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Background: Abortion and fetal death are common in fetuses with holoprosencephaly, so genetic examinations often have to be made in a post-mortem setting. The efficiency of the conventional karyotyping using cultured fibroblasts in these situations is limited due to frequent culture failure. In the current study, archived cases of holoprosencephaly, where post-mortem genetic evaluation was requested and sufficient frozen material was available, were reevaluated using the quantitative fluorescence polymerase chain reaction (QF-PCR) technique., Methods: Testing for aneuploidies of chromosomes 13, 15, 16, 18, 21, 22, X, and Y with the QF-PCR technique was carried out on DNA isolated from archived frozen chorionic villi in seven cases of holoprosencephaly., Results: QF-PCR was successful in all seven cases. Two cases of trisomy 13, two cases of triploidy, and one case of trisomy 18 was found meaning a 71% diagnostic yield. The success rate of QF-PCR (100%, 7/7) was superior compared to conventional karyotyping (43%, 3/7)., Conclusions: Rapid aneuploidy testing using the QF-PCR technique is a simple, reliable, time- and cost-effective method sufficient to conclude the etiologic investigation in the majority of holoprosencephaly cases post-mortem., (© 2024 Wiley Periodicals LLC.)

Recent studies have shown that mitochondria are involved in the pathogenesis of Covid-19. Mitochondria play a role in production of reactive oxygen species and induction of an innate immune response, both important during infections. Common variability of mitochondrial DNA (mtDNA) can affect oxidative phosphorylation and the risk or lethality of cardiovascular, neurodegenerative diseases and sepsis. However, it is unclear whether susceptibility of severe Covid-19 might be affected by mtDNA variation. Thus, we have analyzed mtDNA in a sample of 446 Slovak patients hospitalized due to Covid-19 and a control population group consisting of 1874 individuals. MtDNA variants in the HVRI region have been analyzed and classified into haplogroups at various phylogenetic levels. Binary logistic regression was used to assess the risk of Covid-19. Haplogroups T1, H11, K and variants 16256C > T, 16265A > C, 16293A > G, 16311 T > C and 16399A > G were associated with an increased Covid-19 risk. On contrary, Haplogroup J1, haplogroup clusters H + U5b and T2b + U5b, and the mtDNA variant 16189 T > C were associated with decreased risk of Covid-19. Following the application of the Bonferroni correction, statistical significance was observed exclusively for the cluster of haplogroups H + U5b. Unsurprisingly, the most significant factor contributing to the mortality of patients with Covid-19 is the age of patients. Our findings suggest that mtDNA haplogroups can play a role in Covid-19 pathogenesis, thus potentially useful in identifying susceptibility to its severe form. To confirm these associations, further studies taking into account the nuclear genome or other non-biological influences are needed., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

The authors present a case of 1st trimester miscarriage where an early, complete hydatidiform mole was clinically suspected. Histopathological and immunohistochemical analyses excluded a complete mole, but the histomorphological profile was in concordance with a partial hydatidiform mole. Genetic analysis excluded a partial mole based on biparental genome composition, where further genetic analyses detected trisomy of chromosome 16. Trisomy of chromosome 16 is a frequent cause of 1st trimester abortions and may lead to highly abnormal placental histomorphology mimicking a partial mole. Genetic analyses are crucial for proper differential diagnosis and for the determination of adequate follow-up and prognosis for further pregnancies.

In this review, we summarized the results of experimental and clinical studies about three human endogenous retroviruses and their products-syncytin-1, syncytin-2, and suppressyn in human physiology and pathophysiology. We summed up the described connection with various pathological processes and diseases, mainly with pregnancy-induced hypertensive diseases such as preeclampsia, oncogenesis, gestational trophoblastic disease, and multiple sclerosis. Supposed mechanisms of action and the potential of clinical applications are also described., (© 2023. The Author(s).)

A case of double trisomy 16 and 22 in the second pregnancy loss is presented. DNA analyses (short tandem repeats genotyping) of miscarriage specimen was indicated because of ultrasound suspicion of partial hydatidiform mole. After the partial hydatidiform mole exclusion, further DNA analyses focused on the most common aneuploidies causing pregnancy loss, detected double trisomy 16 and 22 in the product of conception. The couple was referred to clinical genetic consultation and normal parental karyotypes were proved. For further explanatory purposes, archived material from the first pregnancy loss was analyzed and trisomy of chromosome 18 was detected. By comparison of allelic profiles of the mother, father, and both losses, the maternal origin of all aneuploidies was proven what can be attributed to frequent meiosis errors, probably due to advanced maternal age (44 years at the first loss and 45 years at the second loss). In conclusion, aneuploidies can mimic partial hydatidiform mole. Genetic analysis is helpful on the one hand to rule out partial hydatidiform mole and on the other hand to identify aneuploidies and in this way to determine the cause of miscarriage.

