Interleukin-1 receptor antagonist mediates toll-like receptor 3-induced inhibition of trophoblast adhesion to endometrial cells in vitro

STUDY QUESTION Is interleukin-1 receptor antagonist (IL-1RA) involved in the toll-like receptor 3 (TLR 3)-induced inhibition of trophoblast cells' adhesion to endometrial cells in vitro? SUMMARY ANSWER IL-1RA mediates the TLR 3-induced inhibition of trophoblast cells' adhesion to endometrial cells in vitro. WHAT IS KNOWN ALREADY It is well documented that endometrial TLR 3 activation leads to impairment of trophoblast binding to endometrial cells in vitro. IL-1RA is known as an anti-implantation factor, as its injection significantly reduced implantation rates in mice by an effect on endometrial receptivity. STUDY DESIGN, SIZE, DURATION Poly I:C was used as a TLR3 specific ligand and endometrial cells were either treated or not with Poly I:C (treated versus control) in vitro. IL-1RA was applied to block IL-1 signal transduction. IL-1RA was knocked down by Accell Human IL1RN siRNA. Flagellin was used to stimulate TLR 5. SP600125 (JNK) was applied to inhibit the mitogen-activated protein kinases (MAPK) pathway. BAY11 -7082 was used to inhibit the nuclear factor-κB (NF-κB) pathway. The experiments were performed in three replicates on three separate days. PARTICIPANTS/MATERIALS, SETTING, METHODS An in vitro assay was developed using RL95-2 (an endometrial cell line) and JAr (a trophoblast cell line) cells. Initially, the production of IL-1RA in RL95-2 cells in response to TLR 3 activation was measured. To determine whether the TLR 3-induced inhibition of trophoblast binding was mediated through IL-1RA: (i) we evaluated the effect of IL-1RA on the attachment of trophoblast cells to endometrial cells; (ii) we knocked down TLR3-induced IL-1RA gene expression by IL-1RA Small interfering RNA (siRNA) and evaluated trophoblast attachment to endometrial cells. Finally, to clarify through which pathway TLR 3-induced inhibition of trophoblast binding occurs: (i) activation of NF-κB and MAPK was detected by transfecting the cells with secreted placental alkaline phosphatase reporter plasmids bearing promoter sequences for each transcription factor; (ii) the inhibitors for NF-κB and MAPK were used to block signaling; (iii) it was then investigated whether addition of these inhibitors could restore the TLR 3-induced impairment of trophoblast attachment to the endometrial cells. MAIN RESULTS AND THE ROLE OF CHANCE Our results showed that addition of polyinosinic:polycytidylic acid (Poly I:C) to RL95-2 cells significantly increased the production of IL-1RA (P < 0.05). Addition of human recombinant IL-1RA to RL95-2 cells remarkably decreased the adhesion rate of trophoblast cells to endometrial cells (P < 0.05). In addition, suppression of TLR3-induced IL-1RA gene expression in RL95-2 cells significantly restored trophoblast cells attachment to endometrial cells in the presence of Poly I:C (P < 0.05). Only TLR3 and not TLR5 induced MAPK activation (P < 0.05). TLR3 ligation did not affect NF-κB activation. Of NF-kB and MAPK inhibitors, only MAPK's inhibitor could achieve restoration of spheroid adhesion to endometrial cells (P < 0.05). LIMITATIONS, REASONS FOR CAUTION This study has been only done in vitro. Future in vivo studies will confirm our data. WIDER IMPLICATIONS OF THE FINDINGS The findings of this study have a potential clinical application in introducing IL-1RA as one of the diagnostic infertility markers in the endometrium, which can affect the process of embryo adhesion at the time of implantation. Moreover, based on the novel data obtained in the current study, blocking and regulating the MAPK pathway by its inhibitors can be used as a new strategy to prevent and treat virus-induced infertility cases in ART techniques. STUDY FUNDING/COMPETING INTEREST This study was partially funded by a Marie Curie IIF-253948 grant to I.C. and was partially funded by the author's institutions. The authors have no conflict of interest to declare

