Lipid Alterations in Early-Stage High-Grade Serous Ovarian Cancer.

Epithelial ovarian cancer (OC) ranks first in the number of deaths among diseases of the female reproductive organs. Identification of OC at early stages is highly beneficial for the treatment but is highly challenging due to the asymptomatic or low-symptom disease development. In this study, lipid extracts of venous blood samples from 41 female volunteers, including 28 therapy-naive patients with histologically verified high-grade serous ovarian cancer at different stages (5 patients with I-II stages; 23 patients with III-IV stages) and 13 apparently healthy women of reproductive age, were profiled by high-performance liquid chromatography mass spectrometry (HPLC-MS). Based on MS signals of 128 differential lipid species with statistically significant level variation between the OC patients and control group, an OPLS-DA model was developed for the recognition of OC with 100% sensitivity and specificity R 2 = 0.87 and Q2 = 0.80. The second OPLS-DA model was developed for the differentiation between I-II OC stages and control group with R 2 = 0.97 and Q2 = 0.86 based on the signal levels of 108 differential lipid species. The third OPLS-DA model was developed for the differentiation between I-II OC stages and III-IV stages based on the signal levels of 99 differential lipid species. Various lipid classes (diglycerides, triglycerides, phosphatidylchlorines, ethanolamines, sphingomyelins, ceramides, phosphatidylcholines and phosphoinositols) in blood plasma samples display distinctly characteristic profiles in I-II OC, which indicates the possibility of their use as marker oncolipids in diagnostic molecular panels of early OC stages. Our results suggest that lipid profiling by HPLC-MS can improve identification of early-stage OC and thus increase the efficiency of treatment.

[Autoimmune markers for non-invasive diagnosis of endometriosis in women]. / Autoimmunnye markery dlia neinvazivnoi diagnostiki éndometrioza u zhenshchin.

Endometriosis is a common estrogen-dependent chronic disease in women of reproductive age; it is associated with dysregulation of the immune response, local inflammation, and increased formation of autoantibodies. The aim of the study was to investigate the profile of autoantibodies in women with endometriosis and to evaluate their diagnostic value using new modifications of enzyme immunoassay. In women with endometriosis of stage III-IV (n=39), a wide spectrum of autoantibodies was detected, mainly of class G, including antibodies to endometrial antigens (tropomyosin 3, tropomodulin 3), the enzyme α-enolase, steroid (estradiol, progesterone) and gonadotropic hormones. At the same time, the frequency of detection of IgG antibodies to tropomyosin 3, α-enolase, estradiol and human chorionic gonadotropin and their levels in patients with endometriosis were higher than in healthy women (n=26) (p<0.05). IgG-antibodies to tropomyosin 3, α-enolase and estradiol were characterized by higher diagnostic value for endometriosis. The diagnostic value was significantly increased when these antibodies were combined: the AUC reached 0.875 [0.772-0.978] (p<0.0001), the sensitivity and specificity were 83.3% each. Thus, autoantibodies to tropomyosin 3, α-enolase, and estradiol are promising for inclusion in the panel of biomarkers for non-invasive diagnosis of endometriosis.

Enhanced Expression of TLR8 in Placental Tissue in Preeclampsia.

The expression of TLR8 in the placental tissue was studied in specimens from women of reproductive age with early- and late-onset preeclampsia (12 and 8 patients, respectively). The reference groups included 15 women: 10 with uneventful full-term pregnancy and 5 with preterm operative delivery on gestation weeks 28-33. The expression of TLR8 in placental structures was maximum in early-onset preeclampsia (p<0.01) characterized by the gravest clinical course, while the expression of TLR8 in late-onset preeclampsia was comparable with that in full-term pregnancy. This significant increase of TLR8 expression in placental tissue seemed to reflect activation of the key proinflammatory factors of congenital immunity and induction of the systemic inflammatory response. Manifest differences in the expression of TLR8 in late- and early-onset preeclampsia confirmed the hypothesis on different variants of this condition.

