelial cells and smooth muscle cells derived from human embryonic stem cells were implanted in porcine infarcted hearts showing significant engraftment and functional improvement [13]. Also, composite materials based on fibrin and other biopolymers have improved myocardial function in several animal models [14,15]. Encouraging outcomes have been reported from a first clinical trial of transplantation of cardioprogenitor cells incorporated in fibrin patches in patients with severely impaired cardiac function [16,17]. Fibrin scaffolds have also been employed as three-dimensional matrices containing growth factors and embryonic stem cells prone to differentiate into neurons in animal models of spinal cord injury [18]. Tendon regeneration and bone-tendon junction repair have also been accelerated in animal models after the application of mesenchymal stem cells engrafted in fibrin matrices [19]. Accelerated healing of acute and chronic non-healing cutaneous wounds has been achieved in patients treated with fibrin-based sprays containing autologous mesenchymal stem cells [20]. Commercial available scaffolds based on fibrin hydrogels (i.e. BioseedⓇ-C, BioTissue Technologies or CHONDRON™, Sewon Cellontech) have been successfully used in combination with autologous cells in articular cartilage regeneration of knee [21,22].
The so-called “smart” hydrogels, i.e. those which have the ability to respond to environmental changes in temperature, pH, light, magnetic and electric fields, ionic strength, mechanical load, enzymatic activity, antigen concentration, etc., are among the leading candidates to provide controlled delivery of therapeutic agents in regenerative medicine applications [23]. Photothermal heating induced by NIR irradiation has shown great potential as an external stimulus to regulate the activity of “smart” hydrogels. For example, NIR has been used to trigger on-demand cell release upon irradiation of composite hydrogels based in graphene oxide and poly(N-isopropylacrylamide) or to trigger sol-gel transitions in gold nanorod-filled polypeptide hydrogels to modulate drug release [24,25]. Recently, we developed a technological approach which employs “smart” hydrogels based on fibrin to define spatiotemporal patterns of transgene expression in response to incident NIR light [26]. Briefly, during fibrin polymerization catalyzed by soluble thrombin, we included HGNP tailored to show a surface plasmon resonance absorption in the NIR region. These plasmonic nanoparticles (NP) act as phototransducers that convert incident NIR light into heat, increasing the temperature of the surrounding region. The distance over which thermal diffusion and temperature rise occur depend on the exposure time and the thermal diffusion time, defined as the ratio of the thermal conductivity of the irradiated material to the product of the heat capacity and density [27]. Therefore, thermal diffusion distance from the nanoparticulated absorber will depend on the laser operation mode, continuous or pulsed, having longer heat diffusion distances when using continuous lasers. In the NIR region, light scattering prevails over absorption, and light penetrates underneath the subcutaneous tissue at depths that depend on the nature of the irradiated tissue [28]. Thus, NIR light can be used as a non-invasive trigger for activating internally implanted absorbers and generate local hyperthermia. We used these fibrin-based plasmonic hydrogels as scaffolds for genetically modified cells that harbour a heat-activated and ligand-dependent gene circuit that regulates the expression of transgenic growth factors. This regulatory circuit is based on the promoter of the highly heat-inducible HSPA7 gene [29], which drives the expression of a ligand-dependent chimeric transactivator that activates a promoter to which the transgene of interest is linked. Irradiation of photothermal scaffolds with NIR laser can increase the local temperature to trigger, in the presence of ligand, the gene circuit. After implanting NIR-responsive scaffolds populated by stem cells which harbour the gene circuit, transgenic expression of growth factors such as VEGF can be remotely patterned in the host using NIR light [26]. In the present work, we used a covalent coupling to coat HGNP with poly-L-lysine (PLL) through the use of COOH-poly(ethylene glycol)-SH as a linker (COOH-PEG-SH). The thiol group of the heterobifunctional PEG strongly binds to the gold surface while its carboxyl group binds to the amino groups in PLL via amide bond. These functionalized HGNP can efficiently load thrombin, enabling them to polymerize fibrin in situ by mixing the resulting NP and fibrinogen, and therefore simplifying the process of generating plasmonic fibrin matrices. The structural, mechanical and photothermal properties of these matrices were compared to those reported in our previous study [26], which were prepared using three independent components (fibrinogen, HGNP and soluble thrombin) and were used as control in the present study. Interestingly, we found that the metallic core of thrombin-loaded HGNP fragmentates in aqueous medium. This finding represents an exciting advantage for the clinical use of gold nanostructures, whose non-biodegradability has raised concerns regarding long-term toxicity [30], [31], [32].
