Background: Thin endometrium is a significant cause of infertility due to impaired regeneration, reduced receptivity, and insufficient angiogenesis. Mesenchymal stem cell-derived secretome (MSCsec) is a promising cell-free therapy because it contains bioactive molecules that promote tissue repair. This study evaluated the effects of Wharton’s jelly-derived MSC secretome (WJ-MSCsec) on human endometrial stromal cells (EnSCs) and endothelial cells in vitro. Methods: WJ-MSCs were isolated from umbilical cord tissue, characterized, and cultured under serum-free conditions. The concentrated secretome was analyzed using a human angiogenesis proteome array. EnSCs were isolated from endometrial biopsies. EnSC proliferation and migration in the presence of WJ-MSCsec were assessed using CCK-8 and scratch wound-healing assays. Pro-angiogenic activity of WJ-MSCsec was evaluated using a Matrigel tube formation assay with human umbilical vein endothelial cells (HUVECs). Results: Proteomic analysis of WJ-MSCsec revealed the presence of angiogenic, mitogenic, immunomodulatory, and chemotactic factors. Treatment with WJ-MSCsec significantly enhanced EnSC proliferation and accelerated wound closure. Furthermore, WJ-MSCsec markedly promoted endothelial tube formation, increasing total tube length, junction number, and mesh formation. Conclusions: WJ-MSCsec stimulates EnSC proliferation and migration while enhancing angiogenesis in vitro. WJ-MSCsec represents a promising cell-free therapeutic strategy for endometrial regeneration and reproductive disorders associated with impaired endometrial function.

Background/Objectives: Increasing evidence points to hypoxia and inflammation as two major causes of compromised ovarian function. Increased oxidative stress under hypoxic conditions can damage cellular components, leading to the dysfunction and apoptosis of granulosa cells (GCs). The inflammatory response induced by hypoxia may further impair the function of the ovaries and contribute to the development of premature ovarian insufficiency (POI). In animal models of premature ovarian failure, research has demonstrated that the transplantation of mesenchymal stem cells (MSCs) can enhance reproductive outcomes, increase the number of functioning ovarian follicles, and restore estradiol production. However, the specific mechanisms underlying the observed positive results are not well understood. Methods: The present study provides a comparative analysis of how MSCs influence human GC function under inflammatory and hypoxic conditions, using three different experimental approaches: direct co-culture, indirect co-culture with transwell cell culture inserts, and treatment with MSC-derived conditioned medium (MSCcm). Results: Inflammation significantly suppressed GC estradiol secretion and increased apoptosis. MSCs increased estradiol secretion in normal and hypoxic culture conditions when co-cultured directly with GCs. Our results also showed that, under inflammation, MSCs tended to decrease GC proliferation and that hypoxia alone did not have an effect on GC estradiol secretion or proliferation. Conclusions: The study emphasizes the dual nature of MSCs, which largely determines their effects on other cell types, and the need for the condition-specific optimization of MSC therapies for ovarian regeneration.

Background: Fertilization check performed at the 18th hour following classic in vitro fertilization procedure (IVF) or intracytoplasmic sperm injection (ICSI) is a critical stage in assisted reproduction. The success of the treatment is significantly reliant on the quantity of zygotes exhibiting two pronuclei. Consequently, low fertilization rates or complete fertilization failure are highly undesirable outcomes for both patients and reproductive specialists. Applying additional calcium ionophore for oocyte activation subsequent to ICSI may offer benefits and potentially enhance treatment outcomes, particularly for patients who have experienced low or absent fertilization rates (FR) in previous treatment cycles. The aim of the study is to evaluate the efficacy of Ca2+ ionophore application for oocyte activation. Methods: A retrospective analysis of 924 oocytes obtained from 120 patients who underwent ICSI cycles with a history of low or no fertilization as a result of previous unsuccessful treatment rounds. The next ART cycle followed with additional oocyte Ca(2+ )ionophore 2 ionophore activation applied in 57 of the cases in order to optimize the treatment process (Group 1), and 63 patients were included and their outcomes followed as a control group (Group 2).We conducted a comparative analysis of results in both groups. The study's ' s primary outcomes encompassed fertilization, cleavage embryo quality, blastocyst rate, and established clinical pregnancies. Results: At day 1 fertilization check we had 274/386 zygotes (71%FR) in group 1 and 132/410 in group 2 (32.2%FR), (P P < 0.0001). Twenty-two (34.9%) cycles in group 2 resulted in total fertilization failure (TFF). At the cleavage stage top-quality embryos from group 1 were significantly fi cantly higher (P P = 0.0021) in comparison to group 2. Forty-eight embryo transfers (ET) were performed in group 1 resulting in 41.67% clinical pregnancies versus 33 ET and only 4 pregnancies (12.12%) for group 2 (P P = 0.0044). Conclusions: The results confirm the appropriateness of assisted oocyte activation as an additional method in cases of previous fertilization failure cycles.