Objective: The aim of this study is to investigate the potential genetic etiology of cribra orbitalia noted on human skeletal remains., Materials: We obtained and analyzed ancient DNA of 43 individuals with cribra orbitalia. The analyzed set represented medieval individuals from two cemeteries in western Slovakia, Castle Devín (11th-12th century AD) and Cífer-Pác (8th-9th century AD)., Methods: We performed a sequence analysis of 5 variants in 3 genes associated with anemia (HBB, G6PD, PKLR), which are the most common pathogenic variants in present day of European populations, and one variant MCM6:c.1917 + 326 C>T (rs4988235) associated with lactose intolerance., Results: DNA variants associated with anemia were not found in the samples. The allele frequency of MCM6:c.1917 + 326 C was 0.875. This frequency is higher but not statistically significant in individuals displaying cribra orbitalia compared to individuals without the lesion., Significance: This study seeks to expand our knowledge of the etiology of cribra orbitalia by exploring the potential association between the lesion and the presence of alleles linked to hereditary anemias and lactose intolerance., Limitations: A relatively small set of individuals were analyzed, so an unequivocal conclusion cannot be drawn. Hence, although it is unlikely, a genetic form of anemia caused by rare variants cannot be ruled out., Suggestions for Further Research: Genetic research based on larger sample sizes and in more diverse geographical regions., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Background: Lynch syndrome (LS) is an autosomal dominant inherited disorder which causes an increased risk of cancer, especially colorectal and endometrial carcinomas. Recent studies have shown an association between LS and breast cancer as well. The aim of our study is to highlight the possible presence of mutations in genes associated with LS in patients with breast cancer and the need to include the examination of Lynch-associated genes in patients with a family history of breast cancer as well as in patients with recurrent breast cancer, as well as with the occurrence of other Lynch-associated cancer., Materials and Methods: We analyzed tumor tissue samples from 78 patients with primary breast cancer. Our samples were tested with a gene panel associated with the risk of developing breast cancer, while in our study we focused primarily on the occurrence of mutations in mismatch-repair genes. DNA isolated from tumor tissue was sequenced using next generation sequencing (NGS) and analyzed using the Ingenuity Variant Analysis tool. To confirm the germline mutation, we examined the patient's blood sample using NGS sequencing., Results: As a result of our analysis, we managed to identify a mutation in the PMS2 gene in one patient's breast tumor tissue. The presence of this mutation indicates that the resulting cancer may be a consequence of LS. As for pathogenicity, this was probably a pathogenic variant, as we detected deletions in the exon region, which led to frameshift mutation. Moreover, we also identified single-nucleotide pathogenic variants in the TP53 and PIK3CA genes. To definitively establish the diagnosis of LS in the patient, we examined a blood sample, where we also identified a mutation of the PMS2 gene., Conclusion: LS is underdiagnosed in many Lynch-associated cancers. However, in the case of a familial occurrence of breast cancer and other Lynch-associated genes, it is important to think about a possible diagnosis of LS and, if the patient meets the diagnostic criteria, to carry out a genetic examination of Lynch-associated genes.

Currently, when the world is fighting against the rapidly spreading pandemic of COVID-19, the silent epidemic of diabetes should not be set aside. In comparison, while COVID- 19 led to about 6 million deaths in 2021, diabetes caused 6.7 million deaths in the same year. Diabetes mellitus is a serious risk factor for worse outcomes in COVID-19 patients. Moreover, it seems that there is a bidirectional relationship between pre-existing diabetes pandemic and the rapidly spreading COVID-19 pandemic. In this article, we summarize mechanisms by which SARS-CoV-2 infects the host cell and discuss the bidirectional relationship between diabetes and COVID-19. We also focus on clinical variables in which diabetic patients differ from non-diabetic patients and which could have promising predictive value for the course and outcome of diabetic COVID-19 patients' therapy management., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The authors present a case of a partial hydatidiform mole where DNA analysis (STR - short tandem repeat genotyping) showed a triandric monogynic tetraploid genome composition with a XXXY gonosomal complement. This genetic finding clinicopathologically correlates with a partial hydatidiform mole, although it is rare in comparison with the typical, diandric monogynic triploid partial moles. The genetic analysis definitively confirmed the suspected diagnosis of a partial mole. To exclude the possibility that molar pregnancy represented retained products of conception after elective pregnancy termination, STR profiles from molar pregnancy and previous products of conception were compared. Short tandem repeats genotyping is a useful molecular genetic method in the differential diagnosis of partial hydatidiform moles, where clinical-pathological findings are frequently ambiguous.

