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The anticancer antibiotic doxorubicine (DOX) is highly toxic and induces functional complications in vital organs. The effect of DOX on normal cells has not been examined in sufficient detail, and the search for compounds reducing DOX toxicity did not lead to success so far. It has been suggested that DOX induces death of cancer cells via p53-dependent apoptosis, however, the information regarding the role of p73 protein, a member of p53 tumor suppressor family, is scanty. Cytomegalovirus (CMV) induces an antiapoptosis program that allows its replication until death of the target cell. Our objectives were to examine the effect of DOX on normal cells (human fibroblasts), analyze the ability of CMV-induced antiapoptosis program to reduce DOX toxicity, and to evaluate the involvement of p73 protein and its isoforms in the regulation of death of CMV-infected and DOX-treated cells. Within a 24-h time period DOX caused death of about 70% human embryonic lung fibroblasts (HELF) in cell culture, this parameter decreased significantly in CMV-infected DOX-treated HELF cells. TUNEL has shown that the number of cells with DNA fragmentation decreases from 5.2% under the effect of DOX to 3.2% (P < 0.05) after combined CMV-DOX treatment. Analysis of mitotic figures revealed that DOX causes accumulation of mitotic cells, which was not observed in CMV-infected DOX-treated cells. PCR analysis of mRNA of two p73 protein isoforms (TAp73 and dNp73) has shown that in uninfected cells the expression of TAp73 isoform was low, while in CMV-infected cells level of TAp73 was significant and expression of dNp73 was demonstrated for the first time. Expression of TAp73 associated with lack of mitosis block. The activation of caspases 8, 9 and 3 in CMV-infected cells was registered but cell death was not, however, as massive as that caused by DOX. From these findings it can be concluded that CMV attenuates DOX-related damage to normal cells. It can be suggested that induction of TAp73 and dNp73 isoforms provides conditions for reduction of DOX effect which leads to DNA damage and death of normal cells.

Pulsed radiofrequency (PRF) is a percutaneous minimal invasive procedure that can be used when conservative pain therapy methods have been ineffective. The effectiveness of PRF was demonstrated in various good quality randomized control studies, but mechanisms of action are still unclear. The aim of our study is to analyse the histological effects of PRF on the domestic porcine dorsal root ganglion (DRG), and evaluate the expression of biomarkers in gangliocytes. 3 domestic porcines were investigated. Under general anaesthesia and X-ray control, DRG PRF was performed. Four lumbar DRGs (L1, L2, L3, L4) were randomly treated. The opposite side DRGs was used as control. One month after the procedure the animal was euthanized. The lumbar region of the spine was placed in 10% formaldehyde for a month. After this fixation DRG samples were prepared for slide analysis. They were embedded in paraffin in order to obtain 3 microm thick sections, which were then cut by microtome and collected on slide glasses. Using standard immunohistochemical reactions, the materials were tinted to define biomarkers NF, GFAP, Hsp-70 expression and apoptosis by TUNEL kit. The number of cells with NF (26.0 +/- 3.0 vs 16.1 +/- 3.3; p < 0.05), GFAP (12.0 +/- 1.3 vs 3.2 +/- 0.9; p < 0.05) and Hsp-70 (10.0 +/- 1.6 vs 4.2 +/- 1.0; p < 0.05) expression, were larger in the PRF side comparing with the control side. Additionally, glial cells in spinal ganglia of both sides demonstrated immunoreactivity. The instances of apoptosis were not significantly different, in statistical terms, between the control and experimental sides (18.0 +/- 4.0 vs 20.0 +/- 4.0; p = 0.35). PRF in spinal gangliocytes of lumbar region increases neural tissue cytoskeleton factors like NF and GFAP suggesting about active regeneration processes into the cells 1 month after the procedure. Spinal gangliocytes one month after PRF treatment notably increases Hsp-70 expression suggesting about activation of cellular activity and inhibitory role reducing of oxidative stress. Similar number of apoptotic cells in spinal ganglia of lumbar region after PRF and control side suggests about inhibitory role of PRF on programmed cell death and stimulation of cell survival.

The article presents the classification and evaluation of techniques applied in documentation of apoptosis of spermatozoids. The main point of techniques detecting the changes in DNA of apoptosis spermatozoids (SCSA technique, TUNEL test and Comet assay) is revealed The annexin technique, the technique of detection of changes of cells transmembrane potential, the technique of detection of activity of caspase and technique of detection the degree of expression of proteins-regulators of apoptosis are discussed too. The advantages and shortcomings of these techniques are considered.

