TP63-TRIM29 axis regulates enhancer methylation and chromosomal instability in prostate cancer.

BACKGROUND: Prostate adenocarcinoma (PRAD) is the second leading cause of cancer-related deaths in men. High variability in DNA methylation and a high rate of large genomic rearrangements are often observed in PRAD. RESULTS: To investigate the reasons for such high variance, we integrated DNA methylation, RNA-seq, and copy number alterations datasets from The Cancer Genome Atlas (TCGA), focusing on PRAD, and employed weighted gene co-expression network analysis (WGCNA). Our results show that only single cluster of co-expressed genes is associated with genomic and epigenomic instability. Within this cluster, TP63 and TRIM29 are key transcription regulators and are downregulated in PRAD. We discovered that TP63 regulates the level of enhancer methylation in prostate basal epithelial cells. TRIM29 forms a complex with TP63 and together regulates the expression of genes specific to the prostate basal epithelium. In addition, TRIM29 binds DNA repair proteins and prevents the formation of the TMPRSS2:ERG gene fusion typically observed in PRAD. CONCLUSION: Our study demonstrates that TRIM29 and TP63 are important regulators in maintaining the identity of the basal epithelium under physiological conditions. Furthermore, we uncover the role of TRIM29 in PRAD development.

[Changes of a2-macroglobulin activity and endothelin-1 concentration in tears of rabbits after transplantation of retinal pigment epithelium cells derived from the induced pluripotent stem cells]. / Izmenenie urovnia a2-makroglobulina i éndotelina-1 v sleznoi zhidkosti krolikov posle transplantatsii kletok retinal'nogo pigmentnogo épiteliia, differentsirovannykh iz indutsirovannykh pliuripotentnykh stvolovykh kletok.

Retinal diseases accompanied with the dysfunction or death of the retinal pigment epithelial (RPE) cells are widespread, hard to treat, and appear to be a leading case of visual loss and blindness among the persons older than 55 years. Transplantation of RPE cells derived from the induced pluripotent stem cells (IPSC-RPE) is a promising method of therapy for these diseases. To ensure the transplant survival instant follow-up is required. It can be based on biochemical analyses of tear fluid that can be easily non-invasively collected. For the post-transplantation process monitoring we have choosen such polyfunctional bioregulators as α2-macroglobulin (α2-MG) and endothelin-1 (ET-1). RPE atrophy in New Zealand Albino rabbits was modeled via the subretinal injection of bevacizumab. IPSC-RPE in suspension or as a monolayer on the scaffold were transplanted subretinally 1 month after the injection. α2-MG activity and ET-1 concentration in tears were estimated during the first month and after 2, 3 and 7 months after transplantation. On the 7-14 days after transplantation α2-MG activity increased in tears of the both operated and controlateral eye probably as a reaction on the corticosteroid therapy. In 50% rabbits there was one more increase after 2-3 months that could be due to the immune inflammation. Concentration of ET-1 in tears decreased dramatically on the 7-14 days and 7 months after transplantation, and it could have an influence upon the retinal vassal tone. The data obtained show that estimation of bioregulators in tears can help monitoring local metabolic processes after RPE transplantation that is necessary for the opportune, reasonable and focused medicamental correction of post-transplantation process.

Generation of induced pluripotent stem cell line RCPCMi008-A derived from patient with spinocerebellar ataxia 17.

IPSC line RCPCMi004-8 was generated from skin fibroblasts collected from a male patient with spinocerebellar ataxia 17. The patient has expanded trinucleotide CAG repeats in the TBP (TATA-binding protein) gene on chromosome 6q27. The reprogramming of fibroblasts was performed with Sendai viruses containing Oct-4, Sox-2, Klf-4, and c-Myc. Pluripotency was confirmed by immunofluorescence, RT-PCR, and the formation of embryoid bodies. The RCPCMi008-A cell line carries the same trinucleotide CAG repeats in the TBP gene. The RCPCMi008-A cell line can be used to model Spinocerebellar ataxia in vitro.

Generation of induced pluripotent stem cell line RCPCMi004-A derived from patient with Parkinson's disease with deletion of the exon 2 in PARK2 gene.

IPSC line RCPCMi004-A was generated from skin fibroblasts collected from a male patient with early onset Parkinson's disease. The patient carries a heterozygous deletion of the exon 2 of PARK2 gene. The reprogramming of fibroblasts was performed with Sendai viruses containing Oct-4, Sox-2, Klf-4 and c-Myc. Pluripotency was confirmed by immunofluorescence, RT-PCR, and formation of embryoid bodies. The RCPCMi004-A cell line carries the same deletion in PARK2 gene. The RCPCMi004-A cell line can be used to model Parkinson's disease in vitro.

