Search

International audience; Amorphous calcium carbonate nanoparticles (ACC NPs) are promising multifunctional materials for healthcare applications. Due to their instability in aqueous media, pure ACC NPs for the biomedical field are increasingly synthesised in absolute ethanol, using the ammonia diffusion method (ADM). Although this method presents the advantage of providing stable ACC NPs without additives, it requires the use of pure ethanol as solvent. New insights into the formation mechanisms of ACC NPs in ethanol using gas diffusion are presented in this article. The optimisation of the process according to these findings can increase the mass concentration of ACC NPs by a factor of 3.5. As a result, the amount of ethanol required to produce a target mass of particles is significantly decreased, reducing the ecological impact of the process. The stabilisation of the resulting ACC NPs in aqueous media is achieved by a short-time process using phospholipids based on the ethanol injection method. By using the natural electrostatic affinity of negatively charged materials for the positive surface of ACC NPs in ethanol, we reduced the process time from 24 h to 2 minutes, compared with the closest state of the art, decreasing the operating time and corresponding energy consumption. The process does not require the use of synthetic PEGylated lipids for steric stabilisation. In addition, a natural egg-sourced phospholipid was identified as an efficient stabiliser for the first time. The upscaling of our process was successfully demonstrated using a 50 L reactor for bulk synthesis, as well as a continuous flow reactor for industrial continuous flow production.

Dentoalveolar tissue engineering is an emerging yet challenging field, considering the lack of suitable materials and difficulty to produce patient-specific hydrogel scaffolds. The present paper aims to produce a 3D printable and tuneable biomaterial by copolymerizing a synthesized water-soluble chitosan derivative called maleic anhydride grafted chitosan (MA-C) with gelatin using genipin, a natural crosslinking agent. Development and testing of this material for 3D printing, degradation, and swelling demonstrated the ability to fabricate scaffolds with controlled physical properties based on pre-determined designs. The MA-C-gelatin copolymer demonstrated excellent biocompatibility, which was verified by analyzing the viability, growth and proliferation of human dental pulp stem cells seeded on MA-C-gelatin constructs through live/dead, alamar blue and DNA quantification assays. Based on the present findings, the proposed material might be a suitable candidate for dentoalveolar tissue engineering, while further research is required to achieve this goal.

Methacryloyl gelatin (GelMA) is a versatile material for bioprinting because of its tunable physical properties and inherent bioactivity. Bioprinting of GelMA is often met with challenges such as lower viscosity of GelMA inks due to higher methacryloyl substitution and longer physical gelation time at room temperature. In this study, a tunable interpenetrating polymer network (IPN) hydrogel was prepared from gelatin-hyaluronan dialdehyde (Gel-HDA) Schiff’s polymer, and 100% methacrylamide substituted GelMA for biofabrication through extrusion based bioprinting. Temperature sweep rheology measurements show a higher sol-gel transition temperature for IPN (30 °C) compared to gold standard GelMA (27 °C). Furthermore, to determine the tunability of the IPN hydrogel, several IPN samples were prepared by combining different ratios of Gel-HDA and GelMA achieving a compressive modulus ranging from 20.6 ± 2.48 KPa to 116.7 ± 14.80 KPa. Our results showed that the mechanical properties and printability at room temperature could be tuned by adjusting the ratios of GelMA and Gel-HDA. To evaluate cell response to the material, MC3T3-E1 mouse pre-osteoblast cells were embedded in hydrogels and 3D-printed, demonstrating excellent cell viability and proliferation after 10 d of 3D in vitro culture, making the IPN an interesting bioink for the fabrication of 3D constructs for tissue engineering applications.

Cryogels, a subset of hydrogels, have recently drawn attention for cartilage tissue engineering due to its inherent microporous architecture and good mechanical properties. In this study a dual crosslinked pullulan-gelatin cryogel (PDAG) scaffold was synthesized by crosslinking gelatin with oxidized pullulan by Schiff's base reaction followed by cryogelation. Chondrocytes seeded within the PDAG scaffolds and cultured for 21 d

Cryogels, a subset of hydrogels, have recently drawn attention for cartilage tissue engineering due to its inherent microporous architecture and good mechanical properties. In this study a dual crosslinked pullulan–gelatin cryogel (PDAG) scaffold was synthesized by crosslinking gelatin with oxidized pullulan by Schiff’s base reaction followed by cryogelation. Chondrocytes seeded within the PDAG scaffolds and cultured for 21 d in vitro demonstrated enhanced cell proliferation, enhanced production of cartilage-specific extracellular matrix and up-regulated sulfated glycosaminoglycan without altering the articular chondrocyte phenotype. Quantitative reverse transcription-polymerase chain reaction-based gene expression studies, immunofluorescence, and histological studies demonstrated that the PDAG scaffold significantly enhanced the expression of chondrogenic marker genes such as type II collagen, aggrecan, and SOX9. Taken together, these results demonstrated that PDAG scaffold prepared by sequential Schiff’s base reaction and cryogelation would be a promising cell-responsive scaffold for cartilage tissue engineering applications.