Objective: To study apoptosis of individual cellular components of the vascular wall of coronary arteries at different morphological stages of atherosclerosis., Material and Methods: The study was performed on coronary arteries taken from 52 deceased patients with atherosclerosis and coronary heart disease at different stages of atherogenesis. For morphological study prepared paraffin sections, which were stained for morphological studies were prepared paraffin sections, which were stained with hematoxylin and eosin, by Van Gieson, Masson, on lipids with Sudan black B, according to Van Cossu. To determine apoptosis, TUNEL method used in paraffin sections. Apoptotic index (AI) was calculated by TUNEL-positive cells and the average inner shell coronary artery around the perimeter each with increasing microscopic 1000., Results: Investigation showed significant apoptosis (p < 0.05) increase in AI smooth muscle, endothelial cells, macrophages in the coronary arteries affected by atherosclerosis compared to intact control group vascular segments significant reduction AI endothelial, smooth muscle cells and macrophages (p < 0.05) traced from the early stages of atherogenic disorders to atheromatosis., Conclusions: It is established that apoptosis of smooth muscle cells, macrophages and endothelial cells is the most intensive on early stages of atherosclerotic process. In process of progressing of atherosclerosis intensity and prevalence of apoptosis of coronary artery wall cells decreases, and processes of necrosis becomes predominant. Apoptosis of coronary artery wall cells is valuable in increasing the zones of atheromatosis, plaque destabilizations, and also increases the risk of thrombosis and ulcerations.

Apoptosis and differentiation in presumptive neural retina (PNR) and presumptive retinal pigmented epithelium (PRPE) wert investigated during early retina development of toad, Bufo raddei Strauch. TUNEL staining was used to evaluate apoptotic cells and the immunohistochemistry was used to assess the expression levels ofglial fibrillary acidic protein (GFAP), RT97 and tyrosinase (Tyr) during early eye development respectively. The density of apoptotic cells and protein expression were quantitated with Image-Pro Plus 6.0. Apoptosis was found in both PNR and PRPE and the density of apoptotic profiles in PRPE was higher than that in PNR (most P<0.01) at the same stage during early eye development. The expression levels of GFAP and RT97 changed from low to high in PNR, but from high to low in PRPE, whereas the expression level of Tyr, was contrary to those of GFAP and RT97 in both PNR and PRPE. The point of intersection of these, increase and decrease respectively was found at 5-6 h after formation of optic vesicle (FOV). PRPE becomes thinner than PNR, one of the reasons might be due to higher density of apoptosis in PRPE than that in PNR during early eye development. Molecular differentiation, however, occurred after the contact of the optic vesicle outer wall with the overlying ectoderm which promotes the expression of specific molecules and inhibits the expression of non-specific molecules in PNR and PRPE respectively.

Transection of the maxillary nerve initiates apoptosis of the maxillary sinus mucosa cells in rats. Significant activation of apoptosis and proapoptotic factor p53 was found in the epithelium during week 1 after nerve transection. In delayed period after injury, apoptotic cells predominated in the submucosa against the background of Bcl-2 hypoexpression.

We studied the effect of recombinant human erythropoietin on functional activity of skin cells in vitro. It was found that erythropoietin stimulated proliferation of mesenchymal and epithelial cells and effectively protected epidermal HaCaT cells from apoptosis. Insignificant effect of erythropoietin on contraction of collagen gel by mesenchymal cells was revealed. These findings suggest that erythropoietin can be a promising component of wound-healing preparations.

Induced apoptosis in urinary bladder cancer tumor cells of patients was studied using TUNEL reaction. It was shown that increase in induced apoptosis value had a definite correlation between corresponding features of tumor reaction as a response on Gemcitabine-Cisplatin neoadjuvant chemotherapy application. It was found that evaluation of induced apoptosis in urinary bladder cancer tumor cells using TUNEL method allows forecasting the effectiveness of chemotherapy on the cellular level in patients with this type of cancer.

Shown antimicrotrubules activity of new 2,6-dinitroaniline compounds. Investigated their ability on apoptotic processes in a plant cell when used as a composition with inhibitors of acetyl-CoA carboxylase.

