Anti-TPO-mediated specific features of the placenta immunohistochemical profile and possible mechanisms for fetal loss.

Passive transfer of antithyroid antibodies in mice leads to reproductive disorders. The purpose was to assess the placental tissue of experimental animals under the influence of the circulating thyroperoxidase antibodies. We performed an immunohistochemical examination of murine placentae after a passive transfer of thyroperoxidase antibodies. Placentae of mice that passively transferred IgG from healthy donors were used as control samples. For histological examination, 30 placental samples were selected from mice from the anti-TPO group and 40 placental samples were taken from mice from the IgG group. Immunostaining for VEGFR1, THBS 1, Laminin, CD31, CD34, FGF-ß, CD56, CD14, TNF-α, kisspeptin, MCL 1, and Annexin V was performed. There is a significant decrease in the relative area of the expression of VEGFR1 (23.42 ± 0.85 vs. 33.44 ± 0.35, P < 0.01), thrombospondin 1 (31.29 ± 0.83 vs. 34.51 ± 0.75, P < 0.01), CD14 (25.80 ± 0.57 vs. 32.07 ± 0.36, P < .01), CD56 (30.08 ± 0.90 vs. 34.92 ± 0.15, P < 0.01), kisspeptin (25.94 ± 0.47 vs. 31.27 ± 0.57, P < 0.01), MCL 1 (29.24 ± 1.06 vs. 38.57 ± 0.79, P < 0.01) in the labyrinth zone of the placentae of mice from the anti-TPO group compared with control group. A significant increase in the relative expression of laminin and FGF-ß was noted in the group of mice to which antibodies to thyroperoxidase were transferred, compared with the control group (36.73 ± 1.38 vs. 29.83 ± 0.94, P < 0.01 and 23.26 ± 0.61 vs. 16.38 ± 1.01, P < 0.01respectively). Our study exposed an imbalance of pro- and anti-angiogenic factors, decreased representation of placental macrophages and NK cells, abnormal trophoblast invasion processes, and insufficient expression of antiapoptotic factors in the placentae of mice in which anti-TPO antibodies were passively transferred.

Vitamin D significance in pathogenesis of endometriosis.

Genital endometriosis (GE) is a widespread multifactorial disease thus making it necessary to carry on studying its pathogeny in order to work out target therapy techniques. Studying vitamin D role in GE pathogeny is a new promising research trend. PATIENTS AND TECHNIQUE: 25(ОН)D level was determined in peripheral blood (PB) of 440 patients with GE and in peritoneal fluid (PF) - in 49 GE patients; the same test in PB was performed in 30 patients of the control group with the ovulatory menstrual cycle and no gynecologic pathology. 129 patients with GE, as well as 82 women of the control, underwent examination of vitamin D receptor (VDR) polymorphism gene in BsmI, FokI, and TaqI polymorphic locus. We examined vitamin D receptor expression in the eutopic and ectopic endometrium in 32 women with GE and in the endometrium of 20 women from the control group. We also compared the efficacy of combined therapy involving colecalciferol to the standard hormone modulating therapy. RESULTS: It was established that the prevalent GE forms are characterized by lower 25(ОН)D levels both in PB and in PF. It was also found that G/G genotype of VDR BsmI gene polymorphic variant 1.9 times increases GE occurrence risk. VDR expression was authentically lower in the ectopic endometrium compared to the eutopic endometrium. Patients with GE show no VDR expression cyclic variations in the endometrium which is contrary to the control. Therapy in combination with colecalciferol promotes a more expressed decrease of endometriosis-associated pain syndrome and psycho-emotional stabilization of GE patients compared to the standard hormone modulating therapy. CONCLUSION: Vitamin D deficiency plays a significant role in the pathogenesis of GE and Vit D may be applied as its targeted therapy.

Placental expression of endoglin, placental growth factor, leptin, and hypoxia-inducible factor-1 in diabetic pregnancy and pre-eclampsia.

