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N M Pleskach

Publications and source records attributed to N M Pleskach.

At least 19 recordsLinked to original sources
PubMed2005

[Characteristics of p53 protein stabilization in cells of patients with ataxia-telangiectasia after gamma-irradiation].

Stabilization of P53 protein in cells isolated from patients with a grave hereditary disease ataxia-telangiectasia (AT), characterized by strongly enhanced sensitivity to ionizing radiation and impairment of cell cycle control after DNA damage, was studied. The level of expression of these reactions by patients may vary, and it tends to be linked with the severity of the disease. In all AT strains studied, both acquired by the authors and obtained from foreign colleagues, we observed the alteration of timing and character of stabilization of P53 protein, after the action of ionizing radiation in sublethal dosage, as compared to that in cells from healthy donor.

Ataxia Telangiectasia↗
PubMed2004

[Age dymamics of stable chromosome aberration frequency in humans with natural and pathological senescence].

The age dynamics of stable chromosome aberration (SCA) frequency was analysed by fluorescent in situ hybridization (FISH) in human blood lymphocytes derived from donors, irradiated by low doses of ionizing radiation (Chernobyl clean-up workers, nuclear weapon testers, etc.) and patients with hereditary premature aging--Werner's syndrome and Hutchinson-Gilford's syndrome. It was found that the level of SCA was age-dependent and increased in irradiated persons. So, the SCA level may be really an index of a so-called "radiation senescence", and may show a real biological age of irradiated persons. The patients with Werner's syndrome demonstrate increased SCA level in blood lymphocytes, corresponding to the premature aging of the organisms. But in the case of another form of premature aging--Hutchinson--Gilford's syndrome-- no rise of SCA level was found. Some possible reasons of such results are discussed.

Adolescent↗
PubMed2003

[Recognition of internal (TTAGGG)n repeats by telomeric protein TRF1 and its role in maintenance of chromosomal stability in Chinese hamster cells].

Mammalian telomeres contain long tandem (TTAGGG)n repeats, which are protected by a complex of different proteins. Telomeric repeat-binding factors TRF1 and TRF2 play the key role in protection of telomeres through the formation of terminal loops (called T-loop). A T-loop isolates the 3' strand telomeric end and with this mechanism protects telomeres from the influence of enzymes of DNA reparation and telomere fusions and also interferes with the interaction of telomerase with telomeres. Many vertebrate species also contain large blocks of (TTAGGG)n sequences in pericentric and interstitial chromosome bands. The Chinese hamster genome contains a total of 18 arrays of these non-telomeric internal (TTAGGG)n sequences (ITs). Chromosome bands containing these arrays are unstable and should be protected with the help of another mechanism, rather than that using telomeres. In this study we analysed association of Green Fluorescent Protein (GFP)-tagged TRF1 in Chinese hamster V79 cells with ITs. We found that in these cells GFP-TRF1 associates with ITs in the interphase nucleus. We detected a little overlap between IT-associated GFP-TRF1 and random DSB sites visualized after the treatment of V79 cells with ionizing radiation. We found that the treatment of V79 cells with WM significantly increases the frequency of spontaneous chromosome aberrations. These WM effects are possible due to inhibiting phosphorylation of TRF1 by ATM. TRF1 is known to be eliminated from telomeres by overexpression of TANK1, which induces TRF1 poly(ADP-ribosyl)ation. We transfected V79 cells by plasmid encoding TANK1 and found that the frequency of chromosome rearrangements increased in these cells independently of their treatment by IR. Taken together, our results suggest that TRF1 may be involved in the sequence-specific protection of internal non-telomeric (TTAGGG)n repeats.

Animals↗
PubMed2002

[Cytological and molecular changes in the atypical case of accelerated human aging].

A complex research of cells of a patient with unusual form of premature ageing was made. The clinical picture is not typical for any of known forms of hereditary premature aging--progerias. Skin fibroblasts of the patient AG has limited proliferation capacity in vitro. It was shown by fluorescent-immunochemical hybridization (FISH-method), that the level of stable chromosome aberrations in AG blood lymphocytes was characteristic of aged 55-65 years, though as he was only 26 years old. Some characteristic peculiarities, typical for progerias, were found in the reaction of skin fibroblasts of AG to growth factors addition. Some clinical and biochemical peculiarities are results rather, than reasons of the disease. The conclusion is that the premature ageing in this case is a manifestation of Werner's syndrome--one of hereditary forms of accelerated senescence.

Adult↗
PubMed2001

Visualization of focal nuclear sites of DNA repair synthesis induced by bleomycin in human cells.

