[A study of the mechanisms of action of Fertiwell in vivo].

AIM: To investigate the specific mechanisms of action of Fertiwell in a mouse model of D-galactose-induced aging of the reproductive system. MATERIALS AND METHODS: C57BL/6J mice were randomized into four groups: intact mice (control group), a group of mice with artificial accelerated aging treated with D-galactose alone (Gal), D-galactose followed by Fertiwell (PP), and D-galactose followed by a combination of L-carnitine and acetyl-L-carnitine (LC). The artificial accelerated aging of reproductive system was induced by daily intraperitoneal administration of D-galactose at a dose of 100 mg/kg for 8 weeks. After the end of therapy in all groups, the characteristics of sperm, the level of serum testosterone, immunohistochemical parameters, and the expression of specific proteins were evaluated. RESULTS: Fertiwell had a pronounced therapeutic effect on testicular tissues and spermatozoa, restored testosterone levels to normal values, and, in addition, was more effective protector against oxidative stress in the reproductive system compared to L-carnitine and acetyl-L-carnitine, which are widely used in male infertility. Fertiwell at a dose of 1 mg/kg allowed to significantly increase the number of motile spermatozoa to 67.4+/-3.1%, which was comparable to indicators in the intact group. The introduction of the Fertiwell positively affected the activity of mitochondria, which was also expressed in an increase in sperm motility. In addition, Fertiwell restored the intracellular level of ROS to the values of the control group and reduced the number of TUNEL+ cells (with fragmented DNA) to the level of intact control. Thus, Fertiwell, containing testis polypeptides, has a complex effect on reproductive function, leading to a change in gene expression, an increase in protein synthesis, the prevention of DNA damage in the testicular tissue, and an increase in mitochondrial activity in testicular tissue and spermatozoa of the vas deferens, which leads to the subsequent improvement of testicular function.

[Potentiation of nitric oxide-dependent activation of soluble guanylate cyclase by levomycetin, tetracycline, and oxolin].

The influence of antibiotics laevomycetin and tetracycline and the antivirus agent oxolin on the activity of human platelet soluble guanylate cyclase, the stimulation of the enzyme by NO-donors (sodium nitroprusside (SNP) and spermine nanoate (spermine NONO)) and the combination of spermine NONO and YC-1 was investigated. All preparations used in the concentration range 0,1-10 mM had no effect on the basal activity of guanylate cyclase but potentiated the SNP-induced activation of this enzyme. All preparations used synergistically increased (similar to YC-1) spermine NONO-induced activation of soluble guanylate cyclase. At the same time these compounds did not produce the leftward shift of spermine NONO concentration response curve characteristic for YC-1. Moreover, all compounds used did not influence the leftward shift of spermine NONO concentration response curve obtained in the presence of YC-1. This demonstrated that there was no competition between YC-1 and the drugs for interaction with the enzyme. The revealed regulatory phenomen of laevomycetin, tetracycline and oxolin to increase synergistically NO-dependent activation of soluble guanylate cyclase may cause additional pharmacological effects during prolong treatment by these drugs. This fact is necessary taking into account.

[YC-1-like potentiation of nitric oxide-dependent activation of soluble guanylyl cyclase by adrenochrome].

The influence of adrenochrome and YC-1 on spermine NONO-induced activation of human soluble guanylyl cyclase was investigated. Adrenochrome (0.1-10 microM) had no effect on the basal activity, but it potentiated in concentration-dependent manner the spermine NONO-induced activation of this enzyme. Adrenochrome, like YC-1, sensitized guanylyl cyclase towards nitric oxide (NO) and produced the leftward shift of spermine NONO concentration responce curve. Addition of adrenochrome decreased the YC-1-induced leftward shift of spermine NONO concentration response curve. Adrenochrome also inhibited (by 63%) the enzyme activation by YC-1. These data demonstrates the possible competition between adrenochrome and YC-1. Thus, synergistic activation of NO-stimulated guanylyl cyclase activity by adrenochrome represents a new biochemical effect of this compound and indicates that adrenochrome may act as an endogenous regulator of NO-dependent stimulation of soluble guanylyl cyclase. This new property of adrenochrome, similar to YC-1, is necessary taking into account, especially under conditions of overproduction of adrenochrome in organism.

[New approaches to search for antitumoral compounds].

The authors discuss the present-day state of search for antitumoral compounds at Blokhin Russian Oncological Research Center and promising approaches including computer technologies as means of search for new anticancerous targets and drugs.

Induction of P-glycoprotein functional activity and multidrug resistance by a soluble factor(s) produced by some mammalian cells.

In the previous study we have found that Djungarian hamster fibroblasts with high levels of multidrug resistance (MDR) (colchicine-resistance index RI of 1000 to 42000) produce soluble factor(s) communicating MDR to the drug-sensitive cells of the same species by elevating the functional activity of P-glycoprotein (Pgp). Here we have shown that these cells can influence human tumor cells in the same fashion. Rat hepatoma McA RH7777 cells and their colchicine-resistant derivatives are shown to produce a factor with similar effects (induction of MDR and Pgp functional activity in the drug-sensitive cells). These effects seem to depend on the drug resistance level of the donor cells. Our results show that induction of the Pgp-mediated MDR is not species-specific and the tumor cells with intrinsic MDR (arising from the tissue with a high level of Pgp expression) can produce a factor(s) communicating this type of drug resistance to the sensitive cells.