OBJECTIVE: To evaluate a level of expression of endoglin (Eng), leptin (Lep), placental growth factor (PlGF), and hypoxia-inducible factor-1alpha (HIF-1α) in placenta among women with pre-eclampsia and diabetes mellitus (DM), considering the method of glycemia correction and preconception care. MATERIALS AND METHODS: A retrospective cohort study was conducted. A total of 124 women were divided into following groups: type 1 DM (n = 40), type 2DM (n = 31), gestational DM (n = 33), pre-eclampsia without DM (PE) (n = 10) and the control group (n = 10). The histochemical study was performed by using primary monoclonal antibodies to Eng, PlGF, Lep, and HIF-1α (Abcam, UK). RESULTS: The highest level of placental expression of Eng was observed in the PE group (20.34%). The same trend was also typical for T1DM (not planned) and insulin-treated groups: T2DM and GDM. An amount of cell with an PlGF expression was significantly higher in the control group (12.2%), while the lowest was observed in the pre-eclampsia group (1.18%) and T1DM (not planned) (1.26%). The placental leptin expression within each DM group was increased among the patients with unplanned pregnancy and those who received insulin therapy. We observed the lowest Lep expression in the PE group (6.3%). High level of HIF-1α expression was detected in T1DM (not planned) (30.44%) and PE (29.64%) as compared to the control group (11.62%). In T2DM and GDM insulin groups, the HIF-1α expression was significantly higher as compared to diet groups. CONCLUSION: The obtained data show that DM and pre-eclampsia are associated with changes in angiogenic and metabolic placental factor expressions. The degree of changes depends on preconception care and the control of glycemia level during pregnancy.

Cytogenomic Profile of Uterine Leiomyoma: In Vivo vs. In Vitro Comparison.

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.

Uterine Leiomyomas with an Apparently Normal Karyotype Comprise Minor Heteroploid Subpopulations Differently Represented in vivo and in vitro.

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 MED12 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.

Expression of HLA-G and KIR2DL4 receptor in chorionic villous in missed abortion.

OBJECTIVE: Human leukocyte antigen-G (HLA-G) is a molecule that was first known to confer protection to the fetus from destruction by the immune system of its mother. HLA-G expression is mainly restricted to the fetal-maternal interface on the extravillous cytotrophoblast, placenta, amnion.Methods: The purpose of this study is to investigate the HLA-G and KIR2DL4 expression in chorionic villous among 2 groups with missed abortion: group 1 - 27cases with normal karyotype and group 2 - 22 with fetal polyploidy. Criteria of inclusion: abortive material from two groups of women with missed abortion; 6-12 weeks gestational age, singleton pregnancy, cytogenetic of chorionic villous was obligatory - normal fetal karyotype and polyploidy of fetus.Results: During IHC investigations the average relative area of HLA-G expression in trophoblast was counted (in 1st group 33.9 ± 3.5 and in 2nd group 38.6 ± 2.8). Expression of HLA-G the most verified in extravillous chorion stroma. The average relative area of KIR2DL4 receptor was not statistically different among two groups (31.6 ± 2.4 and 32.2 ± 1.7). CONCLUSIONS: These results suggest the role of HLA-G for the progression in early reproductive losses. Low expression of HLA-G is associated with pregnancy complications and can be one of the reasons for spontaneous abortion.

Reproductive History of a Woman With 8p and 18p Genetic Imbalance and Minor Phenotypic Abnormalities.

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.

Expression of vitamin D and vitamin D receptor in chorionic villous in missed abortion.

The contemporary world despite its enough developed medicine and generally highly enlightened population faces a great problem of vitamin, micro-element and nutrient deficiency turning to become the XXI century pandemic. Along with that significant growth of interest can be seen towards vitamin D importance for reproductive physiology. The fact is that vitamin D receptors (VDR) have been detected in women's ovarium tissue, fallopian tubes, decidua and placenta. Some recent years studies have proven that vitamin D may act as immune regulator during implantation. During early pregnancy the trophoblast release vitamin D, which produces anti-inflammatory reaction and also induce decidual tissue growth for successive pregnancy. It was a comparison between the expression of Vitamin D and VDR in chorionic villous in cases of normal pregnancy and missed abortion groups. 64 samples of chorionic villous were taken: 32 from missed abortion and 32 from the induced abortion group. Abortive material was taken from two groups of women residing in North-West region of Russia: missed abortion and pregnancy terminated at woman's wish (induced abortion); 6-12 weeks of gestation, singleton pregnancy. Immune histochemical examination showed homogenous distribution of vitamin D and VDR expression in syncytiotrophoblasts, cytotrophoblasts and chorion villus stroma.Vitamin D expression relative area was 10,3% which is statistically different from the induced abortion group - 15,4% (p<0,01). VDR expression analysis showed its homogenous distribution in chorionic villus structures in both groups. High VDR expression was detected in chorion villus stromal components. In missed abortion group, the morphometry results showed distinctly lower relative area of vitamin D expression against the comparison group (35,9 ± 1,8; 56,1 ± 2,4 p < 0,01). Also in missed abortion group, positively significant correlation has been determined between the level of vitamin D in blood and VDR relative area expression (r = 0,412). In missed abortion group, definite vitamin D and VDR expression decrease was detected compared to the induced abortion group. The results witness vitamin D importance for pregnancy progress.