In this study, we examined DNA repair synthesis in human cells treated with the radiomimetic drug bleomycin, which efficiently induces double-strand breaks (DSBs). Using tyramide-biotin to amplify fluorescent signals, discrete nuclear foci from the incorporation of 5-iododeoxyuridine (IdU) were detected in proliferating human cells treated with bleomycin. We believe this comes from the repair of DSBs. An increase in the number of foci (>5 per nucleus) was detected in a major fraction (75%) of non-S-phase cells labeled for 30 min with IdU 1 h after the end of bleomycin treatment. The fraction of cells with multiple IdU-containing foci was found to decrease 18 h after treatment. The average number of foci per nucleus detected 1 h after bleomycin treatment was found to decrease twofold between 1 and 3.5 h, indicating that the foci may be associated with the slow component of DSB repair. The presence of DSBs in bleomycin-treated cells was confirmed using antibodies against phosphorylated histone H2AX (gamma-H2AX), which is strictly associated with this type of DNA damage. After treatment with bleomycin, non-S-phase cells also displayed heterogeneous nuclear foci containing tightly bound proliferating cell nuclear antigen (PCNA), suggesting an ongoing process of unscheduled DNA synthesis. PCNA is known to be involved in base excision repair, but a fraction of the PCNA foci may also be associated with DNA synthesis occurring during the repair of DSBs.

Antibiotics, Antineoplastic↗
PubMed2000

[UV-induced immobilization of DNA repair protein XPA in different human cell line].

XPA repair protein is absolutely needed for nucleotide excision repair (NER). It preferentially binds UV-irradiated DNA in vitro and possibly takes place in the recognition of pyrimidine dimers, the main type of UV-lesions in DNA. Using immunofluorescent microscopy and immunoblotting technique we have found that XPA protein is fully extractable by Triton X-100 solution from non-irradiated normal human fibroblasts, but after UV-irradiation its extractability decreases in UV-dose dependent manner. UV-induced XPA-immobilization was observed in human cell lines with different types of repair defects, but XPA-extractability from unirradiated cells of these lines was significantly lower in comparison with normal fibroblasts. These data do not permit to make conclusion concerning the distinct connection of this phenomenon with different pathways of NER. Histone deacetylase inhibitor, sodium butyrate, did not change the level of extractability in unirradiated and UV-irradiated normal human cells and CHO cells, defective in global genome repair, that indicated the independence of XPA-immobilization from the level of histone acetylation. It was established with the help of confocal microscopy that XPA-foci in detergent-treated UV-irradiated cell were partially colocalized with the focal sites of PCNA, an auxiliary protein of DNA polymerases delta and epsilon. It may mean that a part of detergent-resistant XPA foci correspond to DNA repair synthesis sites, but the major part of immobilized XPA reflects the early step of repair proteins assembly formation needed for the repair of the lesions.

Cell Line↗
PubMed1999

[Defect of preferential repair of gamma-ray-induced single-strand breaks in transcribed DNA in ataxia-telangiectasia cells].

Transcriptionally activity-dependent heterogeneity of gamma-induced DNA single-strand break repair in ataxia-telangiectasia cells has been investigated. It is found that in AT cells no increased induction of damage in the c-myc gene compares with normal human fibroblasts. At the same time an evident defect of preferential accelerated repair of single-strand breaks in the c-myc gene was observed. DNA single-strand breaks repair in human satellite III and total DNA single-strand breaks in c-myc gene in AT cells correlate with increased radiosensitivity of cells.

Ataxia Telangiectasia↗
PubMed1999

[Features of the radiosensitivity of cells from ataxia-telangiectasia patients].

Cell survival, single and double DNA strand breaks formation and removal, spontaneous and induced chromosome aberrations and sister chromatid exchange (SCE) levels in gamma-irradiated cells of patients with ataxia-telangiectasia (AT) were studied. Except SCE all of the above indexes of AT cells sensitivity, were higher, than in normal human cells, but lower, than it is commonly characteristic of AT cells in literature. A conclusion is that the analysed AT cells belong to the AT-variant form. Possible mechanisms of high radiosensitivity of AT cells, accompanied by radioresistance of DNA replication, are discussed. The authors suppose that the DNA repair defect in AT cells is not primary.

Ataxia Telangiectasia↗
PubMed1999

[Morphofunctional characteristics of fibroblasts in basal cell nevus and Cockayne syndrome].