[The Role of Mutant RNA in the Pathogenesis of Huntington's Disease and Other Polyglutamine Diseases].

Polyglutamine diseases are rare, inherited neurodegenerative pathologies that arise as a result of expansion of trinucleotide CAG repeats in the coding segment of certain genes. This expansion leads to the appearance of mRNA with abnormally long repetitive CAG triplets (mCAG-RNA) and proteins with polyglutamine (PolyQ) tracts in the cells, which is why these pathologies are commonly termed polyglutamine diseases, or PolyQ diseases. To date, nine PolyQ diseases have been described: Huntington's disease, dentatorubral pallidoluysian atrophy (DRPLA), spinal and bulbar muscular atrophy (SBMA), and six different types of spinocerebellar ataxia (SCA 1,2,3,6,7, and 17). PolyQ diseases lead to serious, constantly progressing dysfunctions of the nervous and/or muscular systems, and there currently exists no efficacious therapy for any of them. Recent studies have convincingly shown that mCAG-RNA can actively participate in the pathological process during the development of PolyQ diseases. Mutant RNA is involved in a wide range of molecular mechanisms, ultimately leading to disruption of the functions of transcription, splicing, translation, cytosol structure, RNA transport from the nucleus to the cytoplasm, and, finally, to neurodegeneration. This review discusses the involvement of mutant mCAG-RNA in neurodegenerative processes in PolyQ diseases.

Possibilities for Using Pluripotent Stem Cells for Restoring Damaged Eye Retinal Pigment Epithelium.

The retinal pigment epithelium is a monolayer of pigmented, hexagonal cells connected by tight junctions. These cells compose part of the outer blood-retina barrier, protect the eye from excessive light, have important secretory functions, and support the function of photoreceptors, ensuring the coordination of a variety of regulatory mechanisms. It is the degeneration of the pigment epithelium that is the root cause of many retinal degenerative diseases. The search for reliable cell sources for the transplantation of retinal pigment epithelium is of extreme urgency. Pluripotent stem cells (embryonic stem or induced pluripotent) can be differentiated with high efficiency into the pigment epithelium of the retina, which opens up possibilities for cellular therapy in macular degeneration and can slow down the development of pathology and, perhaps, restore a patient's vision. Pioneering clinical trials on transplantation of retinal pigment epithelial cells differentiated from pluripotent stem cells in the United States and Japan confirmed the need for developing and optimizing such approaches to cell therapy. For effective use, pigment epithelial cells differentiated from pluripotent stem cells should have a set of functional properties characteristic of such cells in vivo. This review summarizes the current state of preclinical and clinical studies in the field of retinal pigment epithelial transplantation therapy. We also discuss different differentiation protocols based on data in the literature and our own data, and the problems holding back the widespread therapeutic application of retinal pigment epithelium differentiated from pluripotent stem cells.

[Genetic Cell Reprogramming: A New Technology for Basic Research and Applied Usage].

Gene function disclosure and the development of modern technologies of genetic manipulations offered the possibility of genetic reprogramming application to alter cell specialization. With the involvement of a gene set that encodes the transcription factors responsible for the pluripotent state, any cell of an adult body could be reprogrammed into the embryonal.state and pluripotency could be induced in this cell. Such reprogrammed cells were called induced pluripotent stem cells (iPSCs), and they are capable of again passing through all developmental stages. This provides new possibilities for studies of the basic mechanisms of developmental biology, the formation of specific cell types, and the whole body. In culture, iPSCs could be maintained permanently in a nontransformed state and permit genetic manipulations while maintaining their pluripotent properties. Such a unique combination of their properties makes them an attractive tool for studies of various pathologies and for the delineation of treatment approaches. This review discusses the basic and applied aspects of iPSCs biology.

No DNA damage response and negligible genome-wide transcriptional changes in human embryonic stem cells exposed to terahertz radiation.

Terahertz (THz) radiation was proposed recently for use in various applications, including medical imaging and security scanners. However, there are concerns regarding the possible biological effects of non-ionising electromagnetic radiation in the THz range on cells. Human embryonic stem cells (hESCs) are extremely sensitive to environmental stimuli, and we therefore utilised this cell model to investigate the non-thermal effects of THz irradiation. We studied DNA damage and transcriptome responses in hESCs exposed to narrow-band THz radiation (2.3 THz) under strict temperature control. The transcription of approximately 1% of genes was subtly increased following THz irradiation. Functional annotation enrichment analysis of differentially expressed genes revealed 15 functional classes, which were mostly related to mitochondria. Terahertz irradiation did not induce the formation of γH2AX foci or structural chromosomal aberrations in hESCs. We did not observe any effect on the mitotic index or morphology of the hESCs following THz exposure.

