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Colorectal cancer is the actual problem in the global health care system, which is associated with high rates of morbidity and mortality. Aim. To compare the expression levels of immunohistochemical markers of proliferation and apoptosis in the colorectal tumor cells and the colorectal tumor-associated stroma; to study the correlations between the proliferation levels, apoptosis levels and clinicopathological parameters of colorectal adenocarcinoma. Materials and methods. The surgical material of 30 patients with distal colon adenocarcinoma (I-IV clinical stages) was analyzed using pathohistological and immunohistochemical methods. Results. It was established that colorectal tumor cells are characterized by the medium proliferation level, the medium p53 expression level and the low apoptosis level, just as colorectal tumor-associated stroma is characterized by the low proliferation, p53 expression and apoptosis levels. The direct strong correlations between the proliferation and apoptosis levels in the tumor cells, and also between the proliferation and apoptosis levels in the tumor-associated stroma were revealed. The direct medium correlations between the p53 expression levels in the tumor cells and the tumor-associated stroma, between the p53 expression level in the tumor cells and the Ki-67 expression level in the tumor-associated stroma, and also between the p53 expression level in the tumor cells and the degree of histological differentiation of the tumor were revealed. Moreover, the reverse strong correlations between the Ki-67 expression levels and the caspase-3 expression levels in the tumor cells and the tumor-associated stroma were revealed. Conclusions. These data indicate the tight association between proliferative and apoptosis processes in the colorectal tumor cells and the colorectal tumor-associated stroma. The interconnections between the p53 accumulation in the tumor cells and the colorectal tumor-associated stroma, the p53 accumulation in the tumor cells and the increase of proliferative activity of the colorectal tumor-associated stroma, and also between the p53 accumulation in the tumor cells and the degree of histological differentiation of the tumor were established. Furthermore, it was revealed that increase of proliferative activity of colorectal adenocarcinoma is associated with decrease apoptotic activity of the tumor.

Objective: To study morphological features of Hassall's corpuscles (HC) and their microenvironment in newborns with increased thymus mass., Material and Methods: The study was carried out on autopsy material of children of the first month of life. Based on the thymic index (TI), 2 groups were identified: with normal (conditional norm) and increased TI value (increased thymus weight). The standard method of histological staining and immunohistochemical methods with antibodies to Pan-CK, CK19, CD68, CD3 and p53 were used in the study. The classification proposed by A.G. Beloveshkin (2013) was used to determine the degree of maturity of HC. Nonparametric Mann-Whitney test was used to determine statistically significant differences in the groups., Results: In the group of children with increased thymus weight, the number of HC decreased by 20%. It was found that the proportion of progressive and mature corpuscles in this group was reduced by 2.3 and 1.6 times, respectively, compared to the group of children with normal thymus weight, while the proportion of regressive corpuscles increased almost 2-fold. In the HC microenvironment, there is an increase in the total number of thymocytes, combined with a decrease in the expression of CD68, CD3 and p53 in them. A sharp decrease in CK19-expressing cells in this group is accompanied by a disruption in the formation of reticular structures characteristic of the comparison group., Conclusion: In the thymus with increased mass, the structural and functional organization changes: along with an increase in the total number of thymocytes in the cortical layer and a decrease in the number of macrophages, epithelial cells and HC (with a predominance of regressive corpuscles), disturbances in the processes of maturation, apoptosis and negative selection of lymphocytes occur, which can lead to development of immunogenesis disorders.