Some morphofunctional characters of fibroblasts in two genetic disorders--Cockayne syndrome (CS) and Basal cell naevus syndrome (BCNS) have been examined. The size of nucleus in BCN1SP line has been shown to be about 1.5 times less as well as the total size of nucleoli per nucleus, while the number of nucleoli was 2 times more compared with other cell lines investigated. Using the method of silver staining numerous nucleoli were shown to contain active loci of the nucleolus organizer regions. With the help of hybridization in situ the number of transcripts of 18S RNA molecules was shown to be 5 times more in BCN1SP cell line, and about 2.8 times more than in the other cell lines tested. The data obtained may be interpreted as a suggestion in favour of a greater activity of the nucleolus organizer regions in BCN1SP cell line followed by the disturbance in protein homeostasis of the cells.

Cell Nucleus↗
PubMed1998

Decreased number of simultaneously operating adjacent clusters of replicons in some human strains with and without X-irradiation.

Several parameters of DNA replicons and replicon clusters have been examined using DNA fiber autoradiography in normal vs. mutant human cell lines showing increased chromosomal sensitivity to ionizing radiation as well as radioresistant DNA synthesis. Rates of synthesis of individual replicons for unirradiated ataxia telangiectasia (AT) AT5MO, basal cell naevus syndrome (BCNS) BCN1SP and Down's Syndrome LCH944 appeared to be in the range seen with two normal fibroblast lines. With the longer labelling times (60-165 min), the average track lengths were longer in normal fibroblasts than mutant cell; after 5 Gy of radiation, normal and mutant cells had similar track lengths for all labelling times (10-165 min), as well as unchanged rates of replicon synthesis. These observations led to the determination of 'chain length', which measures simultaneously active adjacent replicon clusters. The main finding is that 'chain length' in mutant lines was significantly lower than that in the normal fibroblasts; upon 5 Gy irradiation, the values in normal cells were reduced about two-fold while the values for mutant cells remained about the same as controls. Thus, the experiments suggest that in unirradiated mutant cells DNA replication is delayed in a comparable manner as that induced by ionizing radiation in normal cells. A possible relation of the data to the chromosomal radiosensitivity and radioresistant DNA synthesis in the mutant lines is discussed.

Ataxia Telangiectasia↗
PubMed1998

[Defect of preferential repair of gamma-ray-induced single-strand breaks in transcribed and non-transcribed DNA in Cockayne syndrome cells].

The repair of gamma-ray-induced DNA single-strand breaks in transcribed (protooncogene c-myc) and non-transcribed (human satellite III) DNA of normal human fibroblasts and fibroblasts obtained from a patient with Cockayne's syndrome (CS) has been investigated. A method of alkaline sucrose sedimentation was applied besides the Southern hybridization of 32P-DNA, containing sequences analysed with total 3H-DNA distributed through sucrose gradient fractions. No increase in the induction of DNA single-strand breaks was found in gamma-irradiated CS fibroblasts, compared to normal human fibroblasts. At the same time, an evident defect in the preferential repair of single-strand breaks in c-myc gene was observed.

Blotting, Southern↗
PubMed1998

[Organization of DNA replication domains in S-phase nuclei of human cells].

Each chromosome of eukaryotic cells contains multiple units of DNA replication that are activated during S-phase of cell cycle according to a definite program. It is considered at present that the main independent units of replication in mammalian cells represent groups of 20-25 adjacent synchronously activated small (with the average size 100 kbp) replicons. After labelling of nascent DNA with nonradioactive DNA precursors and immunofluorescent staining of incorporated label, discrete replication domains (RDs) are detected in S-phase nuclei. It is assumed that each RD is formed by a single group of synchronously activated small replicons. Since the average rate of replication fork movement is 2 kbp/min, a group of small replicons should finish DNA synthesis within 25 min, and only during this time one RD should incorporate the replicative label. We have studied the duration of DNA synthesis in individual RDs in S-phase human cells using double replicative labelling that can be detected in the nucleus by specific reagents. Our results indicate that in the main fraction of RDs DNA synthesis lasts more than 90 min, that contradicts the generally accepted model of organization of replication units in mammalian cells (Hand, 1978), but is in agreement with an alternative model, according to which the main replication units are single or clustered big replicons more than 300 kbp in size (Liapunova, 1994).

Cell Nucleus↗
PubMed1997

[The mechanism of radioresistant DNA synthesis in ataxia telangiectasia].

Using DNA fiber autoradiography, DNA replication in cells of healthy donors and in those of patients with ataxia telangiectasia (AT) was estimated. A new fact has been demonstrated showing a decreased number of simultaneously operating in tandem groups of replicon clusters in AT cells compared to that in normal cells. The data obtained suggest a reduced frequency of activation in the adjacent replicon clusters in AT cells. It should be noted that the rate of fork movement remained unchanged in AT cells. Besides, the frequency of replication in adjacent replicon clusters remained unaltered after 5 Gy irradiation in AT cells, while the normal cells were radiosensitive to reduction in this replication parameter to the low level seen in both non-irradiated and 5 Gy irradiated AT cells. The relation of the above data to radioresistant DNA synthesis is discussed.