Endometrial carcinoma (ECa) is one of the most common neoplasia of the female genital tract. The phosphatase and tensin (PTEN) homolog is the most frequently mutated tumor suppressor gene in endometrial carcinoma. PTEN encodes a phosphatase, a key regulatory enzyme involved in a signal transduction pathway that regulates cell growth, migration and apoptosis. The study evaluates an association between the morphological appearance of endometrial hyperplasia and ECa, and the presence of PTEN variations, PTEN protein´s level and intracellular localization. A total of 67 archived formalin-fixed and paraffin-embedded human biopsy tissue specimens with normal proliferative and secretory endometrium, endometrial hyperplasia without atypia and endometrial atypical hyperplasia, endometrioid the grade G1 and G3 and serous subtype of ECa were evaluated by sequencing for the presence of mutations in coding regions of PTEN gene of endometrial epithelial cells. The PTEN gene expression and intercellular localization of PTEN protein were evaluated immunohistochemically by immunoreactive score (IRS). PTEN mutation spectrum in endometrial carcinoma was identified for Slovak population. 28 non-silent mutations were identified in PTEN, twelve of them were novel, not annotated in Catalogue of Somatic Mutations in Cancer. Higher frequency of PTEN mutations was observed in serous carcinoma compared to global average. No correlation was observed between samples´ IRS, PTEN cellular localization and identified mutations. PTEN sequencing can be beneficial for patients considering prognosis of disease and sensitivity to treatment.

Colorectal cancer (CRC) is the 3rd most common malignant neoplasm worldwide, with more than two million new cases diagnosed yearly. Despite increasing efforts in screening, many cases are still diagnosed at a late stage, when mortality is high. This paper briefly reviews known genetic causes of CRC (distinguishing between sporadic and familial forms) and discusses potential and confirmed nucleic acid biomarkers obtainable from liquid biopsies, classified by their molecular features, focusing on clinical relevance. We comment on advantageous aspects such as better patient compliance due to blood sampling being minimally invasive, the possibility to monitor mutation characteristics of sporadic and hereditary CRC in a disease showing genetic heterogeneity, and using up- or down-regulated circulating RNA markers to reveal metastasis or disease recurrence. Current difficulties and thoughts on some possible future directions are also discussed. We explore current evidence in the field pointing towards the introduction of personalized CRC management.

Acute myeloid leukemia (AML) is a highly heterogeneous subtype of leukemia, accounting for 25 % of childhood leukemias. By the presence of genetic mutations in hematopoietic/ progenitor stem cells, the bone marrow produces a large number of abnormal undifferentiated leukocytes (blasts), which significantly impairs the proper differentiation of cells. AML is induced by two interventions. Chromosomal translocation during hematopoiesis of intrauterine development is the first intervention. This creates preleukemic fusion genes (PFG), which can later be transformed by a second intervention (point genetic mutation - deletion, insertion ) into a functional malignant clone. Characteristic AML fusion genes include AML1-ETO, PML-RARA or MLL-AF9, which in turn produce hybrid proteins with altered function. Several studies suggest that these PFGs are considered an important prognostic tool in disease assessment. While the incidence of PFG characteristic of acute lymphoblastic leukemia (ALL) has been relatively well studied by several research groups and has been estimated at 1 to 5% in the umbilical cord blood of healthy neonates, PFG relevant to AML are still not sufficiently clarified.

Fighting resistance to antibiotics and chemotherapeutics has brought bioactive peptides to the fore. Peptaibols are short α-aminoisobutyric acid-containing peptides produced by Trichoderma species. Here, we studied the production of peptaibols by Trichoderma atroviride O1 and evaluated their antibacterial and anticancer activity against drug-sensitive and multidrug-resistant bacterium and cancer cell lines. This was substantiated by an analysis of the activity of the peptaibol synthetase-encoding gene. Atroviridins, 20-residue peptaibols were detected using MALDI-TOF mass spectrometry. Gram-positive bacteria were susceptible to peptaibol-containing extracts of T. atroviride O1. A synergic effect of extract constituents was possible, and the biolo-gical activity of extracts was pronounced in/after the peak of peptaibol synthetase activity. The growth of methicillin-resistant Staphylococcus aureus was reduced to just under 10% compared to the control. The effect of peptaibol-containing extracts was strongly modulated by the lipoteichoic acid and only slightly by the horse blood serum present in the cultivation medium. Peptaibol-containing extracts affected the proliferation of human breast cancer and human ovarian cancer cell lines in a 2D model, including the multidrug-resistant sublines. The peptaibols influenced the size and compactness of the cell lines in a 3D model. Our findings indicate the molecular basis of peptaibol production in T. atroviride O1 and the potential of its peptaibol-containing extracts as antimicrobial/anticancer agents.