[The content of cytokines IL-6, IL-8, TNF-α, IL-4 and the level of expression in macrophages CD86 and CD163 in peritoneal fluid has a reverse correlation with the degree of severity of external genital endometriosis]. / Soderzhanie tsitokinovIL-6, IL-8, TNF-α, IL-4 i uroven' ékspressii makrofagami CD86 i CD163 v peritoneal'noi zhidkosti imeet obratnuiu korreliatsiiu so stepen'iu tiazhesti naruzhnogo genital'nogo éndometrioza.

Concentrations of eight different cytokines and the level of expression of CD86 and CD163 macrophages were studied in peritoneal fluid in women with endometriosis. It was found that the concentration of both inflammatory (IL-6, IL-8, TNF-α) and anti-inflammatory cytokines (IL-4) as well as the level of macrophage expression of the proinflammatory marker CD86 and anti-inflammatory marker CD163 increased in women with mild external genital endometriosis (1-2 stage), and did not differ from the control group in women with severe endometriosis (3-4 stage). The content of IL-2, IL-10, CM-CSF and IFN-γ in the peritoneal fluid of women with endometriosis did not differ significantly from the control group. The results of the study indicate that the development of external genital endometriosis may be based on insufficient both inflammatory and anti-inflammatory activity of macrophages in the peritoneal fluid.

Selective Action of N-Arachidonoyl Dopamine on Viability and Proliferation of Stromal Cells from Eutopic and Ectopic Endometrium.

We performed a comparative study of the cytotoxic effect of endocannabinoid N-arachidonoyl dopamine (AA-DA) on cultured stromal cells of ectopic and eutopic endometrium. It was found that AA-DA in the concentration range of 1-20 µM produces more selective cytotoxic effect on the stromal cells of the ectopic endometrium due to interaction with cannabinoid type 1 receptor. In concentrations below 1 µM, AA-DA stimulated the proliferation of stromal cells of the eutopic endometrium and did not affect the division of ectopic endometrium cells. This effect was realized due to its interaction with cannabinoid type 2 receptor.

Correction to: Abstracts : 30th European Congress of Pathology.

In Poster Sessions, the author name G. Khlestova was incorrectly presented as 'K. Galina' in the authorship group for Abstract PS-18-017 (page S151). The name has been corrected in the authorship group shown above.

Proliferation of Peripheral Blood Lymphocytes and Mesenchymal Stromal Cells Derived from Wharton's Jelly in Mixed and Membrane-Separated Cultures.

We studied the effect of mesenchymal stromal cells on proliferation of CFSE-stained T cells in mixed and membrane-separated (Transwell) cultures and in 3D culture of mesenchymal stromal cells from Wharton's jelly. The interaction of mesenchymal stromal cells with mitogen-activated peripheral blood lymphocytes from an allogeneic donor was followed by suppression of T-cell proliferation in a wide range of cell proportions. Culturing in the Transwell system showed the absence of suppression assessed by the fraction of proliferating cells and by the cell cycle analysis. In 3D cultures, contact interaction of mesenchymal stromal cells and lymphocytes was demonstrated that led to accumulation of G2/M phase lymphocytes and G0/G1 phase mesenchymal stromal cells. The suppressive effect of mesenchymal stromal cells from Wharton's jelly is mediated by two mechanisms. The effects are realized within 6 days, which suggests that the therapeutic effects of mesenchymal stromal cells persist until their complete elimination from the body.

Anti-inflammatory and antioxidant effect of cerium dioxide nanoparticles immobilized on the surface of silica nanoparticles in rat experimental pneumonia.