Aim    To study the relationship of blood serum concentration of endocan with indexes of apoptosis and clinical and instrumental characteristics of patients with ischemic heart disease (IHD).Material and methods    The study included 176 subjects (105 men and 71 women). 150 of them were diagnosed with IHD and 26 were healthy volunteers. Anthropometric measurements, coronary angiography, echocardiography, duplex ultrasound scanning of extracranial parts of the brachiocephalic arteries were performed for all patients. Concentrations of endocan (ng / ml), glucose (mmol / l), and apoptotic markers Bcl-2 (ng / ml), Bax (ng / ml), Bcl-2 / Bax, TRAIL (pg / ml), and p53 (ng / ml) were measured in blood serum. Patients were divided into groups based on their SYNTAX scores: group 1 with moderate atherosclerotic lesions of the coronary arteries (CA) (score < 22, 78 patients); group 2 with severe CA atherosclerosis (score 23-32, 37 patients); and group 3 with extremely severe CA lesions (score >33, 35 patients). The control group consisted of healthy volunteers (26 subjects). All groups were age- and sex-matched. Differences were considered statistically significant at p<0.05.Results    A correlation was found between endocan concentration and IHD severity (r=0.32, p<0.001). In group 1, the median endocan concentration was 14.57 ng / ml [8.21; 23.66], in group 2, 19.34 ng / ml [8.425; 26.645], in group 3, 32.13 ng / ml [18.2; 39.12], and in the control group, 6.92 ng / ml [4.62; 9.18]. Correlations of varying strength and significance were observed between the endocan concentration and a number of clinical and instrumental characteristics. Endocan concentrations significantly differed in groups of patients with multifocal atherosclerosis (p<0.01), angina pectoris (p<0.01), a history of myocardial infarction (p<0.001), and obesity (p<0.05) from patients without these signs. Also, a correlation was found between serum endocan concentration and apoptotic markers: TRAIL (r= -0.448, p<0.001); BCL-2 (r= -0.552, p<0.001), Bax (r= -0.519, p<0.001), Bcl-2 / Bax (r= -0.576, p<0.001) and p53 (r= -0.520, p <0.001).Conclusion    The study demonstrated a potential role of endocan as a promising biomarker for risk stratification, prognosis and therapeutic monitoring of IHD patients.

Endometrial cancer occupies the sixth place among oncological morbidity structure in women worldwide. The aim. To compare the expression levels of estrogene receptors – α and progesterone receptors, regulatory proteins p16 and p53, a marker of proliferation Ki-67 and the apoptotic degradation enzyme caspase-3, matrix metalloproteinase 9 (MMP 9) and its tissue inhibitor TIMP-1 in tumor and stromal cells of the invasive non-metastatic endometrioid endometrial carcinoma and also in invasive metastatic endometrioid endometrial carcinoma. Materials and methods: Immunohistochemical and morphometric study of invasive non-metastatic рТ1-4N0Mx endometrioid endometrial carcinoma (n=56; age 42-83 years), invasive metastatic рТ1-4N1-2Mx endometrioid endometrial carcinoma (n=30; age 48-79 years) was performed. Results: Statistically significantly higher expression level of estrogen receptor-α was detected in tumor cells of endometrioid endometrial carcinoma with metastasis to the regional lymph nodes in comparison with the expression level of these receptors in tumor cells of non-metastatic endometrioid endometrial carcinoma. The progesterone receptors expression levels in the tumor and stromal cells of invasive endometrioid endometrial carcinoma with metastasis to regional lymph nodes were lower in comparison with the corresponding expression levels of progesterone receptor in corresponding cells of non-metastatic endometrioid endometrial carcinoma. In stromal cells of endometrioid endometrial carcinoma with metastasis to the regional lymph nodes the expression level of caspase-3 was significantly lower too in comparison with the expression level of this enzyme in stromal cells of non-metastatic endometrioid endometrial carcinoma. In tumor and stromal cells of endometrioid endometrial carcinoma with metastasis to the regional lymph nodes the expression levels of MMP-9 and TIMP-1 are decreased significantly compared to the expression level of these markers in the respective cells of non-metastatic endometrioid endometrial carcinoma. Conclusions. The expression level of estrogen receptor-α in the tumor cells was higher, the expression level of caspase 3 in stromal cells was lower and the expression levels of progesterone receptors, MMP 9 and TIMP-1 in tumor and stromal cells were lower in metastatic endometrioid endometrial carcinoma compared to non-metastatic endometrioid endometrial carcinoma. The obtained data can be used for individual prediction of metastasis to regional lymph nodes in patients with endometrioid endometrial carcinoma.