Endometriosis is one of the most common benign gynecologic disorders affecting reproductive health. There are many theories regarding how endometriosis affects fertility, but the exact mechanisms are still uncertain. We evaluated 100 patients with unilateral ovarian endometriosis and infertility who underwent ovarian cystectomy. The women had at least one patent tube after surgery and were observed for spontaneous pregnancy for up to one year. Patients were divided into two groups according to the endometrioma position. The first group included 67 women with an endometriotic cyst in the left ovary, and the second group included 33 with an endometrioma in the right ovary. Groups were similar according to age, type of infertility, BMI, and size of the endometrioma. The number of patients was significantly higher in the first group. Spontaneous pregnancy after cystectomy occurred in 28.4% of the patients with an endometrioma in left ovary, and in 9.1% with a right endometriotic cyst. The better results regarding spontaneous conception with a left-sided endometrioma in our study are most likely related to the permeability of the fallopian tubes and adhesions in the Douglas pouch as a result of endometriosis. Further investigations are necessary to assess the influence of the laterality of endometriosis on infertility.

Premature ovarian insufficiency (POI) affects 1% of women under 40 years old and represents one of the main causes of infertility in females of childbearing age. POI is a primary ovarian defect resulting from follicular depletion and/or ovarian dysfunction caused by a compromised ovarian somatic cell compartment. Etiologically, ovarian dysfunction could be caused by certain diseases, including infections and autoimmune disorders. There could also be iatrogenic factors such as chemotherapy, radiation, or surgery. The related surgical risks are due to the possibility of disrupting the blood supply to the ovary and/or a subsequent inflammatory reaction. To date, it is unclear whether and how the impaired oxygen delivery and inflammatory environment may influence the basic functions of human ovarian somatic cells. In this study, we used an immortalized human granulosa cell line (COV434) as a model to investigate the effect of exposure to low oxygen levels (hypoxia) and inflammatory conditions on granulosa cell proliferation, apoptosis, and steroidogenesis. The results showed that both hypoxia (5% O2) and proinflammatory cytokine treatment (a combination of TNF-α, IL-1β, IFN-γ and IL-6) significantly increased apoptosis, suppressed proliferation, and affected estradiol secretion in COV434 cells. Our data suggest that granulosa cells could be particularly sensitive to changes in the local environment caused by oxygen deprivation and chronic inflammation.

