(353g) Physically Crosslinked DNA-Based Injectable Hydrogels for Bone Regeneration

Natural polymers such as alginate, hyaluronic acid, chitosan, gelatin or collagen have been extensively investigated for the preparation of injectable hydrogels due to their higher biocompatibility when compared to synthetic polymers. However, in most of the cases, there is a need for an extra functionalization step or the addition of a second polymer to introduce a reactive building block for physical crosslinking. A more advanced strategy to produce injectable hydrogel is by selection of polymers that can self-assemble into defined structures due to the presence of specific molecular recognition motifs. DNA is a natural biopolymer and there is immense potential for treating DNA as a chemical entity instead of a genetic material. DNA-based materials are enriched with inherent qualities such as biocompatibility, programmable sequences and precise recognition properties. The underlying chemical nature of the double stranded DNA facilitates for chemical and physical bonding to other bioactive substances. Moreover, the base-pairing interactions leading to the secondary double stranded structure is highly specific. Given these features, DNA can prove to be an attractive starting material for the fabrication of multifunctional hydrogels and scaffolds for tissue engineering and controllable drug delivery. Herein, we report the first example of an injectable hydrogel with sustained release properties, based on DNA and silicate nanodisks (nSi), that can be prepared by a facile technique of heating and mixing. The proposed strategy presents the advantage of developing physically crosslinked networks without the need of chemical modification of the DNA backbone or design of specific DNA base sequences. A fundamental structural characteristic of DNA is the specific base pairing interactions. These interactions were exploited in the first step of thermal gelation, by promoting the base-pairing among adjacent strands of DNA. As the first step, a DNA solution 4% (w/v) in water was heated at 90°C for 45 seconds followed by a cooling step at 37°C. This process leads to the formation of an interconnected structure, termed as the pre-gel. Thereafter, nSi dispersion was added to the pre-gel at room temperature followed by vigorous vortexing. Nanocomposite hydrogels were developed consisting of 3.5% DNA (w/v) and three different nSi concentrations ranging from 0.1% to 0.5% (w/v). The DNA system without any silicate possesses a highly porous interconnected structure as observed by scanning electron microscopy (SEM). The inclusion of nSi caused a reduction in the porosity of the hydrogel, indicating a more compact network with a higher degree of physical crosslinking. Silicate nanodisks electrostatically interact with the negatively charged phosphate backbone of DNA, thereby enhancing the structural integrity and elasticity. The formulated hydrogels could also function as sustained drug delivery vehicles. We tested the abilityof the nanocomposite hydrogel to modulate the release of a model osteoinductive drug, dexamethasone (Dex), for promoting differentiation of human adipose stem cells (hASCs) to osteogenic lineage. The released drug maintained its activity and modulated gene expression in hASCs. Moreover, the encapsulation of Dex within DNA nanocomposite hydrogels allowed a higher retention of Dex, resulting in significantly increased alkaline phosphatase activity, calcium deposition, and alkaline phosphatase gene expression of hASCs. Overall, the nanoengineered injectable hydrogel obtained from physically crosslinked network is mechanically resilient and can be used for biomedical applications such as bone regeneration, bioactive coatings and therapeutic delivery.