Plasma medicine is a new field focusing on biomedical and clinical applications of cold gas plasmas, including their anticancer effects. Cold plasmas can be applied directly or indirectly as plasma-activated liquids (PAL). The effects of plasma-activated cell growth medium (PAM) and plasma-activated phosphate buffered saline (PAPBS) were tested, using a plasma pen generating streamer corona discharge in ambient air, on different cancer cell lines (melanoma A375, glioblastoma LN229 and pancreatic cancer MiaPaCa-2) and normal cells (human dermal fibroblasts HDFa). The viability reduction and apoptosis induction were detected in all cancer cells after incubation in PAL. In melanoma cells we focused on detailed insights to the apoptotic pathways. The anticancer effects depend on the plasma treatment time or PAL concentration. The first 30 min of incubation in PAL were enough to start processes leading to cell death. In fibroblasts, no apoptosis induction was observed, and only PAPBS, activated for a longer time, slightly decreased their viability. Effects of PAM and PAPBS on cancer cells showed selectivity compared to normal fibroblasts, depending on correctly chosen activation time and PAL concentration, which is very promising for potential clinical applications. This selectivity effect of PAL is conceivably induced by plasma-generated hydrogen peroxide.

Syncytin-1 (gene ERVW-1 ) has been proposed as a marker of pre-eclampsia and malfunctions in placental development. Placenta is heterogeneous tissue, hence the method of biopsy can significantly affect the outcome of analyses. A total of 44 placentae were analyzed by taking 3-30 samples from each. Relative levels of ERVW-1 expression in the placental biopsies were characterized by RT-qPCR. Evaluation of ten biopsies from one placenta individually (not pooling them) is recommended due to the high variability of expression. No significant correlation was found between biopsy localization and level of ERVW-1 expression; therefore, random sampling is recommended. A long cut from the umbilical cord to the edge of the placenta is a convenient approach to placental sampling.

Objective: To summarize the possibilities of the genetic analysis of hydatidiform moles and point out its perspectives in the diagnostics of this disease., Design: Review., Setting: Institute of Medical Biology, Genetics and Clinical Genetics, Faculty of Medicine, Comenius University in Bratislava, Slovak Republic., Methods: Analysis of published literature data from the internet databases PubMed, ScienceDirect, Scopus and printed literature from the period 1963-2019., Results: This review refers on karyotyping, flow cytometry, FISH (Fluorescent in Situ Hybridization), VNTR-RFLP analysis (Variable Number of Tandem Repeats-Restriction Fragment Length Polymorphism), VNTR-PCR analysis (Variable Number of Tandem Repeats-Polymerase Chain Reaction) and STR (Short Tandem Repeat) genotyping of hydatidiform moles. The article summarizes possible application of these methods in the differential diagnostics of molar pregnancy (partial and complete hydatidiform moles) and nonmolar hydropic abortions., Conclusion: Genetic analyses offer precise identification of types of molar pregnancies when histopathological diagnosis is not clear during early stages of pathology.

Background: Statin treatment of hypercholesterolemia is accompanied also with depletion of the mevalonate intermediates, including farnesyl pyrophosphate (FPP) and geranylgeranyl pyrophosphate (GGPP) necessary for proper function of small GTPases. These include Ras proteins, prevalently mutated in pancreatic cancer. In our study, we evaluated the effect of three key intermediates of the mevalonate pathway on GFP-K-Ras protein localization and the gene expression profile in pancreatic cancer cells after exposure to individual statins., Methods: These effects were tested on MiaPaCa-2 human pancreatic cancer cells carrying a K-Ras activating mutation (G12C) after exposure to individual statins (20 μM). The effect of statins (atorvastatin, lovastatin, simvastatin, fluvastatin, cerivastatin, rosuvastatin, and pitavastatin) and mevalonate intermediates on GFP-K-Ras protein translocation was analyzed using fluorescence microscopy. The changes in gene expression induced in MiaPaCa-2 cells treated with simvastatin, FPP, GGPP, and their combinations with simvastatin were examined by whole genome DNA microarray analysis., Results: All tested statins efficiently inhibited K-Ras protein trafficking from cytoplasm to the cell membrane of the MiaPaCa-2 cells. The inhibitory effect of statins on GFP-K-Ras protein trafficking was partially prevented by addition of any of the mevalonate pathway's intermediates tested. Expressions of genes involved in metabolic and signaling pathways modulated by simvastatin treatment was normalized by the concurrent addition of FPP or GGPP. K-Ras protein trafficking within the pancreatic cancer cells is effectively inhibited by the majority of statins; the inhibition is eliminated by isoprenoid intermediates of the mevalonate pathway., Conclusions: Our data indicate that the anticancer effects of statins observed in numerous studies to a large extent are mediated through isoprenoid intermediates of the mevalonate pathway, as they influence expression of genes involved in multiple intracellular pathways.