Response to Comment on (Novel two-dimensional porous graphitic carbon nitride C6N7 monolayer: A First-principle calculations [Appl. Phys. Lett. 2021, 119, 142102])

Recently, reported a comments on the our paper [Appl. Phys. Lett. 119, 142102 (2021)]. With our response, the APL journal rejected their non scientific comments. There are some ambiguities about their claim: 1-They can check the phonon dispersion of their structure to see ZA out-of-plane mode. 2-They report the uniaxial stress-strain responses in Fig 2., which is unrelated to our paper. For a more helpful understanding of the mechanical properties of the novel C6N7 monolayer, they can publish a paper. 3-They mentioned: Using the DFT method and with assuming a thickness of 3.35 A for the C6N7 monolayer based on graphene thickness. Why did they choose this thickness while we know our C6N7 monolayer is at without buckling? The distance of ZA out-of-plane movement of ions in C6N7 is different from Graphene.Comment: 1 pag

Human Trophoblast Cells Modulate Endometrial Cells Nuclear Factor kappa B Response to Flagellin In Vitro

Background: Implantation is a complex process that requires a delicate cooperation between the immune and reproductive system. Any interference in the fine balance could result in embryo loss and infertility. We have recently shown that Toll-like receptor 5 activation results in a decrease of trophoblast cells binding to endometrial cells in an in vitro model of human implantation. However, little is known about the downstream signalling leading to the observed failure in implantation and the factors that modulate this immune response. Methods and Principal Findings: An in vitro model of embryo implantation was used to evaluate the effect of trophoblasts and flagellin on the activation of NF-kappa B in endometrial cells and whether TLR5-related in vitro implantation failure is signalled through NF-kappa B. We generated two different NF-kappa B reporting cell lines by transfecting either an immortalized endometrial epithelial cell line (hTERT-EECs) or a human endometrial carcinoma cell line (Ishikawa 3-H-12) with a plasmid containing the secreted alkaline phosphatase (SEAP) under the control of five NF-kappa B sites. The presence of trophoblast cells as well as flagellin increased NF-kappa B activity when compared to controls. The NF-kappa B activation induced by flagellin was further increased by the addition of trophoblast cells. Moreover, blocking NF-kappa B signalling with a specific inhibitor (BAY11-7082) was able to restore the binding ability of our trophoblast cell line to the endometrial monolayer. Conclusions: These are the first results showing a local effect of the trophoblasts on the innate immune response of the endometrial epithelium. Moreover, we show that implantation failure caused by intrauterine infections could be associated with abnormal levels of NF-kappa B activation. Further studies are needed to evaluate the target genes through which NF-kappa B activation after TLR5 stimulation lead to failure in implantation and the effect of the embryo on those genes. Understanding these pathways could help in the diagnosis and treatment of implantation failure cases

The battle of the sexes starts in the oviduct: modulation of oviductal transcriptome by X and Y-bearing spermatozoa

BACKGROUND:Sex allocation of offspring in mammals is usually considered as a matter of chance, being dependent on whether an X- or a Y-chromosome-bearing spermatozoon reaches the oocyte first. Here we investigated the alternative possibility, namely that the oviducts can recognise X- and Y- spermatozoa, and may thus be able to bias the offspring sex ratio. RESULTS:By introducing X- or Y-sperm populations into the two separate oviducts of single female pigs using bilateral laparoscopic insemination we found that the spermatozoa did indeed elicit sex-specific transcriptomic responses. Microarray analysis revealed that 501 were consistently altered (P-value <0.05) in the oviduct in the presence of Y-chromosome-bearing spermatozoa compared to the presence of X-chromosome-bearing spermatozoa. From these 501 transcripts, 271 transcripts (54.1%) were down-regulated and 230 transcripts (45.9%) were up-regulated when the Y- chromosome-bearing spermatozoa was present in the oviduct. Our data showed that local immune responses specific to each sperm type were elicited within the oviduct. In addition, either type of spermatozoa elicits sex-specific signal transduction signalling by oviductal cells. CONCLUSIONS:Our data suggest that the oviduct functions as a biological sensor that screens the spermatozoon, and then responds by modifying the oviductal environment. We hypothesize that there might exist a gender biasing mechanism controlled by the female