Late replication of the inactive x chromosome is independent of the compactness of chromosome territory in human pluripotent stem cells.

Dosage compensation of the X chromosomes in mammals is performed via the formation of facultative heterochromatin on extra X chromosomes in female somatic cells. Facultative heterochromatin of the inactivated X (Xi), as well as constitutive heterochromatin, replicates late during the S-phase. It is generally accepted that Xi is always more compact in the interphase nucleus. The dense chromosomal folding has been proposed to define the late replication of Xi. In contrast to mouse pluripotent stem cells (PSCs), the status of X chromosome inactivation in human PSCs may vary significantly. Fluorescence in situ hybridization with a whole X-chromosome- specific DNA probe revealed that late-replicating Xi may occupy either compact or dispersed territory in human PSCs. Thus, the late replication of the Xi does not depend on the compactness of chromosome territory in human PSCs. However, the Xi reactivation and the synchronization in the replication timing of X chromosomes upon reprogramming are necessarily accompanied by the expansion of X chromosome territory.

[Cytogenetic examination of nuclear specialists exposed to chronic beta-radiation of tritium].

For the first time the cytogenetic examination of the group of nuclear specialists (79 persons) chronically exposed to tritium beta-radiation for a long period was carried out. The frequencies of unstable (conventional method) and stable (FISH-method) chromosome aberrations have been analyzed. 50 years after the beginning of working under the conditions of the increased radiation level the differences (in comparison with control values) were revealed for all cytogenetic parameters. The frequency of the radiation-specific markers (dicentrics and centric rings) exceeds more than 2-fold the control level. A significant but poor correlation between the frequency of unstable aberrations and the total absorbed dose (during the whole working period) was revealed. A retrospective estimation of irradiation doses for 14 nuclear workers was made by the frequency of stable chromosome aberrations. The obtained dose values ranged from 110 to 1250 mSv.

Generation and characterization of human induced pluripotent stem cells.

Cell biology is one of the most rapidly developing branches in modern biology. The most interesting stages in early embryonic development for cell biology are those when a large number of cells are pluripotent. Inner-cell mass of blastocyst can be cultivated in vitro, and these cells are called embryonic stem cells. They are able to differentiate into different types of cells and tissues. But the greatest interest for practical application is the return (reprogramming) of adult cells into the pluripotent state. In our study for the first time induced pluripotent cells were derived from human umbilical vein endothelial cells by genetic reprogramming. We showed that these cells are similar to embryonic stem cells in their morphology, function, and molecular level. We are the first to show that reprogramming sufficiently changes X-chromosome chromatin state, which is normally inactive in female endothelial cells, towards its activation, providing evidence that endothelial cells are reprogrammed at an epigenetic level.

[Cytogenetic effects on blood lymphocytes of cosmonauts after low doses of space radiation].

Cytogenetic changes in cultures of blood lymphocytes from 37 cosmonauts returned from space flights of varying duration were analyzed. Prolonged stay in space was shown to increase numbers of stable and unstable chromosomal aberrations. Frequency of dicentric and centric rings depends on flight duration, as well as values of accumulated dose from and dose rate of space radiation. Egress into open space leads to an additional growth in the number of chromosomal aberrations. It was found that frequency of chromosomal aberrations in blood lymphocytes remains altered even after several years of cosmonaut's return from space flight. Based on the counts of symmetric translocations obtained with the FISH-technique, mean dose from space radiation following the maiden prolonged space flight amounted to 110 mGy which is in agreement with the biodosimetric data about frequency of dicentric and centric rings (140 mGy).

[Cytogenetic studies of human blood lymphocytes in vivo in a 7-day experiment with immersion].

The investigation had the goal to fulfill in vivo cytogenetic studies of blood lymphocytes of human subjects in a 7-d experiment with "dry immersion". Blood was sampled from 10 subjects prior to and on completion of the experiment. Analysis of chromosomal aberrations in blood lymphocytes after exposure to simulated microgravity did not reveal statistically reliable chromosomal alterations in comparison with spontaneous levels.

Chromosome aberration dosimetry in cosmonauts after single or multiple space flights.