OBJECTIVES Gestational diabetes mellitus (GDM) is one of the most common pregnancy complications. The universal screening for GDM is usually performed between 24-28 weeks' gestation. This often delays the diagnosis and could increase the risk of adverse pregnancy outcomes. Some of the biochemical placental markers - pregnancy associated plasma protein A (PAPP-A) and free-β human chorionic gonadotropin (hCG), probably could provide a diagnostic value for GDM. The aim of our study was to assess if PAPP-A and hCG values were different among pregnant women with and without GDM and respectively, to tested their place in the early GDM screening. MATERIAL AND METHODS We conducted a retrospective, case-control study by reviewing the clinical database records of 662 pregnant women. The analysis includes the data for a two-year period. The patients included in the observation were divided into two groups - GDM group (n = 412) and Euglycemic group (n = 250). Early screening for GDМ between 9-12 weeks' gestation was performed in 173 of the women in the interventional group due to: registered fasting plasma glucose (FPG) above 5.1 mmol/L, obesity, macrosomia in previous pregnancies or family history for diabetes mellitus. The remaining 239 women underwent universal screening at 24-28 weeks' gestation. Mean serum levels of PAPP-A, hCG, FPG, and body mass index (BMI) were measured between 10-13 gestational weeks. Serum levels of PAPP-A and hCG are presented as multiples of the normal median (MoM), adjusted by maternal baseline characteristics and demographics. RESULTS In patients who developed GDM during pregnancy, compared with the control group, we have found significantly lower MoM values of PAPP-A (p < 0.0001), higher levels of FPG (р < 0.0001) and higher BMI (р < 0.0001). Median hCG MoM was similar in both group of pregnant women. CONCLUSION Our findings suggest that low-normal to low reference range values of PAPP-A might be associated with higher risk for GDM. PAAP-A levels can be used as an additional factor to recommend early screening for GDM.

OBJECTIVES:Mesenchymal stem cells (MSCs) exist in almost all tissues. Their unique nature is completed by their immunomodulatory functions, holding promise for the treatment of many diseases. An inflammatory environment precedes the immunosuppressive abilities of MSCs and this study was intended to better understand how umbilical cord MSCs (UCMSCs) react to the process of inflammation, regarding their basic characteristics and behavior when primed with the key pro-inflammatory cytokine, Interferon-γ (IFNγ).MATERIALS AND METHODS:Human MSCs from the umbilical cord were isolated, expanded, and treated with IFNγ. Primed cells were analyzed to define their ability to form colonies, their morphology, differentiation potential, proliferation, and apoptosis rate.RESULTS:UCMSCs treated with IFNγ changed their fibroblast-like morphology and retained the expression of typical MSCs markers. IFNγ treated UCMSCs had significantly higher MFI levels regarding the expression of HLA-I (980.43 ± 556.64) and PD-L1 (598.04 ± 416.90) compared with the control cells (144.97 ± 78.5 and 122.05 ± 103.83, respectively; P<0.01). Under the influence of IFNγ, the cells had a lower population doubling time compared with the control cultures (50.345 ± 9.155 versus 61.135 ± 21.110, respectively; P<0.01) and higher numbers of colony-forming unit-fibroblasts (26.0 ± 12.2 versus 10.2 ± 8.0, respectively; P<0.05). The primed MSCs could not undergo osteogenic and adipogenic differentiation. IFNγ increased the percentage of cells in the apoptotic state on day eight (29.470 ± 6.59 versus 15.708 ± 6.190, respectively; P<0.01).CONCLUSION:The properties of UCMSCs can be influenced by the pro-inflammatory cytokine IFNγ.