Diabetic retinopathy (DR) is a multifactorial disease with complex pathophysiology. It is the main cause of blindness among the people in productive age. The purpose of this literature review is to highlight recent achievements in the genetics of diabetic retinopathy with particular focus on candidate gene studies. We summarized most of the available published data about candidate genes for diabetic retinopathy with the goal to identify main genetic aspects. We conclude that genetic studies reported contradictory findings and no genetic variants meet criteria of a diagnostic marker, or significantly elucidate the root of DR development. Based on these findings it is important to continue with the research in the field of DR genetics, mainly due to the fact that currently new possibilities and approaches associated with utilization of next-generation sequencing are available.

Phosphatase and tensin homolog (PTEN) is a protein that acts as a tumor suppressor by dephosphorylating the lipid second messenger phosphatidylinositol 3,4,5-trisphosphate. Loss of PTEN function has been implicated in the pathogenesis of a number of different tumors, particularly endometrial carcinoma (ECa). ECa is the most common neoplasia of the female genital tract. Our study evaluates an association between the morphological appearance of endometrial hyperplasia and endometrial carcinoma and the degree of PTEN alterations. A total of 45 endometrial biopsies from Slovak women were included in present study. Formalin-fixed and paraffin-embedded tissue samples with simple hyperplasia (3), complex hyperplasia (5), atypical complex hyperplasia (7), endometrioid carcinomas G1 (20) and G3 (5), and serous carcinoma (5) were evaluated for the presence of mutations in coding regions of PTEN gene, the most frequently mutated tumor suppressor gene in endometrial carcinoma. 75% of the detected mutations were clustered in exons 5 and 8. Out of the 39 mutations detected in 24 cases, 20 were frameshifts and 19 were nonsense, missense, or silent mutations. Some specimens harboured more than one mutation. The results of current study on Slovak women were compared to a previous study performed on Polish population. The two sets of results were similar.

Nowadays valid classification of gestational trophoblastic disease, according to the World Health Organisation from the year 2003, divides gestational trophoblastic disease into three groups - molar pregnancies, non-neoplastic non-molar changes of trophoblast and tumours of trophoblast. To the molar pregnancies belong complete, partial, invasive and metastatic hydatidiform mole. In the differential diagnosis it is important to distinguish the complete hydatidiform mole from other forms of gestational trophoblastic disease, because there is an increased risk of malignant transformation of trophoblast cells in complete hydatidiform mole. 10 cases of genetically confirmed diploid complete mole and 10 cases of genetically confirmed triploid partial mole were included into our retrospective study. All cases were examined microscopically in the basic haematoxillin and eosin staining and immunohistochemically with the use of antibodies against human choriogonadotropin hormone, placental alkaline phosfatase and protein p57. Villous cytotrophoblast, stromal villous cells, extravillous trophoblast and decidual cells were p57 positive in all cases of partial hydatidiform mole. All 10 cases of complete hydatidiform mole were p57 negative in stromal villous cells and villous cytotrophoblast. P57 protein is a marker distinguishing complete hydatidiform moles from partial moles.

Backgrounds: AML is an aggressive, phenotypically and genetically heterogenous clonal disease of hematopoietic progenitor cells with a great molecular variability. New WHO classification 2008 divides de novo AML according to cytogenetic and molecular prognostic and predictive markers. Recently, it is increasingly possible to identify a subgroup of poorer prognosis patients among those with normal karyotype AML. The aim of our study was to identify prognostically important molecular markers in children with AML, to stratify patients with normal karyotype and to monitor the disease according the genetic findings., Material and Methods: In 2008-2010, we analyzed bone marrow and peripheral blood samples of 20 children with de novo AML by conventional cytogenetic analysis, fluorescence in situ hybridisation and molecular diagnostics. The molecular analysis was performed on the cDNA level, with the restriction analysis of PCR products (FLT3-TKD), conventional PCR (MLL-PTD, NPM1mut, FLT3-ITD) and quantification RT-PCR method (expression of fusion transcripts, BAALC, WT1)., Results: Samples from 20 children with AML were analyzed using the conventional cytogenetics, FISH and molecular methods. Abnormal karyotype was identified in 13 patients (65%). Further analysis revealed FLT3-ITD in 5/20 (25%), FLT3-TKD in 3/20 (15%), NPM1mut in 2/20 (10%) and MLL-PTD in 1/20 (5%), overexpression of WT1 gene in 15/20 (75%) and overexpression of BAALC in 13/20 (65%) patients., Conclusion: Wide cytogenetic and molecular screening helped to find at least one genetic marker in all 20 patients for later follow-up and risk stratification. 4/20 (20%) patients died of the disease progression.