The therapeutic efficacies of the majority of the biopharmaceutics were altered due to their short half-life and fast clearance from circulation. Recent studies revealed that fusion of biopharmaceutics with conformationally disordered polypeptide chains, produced by genetic methods (PAS and XTEN) can enhance the half-life of the therapeutic agent. Similar to polyethylene glycol, the genetically modified polypeptide alternatives such as PAS and XTEN are hydrophilic, nontoxic, nonimmunogenic, neutral polymers, and possess expanded hydrodynamic volume. In addition, they are biodegradable and exhibit very good stability in serum proteases while undergoing degradation upon accumulation in kidney counterparts. This chapter describes in detail the preparation of PAS and XTEN sequences and corresponding gene cassettes, the production of PAS fusion proteins, and their physicochemical and biological properties and applications.

Asymmetric flow field-flow fractionation (AF4) coupled with UV-Vis spectroscopy, multi-angle light scattering (MALS) and refractive index (RI) detection has been applied for the characterization of MIL-100(Fe) nanoMOFs (metal–organic frameworks) loaded with nucleoside reverse transcriptase inhibitor (NRTI) drugs for the first time. Empty nanoMOFs and nanoMOFs loaded with azidothymidine derivatives with three different degrees of phosphorylation were examined: azidothymidine (AZT, native drug), azidothymidine monophosphate (AZT-MP), and azidothymidine triphosphate (AZT-TP). The particle size distribution and the stability of the nanoparticles when interacting with drugs have been determined in a time frame of 24 h. Main achievements include detection of aggregate formation in an early stage and monitoring nanoMOF morphological changes as indicators of their interaction with guest molecules. AF4-MALS proved to be a useful methodology to analyze nanoparticles engineered for drug delivery applications and gave fundamental data on their size distribution and stability. [Figure not available: see fulltext.].

Versatile derivatization feasible for cardanol to generate various amphiphilic structure-directing molecules bestows this naturally occurring phenol a great promise in the field of biomedical research. This chapter discusses the use of cardanol and cardanol derivatives for cancer therapy, drug delivery, and imaging applications. Cardanol has proven anti-proliferative effects on cancer cells, which could help its derivatives to evolve as therapeutic alternatives for anticancer treatment. At the same time, the structure-directing capacity of cardanol derivatives has been successfully used to obtain various gels and nanoparticles for drug delivery and imaging applications. These findings are summarized in three sections namely cardanol as an anticancer drug, cardanol gels, and cardanol-based nanomorphologies for drug delivery applications. The chapter also discusses prospective biomedical applications of cardanol.

Metal–organic frameworks (MOFs) are coordination polymers of interest for biomedical applications. Of particular importance, nanoparticles made of iron(III) trimesate (MIL-100, MIL standing for Material Institut Lavoisier) (nanoMOFs) can be conveniently synthesised under mild and green conditions. They were shown to be biodegradable, biocompatible and efficient to encapsulate a variety of active molecules. We have addressed here the challenges to encapsulate a highly hydrophilic anticancer prodrug, phosphated gemcitabin (Gem-MP) known for its instability and inability to bypass cell membranes. MIL-100 nanoMOFs acted as efficient “nanosponges”, soaking Gem-MP from its aqueous solution with almost perfect efficiency (>98%). Maximal loadings reached ∼30 wt% reflecting the strong interaction between the drug and the iron trimesate matrices. Neither degradation nor loss of crystalline structure was observed after the loading process. Storage of the loaded nanoMOFs in water did not result in drug release over three days. However, Gem-MP was released in media containing phosphates, as a consequence to particle degradation. Drug-loaded nanoMOFs were effective against pancreatic PANC-1 cells, in contrast to free drug and empty nanoMOFs. However, an efflux phenomenon could contribute to reduce the efficacy of the nanocarriers. Size optimization and surface modification of the nanoMOFs are expected to further improve these findings.

Doxorubicin (DOX) entrapment in porous Fe(III)-trimesate metal organic frameworks (MIL-100(Fe)) nanoparticles was investigated in neutral Tris buffer via UV-vis absorption, circular dichroism (CD), and fluorescence. The binding constants and the absolute spectra of the DOX-MIL-100(Fe) complexes were determined via absorption and fluorescence titrations. A binding model where DOX associates as monomer to the dehydrated Fe3O (OH)(H2O)2 [(C6H3)(CO2)3]2 structural unit in 1:1 stoichiometry, with apparent association constant of (1.1 to 1.8) × 10(4) M(-1), was found to reasonably fit the experimental data. Spectroscopic data indicate that DOX binding occurs via the formation of highly stable coordination bonds between one or both deprotonated hydroxyl groups of the aglycone moiety and coordinatively unsaturated Fe(III) centers. Complete quenching of the DOX fluorescence and remarkable thermal and photochemical stability were observed for DOX incorporated in the MIL-100(Fe) framework.