Ectopic neurotransplantation can be used as an in vivo culture method for evaluation of the effect of various environmental factors (neurotransmitters, cytokines, and other bioactive substances) on histogenesis of the developing brain. The study was performed 1 day after allotransplantation of neocortical primordia from embryos of rats receiving intraperitoneal injection of p-chlorophenylalanine on day 11 of pregnancy. Under these experimental conditions the number of degenerating cells increased, while the count of mitotic neuroepithelial cells was 2.5-fold lower compared to neurotransplants of intact embryos at the same stage of development. Incubation of neocortical primordia in serotonin-containing medium before transplantation prevented cell death and promoted division of transplanted cells. Serotonin plays a role in the regulation of neuroepithelial cell proliferation and prevents cell death in the developing neocortex.

36 specimens of the myocardium were obtained during necropsy of patients who had died of acute cardiac failure. Bcl-2, caspase 3 and p53 were studied immunohistochemically. TUNEL method was used to study apoptic DNA fragmentation in cardiomyocytes (Cms). The results suggest that apoptosis contributes to the development of cardiomyocyte injury in ishemia. Apoptotic death of Cms may lead to electric instabilly of the myocardium in early ishemia and cause sudden cardiac death. Apoptosis of the Cms may precede extension of the necrotic area. Further studies of apoptotic death of Cms in ishemic heart disease may help to renew therapeutic strategies aimed at regulation of apoptosis.

The aim of the study was assessment of hepatocyte apoptosis depending on expression of Fas and FasL proteins by various liver cells in patients with chronic viral hepatitis B (CVHB) or chronic viral hepatitis C (CVHC). The symptoms of hepatocyte apoptosis were observed in 3 of 12 patients with CVHB and in 9 of 14 patients with CVHC, the proportion of apoptotic cells being 12-65%. Hepatocytes of healthy people and patients with hepatitis B or C express Fas protein in the cytoplasm diffusely, as granules or on cell membrane. In health, hepatocytes do not express FasL, but in CVH they do. The highest apoptosis was observed in Fas protein location as granules in cytoplasm or in their preferable location on the cell membrane. The severity of hepatocyte apoptosis in CVH directly correlated with FasL expression by the cells of the lymphoid-histiocytic infiltrate in the liver and inversely correlated with FasL expression by hepatocytes. Thus, a great part of hepatocytes in CVH are killed by the virus; Fas/FasL interaction is leading in damage to hepatocytes in CVH.

Endometrial carcinoma is the most incident cancer of human reproductive system. There are unequivocal evidences of relationship between complex and atypical hyperplasia and development of cancer. Apoptosis plays a significant role in the maintenance of equilibrium between cell death and proliferation and contributes to prevention of tumorigenesis. Internucleosomal DNA fragmentation known as one of the most important criteria of apoptosis cannot be used for evaluating the risk of cancer development because it reflects the current level of apoptosis but is useless for evaluating the real limits of apoptosis intensity in certain types of tissue. For estimating the possibility of apoptosis development in endometrial tissues, a new method of quantitation of nuclear Ca(2+)/Mg(2+)-dependent endonuclease (NCME) activity has been developed. Fifteen samples of normal endometrial tissues at middle proliferative, secretory, premenstrual, and menstrual phases, 43 samples of hyperplastic endometrial tissues, 13 samples of endometrial polyps, and 17 samples of endometrial adenocarcinoma were collected by diagnostic curettage of the uterine cavity and by hysterectomy (carcinomas). The material was examined by 1) TUNEL method and 2) agarose gel electrophoresis of DNA cleaved by nuclear CME in isolated cell nuclei in the presence of Ca(2+) and Mg(2+) ions, followed by quantitation of CME activity. The activity of NCME was found to decrease from normal endometrium (1.1 +/- 0.12 U, without significant changes throughout the menstrual cycle) through polyps (0.9 +/- 0.15 U), cystic hyperplasia (0.45 +/- 0.06 U, p < 0.01), and adenomatous hyperplasia (0.32 +/- 0.08 U, p < 0.01) to adenocarcinoma (0.37 +/- 0.11 U, p < 0.01 for well differentiated, 0.16 +/- 0.08 U, p < 0.01 for moderately differentiated, and 0.03 +/- 0.02 U, p < 0.01 for poorly differentiated samples). The TUNEL-specific staining pattern in normal endometrium varied in a wide range during the menstrual cycle (from poorly stained individual cells in the proliferative phase to intensely stained cell clusters in the premenstrual phase). At the same time the difference between the normal endometrium in the proliferative phase and pathologically changed endometrium (hyperplasia or cancer) could not be detected by the TUNEL technique. Hence, TUNEL is useless for predicting cancer development in hyperplasia and precancer. By contrast, evaluation of NCME activity helps detect the early disorders in the proliferative processes coursing in endometrial tissues and thus prevent tumor development.