Level Crossing Analysis of the Stock Markets

We investigate the average frequency of positive slope $\nu_{\alpha}^{+}$, crossing for the returns of market prices. The method is based on stochastic processes which no scaling feature is explicitly required. Using this method we define new quantity to quantify stage of development and activity of stocks exchange. We compare the Tehran and western stock markets and show that some stocks such as Tehran (TEPIX) and New Zealand (NZX) stocks exchange are emerge, and also TEPIX is a non-active market and financially motivated to absorb capital.Comment: 6 pages and 4 figure

Drug-related mutational patterns in hepatitis B virus (HBV) reverse transcriptase proteins from Iranian treatment-Naïve chronic HBV patients

Background: Immunomodulators and Nucleotide analogues have been used globally for the dealing of chronic hepatitis B virus (HBV) infection. However, the development of drug resistance is a major limitation to their long-term effectiveness. Objectives: The aim of this study was to characterize the hepatitis B virus reverse transcriptase (RT) protein variations among Iranian chronic HBV carriers who did not receive any antiviral treatments. Materials and Methods: Hepatitis B virus partial RT genes from 325 chronic in active carrier patients were amplified and directly sequenced. Nucleotide/amino acid substitutions were identified compared to the sequences obtained from the database. Results: All strains belonging to genotype D.365 amino-acid substitutions were found. Mutations related to lamivudine, adefovir, telbivudine, and entecavir occurred in (YMDD) 4% (n = 13), (SVQ) 17.23% (n = 56), (M204I/V + L180M) 2.45% (n = 8) and (M204I) 2.76% (n = 9) of patients, respectively. Conclusions: RT mutants do occur naturally and could be found in HBV carriers who have never received antiviral therapy. However, mutations related to drug resistance in Iranian treatment-naïve chronic HBV patients were found to be higher than other studies published formerly. Chronic HBV patients should be monitored closely prior the commencement of therapy to achieve the best regimen option. © 2013, KOWSAR Corp

Wpływ przedoperacyjnego stosowania kwasu acetylosalicylowego na występowanie krwawienia pooperacyjnego i okołooperacyjnego zawału serca u osób poddawanych pomostowaniu aortalno-wieńcowemu