Partial restoration of the actin cytoskeleton in transformed Syrian hamster fibroblasts selected for low levels of 'typical' multidrug resistance.

Two independent colchicine (CLC)-resistant sublines of Rous sarcoma virus-transformed Syrian hamster fibroblasts were isolated. Each subline represented variants with 11- and 12.4-fold resistance, respectively, their 23- and 23.7-fold resistant descendants, as well as variants cultured in CLC-free medium for 10 months without loss of resistance. All variants demonstrated 'typical' multidrug resistance. The parental cells contained actin in dispersed form, as determined by rhodamine-phalloidin staining. In contrast, already in 11- and 12.4-fold resistant sublines up to 30% of cells demonstrated restored stress fibers. Cultivation in CLC-free medium leads to the accumulation of cells with a partially restored actin cytoskeleton. Putative mechanisms of up-regulation of stress fiber assembly in cells with P-glycoprotein-mediated multidrug resistance are discussed.

[The use of verapamil in tests for determining the drug resistance of leukemic blasts]. / Ispol'zovanie verapamila v testakh dlia opredeleniia lekarstvennoi ustoichivosti leikoznykh blastov.

To determine prognostically unfavourable groups of acute leukemia patients, the authors studied the in vitro accumulation of 3H-vincristine (Vcr) and adriamycin (ADR) as well as inclusion of 3H-cytosar (Ara-C) into marrow blast DNA from patients showing different effects of treatment. It was found resistant to induction chemotherapy increases with verapamil addition to culture medium (Vrp+ cells). ADR inclusion into Vrp+ cells was the same as that into Vrp- cells. The inclusion of 3H-Ara-C into S-phase cell DNA in the cells Vrp+ was 3 time that in the cells Vrp-. All the responders to cytosar treatment had Vrp- blasts. It is evident that evaluation of Vrp effect on 3H-Vcr accumulation under short-term culturing is able to indicate groups of patients with low probability of achieving complete remissions in response to standard regimens of Vcr, Ara-C and anthracyclines treatment.

[Biochemical indicators of anthracycline sensitivity of blast cells from patients with hemoblastoses]. / Biokhimicheskie pokazateli chuvstvitel'nosti k antratsiklinam blastnykh kletok bol'nykh gemoblastozami.

It was shown that accumulation of adriamycin (ADR) and its impact on DNA synthesis in the blast cells of patients with hemoblastosis studied under conditions of short-term incubation correlated with the antileukemic activity of anthracyclines. The combination of the two biochemical and pharmacological indices, i.e. the high levels of ADR accumulation and inhibition of DNA synthesis, is may useful in solving the problem on inclusion of the antibiotics to the schemes of combined chemotherapy of patients with hemoblastosis.

[The correlation of the in vitro 14C-glycine absorption by murine and human leukemic cells with the antitumor effect of vincristine]. / Korreliatsiia pogloshcheniia 14C-glitsina in vitro leikoznymi kletkami myshei i cheloveka s protivoopukholevym éffektom vinkristina.

Vincristine (VCR) studied for its in vitro effect on glycine accumulation by mouse leukemic cells and blood blast cells of patients with hemoblastosis. It is found that the value of glycine cell accumulation and a degree of inhibition of the initial level of amino acid accumulation in a cell by alkaloid correlate with the antitumour action of the preparation. The glycine accumulation by blood blast cells (less than 40 nmol/min/10(9) cells), absence of the VCR effect on this process are found to be unfavourable prognostic factors for successful chemotherapy of patients by VCR. The possible role of the cellular membrane in the mechanism of the VCR action is discussed.

[In vitro 3H-vincristine accumulation as a possible marker of the sensitivity of animal and human leukemic cells to the preparation]. / Nakoplenie 3H-vinkristina in vitro kak vozmozhnyi marker chuvstvitel'nosti leikoznykh kletok zhivotnykh i cheloveka k preparatu.

The uptake of 3H-vincristine into the leukemic cells of animals and blast cells of peripheral blood of patients with leukemia has been studied. The capacity of cells to accumulate the drug is found to be of significant value for their sensitivity to vincristine. The in vitro low-alkaloid accumulation (less than 18 pmol/mg protein) by cells is an unfavourable prognostic sign for vincristine chemotherapy.

[Comparison of free ribonucleotide pool in the spleens of C57BL and DBA/2 mice and in leukemia cells sensitive and resistant to 5-fluorouracil]. / Sravnenie fonda svobodnykh ribonukleotidov v selezenke myshei C57BL i DBA/2 i v kletkakh leikozov, chuvstvitel'nykh i ustoichivykh k 5-ftoruratsilu.

The amount of free purine and pyrimidine ribonucleotides in the spleens of mice (C57Bl and DBA/2) and in lympholeukemia cells (La and L1210), sensitive and with induced resistance to 5-fluorouracil, was determined by chromatography on a column with DEAE-cellulose. It was found that the cytidine ribonucleotide pool in the spleens of DBA/2 mice is 2 times lower as compared to C57Bl mice. The lympholeukemia cells (La and L1210) isolated from the animals also differed in their uridine nucleotide pools. The development of leukemia was accompanied by a decrease in ATP and GTP. No significant changes in the total amount of pyrimidine nucleotides under developing resistance to 5-fluororuacil were observed.