Section snippets
Nanoparticle synthesis and characterization
HGNP were prepared following a new variant from a previously reported method [33]. Briefly, 800 µL of 0.4 M cobalt chloride hexahydrate and 1.6 mL of 0.1 M sodium citrate dihydrate were mixed in a round-bottom flask with 400 mL of deionized water. The solution was de-aired by bubbling with argon for 45 min. Then 4 mL of a 1 wt% solution of poly(vinylpyrrolidone) (PVP; MW = 55 KDa) and 900 µL of 0.1 M sodium borohydride were injected under magnetic stirring. The solution turned from pale pink to
Characterization of HGNPT
Due to electrostatic repulsive forces, the direct coupling between negatively charged HGNP and thrombin cannot be achieved directly. In a previous work, we reversed the electrokinetic charge of HGNP thought a conjugation strategy that involved a cationic polymer to promote supramolecular electrostatic interactions. Layer-by-layer surface functionalization of HGNP using positively charged poly(allylamine hydrochloride) resulted in a high local concentration of amines which led to the drastic
Conclusion
The functionalization of HGNP surface with PLL through the use of heterobifunctional PEG as a linker allowed the efficient grafting of thrombin onto the plasmonic nanomaterial. The surface modification of HGNP did not change significantly the resonance plasmon in the NIR region but fostered the destabilization of the metallic shell at body temperature. Thrombin-conjugated HGNP released protease content to the aqueous phase when dispersed in cell culture medium, which induced the cleavage of
Acknowledgments
The authors thank to D. Morales (Confocal Microscopy Laboratory, Universidad Autónoma de Madrid, Spain) for excellent technical assistance. This work was supported by grant PI15/01118 from Instituto de Salud Carlos III (ISCIII)-Fondos FEDER, Ministry of Economy and Competitiveness (MINECO), Spain, grants RTI2018-095159-B-I00 and SAF2013-50364-EXP from MINECO, grant Roche-IdiPAZ from the intramural funding program of Foundation for Biomedical Research of La Paz University Hospital-IdiPAZ, grant
Disclosure
The work described herein was partially supported by contracts from HSF Pharmaceuticals S.A. to N.V.
References (52)
Thrombin generation and fibrin clot structure
Blood Rev.
(2007)Fibrinogen and fibrin structure and functions
J. Thrombosis Haemostasis JTH
(2005)- et al.
Fibrin microbeads (FMB) as biodegradable carriers for culturing cells and for accelerating wound healing
J. Invest. Dermatol.
(1999) - et al.
Elastic sealants for surgical applications
Eur. J. Pharm. Biopharm. Offic. J. Arbeitsgemeinschaft Pharmazeutische Verfahrenstechnik eV
(2015) - et al.
Biomaterials for the treatment of myocardial infarction: a 5-year update
J. Am. Coll. Cardiol.
(2011) - et al.
Cell delivery: intramyocardial injections or epicardial deposition? a head-to-head comparison
Ann. Thorac. Surg.
(2009) - et al.
Cell- and gene-based approaches to tendon regeneration
J. Should. Elbow Surg.
(2012) - et al.
Temporal and spatial patterning of transgene expression by near-infrared irradiation
Biomaterials
(2014) - et al.
High-level, heat-regulated synthesis of proteins in eukaryotic cells
Gene
(1986) - et al.