The aim of our study was to verify possible utilization of RT-PCR method (Reverse Transcriptase-Polymerase Chain Reaction) as a diagnostic and prognostic modality of the progression of prostate cancer. This approach is commonly used for the detection of circulating carcinomatous cells in peripheral blood of patients with malignant breast tumors, and our ambition was to adopt this method for patients with prostate cancer. The contribution of this method consists in its ability to detect early stages of the dispersion of carcinomatous cells, so called micrometastases, in the peripheral circulation of patients. The estimation of the progression of the disease is especially important for the selection of appropriate therapy for individual patients. Using this method we analyzed 50 men: 28 patients with clinically localized or locally advanced prostate cancer, 7 patients with clinically proven metastases, 8 patients with benign prostatic hyperplasia, and 7 healthy young men.

Evaluation of the frequency index for mtDNA particular sequences in the Caucasian population is crucial for forensic practice. There are two hypervariable regions in the mtDNA D-loop, HV1 and HV2. Both of them, 610 bp altogether, were sequenced, 342 bp from the hypervariable region HV1 (16024-16365) and 268 bp from the hypervariable region HV2 (73-340). We have analyzed 374 randomly selected non-related individuals of th e Caucasian population and 192 individuals of the Roma subpopulation living in Slovakia. The main goals of the work were to introduce and standardize methods of analysis of variability of the HV1 and HV2 regions of mtDNA for forensic use; to characterize the variability of mtDNA in the Slovakian population taking into account the subpopulations; classification of mtDNA profiles into haplotype groups, and comparison with other haplotype groups in Europe in the frame of phylogenetic studies.

The study of molecular markers in various types of human carcinomas, as well as in carcinoma of prostate, is focused on genes responsible for the formation of carcinoma. Mutation, amplification or other changes in these genes or in their protein products are being examined and compared with traditional prognostic markers. These genes can be characterized as oncogenes, tumor suppressor genes or genes for other significant cell functions. However, studies are often limited by heterogenity and multifocality of tumors, especially in prostate cancer. In this review, we offer a survey of some of the most frequent diagnostic and prognostic parameters of molecular biology research in relation to prostate cancer.

Objective: Study on Y chromosomal AZF region deletions in Slovak population, application of DNA technique., Design: Genetic-prospective study., Setting: Institute of Medical Biology, Genetics and Clinical Genetics, Faculty of Medicine, Comenius University in Bratislava., Methods: For detecting microdeletions in the Y-chromosomal AZF region in men with fertility disorders and for identifying Y-specific sequences we used the method of polymerase chain reaction (PCR) with using three different sets of sY sequences. For a verification of the specific type of deletion we used also fluorescently labeled kit., Results: Diagnoses of referred patients were divided into 2 groups: azoospermia, oligospermia. In the followed-up group of 822 patients there were 349 patients with azoospermia, 473 patients with oligospermia. Globally we reported 38 cases of deletions in the AZF region of the Y chromosome, i.e. 4.62%. 24 patients with deletion are from the group of patients with azoospermia, i.e. 6.88%, 14 patients are from the group of patients with oligospermia (2.95%). Considering particular types of deletions we recorded deletions in each region, AZFa, AZFb and AZFc, combinated AZFbc deletion, but also a complete deletion of the whole AZF region., Conclusion: The study confirmed that detection of microdeletions of the AZF region is significant from diagnostic and prognostic view and it pointed out the importance of selection criteria for selecting patients.

Background: It is well known that the frequency of multiple gestation increases after in vitro fertilization in which more than one embryo is transferred. The aim is to report an octuplet pregnancy following intracytoplasmic sperm injection and cryo embryo transfer., Case Report: A 31-year-old woman and her 35-year-old husband, both Caucasian, complained of primary infertility. The intracytoplasmic sperm injection and cryo embryo transfer procedure was recommended as treatment. Ovarian stimulation was performed using recombinant follicle-stimulating hormone and human menopausal gonadotropin. Two embryos were transferred to the uterus. This fertilization cycle was unsuccessful. Three months later, cryo embryo transfer of two frozen/thawed embryos was performed, which resulted in pathological monozygotic octuplet anembryonic pregnancy. Two, and later eight, empty sacs were detected by sonographic examination. Cytogenetic examination revealed normal male karyotype. DNA analysis confirmed identical polymorphisms in all the gestation sacs, i.e. a monozygotic origin of all eight sacs., Conclusions: We report a rare case of octuplet pregnancy after intracytoplasmic sperm injection and cryo embryo transfer.