Aim. Comparison analysis of expression levels of Ki-67, p53, and caspase 3 was performed between the tumor cells and the stromal cells of non-invasive intestinal-type gastric adenocarcinoma on the one hand and the tumor cells and the stromal cells of invasive intestinal-type gastric adenocarcinoma on the other hand. Materials and methods. For this purpose, pathohistological and immunohistochemical studies were used. The surgical material of 10 patients with non-invasive (pTis, pT1) intestinal-type gastric adenocarcinoma and 10 patients with invasive (pT2, pT3, pT4) intestinal-type gastric adenocarcinoma were analyzed. Results. It was established that intestinal-type gastric adenocarcinoma is characterized by the medium Ki-67 labeling index, the low expression level of p53, the low expression level of caspase 3 in the tumor cells, and also the low Ki-67 labeling index and the extremely low expression level of caspase 3 in the stromal cells. The tumor cells of invasive intestinal-type gastric adenocarcinoma are characterized by the 2 times higher expression level of p53, than the tumor cells of non-invasive intestinal-type gastric adenocarcinoma. Moreover, direct correlation between the medium Ki-67 labeling index of the tumor cells and the low Ki-67 labeling index of the stromal cells, and also direct correlation between the medium Ki-67 labeling index in the tumor cells of intestinal-type gastric adenocarcinoma and the low expression level of p53 in these cells were identified. Conclusions. These data demonstrate a significantly more activity of proliferation and apoptosis in the tumor cells of intestinal-type gastric adenocarcinoma, in comparison with the stromal cells of this tumor. Invasive intestinal-type gastric adenocarcinoma differs by the 2 times higher expression level of p53, in comparison with the same index for non-invasive carcinoma. Furthermore, a tight association between proliferative processes in the tumor cells and in the stromal cells of intestinal-type gastric adenocarcinoma, and also an association between proliferative processes and accumulation of mutant protein p53 in the tumor cells of intestinal-type gastric adenocarcinoma were identified.

Endometrial cancer occupies the sixth place among newly diagnosed neoplasms in women. In late stage it has unpredictable prognosis. Immunohistichemical studies of the tumor are very important for prognosis of tumor course. The aim. To characterize the expression of the estrogen receptors-alpha, progesterone receptors, p53, p16, ki-67 and caspase 3 in the invasive endometrioid endometrial adenocarcinoma compared with the normal proliferative endometrium. Materials and methods. The uteruses of 56 women with the verified diagnosis of endometrial cancer (stage pT1-4MxNx) and biopsies of 30 women with normal proliferetive endometrium were analyzed. For this purpose, immunohistochemical and morphometrical studies were used. Results: It was established that level of expression of estrogen receptors-alpha was significantly lower in tumor glands and stroma compared with normal proliferative endometrium: the median was 6.0 (5.0; 7.0) points for tumor glands and 7.0 (6.0; 8.0) points for glands of normal proliferative endometrium(р

Introduction: There is no convincing evidence of the persistence of acute or the development of chronic bacterial-induced prostatic inflammation in the long term when infected with various titers of the uropathogen. Along with this, controversial data are presented on the relationship between post-infectious chronic inflammation and neoplastic changes in prostate tissues., Objective: To carry out, based on the experimental data: 1) assessment of the degree of bacterial contamination and the severity of histological changes in prostate tissues on the 60th follow-up day in case of transurethral infection with various uropathogens in titers of 102,3,5 CFU/ml; 2) fundamental comparative analysis between the indicators of the inoculated test-titer and microbial load with the severity of histological changes in prostate tissues; 3) verification of neoplastic transformations in the prostate tissues during a long-term persistent bacterial-induced inflammatory process., Materials and Methods: Animal studies were conducted using FELASA protocols. Laboratory animals: 14 New Zealand rabbits. Tested uropathogens: aerobes - E. coli, S. haemolyticus, anaerobes - P. niger. Titers: 102,3,5 CFU/ml. Uropathogen inoculation technique: topical transurethral., Randomization: all laboratory animals were divided into 5 groups according to the uropathogen (4 experimental, 1 control). Follow-up period: 60 days. Sacrification and autopsy of the animals were performed on day 60. Biopsies were taken from various parts of the prostate, as well as from the bladder neck and the edge of the membranous urethra. Cultural, histological and immunohistochemical (expression of p53 and Ki-67) studies of prostate tissues were conducted. Statistical data processing was performed using the GraphPad Prism 9.0 program (GraphPad Software Inc., Graphpad Holdings LLC, San Diego, CA, USA) applying descriptive and non-parametric statistics., Results: Two individuals infected with S. haemolyticus + P. niger had a lethal outcome. The contamination of prostate tissue was determined in all cases of infection. In 88.9% of the cases, an increase in tissue microbial load was determined compared to the initial titer. Multivariate analysis of culture study values revealed the presence of intragroup differences in prostate contamination only between infection with E. coli 103 CFU/ml and E. coli 105 CFU/ml (p=0.006), as well as intergroup differences between infection with E. coli 105 CFU/ml and P. niger 105 CFU/ml (p=0.013). The histological study revealed moderate proliferative inflammation after inoculation with 102,3,5 CFU/ml in the E. coli and S. haemolyticus groups. In the case of S. haemolyticus, it was more pronounced due to the presence of persistent alterative lesion foci; in the P. niger group, mild proliferative transformations were observed in prostate tissues in all cases. The immunohistochemical study of changes determined p53 expression (10.0%) in some areas of the glandular epithelium of prostate glands (but without a positive internal control) only in case of infection with E. coli 105 CFU/ml. Areas of glandular epithelium with Ki-67 expression ( less or equal 25.0%) were visualized in all tested groups, mainly at titers of 103 and 105 CFU/ml, but the severity of proliferative activity was not high (1+). There were no foci of prostate tissue with simultaneous nuclear activity of p53 and Ki-67., Conclusion: Proliferative inflammation of different intensity in prostate tissues was observed after sixty days. Its severity was mainly determined by the type of infecting agent (S. haemolyticus > E. coli > P. niger) and was not dependent on the inoculated titer and the subsequent microbial load of prostate tissues. No areas of neoplastic transformation of prostate tissues were reliably identified in the case of a bacterial-induced inflammatory process in the estimated follow-up period.