Biodistribution of gold nanoparticles and gene expression changes in the liver and spleen after intravenous administration in rats
Biomaterials
(2010)
Protracted elimination of gold nanoparticles from mouse liver
Nanomed. Nanotechnol. Biol. Med.
Purification, characterization and insolubilization of bovine thrombin
Thromb. Res.
A negative staining method for high resolution electron microscopy of viruses
Biochim. Biophys. Acta
Computer modeling of fibrin polymerization kinetics correlated with electron microscope and turbidity observations: clot structure and assembly are kinetically controlled
Biophys. J.
Native fibrin gel networks observed by 3D microscopy, permeation and turbidity
Biochim. Biophys. Acta
Gelation dynamics and gel structure of fibrinogen
Colloids Surf. B Biointerfaces
Fibrinopeptides and fibrin gel structure
Biophys. Chem.
Role of pore size and morphology in musculo-skeletal tissue regeneration
Mater. Sci. Eng. C Mater. Biol. Appl.
Naturally and synthetic smart composite biomaterials for tissue regeneration
Adv. Drug Deliv. Rev.
Pro-angiogenic near infrared-responsive hydrogels for deliberate transgene expression
Acta Biomater.
Roles for thrombin and fibrin(ogen) in cytokine/chemokine production and macrophage adhesion in vivo
Blood
Fibrin clots are equilibrium polymers that can be remodeled without proteolytic digestion
Sci. Rep.
Biodegradable synthetic polymers for tissue engineering
Eur. Cell Mater.
Role of fibrin sealants in liver surgery
Dig. Surg.
Reduction of intraoperative air leaks with progel in pulmonary resection: a comprehensive review
J. Cardiothorac Surg.
Fibrin adhesive is better than sutures in pterygium surgery
Cornea
Cited by (10)
Cationic Cu(I)-covalent organic framework as self-enhanced synergetic Photothermal/Phtodynamic/Cationic/Enzymatic antibacterial agent
2023, Reactive and Functional PolymersRemote control of transgene expression using noninvasive near-infrared irradiation
2023, Journal of Photochemistry and Photobiology B: BiologyLight activated pulsatile drug delivery for prolonged peripheral nerve block
2022, BiomaterialsCitation Excerpt :In this work, a new triggerable drug delivery system (POEGMA@HGNPs NGs) based on a thermoresponsive copolymer nanogel (poly(oligo(ethylene glycol) methyl ether methacrylate (POEGMA) NG) decorated with hollow gold nanoparticles (HGNPs) and loaded with bupivacaine (BPV) has been developed. These gold nanoparticles, with plasmonic absorption in the near-infrared (NIR) region, induced photothermal heating which was able to produce the collapse of the polymeric nanogel only when illuminated, promoting the release of the drug on-demand [18,19]. BPV was chosen as anesthetic drug model since it is indicated for local infiltration, peripheral nerve block, sympathetic nerve block, and epidural and caudal blocks [20,21].
Local delivery of bone morphogenetic protein-2 from near infrared-responsive hydrogels for bone tissue regeneration
2020, BiomaterialsCitation Excerpt :Data herein show that same approach can be used to control the secretion of BMP-2 in bone defects. In our previous studies, we explored plasmonic fibrin scaffolds polymerized using 10 mg mL−1 fibrinogen and 2 U ml−1 thrombin, which provided a suitable environment for the incorporation of cells harbouring a heat-activated and dimerizer-dependent gene switch [32,37,51]. In the present study, we increased fibrinogen concentration to 20 mg mL−1 to generate a fibrin scaffold, aimed to bone repair, with improved structural and mechanical properties while allowing cellular functions.
An Intrinsic Photothermal Supramolecular Hydrogel with Robust Mechanical Strength and NIR-Responsive Shape Memory
2024, Macromolecular Rapid CommunicationsNanomedical research and development in Spain: improving the treatment of diseases from the nanoscale
2023, Frontiers in Bioengineering and Biotechnology
- 1
These authors contributed equally to this work.