RAG belongs to appropriate inhibitors of protein glycation, i.e. formation of advanced glycation end products, which are thought to be responsible for some complications of DM, including neuropathy, angiopathy, retinopathy and nephropathy. In the present study authors have evaluated the genotoxic effect of RAG on the cell culture of human neonatal fibroblasts (B-HNF-1) in regard to its potential clinical application as inhibitor of advanced glycation end products in relationships to the pathogenesis of chronic diabetic complications. The direct contact cytotoxicity assay and micronucleus test were performed. The results showed that RAG in the concentration range of 1 x 10-4 to 1 x 10-6 mol.l-1 did not induce any changes in the morphology of exposed B-HNF-1 cells. The frequency of micronuclei was not significantly increased as well. The inhibitive effect of resorcylidene aminoguanidine was directly proportional to its concentration. It can be concluded that RAG at the selected concentrations has an inhibitive effect on proliferation of the treated cells and, at the same time, does not display any genotoxic effects on B-HNF-1 cells.

Objective: DNA analysis of different forms of gestational trophoblastic disease., Design: Retrospective clinical study., Setting: Slovak Center of Trophoblastic Disease, Bratislava, Slovak Republic., Methods: In the period of September 1993 to April 2003, eighty-nine cases of gestational trophoblastic disease were analysed. There were 22 cases of partial hydatidiform moles, 58 cases of complete hydatidiform mole, 5 cases of invasive mole and 4 cases of gestational choriocarcinomas. Southern hybridization and polymerase chain reaction were used for DNA analysis., Results: From 22 analyzed cases of partial hydatidiform moles 19 (86.4%) were triploid and 3 (13.6%) diploid ones. There were 58 cases of complete hydatidiform mole and out of them 29 (50%) were homozygous, 28 (48.3%) heterozygous, and in one case (1.7%) both paternal and maternal genome was detected. In 8 cases of heterozygous and in one case of homozygous complete hydatidiform mole occurred a malignant transformation to gestational choriocarcinoma., Conclusions: Molecular analysis can determine the nuclear DNA origin of complete hydatidiform mole and allow us to define the patients with higher risk of malignant transformation usually to gestational choriocarcinoma.

During assessment of the progression of several types of carcinomas, such as cancer of the breast, lungs, prostate or urinary bladder world-wide the presence of circulating cells (micrometastases) in the circulation is followed up. These methods are gradually introduced also in Slovakia. First we tried in the Institute of Medical Biology and Genetics Medical Faculty Comenius University in collaboration with the Urological Clinic of Dérers the Faculty Hospital with policlinic to apply this method in carcinoma of the prostate (CaP). We detected the presence of epithelial prostate cells in the peripheral blood stream of patients with advanced prostate cancer where before secondaries were not detected.

The potential genotoxic activity of chemical substances in vitro is usually assessed by the micronucleus test and by karyological analysis. Use of the fluorescent plus Giemsa (FPG) technique is also recommended in the event that positive results are found in the micronucleus test, or if there is an increased rate of structural and numerical chromosome aberrations compared with controls. The tested substance, aminoguanidine (AG), has a marked ability to inhibit the toxic effects of carbonyl products (carbonyl stress) that arise during the end-phases of non-enzymatic protein glycation both in vitro and in vivo. The importance of this ability follows the finding that the production of advanced glycation end-products (AGE) is a part of the molecular mechanism of the pathogenesis of chronic diabetic complications. The aim of this study was to test the cytotoxic and clastogenic effects of AG on cells of the diploid cell line B-HEF-2, derived from a three-month-old male fetus. The results of the test did not reveal any induction of micronucleus production in the analyzed cells at AG concentrations ranging between 1 x 10(-2) and 1 x 10(-4) mol.L-1. Karyological analysis showed no clastogenic effect of the tested substance nor any increased rate of structural chromosome aberrations. The positive properties of AG and to its potential use as a glycoxidation inhibitor and AGE production are somewhat dimmed by its ionic nature, which hampers hydrophobic interaction with the nonpolar components of biological membranes. For this reason, the authors will further study the cytotoxicity and cytogenetic analysis of Schiff bases of AG synthesis on the basis of natural aldehydes (resorcine aldehyde, pyridoxal, etc.) in which antiglycation activity has been detected.

Human high-risk papillomaviruses (HR-HPVs) are involved in the induction of invasive cervical cancer. The aim of this study was to introduce a simple, semi-automated and reproducible approach suitable for HR-HPV detection in clinical practice. The procedure is based on DNA isolation, nested polymerase chain reaction, single strand conformational polymorphism and evaluation of HR-HPV genotypes with Gel-Pro software. The clinical performance of the new approach was assessed in two different patient materials: 1) cervical smears with cytological classification Pap2-3 or Pap3 lacking nuclear atypia (anisonucleosis and polychromasia) or koilocytotic atypia and without any previous therapy 2) formalin-fixed, paraffin-embedded cervical carcinoma and lymph node sections. Using the new approach we detected HR-HPV DNA in 64% patient samples cytologically classified as Pap2-3 or Pap3 respectively and in 80% formalin-fixed, paraffin-embedded lymph node sections histologically classified as lymph nodes without carcinoma cell infiltration. The combination of methods described in this study results in increased sensitivity of HR-HPV identification allowing detection of HPV DNA in a very small amount of target DNA so that it can be widely used in distinguishing the pre- malignant lesions and in determination of invading carcinoma cells to lymph nodes in patients with advanced cervical cancer. The new approach is useful in unambiguous HR-HPV genotyping even in double-HPV infection.