Wstęp: Podjęto próbę oceny wyników klinicznych (śmiertelność, występowanie krwawienia pooperacyjnego i okołooperacyjnego zawału serca) u pacjentów, których poddano pierwszej operacji pomostowania aortalno-wieńcowego, otrzymujących w okresie przedoperacyjnym kwas acetylosalicylowy. Metoda: Do prospektywnego, randomizowanego badania przeprowadzonego metodą ślepej próby włączono 200 pacjentów, których podzielono na dwie grupy. Osoby z jednej z nich otrzymywały kwas acetylosalicylowy w dawce 80&#8211;160 mg, natomiast chorzy z drugiej grupy przyjmowanie tego leku zakończyli przynajmniej 7 dni przed operacją. Pierwotnymi punktami końcowymi badania były: zgon w trakcie hospitalizacji i powikłania związane z krwawieniem (pooperacyjna utrata krwi na oddziale intensywnej opieki medycznej, reoperacja z powodu krwawienia oraz konieczność przetoczeń koncentratu krwinek czerwonych lub innych preparatów krwiopochodnych). Za wtórny punkt końcowy przyjęto występowanie okołooperacyjnego zawału serca. Wyniki: Pacjenci leczeni kwasem acetylosalicylowym nie różnili się w zakresie charakterystyki od osób, u których nie wdrożono tej formy terapii. Stwierdzono natomiast istotną różnicę w wielkości pooperacyjnej utraty krwi (608 &#177; 359,7 ml vs. 483 &#177; 251,5 ml; p = 0,005) i konieczności przetoczeń masy erytrocytarnej (1,32 &#177; 0,97 j. vs. 0,94 &#177; 1,02 j.; p = 0,008). Grupy nie różniły się w zakresie zapotrzebowania na płytki krwi i liczby zgonów szpitalnych oraz częstości reoperacji z powodu krwawienia. Nie wykazano również istotnych statystycznie różnic między grupami w występowaniu rzeczywistego i prawdopodobnego zawału serca (odpowiednio p = 0,24 i p = 0,56) oraz śmiertelności wewnątrzszpitalnej. Wnioski: Stosowanie kwasu acetylosalicylowego przed operacją zwiększało krwawienie pooperacyjne i konieczność przetoczeń masy erytrocytarnej, nie wpływając na liczbę zgonów, częstość reoperacji i występowanie okołooperacyjnego zawału serca. Zaleca się odstawienie kwasu acetylosalicylowego na 7 dni przed operacją (Folia Cardiologica Excerpta 2008; 3: 35&#8211;39

Distribution of shallow NV centers in diamond revealed by photoluminescence spectroscopy and nanomachining

We performed nanomachining combined with photoluminescence spectroscopy to understand the depth distribution of nitrogen-vacancy (NV) centers formed by low energy nitrogen ion irradiation of the diamond surface. NV− and NV0 fluorescence signals collected from the surface progressively machined by a diamond tip in an atomic force microscope (AFM) initially rise to a maximum at 5 nm depth before returning to background levels at 10 nm. This maximum corresponds to the defect depth distribution predicted by a SRIM simulation using a 2.5 keV implantation energy per nitrogen atom. Full extinguishing of implantation produced NV− and NV0 zero phonon line peaks occurred beyond 10 nm machining depth, coinciding with the end of easy surface material removal and onset of significant tip wear. The wear ratio of for NV active, ion irradiated diamond compared to the single-crystal diamond tip was surprisingly found to be 22:1. The reported results constitute the first integrated study of in-situ machining and wear characterization via optical properties of the diamond surface containing shallow formed NV centers. We discuss possible metrology applications for diamond tools used in precision manufacturing

Cellular, extracellular and extracellular vesicular miRNA profiles of pre-ovulatory follicles indicate signaling disturbances in polycystic ovaries

Cell-free RNAs have the potential to act as a means of gene expression regulation between cells and are therefore used as diagnostic markers describing the state of tissue environment. The origin and functions of such RNAs in human ovarian follicle, the environment of oocyte maturation, are unclear. The current study investigates the difference in the microRNA profiles of fertile women and polycystic ovary syndrome (PCOS) patients in three compartments from the same preovulatory follicle: mural granulosa cells (MGC), cell-free follicular fluid (FF), and extracellular vesicles (EV) of the FF by small RNA sequencing. In silico analysis was used for the prediction and over-representation of targeted pathways for the detected microRNAs. PCOS follicles were distinguished from normal tissue by the differential expression of 30 microRNAs in MGC and 10 microRNAs in FF (FDR < 0.1) that commonly regulate cytokine signaling pathways. The concentration of EV-s was higher in the FF of PCOS patients (p = 0.04) containing eight differentially expressed microRNAs (p < 0.05). In addition, we present the microRNA profiles of MGC, FF, and EV in the fertile follicle and demonstrate that microRNAs loaded into EVs target mRNAs of distinct signaling pathways in comparison to microRNAs in FF. To conclude, the three follicular compartments play distinct roles in the signaling disturbances associated with PCOS