Reserve‐adaptive potential of sperm plasma in stallions of different ages

Abstract

During the aerobic metabolism of sperm, various reactive oxygen species are produced. Sperm cells are extremely vulnerable to oxidative stress (OS), which causes loss of membrane integrity, resulting in increased cell permeability, enzyme inactivation, structural DNA damage and cellular apoptosis. The aim of this study was to assess the reserve‐adaptive potential (RAP) of sperm plasma in stallions of various ages. RAP is the difference between the level of spontaneous and induced oxidative protein modification (OMP), reflecting the degree of OS. Spontaneous OMP reflects the accumulation of OS products in vivo. To determine the level of spontaneous OMP, a sperctrophotometric analysis of 2,4‐dinitrophenylhydrazones (DNPH) formed during the interaction of carbonyl derivatives of sperm plasma proteins with 2,4‐dinitrophenylhydrazine was carried out (spectrophotometer “SF‐2000”, Russia). R.L. Levine, modified by E.E. Dubinina. Measurements of DNPH derivatives were carried out at 14 wavelengths: in the range 260‐280nm for the registration of aldehyde dinitrophenylhydrazones of a neutral nature (ADNPHn), of a basic nature (ADNPHb) – 258‐264 and 428‐520nm, for the determination of ketone‐dinitrophenylhydrazones of a neutral nature (CDNPHn) 363‐370nm, basic character (CDNFGb) – 430‐434 and 524‐535nm. After that, the total amount of carbonyl derivatives was found. Induced OMP was determined in a similar way, but after adding iron (II) sulfate and hydrogen peroxide solutions to the reaction mixture. This provoked the formation of new carbonyl groups. The more RAP, the higher the ability of the biological material to resist the OS.Two groups of stallions were formed: the first group consisted of 15 stallions aged 13 to 20 years (average age 15.2±1.8 years), the second group consisted of 15 stallions aged 3 to 6 years (average age 4.0±0.5 years). The spontaneous OMP of sperm plasma in aged stallions was 495.6 Eod/ g protein, the induced one – 573.4 Eod / g protein. In the group of young stallions, spontaneous OMP – 347.2 Eod / g protein, induced – 485.3 Eod / g protein. The OMP level of sperm plasma in older stallions is initially higher than in young stallions (p = 0.04), which indicates the accumulation of carbonyl derivatives in vivo as a result of the prevalence of OS over the antioxidant capabilities of the organism. But when additional oxidizing components are added to the reaction mixture, the OMP of aged stallions does not increase as significantly as in young ones. RAP of sperm plasma of old stallions is 13.56%, of young stallions – 28.75%. The ability to withstand OS, the reserve of antioxidant capabilities of sperm plasma in young stallions is 2.1 times higher than in older stallions.