We performed a comparative cytogenomic analysis of cultured and uncultured uterine leiomyoma (UL) samples. The experimental approach included karyotyping, aCGH, verification of the detected chromosomal abnormalities by metaphase and interphase FISH, MED12 mutation analysis and telomere measurement by Q-FISH. An abnormal karyotype was detected in 12 out of 32 cultured UL samples. In five karyotypically abnormal ULs, MED12 mutations were found. The chromosomal abnormalities in ULs were present mostly by complex rearrangements, including chromothripsis. In both karyotypically normal and abnormal ULs, telomeres were ~40% shorter than in the corresponding myometrium, being possibly prerequisite to chromosomal rearrangements. The uncultured samples of six karyotypically abnormal ULs were checked for the detected chromosomal abnormalities through interphase FISH with individually designed DNA probe sets. All chromosomal abnormalities detected in cultured ULs were found in corresponding uncultured samples. In all tumors, clonal spectra were present by the karyotypically abnormal cell clone/clones which coexisted with karyotypically normal ones, suggesting that chromosomal abnormalities acted as drivers, rather than triggers, of the neoplastic process. In vitro propagation did not cause any changes in the spectrum of the cell clones, but altered their ratio compared to uncultured sample. The alterations were unique for every UL. Compared to its uncultured counterpart, the frequency of chromosomally abnormal cells in the cultured sample was higher in some ULs and lower in others. To summarize, ULs are characterized by both inter- and intratumor genetic heterogeneity. Regardless of its MED12 status, a tumor may be comprised of clones with and without chromosomal abnormalities. In contrast to the clonal spectrum, which is unique and constant for each UL, the clonal frequency demonstrates up or down shifts under in vitro conditions, most probably determined by the unequal ability of cells with different genetic aberrations to exist outside the body.

We report on the phenotype and the reproductive history of an adult female patient with an unbalanced karyotype: 8p23 and 18p11.3 terminal deletions and 8p22 duplication. The indication for karyotyping of the 28-year-old patient was a structural rearrangement in her miscarriage specimen: 45,ХХ,der(8;18)t(8;18)(p23;p11.3). Unexpectedly, the patient had the same karyotype with only one normal chromosome 8, one normal chromosome 18, and a derivative chromosome, which was a product of chromosomes 8 and 18 fusion with loss of their short arm terminal regions. Fluorescence in situ hybridization revealed that derivative chromosome was a pseudodicentric with an active centromere of chromosome 8. Array comparative genomic hybridization confirmed 8p and 18p terminal deletions and additionally revealed 8p22 duplication with a total of 43 OMIM annotated genes being affected by the rearrangement. The patient had minor facial and cranial dysmorphia and no pronounced physical or mental abnormalities. She was socially normal, had higher education and had been married since the age of 26 years. Considering genetic counseling, the patient had decided to conceive the next pregnancy through in vitro fertilization (IVF) with preimplantation genetic testing for structural chromosomal aberrations (PGT-SR). She underwent four IVF/PGT-SR cycles with a total of 25 oocytes obtained and a total of 10 embryos analyzed. Only one embryo was balanced regarding chromosomes 8 and 18, while the others were unbalanced and demonstrated different combinations of the normal chromosomes 8 and 18 and the derivative chromosome. The balanced embryo was transferred, but the pregnancy was not registered. After four unsuccessful IVF/PGT-SR cycles, the patient conceived naturally. Non-invasive prenatal testing showed additional chromosome 18. The prenatal cytogenetic analysis of chorionic villi revealed an abnormal karyotype: 46,ХХ,der(8;18)t(8;18)(p23;p11.3)mat,+18. The pregnancy was terminated for medical reasons. The patient has a strong intention to conceive a karyotypically normal fetus. However, genetic counseling regarding this issue is highly challenging. Taking into account a very low chance of balanced gametes, emotional stress caused by numerous unsuccessful attempts to conceive a balanced embryo and increasing age of the patient, an IVF cycle with a donor oocyte should probably be considered.