Кріоконсервування гамет та ембріонів – важливий технологічний етап у лікуванні безпліддя з використанням допоміжних репродуктивних технологій (ДРТ) [8]. Різні дослідження продемонстрували високі показники запліднення, настання вагітності та пологів після кріоконсервування ооцитів (КО), які збігаються з таким після використання свіжовиділених ооцитів [4]. Деякі автори вважають, що селективне заморожування ооцитів потребує більш детального вивчення, оскільки на сьогодні відсутні чіткі докази щодо його безпеки, ефективності та економічної доцільності [2]. Слід зазначити, що на результат КО впливають численні фактори: вік та стан здоров’я жінки, якість ооцитів, режими їх вітрифікації, відігріву, відповідні умови зберігання та техніка виконання інтрацитоплазматичного введення сперматозоїдів (ICSI) в ооплазму ооцита [7]. Відомо, що здатність жінки народити дитину знижується з віком. Цей спад починається у 32 роки і стає критичним після 37 років [6]. Результати мета-аналізу показали зниження частоти настання вагітності у жінок старших за 36 років після використання вітрифікованих ооцитів [3]. Пацієнтки, які бажають кріоконсервувати автологічні ооцити для планування відстроченої вагітності, повинні бути проінформовані про реальний шанс народити дитину з урахуванням віку та оваріального резерву. У разі використання вітрифікованих донорських ооцитів важливо знати необхідну кількість для одного реципієнта з метою виключення редукції зайвих ембріонів. Cryopreservation of gametes and embryos has become an important biotechnological component of infertility treatment by means of assisted reproductive technologies (ART) [8]. Diff erent studies show good fertilization, pregnancy and delivery rates after oocyte cryopreseravtion (OC) similar to the rates from fresh oocytes [4]. Some authors think that selective oocyte freezing remains controversial because there are no clear evidence-based guidelines regarding its safety, effi cacy and cost-eff ectiveness [2]. Most important parameters aff ecting the OC success are age and health conditions of a woman, quality of the oocytes, reliable vitrifi cation and warming regimens, appropriate storage conditions and precise intracytoplasmic sperm injection (ICSI) after warming [7]. It is well known that the ability of a woman to have her own child decreases with the age. This decline usually starts at age of 32 years and becomes very signifi cant after 37 years’ age [6]. One meta-analysis showed a decrease in the pregnancy rate in women after the age of 36 after using vitrifi ed oocytes [3]. The women who cryopreserve autologous oocytes for delayed pregnancy have to be informed about the real chance to give birth of a baby depending on their age and number of vitrifi ed oocytes. Concerning the vitrifi ed donor oocytes, it is important to have data about the number of oocytes needed for one recipient to get enough embryos, and at the same time without exceeding some limit because of ethical aspects due to the probable elimination of not necessary embryos.

Hip replacement is one of the most effective surgical procedures in medicine. Despite the fact that the clinical results of arthroplasty are usually excellent, some implants loosen with time and require revision. To help resolve this problem, research has focused on providing conditions that have a positive biological effect on bone building elements, resulting in a fast, seamless proliferation and differentiation of mesenchymal stem cells (MSC) into osteoblasts. That is why developing an in vitro model for studying MSC functions at the implant–tissue contact zone is essential. The aim of this study was to investigate the proliferation and differentiation capacity of human bone-marrow derived MSC in contact with medical implant-grade Ti-6Al-4V alloy and β-tricalcium phosphate (β-TCP). Human MSC were isolated from bone marrow aspirates obtained from the femoral canal during hip replacement surgeries and seeded on Ti-6Al-4V alloy squared specimens and β-TCP granules. The results indicate that MSC were able to effectively adhere to both biomaterial surfaces and, at the same time, retain their proliferative and osteogenic/adipogenic differentiation potential. We suggest that both Ti-6Al-4V alloy and β-TCP surfaces provide suitable conditions for the proliferation and differentiation of bone marrow MSC and can be used as in vitro models to explore MSC functions at the bone–implant interface.

Purpose Along with comparative investigation of the decidualization potential and IL-6 secretion by fresh and frozen ESCs, we also aimed to evaluate the effectiveness of co-culture systems based on fresh or frozen ESCs in terms of clinical pregnancy rates. Methods Outcome analysis of a total of 215 IVF cycles with co-culture with fresh or frozen ESCs was performed. Endometrial tissue was obtained from 17 healthy donors. Concentrations of secreted prolactin, IGFBP-1, and IL-6 in conditioned media from cultured fresh and frozen ESCs (decidualized or not) were measured using ELISA or ECLIA. Results Embryo co-culture with frozen ESCs resulted in a much lower pregnancy rate compared to the alternative system using fresh ESCs. Furthermore, cultivated frozen ESCs showed considerably decreased release of prolactin, IGFBP-1, and IL-6 compared to fresh ESCs, indicating that cryopreservation negatively affects their decidualization potential and cytokine production. Conclusions Altogether, this data illustrates the need for optimization and improvement of the existing autologous endometrial co-culture systems.

