Abstract
Aims: The aim of the project was to verify whether high molecular weight hyaluronic acid (HA) could be used as a cryoprotective in the cryopreservation of dental pulp stem cells (DPSC) in an attempt to reduce the concentration of dimethylsulfoxide (DMSO). Conclusion: During the experiment we verified the possibility of using high molecular weight HA as a cryoprotective agent in an effort to reduce the concentration of cytotoxic DMSO.
During cryopreservation, DPSC are affected by the formation of intracellular and extracellular ice crystals. Using combination of high-molecular substances that do not penetrate the intracellular space is possible to prevent the formation of extracellular ice crystals.
In contrast, DMSO, as a low molecular weight substance, enters the interior of the cells and protects them from the formation of intracellular ice crystals. In addition, DMSO also affects the melting and freezing temperatures, thus preventing or limiting the formation of ice crystal seed centers.
When measuring viability of DPSC immediately after thawing, cells best protected 10% DMSO with an average viability of 78.6% +- 11.3%. However, even with the combination of HA and DMSO at a concentration of 10% or 5%, the viability was above 70%.
The viability decreased below 70% only when the DMSO concentration fell below 2%. The use of HA as a cryoprotectant alone does not appear to provide sufficient protection for DPSC during cryopreservation.
Surprisingly, after a week of culture, the highest viability was measured in subgroups 3 (10% DMSO + 2% HA - 96.0% +- 1.7%), 4 (5% DMSO + 1% HA - 96.2% +- 1.5) %) and 5 (5% DMSO + 2% HA - 95.8% +- 1.7%). The standard cryopreserved group (10% DMSO) showed an average viability of 95.9% +- 2.0%.
Conversely, lower viability was again seen in KBZD frozen in medium using hyaluronic acid and DMSO at a concentration of 2% or less.