In the present study, we aimed to check whether uterine leiomyomas (ULs) with an apparently normal karyotype in vitro comprise “hidden” cell subpopulations with numerical chromosome abnormalities (heteroploid cells). A total of 32 ULs obtained from 32 patients were analyzed in the study. Each UL was sampled for in vivo and in vitro cytogenetic studies. Karyotyping was performed on metaphase preparations from the cultured UL samples. A normal karyotype was revealed in 20 out of the 32 ULs, of which 9 were selected for further study based on the good quality of the interphase preparations. Then, using interphase FISH with centromeric DNA probes, we analyzed the copy number of chromosomes 7 and 16 in 1,000 uncultured and 1,000 cultured cells of each selected UL. All of the ULs included both disomic cells representing a predominant subpopulation and heteroploid cells reaching a maximum frequency of 21.6% (mean 9.8%) in vivo and 11.5% (mean 6.1%) in vitro. The spectrum of heteroploid cells was similar in vivo and in vitro and mostly consisted of monosomic and tetrasomic cells. However, their frequencies in the cultured samples differed from those in the uncultured ones: while the monosomic cells decreased in number, the tetrasomic cells became more numerous. The frequency of either monosomic or tetrasomic cells both in vivo and in vitro was not associated with the presence of <i>MED12</i> exon 2 mutations in the tumors. Our results suggest that ULs with an apparently normal karyotype consist of both karyotypically normal and heteroploid cells, implying that the occurrence of minor cell subpopulations with numerical chromosome abnormalities may be considered a characteristic of UL tumorigenesis. Different frequencies of heteroploid cells in vivo and in vitro suggest their dependence on microenvironmental conditions, thus providing a pathway for regulation of their propagation, which may be important for the UL pathogenesis.

Polycystic ovary syndrome (PCOS) is one of the most common endocrine disorders in women of reproductive age. It is known that kisspeptin stimulates activity of GnRH neurons and secretion of FSH and LH, thus disruption of interaction between kisspeptin and its receptor leads to anovulation. The aim of the study was to investigate the role kisspeptin in the pathogenesis of polycystic ovary syndrome. Materials and methods. The study included 14 patients with classic phenotype of PCOS and 11 healthy women of the control group. All the patients underwent laparoscopy and hysteroscopy with histological examination of ovarian tissue and endometrium. Determination of kisspeptin, FSH, LH, prolactin, AMH, estradiol, estrone, androgens (free testosterone and dehydroepiandrosterone sulfate) levels in peripheral blood in healthy women and patients with PCOS was performed by ELISA on the 2d and 8th days of menstrual cycle. Progesterone levels were investigated on the 18th-22d days of menstrual cycle. Expression of kisspeptin and its receptor in ovarian tissue and endometrium was estimated using immunohistochemical method. Results. Level of kisspeptin in peripheral blood of patients with PCOS tends to increase compared to its level in the control group, but the found difference was not reliable. Direct correlation between serum level of kisspeptin and levels of LH, free testosterone and DHEA-S was revealed in patients with PCOS. Immunohistochemical study in patients with PCOS showed a significant increase in the area of expression of КІЅЅ1 and KISS1R receptor in endometrium and in ovarian biopsies compared to these values in the control group. Conclusion. The obtained data show a definite role of kisspeptin in pathogenesis of polycystic ovary syndrome, but further research is needed.

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Background. The comparative evaluation of morphological criteria of endometrial dysfunction in patients with primary infertility associated with pelvic inflammatory disease, external genital endometriosis and uterine myoma revealed common morphological manifestations in the form of an imbalance of the endometrium receptor profile on the background of the high frequency of chronic endometritis.The aim of the study was the comparative evaluation of ER and Pg receptors expression, inflammatory markers and an inhibitor of cyclin-dependent kinases p16ink4a in the endometrium of women with infertility associated with pelvic inflammatory disease, endometriosis and uterine myoma.Materials and methods. The study was included 298 patients: 95 patients with infertility associated with pelvic inflammatory disease, 73 women with infertility associated with external genital endometriosis degree I-II, 70 patients with infertility associated with uterine myoma and 60 patients with infertility associated with male factor. All patients were examined and treated in FSBSI “The Research Institute of Obstetrics, Gynecology and Reproductology named after D.O. Ott”. Endometrial biopsies were performed on 19-24 day of the cycle. Histological and immunohistochemical study of endometrial biopsies were performed by standard methods. The expression of ER and PgR receptors, inflammatory markers (CD8+, CD20+, CD4+, SD138+) and cyclin-dependent kinase inhibitor p16INK4a was studied by the immunohistochemical method. The evaluation of the markers expression was performed by semiquantitative method H-Score, as well as qualitative and quantitative methods of computer image analysis system “Morphology 5.0” (VideoTest, Russia). Statistical processing of the results was performed using statistical packages (STATGRAPHICS v.6.0).Results. There is a violation of the endometrium secretory transformation in patients with infertility associated with pelvic inflammatory disease, uterine myoma and external genital endometriosis on the background of the high frequency of chronic endometritis. Receptor imbalance is characterized by impaired ER and PgR receptor ratio in the glands and in endometrial stromal component as a result of chronic inflammation.