Objective: To evaluate subsequent pregnancy experience in patients following chemotherapy for malignant form of gestational trophoblastic disease., Design: Retrospective clinical study., Setting: Slovak Center of Trophoblastic Disease, Bratislava, Slovak Republic., Methods: There were evaluated subsequent pregnancy experiences in 38 patients after chemotherapy for malignant form of gestational trophoblastic disease registered in Slovak center of gestational trophoblastic disease. Histological and cytogenetical analysis of all placentas after deliveries and material from curettage specimens after miscarriages, abortions and ectopic pregnancies were performed., Results: 11 women conceived following successful chemotherapy of gestational trophoblastic disease became pregnant a total 19 times. Out of them there were 9 full-term deliveries, 2 spontaneous abortion, 1 ended in ectopic pregnancy and 7 pregnancies were terminated in therapeutic abortion. Cytogenetical analysis was successful in 7 to 10 reproductive losses with normal karyotype in all analysed cases., Conclusion: Patients after successful chemotherapy of gestational trophoblastic disease have a normal reproductive outcome.

Invasive hydatidiform mole is a relative rare form of gestational trophoblastic disease (GTD). Most of hydatidiform moles remit after evacuation but some of them have the tendency to invade the myometrium. In some rare cases the trophoblastic tissue can be found in other tissues like lungs, vulva, vagina or broad ligament. The aim of the study was to demonstrate some of clinical, immunohistochemical and DNA analysis findings of a patient with a previous diagnosis of a complete hydatidiform mole.

In this paper a set of medical and meteorological data for the period of years 1985-1987 is elaborated and statistically evaluated in order to find correlations between daily occurrence of ischemic and haemoragic forms of cerebral stroke and particular biotropic types of weather. For tracing we gathered clinical medical data from eight health-service institutions in Bratislava. We compared both annual and seasonal courses of biometeorological types of weather in the investigated period with annual and seasonal courses of the followed sickness rate. We have found that the most unfavourable weather type is the central low pressure area with transition of the frontal system. The most favourable atmospheric type is a sunny weather in the warm high pressure area without surface inversion. (Tab. 2, Fig. 4, Ref. 10.)

The aim of our study was to confirm the diagnosis of partial hydatidiform mole (PMH) and to determine mechanism of PMH development by chromosomal DNA analysis. In our study we analysed 8 cases of morphologically and histologically confirmed PMH. Their karyotype was determined by using methods of direct, 24-hour and a long-term tissue culture. The restriction fragment length polymorphisms (RFLP) method was used for DNA analysis of PMH chorionic villi and from progenitor lymphocytes in peripheral blood. All cases were triploid. DNA analysis revealed in six cases one maternal and two paternal RFLP bands, and in two cases two maternal and one paternal RFLP bands.

In our study we analyzed 13 cases of histologically defined complete hydatidiform mole (CHM), by using cytogenetic and molecular genetic methods. There were seven homozygous and six heterozygous CHM. In one case of heterozygous CHM we found a biparental contribution to genomic DNA. This is evidence of a more heterogeneous etiopathogenesis of hydatidiform mole.

Patients with familial adenomatous polyposis coli (FAP) reveal numerous colorectal adenomas as well as benign and malignant extracolonic lesions. Adenomatous polyposis coli (APC) gene mutations are the crucial genetic defect in FAP. The APC mutation molecular analysis of 20 FAP families was performed using the novel and effective method of the heteroduplex analysis (HDA). All of these families were screened for mutations in APC exon 15. APC mutations were identified in 4 individuals of two families. These two families were also screened by the protein truncation test (PTT). The PTT results confirmed previous findings obtained by HDA. The results of molecular analysis were correlated with the clinical manifestations of extracolonic lesions and congenital hypertrophy of retinal pigment epithelium (CHRPE). Positive correlation of all clinical examinations and mutations of APC gene was observed in all 4 FAP patients.

A 17-year old woman had in a short time period (seven months) a very exciting reproduction history. Molar pregnancy in December 1993, choriocarcinoma in January 1994 and induced abortion in June 1994. DNA analysis proved the origin of the choriocarcinoma in the previous molar pregnancy.