A massage with the potent counter-inflammatory material, cerium dioxide nanoparticles, is promising and the antioxidant properties of CeO2 are considered the main, if not the only, mechanism of this action. Nevertheless, the elimination of ceria nano-particles from the organism is very slow and there is a strong concern for toxic effect of ceria due to its accumulation. To overcome this problem, we engineered a combined material in which cerium nanoparticles were immobilized on the surface of silica nanoparticles (CeO2 NP), which is shown to be easily removed from an organism and could be used as carriers for nano-ceria. In our study particle size was 220±5nm, Zeta-potential -4.5mV (in water), surface charge density -17.22µC/cm2 (at pH 7). Thirty-six male Wistar rats, 5 months old and 250-290g were divided into four groups: 1) control; 2) CeO2 NP treatment; 3) experimental pneumonia (i/p LPS injection, 1mg/kg); and 4) experimental pneumonia treated with CeO2 NP (4 times during the study in dosage of 0.6mg/kg with an orogastric catheter). Gas exchange and pulmonary ventilation were measured four times: 0, 1, 3 and 24h after LPS injection in both untreated and CeO2 NP-treated animals. The mRNA of TNF-α, Il-6, and CxCL2 were determined by RT-PCR. ROS-generation in blood plasma and lung tissue homogenates were measured by means of lucigenin- and luminol-enhanced chemiluminescence. Endotoxemia in the acute phase was associated with: (1) pathological changes in lung morphology; (2) increase of ROS generation; (3) enhanced expression of CxCL2; and (4) a gradual decrease of VO2 and VE. CeO2 NP treatment of intact animals did not make any changes in all studied parameters except for a significant augmentation of VO2 and VE. CeO2 NP treatment of rats with pneumonia created positive changes in diminishing lung tissue injury, decreasing ROS generation in blood and lung tissue and decreasing pro-inflammatory cytokine expression (TNF-α, Il-6 and CxCL2). Oxygen consumption in this group was increased compared to the LPS pneumonia group. In our study we have shown anti-inflammatory and antioxidant effects of CeO2 NP. In addition, this paper is the first to report that CeO2 NP stimulates oxygen consumption in both healthy rats, and rats with pneumonia. We propose the key in understanding the mechanisms behind the phenomena lies in the property of CeO2 NP to scavenge ROS and the influence of this potent antioxidant on mitochondrial function. The study of biodistribution and elimination of СеО2NP is the purpose of our ongoing study.

Ultrastructural Characteristics of Placental Telocytes.

Telocytes of placental villi were studied by electronic microscopy during physiological pregnancy. Ultrastructural features of telocytes indicating their heterogeneity and presence of at least three types of villi depending on their localization and kind were observed. All placental telocytes were characterized by small amount of organelles including mitochondria. Presence of long thin processes, which generated a branching network by contacting with each other, served as a typical feature of telocytes including telocytes of the stroma and intermediate villi. Telocytes were absent in the terminal villi.

Conditions for Collection of Placental Tissue Samples for Culturing of Multipotent Mesenchymal Stromal Cells.

Placentas from women aged 25-32 years with normal course of gestation were studied. It is essential to stick to certain methodological approaches for preparing viable multipotent mesenchymal stromal cell culture and to carry out morphological (macro and micro) evaluation of the chorionic villi, umbilical cords, and placentas. At stage I of the study, patients' histories, labor course, and examinations of the newborns should be analyzed to exclude women with genital and extragenital diseases. At stage II, it is essential to stick to special regulations and methods for collection of specimens of the cord, amnion, and placental tissue proper. Histological control of the placental structures collected for multipotent mesenchymal stromal cell culturing is obligatory.

[Comparison of the expression profile of surface molecular markers on mesenchymal stromal cell cultures isolated from human endometrium and umbilical cord]. / Sravnenie profilia ékspressii poverkhnostnykh molekuliarnykh markerov na mezenkhimnykh stromal'nykh kletkakh kul'tur, poluchennykh iz éndometriia i pupoviny cheloveka.

Endometrial mesenchymal stromal cells (eMSCs), along with mesenchymal stromal cells (MSCs) isolated from other tissues, are promising for use in regenerative medicine. The benefits of eMSCs include their presence in adults, simplicity of isolation, high proliferative and differentiation capacity. In this study, we have employed the flow cytometry technique to assess expression of 28 molecular markers on the surface of two eMSCs cultures. The culture of MSCs isolated from Wharton's jelly of the umbilical cord (uMSCs) was used as a reference, because uMSCs were studied in details earlier and demonstrated their effectiveness in vivo. Both types of MSCs demonstrated similar expression profiles. They included stem cells surface molecules, cell adhesion molecules and their ligands, some receptor molecules responsible for cell metabolism and proliferation, as well as immunological response molecules.