The aim of this study is to describe the morphological and functional characteristics of the cardiac in sudden infants death syndrome (SIDS) in children under one year of age. Twenty eight cases of SIDS were studied histologically and immunohistochemically. Histological examination of the sectional material was carried out using standard and additional stains. The study of cardiac muscle tissue with routine staining with haematoxylin-eosin revealed interstitial oedema, uneven plethora of blood vessels: dystonia and weak plethora of part of the coronary arteries, excess of red blood cells in the veins as well as microcirculatory plethora with stasis of the erythrocytes. An immunohistochemical study revealed a mild over expression of p53 in cellular elements, small-focal expression of CD68 in cardiomyocytes apoptosis intramural areas, activation of mast cells (CD117), expression of ki-67 in macrophages, proliferation of fibroblasts. Additional forensic criteria for the diagnosis of SIDS were determined in the form of atrial endocardial fibrosis and interventricular septum; the expression of CD68, CD117 in fibroblasts, mast cells and lymphocytes; apoptosis of cardiomyocytes (expression of p53), proliferation of fibroblasts and remodeling of the heart (expression of ki-67).

Objective: Identification of the relationship between clinical and neuroimaging data and the content of serum biomarkers (damage marker - neuron-specific enolase (NSE), apoptosis - p53 protein and neuroplasticity - brain-derived neurotrophic factor (BDNF)) in acute, early and late recovery periods of ischemic stroke., Material and Methods: Eighty patients in the acute, early and late recovery periods of ischemic stroke, aged from 49 to 75 years, were examined. The comparison group included 20 patients with chronic cerebral ischemia, comparable to the study group. A clinical assessment was carried out on the following scales: The National Institutes of Health Stroke Scale (NIHSS), the European Stroke Scale (ESS), the modified Rankin scale, the Bartel Index and SS-QOL. In the acute, early and late recovery periods of ischemic stroke, the dynamics of biomarker levels (NSE, p53 protein, BDNF) in blood serum and brain MRI in T1, T2, FLAIR, DWI modes were quantified., Results: In patients with a favorable course in the acute period of stroke, low values of the NSE level and a trend towards increased levels of serum BDNF by the 10 th day of the disease were noted. For the first time, it was found that high NSE and stable BDNF levels or a tendency to their decrease were observed in the acute period in patients with an unfavorable course of ischemic stroke. In the early recovery period of ischemic stroke, a strong correlation was established between the degree of independence and disability of patients, and the levels of serum NSE and BDNF. A significant inverse correlation was proved between the severity of neurological deficit, assessed on the ESS, in the late recovery period of ischemic stroke (12 months) and the level of BDNF in blood serum ( r =-0.464)., Conclusion: The selected complex of biochemical methods allowed us to assess the severity of damage, the activity of apoptotic and compensatory neurotrophic capabilities of brain tissue in ischemic stroke, as well as their relationship with clinical and neuroimaging data. We have shown the prognostic significance of the selected markers, which will allow, with an integrated approach, more accurate prediction of the further course and outcome of stroke.