Adult↗
PubMed1997

[Decreased survivability and a DNA repair defect in the cells of patients with xeroderma pigmentosum and Cockayne syndrome under the action of radiation and chemical mutagens].

The action of ionizing radiation and chemical mutagens--epoxides (ethylene oxide, propylene oxide, epichlorohydrin)--upon survival and repair processes in xeroderma pigmentosum (XP2SP) and Cockayne syndrome (CS1SP) patients' cells was studied, compared to healthy donor's cells VH-10 and C5RO. Ionizing radiation was demonstrated to enhance significantly higher survival decrease of XP2SP and CS1SP fibroblasts, compared to healthy donor's cells, according to the cloning efficiency criterion. In contrast to this, no significant difference between XP2SP and healthy donor's cells was found, according to cells' ability to replicative DNA synthesis after gamma irradiation. Differences in survival of mutant cells and healthy donor's cells after treatment by epoxides were found significant only following XP2SP being treated by ethylene oxide. DNA single-string breaks in XP2SP and in CS1SP cells treated by mutagens studied were proved to occur with the same frequency as in the DNA of the control cells; however the DNA repair according to this criterion was significantly suppressed in mutant cells.

Cell Survival↗
PubMed1997

[The isolation and analysis of lymphoblastoid cell lines from patients with xeroderma pigmentosum and progeria].

Lymphoblastoid cell lines from patients with xeroderma pigmentosum (2 forms) and progeria (unusual form) were established using transformation of peripheral blood lymphocytes by Epstein--Barr virus. The influence of different UV doses on cell vitality, proliferation and cell cycle progression was studied by means of flow cytometry. The cell vitality was determined after incubation of cells with etidium bromide and FDA. We used cytograms with two logarithmic signals (log green/log red) to discriminate the cell cycle status. Cell cultures were used with density of 500,000 cells per 1 ml, previously synchronized at G-phase by the incubation in a medium with low serum content. The effect of UV irradiation was followed during 72 h. Among four analysed cell lines only line XP2SP demonstrated enhanced UV sensitivity, expressed by decreasing of the amount of living cells after the UV dose of 2.5 J/m2 and higher. The cell cycle studies showed that cells were blocked in S-phase and simultaneously the amount of apoptotic cells with both reduced DNA content and ability to bind FDA was seen increased. Similar events were observed in the control line only after the dose of 20 J/m2 and higher.

Cell Line↗
PubMed1996

[The determination of the complementation groups for the cells of patients with xeroderma pigmentosum and the Cockayne syndrome found in Russia].

Complementation groups for xeroderma pigmentosum (XP) and Cockayne's syndrome (CS) cells have been first determined for patients encountered in the former Soviet Union. The determination was carried out using fusion of fibroblasts to be examined with those of already known complementation groups, and subsequently registering the level of DNA unscheduled synthesis (for XP cells) and RNA synthesis recovery (for CS cells) after UV-irradiation. The evidence of the complementation was normalization of these indexes. Cells of XP2SP and XP4SP patients are shown to fall under the XPC complementation group, whereas CS1SP cells are classified within the CSA complementation group.

Cell Fusion↗
PubMed1995

Slower synthesis of individual replicons and adjacent replicon clusters in a radiosensitive xeroderma pigmentosum strain with and without X-irradiation.

Two replication parameters, synthesis of individual replicons and adjacent replicon clusters, were measured using DNA fiber autoradiography in a radiosensitive form of group C xeroderma pigmentosum (XP) XP2SP, in group C XP4SP, in group A Cockayne syndrome (CS) CS1SP and two normal human fibroblast strains. The novel observation here is that in non-irradiated XP2SP cells synthesis of individual replicons was significantly retarded as compared with all other cell lines tested and remained unchanged after 5-Gy X-rays. Also the number of simultaneously operating adjacent replicon clusters was uniquely reduced in only non-irradiated XP2SP cells and remained unaltered after 5-Gy irradiation. While the normal, XP4SP and CS1SP cells are radiosensitive to reduction in this replication parameter to a low level seen in both non-irradiated and 5-Gy irradiated XP2SP cells. Thus, non-irradiated XP2SP cells mimic irradiated normal, XP4SP and CS1SP cells. A possible relation of the above abnormalities in individual replicons and adjacent replicon clusters to a high incidence of spontaneous sister-chromatid exchanges and X-irradiation-induced chromosomal aberrations in XP2SP cells is discussed.

Cells, Cultured↗