Carefully designed micro-and nanocarriers can provide significant advantages over conventional macroscopic counterparts in biomedical applications. The set of requirements including a high loading capacity, triggered release mechanisms, biocompatibility, and biodegradability should be considered for the successful delivery realization. Porous calcium carbonate (CaCO3) is one of the most promising platforms, which can encompass all the beforehand mentioned requirements. Here, we study both the particles formation and biological applicability of CaCO3. In particular, anisotropic differently shaped CaCO3 particles were synthesized using green sustainable approach based on co-precipitation of calcium chloride and sodium carbonate/bicarbonate at different ratios in the presence of organic additives. The impact of salts concentrations, reaction time, as well as organic additives was systematically researched to achieve controllable and reliable design of CaCO3 particles. It has been demonstrated that the crystallinity (vaterite or calcite phase) of particles depends on the initial salts' concentrations. The loading capacity of prepared CaCO3 particles is determined by their surface properties such as specific surface area, pore size and zetapotential. Differently shaped CaCO3 particles (spheroids, ellipsoids, toroids) were used to evaluate their uptake efficiency on the example of C6 glioma cells. The results show that the ellipsoidal particles possess a higher probability for internalization by cancer cells. All tested particles were also found to have a good biocompatibility. The capability to design physicochemical properties of CaCO3 particles has a significant impact on drug delivery applications, since the particles geometry substantially affects cell behavior (internalization, toxicity) and allows outperforming standard spherical counterparts.

CRISPR-Cas9 is a revolutionary genome-editing technology that has enormous potential for the treatment of genetic diseases. However, the lack of efficient and safe, non-viral delivery systems has hindered its clinical application. Here, we report on the application of polymeric and hybrid microcarriers, made of degradable polymers such as polypeptides and polysaccharides and modified by silica shell, for delivery of all CRISPR-Cas9 components. We found that these microcarriers mediate more efficient transfection than a commercially available liposome-based transfection reagent (>70% vs. <50% for mRNA, >40% vs. 20% for plasmid DNA). For proof-of-concept, we delivered CRISPR-Cas9 components using our capsules to dTomato-expressing HEK293T cells-a model, in which loss of red fluorescence indicates successful gene editing. Notably, transfection of indicator cells translated in high-level dTomato knockout in approx. 70% of transfected cells. In conclusion, we have provided proof-of-principle that our micro-sized containers represent promising non-viral platforms for efficient and safe gene editing.

The incorporation of bioactive compounds onto polymer fibrous scaffolds with further control of drug release kinetics is essential to improve the functionality of scaffolds for personalized drug therapy and regenerative medicine. In this study, polymer and hybrid microcapsules were prepared and used as drug carriers, which are further deposited onto polymer microfiber scaffolds [polycaprolactone (PCL), poly(3-hydroxybutyrate) (PHB), and PHB doping with the conductive polyaniline (PANi) of 2 wt % (PHB-PANi)]. The number of immobilized microcapsules decreased with increase in their ζ-potential due to electrostatic repulsion with the negatively charged fiber surface, depending on the polymer used for the scaffold's fabrication. Additionally, the immobilization of the capsules in dynamic mechanical conditions at a frequency of 10 Hz resulted in an increase in the number of the capsules on the fibers with increase in the scaffold piezoelectric response in the order PCL < PHB < PHB-PANi, depending on the chemical composition of the capsules. The immobilization of microcapsules loaded with different bioactive molecules onto the scaffold surface enabled multimodal triggering by physical (ultrasound, laser radiation) and biological (enzymatic treatment) stimuli, providing controllable release of the cargo from scaffolds. Importantly, the microcapsules immobilized onto the surface of the scaffolds did not influence the cell growth, viability, and cell proliferation on the scaffolds. Moreover, the attachment of human mesenchymal stem cells (hMSCs) on the scaffolds revealed that the PHB and PHB-PANi scaffolds promoted adhesion of hMSCs compared to that of the PCL scaffolds. Two bioactive compounds, antibiotic ceftriaxone sodium (CS) and osteogenic factor dexamethasone (DEXA), were chosen to load the microcapsules and demonstrate the antimicrobial properties and osteogenesis of the scaffolds. The modified scaffolds had prolonged release of CS or DEXA, which provided an improved antimicrobial effect, as well as enhanced osteogenic differentiation and mineralization of the scaffolds modified with capsules compared to that of individual scaffolds soaked in CS solution or incubated in an osteogenic medium. Thus, the immobilization of microcapsules provides a simple, convenient way to incorporate bioactive compounds onto polymer scaffolds, which makes these multimodal materials suitable for personalized drug therapy and bone tissue engineering.