To date, no modern methods of treatment allow overcoming malignant potential of glial neoplasms and significant increase of survival. Analysis of glioblastoma radioresistance using cancer cell cultures is one of the perspective directions, as radiotherapy is standard and available treatment method for these neoplasms. This review summarizes current studies identifying many factors of radioresistance of glial tumors, such as hypoxia, microenvironment and metabolic features of tumor, stem cells, internal heterogeneity of tumor, microRNA, features of cell cycle, DNA damage and reparation. We obtained data on involvement of various molecular pathways in development of radioresistance such as MEK/ERK, c-MYC, PI3K/Akt, PTEN, Wnt, JAK/STAT, Notch, etc. Changes in activity of RAD51 APC, FZD1, LEF1, TCF4, WISP1, p53 and many others are determined in radioresistant cells. Further study of radioresistance pathways will allow development of specific target aptamers and inhibitors.

Seborrheic keratosis (SK) is a benign skin tumor of unknown etiology and pathogenesis. Many details remain unclear despite that there have been a number of studies of cell-cycle abnormalities., Aim: to investigate the expression of the cell-cycle regulatory proteins p53 and p16 and the cell proliferation marker Ki-67 in SK., Subjects and Methods: The investigation used intraoperative SK material obtained from 130 patients. Tumors were removed from UV-exposed parts of the body in 63 (48%) patients and from the places that were more often closed in 67 (51.5%). An immunohistochemical (IHC) study was performed using monoclonal antibodies to p53, p16, and Ki-67., Results: A positive reaction with monoclonal antibodies to p53 was recorded in 66 (50.7%) SK samples. In 92.1% of cases, the expression of p53 was found in SK located at the sites that were most exposed to UV radiation (p=0.00001). A positive reaction with monoclonal antibodies to p16 was observed in all SK cases as cytoplasmic staining of more than 50% of the tumor cells: a strong staining in 63 SK samples (overexpression) and a weak staining in 67 SK ones. The level of p16 expression correlated with age (R=0.21; p=0.019) and SK location at the sites exposed to increased insolation (R=0.35; p=0.000038). Overexpressions of p53 and p16 were significantly more commonly recorded in irritated SK. The tumor proliferative activity by the level of Ki-67 expression was low (3.0 to 11.3%). The largest number (8.5±4.8%) of proliferating cells was recorded in irritated SK (p=0.0000001)., Conclusion: The found disorders in the expression of cell-cycle regulatory proteins in SK are suggestive of tumor suppressor activation and keratinocyte senescence. There may be malignant tumor transformation in irritated SK in terms of the significant increase in the expression of p53, p16 in the presence of high cell proliferative activity.

Objective: To identify the expression of the molecular markers p53 and p16INK4A in head and neck squamous cell carcinoma (HNSCC) and to assess their impact on its clinical and morphological characteristics and overall survival (OS) rates in patients with HNSCC., Material and Methods: Histological blocks were immunohistochemically studied using anti-p16 and p53 monoclonal antibodies in Krasnodar Clinical Oncology Dispensary One in 2011 to 2016. Overexpression of p16INK4A was established in the presence of 3 and 4 staining points (nuclear and/or cytoplasmic staining in 40% or more tumor cells). That of p53 was determined in the presence of nuclear staining (3+) in more than 50% of tumor cells., Results: Overexpression of p16 was found in 15 (27%) patients (9 (60%) men and 6 (40%) women). The p16-positive tumor status was associated with the female sex (p=0.023), which was characteristic of tonsil cancer (p<0.001) and represented by the nonkeratinizing type (p=0.008). Overexpression of p16 was associated with more frequent regional lymph node metastases (p=0.029). Overexpression of p53 was related to G2 tumor (p=0.021) and expression of p53 was less than 50% associated with tongue body cancer (p=0.004). Kaplan-Meier analysis showed that the 3-year OS in p16-positive HNSCC patients was significantly higher than that in p16-negative ones (p=0.048). Significantly higher OS rates were observed in p16-positive HNSCC patients than in p16INK4A-negative ones for Stage III-IV (p=0.021). OS rates in HNSCC patients with co-expression of p16INK4A (3 and 4 points) and p53 (3+) were significantly higher than in the absence of a combination of these molecular markers (p=0.049). At the same time, OS in HNSCC patients with co-expression of p16INK4A (3 and 4 points), p53 (3+) was significantly higher than in the absence of a set of these molecular markers for stage III-IV (p=0.01). OS in patients with Stages I-II HNNSCC and co-expression of p16INK4A (3 and 4 points) and p53 (3+) did not significantly differ from that in the absence of a set of these molecular markers (p=0.960)., Conclusion: Overexpression of p16INK4A (3 and 4 points) can be used as a prognostic marker to divide patients into subgroups with different clinical and morphological characteristics. The data on the correlation of p53 overexpression as a marker of mutations in the TP53 gene are contradictory and the study has not revealed the worst overall survival rates. A set of markers for the expression of p16 (≥40%) and p53 (≥50%) has been proposed for use as a favorable prognostic sign.

