Anticlastogenic activity of aminoseleton under the effect of cyclophosphamide on the bone marrow of mice

The tissue drug aminoseleton, designed at the All-Russian Veterinary Research Institute of Pathology, Pharmacology and Therapy, was obtained from the spleen of cattle by cryogenic fractionation. Adaptogenic, membrane stabilizing, stress-protective, antioxidant and immunomodulatory properties of aminoseleton have been already shown. In this work, the anticlastogenic effect of the tissue drug aminoseleton on the bone marrow cells of mice exposed to the experimental mutagen was revealed for the first time. In addition, the preservation of cytogenetic stability and mitotic activity in bone marrow cells of healthy animals was shown when using the study drug. The objective of this work was to assess the effect of aminoseleton on the cytogenetic stability of bone marrow cells in healthy mice and mice exposed to the experimental mutagen, as well as to identify the antimutagenic properties of the drug in relation to the genotoxic effect of cyclophosphamide (CP) using a micronucleus test. The experiments were carried out on outbred white mice ( Mus albus officinarum ), which were divided into six groups subjected to the following treatments: i) intramuscular administration of sterile isotonic sodium chloride solution in a volume of 0.2 ml (negative control, n = 12); ii) intraperitoneal injection of 0.2 ml of CP (Baxter Oncology GmbH, Germany) at a dose of 20.0 mg/kg of body weight (positive control, n = 12); iii) intramuscular single injection of 0.2 ml of aminoseleton at a therapeutic dose of 0.5 ml/kg ( n = 12); iv) intramuscular single injection of 0.2 ml of aminoseleton at a tenfold therapeutic dose of 5.0 ml/kg ( n = 12); v) intramuscular single injection of 0.2 ml of aminoseleton at a dose of 0.5 ml/kg with intraperitoneal injection of 0.2 ml of CP at a dose of 20.0 mg/kg in 72 h ( n = 6); vi) intramuscular fivefold injection of 0.2 ml of aminoseleton at a dose of 0.5 ml/kg with 24 h intervals and intraperitoneal administration of CP is similar to animals of other groups 72 h after the fifth injection ( n = 6). To determine the amount of chromosomal aberrations in the bone marrow, 2.5 h before euthanasia, mice were injected intraperitoneally with 0.025 % colchicine (PanEco, Russia). Bone marrow cells were washed out of the femurs using Hanks' buffer solution (pH 7.4), the cell suspension was incubated in 0.075 molar hypotonic KCl solution, then the cells were fixed with acetoalcohol cooled to 4 °C and stained by Romanowsky-Giemsa procedure. The mitotic index (MI) was assessed by the number of dividing cells per 1000 bone marrow cells. The number of cells with chromosomal aberrations was counted in 100 metaphase plates per animal. Single and paired fragments, exchanges and achromatic gaps, as well as cells with multiple pathologies were counted. To study the frequency of micronuclei (micronucleus test) of polychromatophilic erythrocytes (PCE), the obtained bone marrow cells were added to 1 % albumin solution in Hanks' buffer solution (pH 7.4) and applied to glass slides, then the samples were dried, fixed with methanol and stained by Romanowsky-Giemsa protocol. The frequency of micronuclei per 1000 PCE was determined; a total of 2000 PCE per animal was studied. The proportion of PCE per 500 normochromic erythrocytes (NE) and PCE was also calculated. The frequency of chromosomal aberrations and micronuclei when administering the drug at the studied doses did not statistically significantly differ from that in animals of their negative control group that was 1.0±0.40 and 0.2±0.06 %, respectively. The administration of aminoseleton also had no effect on the mitotic index of bone marrow cells in experimental animals. The course administration of aminoseleton reduced the clastogenic effect of cyclophosphamide, assessed by the number of micronuclei in polychromatophilic erythrocytes of the bone marrow, from 2.3±0.21 % in mice from the positive control group to 1.0±0.40 % in animals after a course of aminoseleton injections. Thus, the cla