According to the minimal criteria of the International Society of Cellular Therapy, mesenchymal stem cells (MSCs) are a population of undifferentiated cells defined by their ability to adhere to plastic surfaces when cultured under standard conditions, express a certain panel of phenotypic markers and can differentiate into osteogenic, chondrogenic and adipogenic lineages when cultured in specific inducing media. In parallel with their major role as undifferentiated cell reserves, MSCs have immunomodulatory functions which are exerted by direct cell-to-cell contacts, secretion of cytokines and/or by a combination of both mechanisms. There are no convincing data about a principal difference in the profile of cytokines secreted by MSCs isolated from different tissue sources, although some papers report some quantitative but not qualitative differences in cytokine secretion. The present review focuses on the basic cytokines secreted by MSCs as described in the literature by which the MSCs exert immunodulatory effects. It should be pointed out that MSCs themselves are objects of cytokine regulation. Hypothetical mechanisms by which the MSCs exert their immunoregulatory effects are also discussed in this review. These mechanisms may either influence the target immune cells directly or indirectly by affecting the activities of predominantly dendritic cells. Chemokines are also discussed as participants in this process by recruiting cells of the immune systems and thus making them targets of immunosuppression. This review aims to present and discuss the published data and the personal experience of the authors regarding cytokines secreted by MSCs and their effects on the cells of the immune system.

Gonadotropin releasing hormone (GnRH) analogues are used routinely in ART to prevent a premature luteinizing hormone (LH) surge in women undergoing controlled ovarian hyperstimulation. The impact of GnRH analogues used in IVF protocols on endometrial function and receptivity is still relatively unclear, aside from their main role in pituitary suppression. Decidualization of human endometrium in the late luteal phase of the menstrual cycle is a profound morphological and biochemical alteration process critical for implantation and placental development. The purpose of this study was to examine whether GnRH analogues Ganirelix and Triptorelin directly affect the process of human endometrial stromal cell decidualization. Our results suggest that under the experimental conditions used, Ganirelix and Triptorelin do not influence the in vitro decidualization capacity of human endometrial stromal cells.

Glioblastoma multiforme (GBM) is the most common and malignant tumor in the central nervous system. One of the contemporary hypotheses postulates that its pathogenesis is associated with the cancer stem cells (CSCs) which originate from mutations in the normal neural stem cells residing in their specific "niches." Simultaneously with its aggressive development the tumor suppresses the local immune system by different secreted and/or cell expressed factors. Progesterone-induced blocking factor (PIBF) is an immunomodulatory protein with known role in the regulation of the immune response in the reproductive system. Expression of PIBF has been described in some tumors as one of the factors suppressing the anti-tumor immunity. The aim of the present study was to check for the expression of PIBF from cells isolated from six GBMs. To characterize the cultured cells and to study the PIBF expression confocal microscopy, flow cytometry, ELISA, and real-time PCR were used. The results obtained showed expression of markers typical for cancer CSCs and secretion of interleukin 6 by the GBM-derived cultured cells. The results convincingly prove that PIBF is intracellularly expressed by the cultured cells from the all six GBM samples, and this fact is confirmed by three different methods-flow cytometry, confocal microscopy, and real-time PCR. This paper reports for the first time the expression of PIBF by GBM-derived cells cultured in vitro and reveals a new aspect of the immunosuppressive mechanism used by GBM in escaping the immune control.

The aim of this study was to investigate the proliferation and osteogenic differentiation of mesenchymal stern cells in contact with porous coated titanium implant. Human mesenchymal stem cells (BM-MSCs) were obtained from bone marrow aspirate from the femoral canal during elective hip replacement. The MSCs were isolated and cultured. The resulting cells were seeded on porous coated titanium plates obtained from prosthesis blanks that had all the characteristics of the final product. The same cells were grown on classic plastic polystyrene plates as a control specimens. The proliferation and osteogenic differentiation of the isolated and cultured mesenchymal stem cells were examined Our results show that under the same conditions there were no significant differences between the study and control groups. Titanium implants with porous coated surface provide favorable conditions for the proliferation and differentiation of bone marrow stem cells and did not inhibit their development. In conclusion, it may be possible to advance this methodology to stimulate healing of bone defects and particularly massive osteolysis in the revision arthroplasty setting. MSCs may provide an alternative to the use of autologous and allogenic bone-grafts in orthopedic surgery.