Objective: Our aim was to study the mechanisms of hypoxia development at pregnancy associated with cytomegalovirus infection (CMVI)., Methods: 30 parturient women with CMVI relapse at the 25-28 weeks of pregnancy and their newborns were examined. Cytochrome C, Hsp-70, p53, Bcl-2 and caspase-3 in placenta homogenate were found out with serologic methods, the morphology of erythrocytes with cytophotometry, erythrocytes membrane proteins with disc-electrophoresis method, TBA-active products with V.B. Gavrilov's method, superoxide dismutase activity with spectrophotometry, 2.3-diphosphoglyceric acid (2.3 DPG) with I.S. Luganov's method, erythrocytes membrane microviscosity with fluorimethric method, oxyhemoglobin and methemoglobin with Evelyn and Malloy' method, and erythrocytes deformability with M. T. Lutsenko's method., Results: In blood erythrocytes of CMV-seropositive parturient women there was the decrease of cytoskeleton proteins: α- and β-spectrine was 1.14 times less, ankyrin was 1.62 times less, band 4. 1 protein was 1.29 times less; there was 1.87 times increase of antigen-binding glycophorin, 1.37 times growth of TBA-active products and 1.35 times drop of superoxide dismutase activity; the deformability index was 9.5 times less, 2.3 DPG was 1.22 times less and oxyhemoglobin was 1.06 times less. In placenta homogenate Bcl-2 was 1.5 times less, Hsp-70 was 2.5 times more, p53 was 6.1 times more, cytochrome C was 1.76 times more, caspase-3 was 3.86 times more. In umbilical cord blood erythrocytes 2.3 DPG was 1.3 times more and oxyhemoglobin was 1.06 times less., Conclusion: The obtained data proves that CMVI relapse at 25-28 weeks of pregnancy causes the disorder of morphofunctional state of mother's blood erythrocytes and their ability to oxygenation, the development offetoplacental barrier, the decrease offetus oxygen blood supply and the development of intrauterine hypoxia.

The effects of recombinant IL-2 on apoptosis of lymphocytes of healthy donors were studied in in vitro experiments. It was shown that the inductive and inhibitory effects of IL-2 on apoptotic process depend on the dose of the cytokine and cell microenvironmental conditions. Culturing of lymphocytes with recombinant IL-2 increases the percent of cells with reduced transmembrane potential, reduces the content of intracellular proteins Bcl-2, Bcl-X(L) and и Bax, and increases the level of Bad. The proapoptotic effect of this cytokine is realized with participation of nuclear transcription factor NF-κB and transcription factor P53.

The effect of 20 amino acids and the synthetic tetrapeptide cardiogen in concentration 10(-12) M was investigated in organotypic tissue culture on the cell proliferation and apoptosis development in myocard tissue explant in 3- and 24-months old rats. 7 of 20 amino acids stimulated the cell proliferation in the young rats. In the same time, only 2 amino acids were active in myocard culture from the old rats. The tetrapeptide cardiogen demonstrated the great stimulating effect on the proliferation both in tissues from young and old rats. The immunohistochemical study demonstrated a decrease of the p53 protein expression by cardiogen action. This fact can testify that cardiogen inhibits the apoptosis process in the myocard tissue.

