Macroporous chitosan/alginate hydrogels crosslinked with genipin accumulate and retain glioblastoma cancer cells

Abstract

Grade IV multiforme glioblastoma (GBM) is an aggressive cancer that remains incurable due to the GBM cells invading and proliferating in the surrounding healthy tissues, even after tumor resection. A new therapeutic paradigm to treat GBM is to attract and accumulate GBM cells in a macroporous hydrogel inserted in the surgical cavity after tumor resection, followed by a targeted high dose of radiotherapy. This work presents a molding-based method to prepare macroporous hydrogels composed of sodium alginate and chitosan, homogeneously mixed in solution using sodium bicarbonate, and subsequently crosslinked with genipin and calcium chloride. The gels display a blue color, the result of chitosan crosslinking with genipin, fully interconnected pores with an average diameter of 180 μm (and tunable over a wide range), with a compression modulus of 10 kPa, close to the value of brain tissues. The gels are stable in cell culture media and keep their integrity after radiation doses comparable to current GBM treatment levels. Finally, F98 GBM cells accumulate relatively homogeneously and are retained within the gels.

Compression tests were performed with a Mach-1 mechanical tester (Biomomentum, Laval, Canada, Actuator A400.25, load cell of 150 g, 10 mm diameter indenter) on bulk and porous gels. The indenter diameter was chosen to be slightly larger than the diameter of the gels. In addition to the porous gels, bulk gels (without pores) were cut with a 5 mm diameter biopsy punch. All gels were fully covered with water in the instrument testing cell to avoid the effects of capillary forces. Unconstrained compression tests were carried out in two steps: (1) the indenter established contact with the gel (speed = 25 μm s−1, stop criteria: 0.25 g), at the end of which the testing cell reservoir was filled with water; (2) a first pre-compression (displacement of 50 μm) at the same speed was applied to make sure that the probe was fully in contact with the gel surface, followed by two compressions, each totaling a 25 μm displacement at 20 μm s−1 on the gels. The compression modulus was calculated as follow: the Mach-1 software records the displacement Δh in μm and the applied load G in kg at an acquisition rate of 0.1 s^-1.