"Nanobiotechnology for health-care"
Effect of Ag nanoparticles on cryopreserved mesenchymal stem cells from cartilage tissue
Volkova N.O., Stepanyuk L.V., Goltsev A.M.
Institute for Problems of Cryobiology and Cryomedicine, NAS of Ukraine.
Pereyaslavska str.23, Kharkiv-61016, Ukraine.
E-mail: volkovana781@gmail.com

The nanoscale materials have a dose-dependent effect on biological objects. The finding of the presence/absence of Ag nanoparticles (AgNPs) cytotoxic action is an important direction of nanobiotechnological research. The effect of AgNPs on morphological characteristics, proliferative and antioxidant potentials of cryopreserved mesenchymal stem cells (CrMSCs) from cartilage tissue was investigated.
The AgNPs (Sigma-Aldrich, USA) with an average size of 15 nm were used in the study. CrMSCs from cartilage tissue were incubated with AgNPs at final concentrations of 4, 6, 10, 20 μg/mL for 1 hour. The control was cells without the influence of AgNPs (0 μg/mL). On the 1, 3, 7, and 10 days of cultivation the number of viable cells was determined by MTT test. Analysis of total antioxidant status (TAS) was performed using Randox test kit (UK). On the 10th day, the CrMSCs were fixed in a 4 % solution of paraformaldehyde followed by azure-II and eosin staining by Romanovsky-Giemsa. The results were processed with Student's t-test using Excel software.
The use of 4 and 6 μg/ml AgNPs did not influence on morphological characteristic, proliferative and TAS activity in CrMSCs from cartilage tissue. AgNPs at concentrations of 10 and 20 μg/mL decreased the TAS index (1.3 and 1.5-fold respectively) and proliferative potential (1.2 and 1.6-fold respectively) if compare with the control. The use of AgNPs at concentrations of 10 and 20 μg/ mL led to changes in the morphological characteristics of CrMSCs from cartilage tissue in relation to control samples, namely signs of cytoskeletal dystrophy, cytoplasmatic granularity and vacuolization of nuclei. It was found that AgNPs in concentrations of 4 and 6 µg/mL are safe for CrMSCs from cartilage tissue, while increase up to 10 µg/mL has a toxic effect manifested by the change of proliferative, antioxidant activities and morphological characteristic. 
Thus, the presented results are belonged to the field of applied nanobiotechnology, that spreads to field restorative medicine, mostly in creation of cartilage bioimplants.
The work was carried out within the framework of the target research program of the NAS of Ukraine "Genomic, molecular and cellular bases of development of innovative biotechnologies", agreement No. 2.2.6.132/21.