EcoRI fragments of DNA isolated from the different mouse organs were hybridized to radioactivity labelled probe specific for the gene of oncoprotein p53. The analysis of the blot-hybridization points to the existence of the specific blockage of an EcoRI site flanking a 3.3 kb fragment of DNA including the pseudogene p53, isolated from the skin tissue. The existence of a polymorphous EcoRI site localized distally to the pseudogene p53 has been demonstrated in the DNA of mice of different lines.

It has been shown that phosphoprotein p53 may participate in the DNA-synthesis stimulation in the mixed culture of tumour cells and syngeneic embryonic mouse cells (10 days of pregnancy). This effect has been eliminated by the preliminary treatment of embryonic cells in vitro by the monoclonal antibodies to p53 (pAb421) in the model system. A conclusion is drawn that p53-epitopes of embryonic cell surface membrane take part in the formation of the determinant which is recognized by receptors of tumour cells. The stimulation of DNA-synthesis in tumour cells is resulted by this process.

The detailed analysis of mRNA structure coding for p53 protein from the intact and KonA stimulated lymphocytes from mouse spleen has shown both matrices to be identical. mRNAs have been analyzed by S1 mapping. Both mRNAs directed in vitro the synthesis of p53 protein reacting with monoclonal antibodies PAb421 and PAb248 both of which are specific for one of p53 forms synthesized by lymphocytes in vivo. Thus, the phenomenon of epitopes exclusion on p53 surface from intact or activated B-lymphocytes might be explained by peculiarities of posttranslational step of protein synthesis.

S1 mapping and nucleotide sequencing were used for localization of exon-intron junctions of the human p53 gene. The 3'-end of the gene was localized 12 nucleotides downstream AATAAA sequence, though an mRNA, 94 nucleotides shorter, was observed in one tumor. No typical promoter sequences were found at the 5'-end of the gene. In cell-free HeLa extracts transcription of RNA initiates within the sequence of possible hairpin-loop structure. Organization of the promoter region of the human p53 gene is discussed.

43 cDNA clones specific for murine cellular tumor antigen p53 were isolated from a library constructed using 17S fraction of mRNA from the SV40 transformed murine fibroblasts (SVT2). These clones contain the whole coding region of p53 mRNA and the most of non-translated sequences. A plasmid containing 1.8 kb insert of p53 cDNA was constructed. The p53 specific insert in this plasmid was colinear with p53 mRNA, as revealed by S1 nuclease analysis. 5'-region of the p53 gene comprising non-translated and promoter areas was cloned from the mouse genomic library. A combined clone containing promoter and the whole region, corresponding to p53 mRNA has been constructed.

Two allelic p53 genes were investigated. The allelic gene with BgIII site in the first intron codes for faster p53 protein variant, the other allelic gene coding for slower p53 protein variant (according to the mobility in SDS-PAAG). Exons and flanked regions of introns were sequenced. In coding regions allelic genes only differ by second nucleotides of codon 72 (G----C), which leads to the change in amino acid sequence (Arg----Pro). The relationship of these changes and the function of p53 is discussed.

p53 anti-sense sequences were introduced into normal NIH3T3 and transformed CMS 4 cells by infection with the recombinant retrovirus carrying a repeat of the 5'-terminal fragment of p53 cDNA. Clones selected for G418 resistance showed a marked inhibition of proliferative capacity and a reduced ability to enter DNA replication after stimulation of quiescent cells with serum. Clones showing moderate inhibition of proliferation were shown to contain truncated anti-sense DNA integrated into the genome. The anti-sense DNA was transcribed and it correlated with the reduction of the p53 protein level in the cell clones studied. We conclude that the appropriate expression of p53 appears to be required for cell proliferation.

Protein (m.v. 53 kilodalton) with electrophoretic mobility identical to that of protein detected in leukocytes of patients with Down's syndrome and of protein p53 obtained from mouse ascites carcinoma was demonstrated by polyacrylamide gel electrophoresis in healthy donors' leukocytes cultured with PHA and in peripheral blood of patients with chronic myeloleukemia. Appearance of the class p53 proteins in leukocytes of patients may be connected with the presence in their blood of the population of immature or transformed, proliferating or merely DNA-synthesizing leukocytes, particularly those amplifying the gene that codes protein p53.

Nucleotide sequencing of cDNA clone specific for murine oncoprotein p53 from the SVTZ all line has revealed nucleotide changes as compared with previously published structures of p53 cDNA originated from other cell lines.