Institute of Fundamental Technological Research

Current treatments of degenerated intervertebral discs often provide only temporary relief or address specific causes, necessitating the exploration of alternative therapies. Cell-based regenerative approaches showed promise in many clinical trials, but
limitations such as cell death during injection and a harsh disk environment hinder their effectiveness. Injectable microscaffolds offer a solution by providing a supportive microenvironment for cell delivery and enhancing bioactivity. This study evaluated the
safety and feasibility of electrospun nanofibrous microscaffolds modified with chitosan (CH) and chondroitin sulfate (CS) for treating degenerated NP tissue in a large animal model. The microscaffolds facilitated cell attachment and acted as an effective delivery system, preventing cell leakage under a high disc pressure. Combining microscaffolds with bone marrow-derived mesenchymal stromal cells demonstrated no cytotoxic effects and proliferation over the entire microscaffolds. The administration of cells attached to microscaffolds into the NP positively influenced the regeneration process of the intervertebral disc. Injectable poly(L-lactide-co-glycolide) and poly(L-lactide) microscaffolds enriched with CH or CS, having a fibrous structure, showed the potential to promote intervertebral disc regeneration. These features collectively address critical challenges in the fields of tissue engineering and regenerative medicine, particularly in the context of intervertebral disc degeneration.

microscaffolds,cell carriers,injectable biomaterials,intervertebral disc,laser micromachining,electrospinning

The use of injectable biomaterials for cell delivery is a rapidly expanding field which may revolutionize the medical treatments by making them less invasive. However, creating desirable cell carriers poses significant challenges to the clinical implementation of cell-based therapeutics. At the same time, no method has been developed to produce injectable microscaffolds (MSs) from electrospun materials. Here the fabrication of injectable electrospun nanofibers is reported on, which retain their fibrous structure to mimic the extracellular matrix. The laser-assisted micro-scaffold fabrication has produced tens of thousands of MSs in a short time. An efficient attachment of cells to the surface and their proliferation is observed, creating cell-populated MSs. The cytocompatibility assays proved their biocompatibility, safety, and potential as cell carriers. Ex vivo results with the use of bone and cartilage tissues proved that NaOH hydrolyzed and chitosan functionalized MSs are compatible with living tissues and readily populated with cells. Injectability studies of MSs showed a high injectability rate, while at the same time, the force needed to eject the load is no higher than 25 N. In the future, the produced MSs may be studied more in-depth as cell carriers in minimally invasive cell therapies and 3D bioprinting applications.

Intrinsically conducting polymers (ICPs) are widely used to fabricate biomaterials; their application in neural tissue engineering, however, is severely limited because of their hydrophobicity and insufficient mechanical properties. For these reasons, soft conductive polymer hydrogels (CPHs) are recently developed, resulting in a water-based system with tissue-like mechanical, biological, and electrical properties. The strategy of incorporating ICPs as a conductive component into CPHs is recently explored by synthesizing the hydrogel around ICP chains, thus forming a semi-interpenetrating polymer network (semi-IPN). In this work, a novel conductive semi-IPN hydrogel is designed and synthesized. The hybrid hydrogel is based on a poly(N-isopropylacrylamide-co-N-isopropylmethacrylamide) hydrogel where polythiophene is introduced as an ICP to provide the system with good electrical properties. The fabrication of the hybrid hydrogel in an aqueous medium is made possible by modifying and synthesizing the monomers of polythiophene to ensure water solubility. The morphological, chemical, thermal, electrical, electrochemical, and mechanical properties of semi-IPNs were fully investigated. Additionally, the biological response of neural progenitor cells and mesenchymal stem cells in contact with the conductive semi-IPN was evaluated in terms of neural differentiation and proliferation. Lastly, the potential of the hydrogel solution as a 3D printing ink was evaluated through the 3D laser printing method. The presented results revealed that the proposed 3D printable conductive semi-IPN system is a good candidate as a scaffold for neural tissue applications.

Intervertebral disc diseases are a significant medical problem affecting many people around the world. In Poland, the statistics of the Social Insurance Institution (Medical Abuse in 2016) indicate that low back pains and other intervertebral disc diseases constitute 17% of the total number of days of sick leave. In connection with the above, current work describes design of a composite scaffold as a carrier in cell therapy, which will contribute to the regeneration of the intervertebral disc, including the increase of its height. Our composite scaffold include nanofibers that were prepared with the use of the electrospinning method. This method is a simple but powerful technique for fabricating desirable nano- and microfibers by using a high potential electric field. Human Mesenchymal stem cells (MSCs) were cultured on the scaffold from poly(L-lactide). Proliferation kits and fluorescence microscopy were used to asses cells’ viability and adherence to the nanofibers’ surface. hMSCs were efficiently cultured on the nanofibrous scaffold. Cells could be readily detected in porous structure of the scaffold after 7 and 14 days of culture. Viability and proliferation kits proved that the material is not toxic. Drug release from nanofibrous material of model growth factor was conducted with pharmacopeia protocols. Drug release of the 14 kDa growth factor was achieved for 14 days without burst release. Nanofibrous biomaterials prove their advances in many tissue engineering applications. Adjustable porosity of the scaffold and the biocompability of biomaterial make it perfect candidate for cells’ scaffold in many medical procedures and also as a drug release carrier. With the use of single nanofibers, such biomaterials can also be readily used in minimally invasive procedures to regenerate IVD.

nanofibers, IVD, MSC