A nitro-benzofurazan-triazolyl carboxymethylated β-cyclodextrin (NBFT-CMβCyD) and an oligomer of carboxymethyl β-cyclodextrin (sodium salt), crosslinked with epichlorohydrin and labeled with rhodaminyl groups (pβCyD-Rh), exhibit very high affinity in aqueous solution for octacationic photosensitizer [(CH3)8LZn]8+ neutralized by I− ions (L = tetrakis-2,3-[5,6-di(2-(pyridiniumyl)pyrazino]porphyrazinato dianion). The photosensitizer (PS) forms complexes with 1 : 2 and 2 : 2 CyD:PS stoichiometry, which were characterized as binding constants and UV-Vis absorption and fluorescence properties. The self-association tendency of [(CH3)8LZn]8+, leading to a monomer–dimer equilibrium shift towards the dimer even at very low concentrations (≈10−6 M), is not contrasted by either NBFT-CMβCyD or the pβCyD-Rh oligomer, both of which completely convert the [(CH3)8LZn]8+ monomer fraction to the dimer form in the bound state. Quenching of fluorescence observed in [(CH3)8LZn]8+ upon binding with either hosts is consistent with the conversion of the monomer to the negligibly fluorescent dimer. The complexes formed with the CMβCyD units of the pβCyD-Rh oligomer have average association constants, which are larger by 6–7 orders of magnitude than those with the CMβCyD monomer in the NBFT-labeled derivative.

Two citric acid crosslinked gamma-cyclodextrin oligomers (p gamma-CyD) with a MW of 21-33 kDa and 10-15 gamma-CyD units per molecule were prepared by following green chemistry methods and were fully characterized. The non-covalent association of doxorubicin (DOX) with these macromolecules was investigated in neutral aqueous medium by means of circular dichroism (CD), UV-vis absorption and fluorescence. Global analysis of multiwavelength spectroscopic CD and fluorescence titration data, taking into account the DOX monomer-dimer equilibrium, evidenced the formation of 1 : 1 and 1 : 2 p gamma-CyD unit-DOX complexes. The binding constants are 1-2 orders of magnitude higher than those obtained for gamma-CyD and depend on the characteristics of the oligomer batch used. The concentration profiles of the species in solution evidence the progressive monomerization of DOX with increasing oligomer concentration. Confocal fluorescence imaging and spectral imaging showed a similar drug distribution within the MCF-7 cell line incubated with either DOX complexed to p gamma-CyD or free DOX. In both cases DOX is taken up into the cell nucleus without any degradation.

The association of doxorubicin (DOX) with gamma-cyclodextrin (gamma-CyD) was studied in phosphate buffer of pH 7.4, at 22 degrees C by performing titration experiments monitored with circular dichroism (CD), UV-vis absorption, and fluorescence. Global analysis of multiwavelength spectroscopic data obtained at different DOX concentrations was performed by taking into account the DOX monomer-dimer equilibrium. Formation of gamma-CyD: DOX 1 : 1, 2 : 1, 1 : 2 and 2 : 2 complexes was evidenced. The stability constants and the absolute CD, UV-vis absorption and fluorescence spectra of all the complexes were determined. gamma-CyD did not prove to be able to disrupt the DOX dimer when the latter is the predominant form in solution. The triplet state absorption and kinetic properties of DOX in the presence of gamma-CyD and in ethanol were also determined by laser flash photolysis. The excited singlet and the triplet features indicated the environment experienced by DOX in the CyD complexes is ethanol-like. The structure of the gamma-CyD: DOX 1 : 1 complex was investigated by Molecular Mechanics (MM) and Molecular Dynamics (MD) for both the neutral and the positively charged (-NH3+ in the daunosamine moiety) DOX forms. The possibility of interaction of both the aglycone and the daunosamine parts with the gamma-CyD cavity was evidenced.

The association of doxorubicin (DOX) and artemisinin (ART) to a beta-CyD-epichlorohydrin crosslinked polymer (p beta-CyD), organized in nanoparticles of ca. 15 nm size, was investigated in neutral aqueous medium by circular dichroism (CD), UV-vis absorption and fluorescence. The stability constants and the absolute CD spectra of the drug complexes were determined by global analysis of multiwavelength data from spectroscopic titrations. The polymer p beta-CyD proved able to disrupt the DOX dimer when the latter is the predominant form of DOX in solution. The spectroscopic and photophysical properties of the complexes evidenced an alcohol-like environment for ART and an improved inherent emission ability for DOX in the nanoparticle frame.