BACKGROUND AND AIMS: Cosmic radiation is one of the main hazards for manned space exploration. Uncertainty in radiation risk estimates for crews of long-term missions are very high, and direct biological measurements are necessary. We measured chromosomal aberrations in peripheral blood lymphocytes from 33 cosmonauts involved in space missions during the past 11 years. METHODS: Blood lymphocytes from the cosmonauts were stimulated to grow in vitro and were harvested at their first mitosis. Slides were either stained with Giemsa stain for dicentrics analysis, or painted with whole-chromosome DNA probes for translocation analysis (FISH). RESULTS: A statistically significant increase in the yield of chromosomal aberrations was measured following long-term space missions in lymphocytes from cosmonauts at their first flight. No significant changes in aberration frequencies were observed for short-term taxi flights. The increase in long-term missions was consistent with the values calculated from physical dosimetry data. However, for cosmonauts involved in two or more space flights, the yield of interchromosomal exchanges was not related to the total duration of space sojourn or integral absorbed dose. Indeed, the yield of aberrations at the end of the last mission was generally in the range of background frequencies measured before the first mission. CONCLUSIONS: Chromosome aberration dosimetry can detect radiation damage during space flight, and biological measurements support the current risk estimates for space radiation exposure. However, for cosmonauts involved in multiple space missions the frequency of chromosomal aberrations is lower than expected, suggesting that the effects of repeated space flights on this particular endpoint are not simply additive. Changes in the immune system in microgravity and/or adaptive response to space radiation may explain the apparent increase in radioresistance after multiple space flights.

[Effect of age and radiation exposure on the frequency of translocations and dicentrics detected by FISH method in human lymphocytes]. / Vliianie vozrasta i oblucheniia na chastotu translokatsii i ditsentrikov, opredeliaemykh metodom FISH, v limfotsitakh cheloveka.

The frequencies of translocations and dicentrics detected by "chromosome painting" in lymphocytes were estimated in 115 healthy donors and in 273 people exposed to uncontrolled irradiation at low doses 1-4 years ago. Age responses of both types of exchanges at the age range from 3 to 85 years fit to quadratic model. The frequency of translocations grew faster with age than the frequency of dicentrics. The yields of stable exchanges in exposed people was significantly higher than those in control donors of corresponding ages.

[Tuberculosis epidemiological situation in the Republic of Belarus]. / Epidemiologicheskaia situatsiia po tuberkulezu v Respublike Belarus'.

The paper gives data on the spread of tuberculous infection in Byelarus. More preventive measures are shown to improve major epidemic indices, by stabilizing the situation primarily among children and adolescents. The epidemiological situation remains to be alarming among the contingents of the boarding facilities of the Ministry of Social Welfare and convicts at reformatories.

[Clinical and epidemiological characteristics of patients with drug-resistant tuberculosis]. / Kliniko-épidemiologicheskaia kharakteristika bol'nykh s lekarstvenno-ustoichivym tuberkulezom.

The paper analyzes the drug-resistant causative agent of tuberculosis. The bacterial isolation rates were studied among the cohorts registered at tuberculosis control facilities in the Republic of Byelarus in 1997. The epidemiological significance of patients isolating polyresistant Mycobacterium tuberculosis strains is shown.

[The dependence of the frequency of stable and unstable chromosome aberrations on the dose of the irradiation of human lymphocytes in vitro]. / Zavisimost' chastoty stabil'nykh i nestabil'nykh aberratsii khromosom ot dozy oblucheniia limfotsitov cheloveka in vitro.

Two methods (FISH and Giemsa) were used to estimate the frequency of stable and unstable chromosome aberrations in human lymphocytes exposed to gamma-irradiation in vitro in the dose range of 0.1 to 1.5 Gy. The DNA-probes specific to the chromosomes 1, 4, and 12 were used in combination with a pancentromeric probe. It was revealed that the dose-effect dependences for translocations (FISH) and dicentrics (FISH and Giemsa) were similar and could be adequately described by a linear-quadratic function.

[Stable chromosome aberrations in the lymphocytes of the peripheral blood in persons who suffered as a result of the accident at the Chernobyl Atomic Electric Power Station]. / Stabil'nye khromosomnye aberratsii v limfotsitakh perifericheskoi krovi lits, postradavshikh v rezul'tate avarii na ChAES.

The people suffered from Chernobyl accident exhibited the increased frequency of stable chromosome aberrations in peripheral lymphocytes as it was estimated by two-colour fluorescence in situ hybridisation with whole chromosome specific DNA probes for chromosomes 1, 4 and 12. However, the age dependence of this parameter in control group has to be estimated before the connection of this effect with radiation factor will be possible.