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It is of high clinical relevance in bone tissue engineering that scaffolds promote a high seeding efficiency of cells capable of osteogenic differentiation, such as human bone marrow-derived mesenchymal stem cells (hMSCs). We evaluated the effects of a novel polycaprolactone (PCL) scaffold on hMSC seeding efficiency, proliferation, distribution and differentiation. Porous PCL meshes prepared by fused deposition modeling (FDM) were embedded in matrix of hyaluronic acid, methylated collagen and terpolymer via polyelectrolyte complex coacervation. Scaffolds were cultured statically and dynamically in osteogenic stimulation medium for up to 28 days. Compared to naked PCL scaffolds, embedded scaffolds provided a higher cell seeding efficiency (t-test, P<0.05), a more homogeneous cell distribution and more osteogenically differentiated cells, verified by a more pronounced gene expression of the bone markers alkaline phosphatase, osteocalcin, bone sialoprotein I and bone sialoprotein II. Dynamic culture resulted in higher amounts of DNA (day 14 and day 21) and calcium (day 21 and day 28), compared to static culture. Dynamic culture and the embedding synergistically enhanced the calcium deposition of hMSC on day 21 and day 28. This in vitro study provides evidence that hybrid scaffolds made from natural and synthetic polymers improve cellular seeding efficiency, proliferation, distribution and osteogenic differentiation.

Microstructured 20 μm thick polymer filaments used as nerve implants were loaded with chitosan/siRNA nanoparticles to promote nerve regeneration and ensure local delivery of nanotherapeutics. The stable nanoparticles were rapidly internalized by cells and did not affect cell viability. Target mRNA was successfully reduced by 65-75% and neurite outgrowth was enhanced even in an inhibitory environment. This work, thus, supports the application of nanobiofunctionalized implants as a novel approach for spinal cord and nerve repair.

Harnessing the RNA interference pathway offers a new therapeutic modality; however, solutions to overcome biological barriers to small interfering RNA (siRNA) delivery are required for clinical translation. This work demonstrates, by direct northern and quantitative PCR (qPCR) detection, stability, gastrointestinal (GI) deposition, and translocation into peripheral tissue of nonmodified siRNA after oral gavage of chitosan/siRNA nanoparticles in mice. In contrast to naked siRNA, retained structural integrity and deposition in the stomach, proximal and distal small intestine, and colon was observed at 1 and 5 hours for siRNA within nanoparticles. Furthermore, histological detection of fluorescent siRNA at the apical regions of the intestinal epithelium suggests mucoadhesion provided by chitosan. Detection of intact siRNA in the liver, spleen, and kidney was observed 1 hour after oral gavage, with an organ distribution pattern influenced by nanoparticle N:P ratio that could reflect differences in particle stability. This proof-of-concept work presents an oral delivery platform that could have the potential to treat local and systemic disorders by siRNA.

RNAi-based strategies provide a great therapeutic potential for treatment of various human diseases including kidney disorders, but face the challenge of in vivo delivery and specific targeting. The chitosan delivery system has previously been shown to target siRNA specifically to the kidneys in mice when administered intravenously. Here we confirm by 2D and 3D bioimaging that chitosan formulated siRNA is retained in the kidney for more than 48 hours where it accumulates in proximal tubule epithelial cells (PTECs), a process that was strongly dependent on the molecular weight of chitosan. Chitosan/siRNA nanoparticles, administered to chimeric mice with conditional knockout of the megalin gene, distributed almost exclusively in cells that expressed megalin, implying that the chitosan/siRNA particle uptake was mediated by a megalin-dependent endocytotic pathway. Knockdown of the water channel aquaporin 1 (AQP1) by up to 50% in PTECs was achieved utilizing the systemic i.v. delivery of chitosan/AQP1 siRNA in mice. In conclusion, specific targeting PTECs with the chitosan nanoparticle system may prove to be a useful strategy for knockdown of specific genes in PTECs, and provides a potential therapeutic strategy for treating various kidney diseases.

Chitosan is a natural cationic copolymer of N-acetyl glucosamine and D-glucosamine, varying in composition, sequence and molecular chain length. Because of its bio- and cyto-compatibility, biodegradability and bioresorbability, chitosan has been investigated for application in various biomedical fields such as drug and gene delivery, tissue engineering, wound healing, and for use in antimicrobial, antiviral and immunoadjuvant strategies. With the rise of nanotechnology, chitosan together with bioactive nanoparticles are fabricated into various bionanocomposites, providing alternatives to new era of regenerative medicine and drug delivery vesicles. The present paper will review the preparations and biomedical applications of such chitosan composites, their current achievements, limitations and future perspectives. In this respect, the effect of chitosan properties on the interaction with nanoparticles and its consequences for applicability of the resulting composites will be discussed.