Biopolym. Cell. 2021; 37(1):14-22.
http://dx.doi.org/10.7124/bc.000A48
Molecular and Cell Biotechnologies
The impact of cryopreservation with polyvinyl alcohol on the survival and functional activity of rat testis interstitial cells
1, 2, 3Pakhomov O. V., 1, 4Sidorenko O. S.
  1. Institute for Problems of Cryobiology and Cryomedicine, NAS of Ukraine
    23, Pereyaslavskaya Str., Kharkiv, Ukraine, 61015
  2. V. N. Karazin Kharkiv National University
    4, Svobody Ave., Kharkiv, Ukraine, 61022
  3. Kharkiv National Medical University
    4, Nauki Ave., Kharkiv, Ukraine, 61022
  4. Institute for Scintillation Materials, NAS of Ukraine
    60, Lenin Ave., Kharkiv, Ukraine, 61001

Abstract

Aim. To investigate the impact of polyvinyl alcohol (PVA) in cryoprotective media on the survival and proliferation of rat testis interstitial cells (ICs) after cryopreservation. Methods. Rat ICs were obtained by enzymatic processing of testis with collagenase (type I) and DNase I. The obtained cells were cryopreserved by three methods. Methods 1 and 2 implied the use of cryoprotective medium (CM) based on 1.4 M dimethylsulfoxide (DMSO) and 10 % fetal cow serum (FCS) but utilized different cooling rates to –70°C. Method 1: ICs were cooled with 1°C/min. Method 2: after initiation of crystallization the cells were cooled with 15°C/min to –40°C and with 20°C/min from –40 to –70°C. Method 3 implied the cooling rate of 1°C/min but FCS was replaced by 20 mg/ml PVA. When the temperature –70°C was reached the samples were plunged into liquid nitrogen (–196°C). After warming and removal of CM the ICs were cultured in Ham’s/F12 with and without human chorionic gonadotropin (hCG). Results. The use of method 2 improved the survival of Leydig cells in samples to 79.5 (70.0; 90.0) % comparing with method 1 (42.0 (37.0; 47.0) %). The use of PVA in method 3 did not have an effect on ICs and Leydig cell survival comparing with method 1. Cell culturing showed that the number of Leydig cells in wells rose to 17 (0.09; 0.39) and 0.12 (0.08; 0.14) ×106 after cryopreservation by method 2 and 3, respectively, when the cells were stimulated with hCG. These values are several times higher than the initial number of cells in the wells (0.65 (0.57;0.71) ×104). Conclusion. PVA in combination with other components of cryoprotective medium promoted the survival of Leydig cells capable of further proliferation in culture, especially, in the presence of hCG.
Keywords: testis interstitial cells, Leydig cells, cryopreservation, polyvinyl alcohol, cell proliferation, human chorionic gonadotropin
Full text: (PDF, in English)

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