Effect of Fibroblast-Like Cells of Mesenchymal Origin of Cytotoxic Activity of Lymphocytes against NK-Sensitive Target Cells.

We studied immunosuppressive properties of skin fibroblasts and mesenchymal stromal cells against NK cells. In vitro experiments showed that mesenchymal stromal cells isolated from human umbilical cord and human skin fibroblasts can considerably attenuate cytotoxic activity of NK cells against Jurkat cells sensitive to NK-mediated lysis. NK cells cultured in lymphocyte population exhibited higher cytotoxic activity than isolated NK cells. Mesenchymal stromal cells or fibroblasts added 1:1 to lymphocyte culture almost completely suppressed NK cell cytotoxicity. This suggests that fibroblast-like cells can suppress not only isolated NK cells, but also NK cells in natural cell microenvironment.

[Morphological Features of Mesenhymal Stroma Cells of Chorionic Villi].

Background: Nowadays autologous mesenchymal placental stromal cells (MSCs) may use to treat for various diseases both of the mother and the child. Stroma of the placenta villi is appropriated origin for cell culture isolation. Aim: of the study was to evaluate the possibility for selection and use of placental tissue for mesenchymal stromal cells. Materials and methods: The present study was based on 45 placental samples of women aged 27−38 yy. who underwent surgical delivery at 36−40 weeks of gestation. 30 of these women have been enrolled in the basic group including children with congenital abnormalities (CA). The comparison group consisted of 15 patients with physiological pregnancy. We performed histological examination (with hematoxylin and eosin staining), immunohistochemical examination (with use monoclonal antibodies CD90 (1:25; Abcam, UK), СD105 (1:500; Abcam, UK), CD44 (1:25; Dako), СD73 (1:200, Abcam, UK), and electron microscopy (by microscope Philips/FEI Corporation, Eindhoven, Holland). Eclipse 80i microscope (Nikon Corporation, Japan) was used to examine the immunohistochemical reactions as a brown staining. The evaluation of the intensity of reaction was conducted by NIS-Elements Advanced Research 3.2 program (Czech Republic). Student's t-test and analysis of variance were used to compare the mean values. Differences were considered statistically significant at p<0.05. Results: Interstitial cells of the stroma of the villi with CA had fibroblastic differentiation as revealed degenerative changes of the cells. The histologic examination with hematoxylin and eosin staining revealed significant fibrosis of the stroma of the placenta villi in CA group (p<0,01). Immunohistochemical study of stem and intermediate chorionic villi revealed no significant differences in staining of CD44+, СD90+, СD73+, and CD105+ cells if compared to the control group (p>0.05). Although CD105 expression was significantly lower in the CA group (0.058±0.0049) than in the control group (0.088±0.0039) (p<0.05). However, electron microscopy detected the villi interstitial stromal cells with fibroblastic differentiation in CA group. Conclusions: Thus, it is necessary to exclude placenta with obstetrical history, somatic, and congenital pathology of the mother and the child when selecting the placental cell culture. Moreover, choosing a sample the morphological structure of the placenta should be taken into consideration. However, congenital malformations of the fetus, pathology of the mother cultivate mesenchymal stromal cells of placentas is inappropriate and should be taken advantage of the donor cells.

Hyaluronic Acid in Vascular and Immune Homeostasis during Normal Pregnancy and Preeclampsia.

Preeclampsia (PE) is a multisystem pathologic state that clinically manifests itself after the 20th week of pregnancy. It is characterized by high maternal and perinatal morbidity and mortality. According to modern concepts, the impairment of trophoblast invasion into maternal spiral arteries, leading to the development of ischemia in placenta, is considered to be the major pathogenetic factor of PE development. Ischemic lesions initiate the development of a systemic inflammatory response (SIR) and endothelial dysfunction, which is the main cause of the multiple organ failure in PE. Some data has appear indicating the importance of a glycans-forming endothelial glycocalyx and extracellular matrix (ECM) for placenta morphogenesis, as well as their role in the regulation of vascular permeability and vascular tone in hypertension disorders and, in particular, PE. Since intact glycocalyx and ECM are considered to be the major factors that maintain the physiological vascular tone and adequate intercellular interactions, their value in PE pathogenesis is underestimated. This review is focused on hyaluronic acid (HA) as the key glycan providing the organization and stabilization of the ECM and glycocalyx, its distribution in tissues in the case of presence or absence of placental pathology, as well as on the regulatory function of hyaluronic acids of various molecular weights in different physiological and pathophysiological processes. The summarized data will provide a better understanding of the PE pathogenesis, with the main focus on glycopathology.

Comparison of the Expression Profiles of Immune Response Gene mRNA in Umbilical and Venous Blood of Newborns of the First Day of Life.

The expression of immune response gene mRNA in the umbilical and venous blood were compared in newborns of the first day of life with and without signs of infection. The expression of il1b, il6, il8, il10, il12a, il15, il18, tnfa, tgfb1, tbx21, gata3, foxp3, rorc2, cd45, cd68, cd69, tlr2, tlr4, tlr9, and mmp8 mRNA was evaluated in umbilical and venous blood cells of newborns by reverse transcription real time PCR. In full-term newborns without signs of infection, the expression of il8, tlr2, tlr4, and mmp8 in venous blood was higher than in umbilical blood, while in preterm newborns, the levels of mmp8 transcript were elevated while the levels of tlr9, cd45, and gata3 were reduced. The expression of some markers differed in the umbilical and venous blood and in newborns with congenital infectious disease and without signs of infection.

Mesenchymal stem cells from human dental pulp: isolation, characteristics, and potencies of targeted differentiation.

We studied cell cultures isolated from the pulp of third molar germ of an adult human and from the skin of a human fetus on gestation day 10. Both cultures expressed similar repertoire of surface markers typical of multipotent mesenchymal cells (CD44, CD90, and CD105). Under in vitro conditions, dental pulp cells were more susceptible to factors inducing their differentiation into adipogenic, chondrogenic, and osteogenic lineage cells.

Isolation of mesenchymal stromal cells from extraembryonic tissues and their characteristics.

We describe a method of isolation of human mesenchymal stromal cells from the umbilical cord (Wharton's jelly) and human placenta: amnion, placental villi, and trophoblast. Morphology, immunophenotypic characteristics, and differentiation potencies of isolated cells were studied. The capacity of mesenchymal stromal cells from extraembryonic tissues to osteogenic, adipogenic, and chondrogenic differentiation was demonstrated and the dynamics of this process was described. The isolated cells met the criteria for multipotent mesenchymal stem cells.

Serum metabolic profiles of pregnant women with burdened obstetrical history.

The content of low-molecular-weight components in blood serum was studied by tandem mass-spectrometry in pregnant women. Serum metabolic profiles of patients with a grave obstetrical history were detected. The most significant changes were observed for the concentrations of low-molecular-weight substances involved in glucogenesis and ß-oxidation processes and in metabolic chains involving carbohydrates, carnitines, amino acids, and lipids.

Analysis of purine metabolites in maternal serum for evaluating the risk of gestosis.

Metabolome analysis of the serum from pregnant patients aimed at detection of low-molecular-weight biomarkers of gestation process disorders indicated a relationship between the metabolic profile of maternal serum and risk of gestosis. In women with pre-eclampsia or preterm delivery, analysis of serum purine metabolites revealed changes in the metabolite concentrations, associated with pregnancy complications.

Neuroprotective effect of mesenchymal and neural stem and progenitor cells on sensorimotor recovery after brain injury.

We studied the effect of systemic administration of multipotent stem cells on impaired neurological status in rats with brain injury. It was found that transplantation of multipotent mesenchymal stromal cells of the bone marrow or human neural stem and progenitor cells to rats with local brain injury promoted recovery of the brain control over locomotor function and proprioceptive sensitivity of forelegs. The dynamics of neurological recovery was similar after transplantation of fetal neural stem and progenitor cells and multipotent mesenchymal stromal cells. Transplantation of cell cultures improved survival of experimental animals. It should be noted that administration of neural stem and progenitor cells prevented animal death not only in the acute traumatic period, but also in delayed periods.