Action Of IL Research Articles

P–689 IL–6/IL–10 ratio as predictor of poor ovarian response in women undergoing in-vitro fertilization

Abstract Study question Are serum cytokines levels associated with ovarian response in IVF cycles? Summary answer The IL–6/IL–10 ratio is higher in patients with low ovarian response. What is known already Previous studies reported differences in the levels of IL–2, Il–6, IL–8, IL–10 and VEGF in follicular fluid between young patients with low ovarian response and normoresponder women. In addition, it is known that IL–6 plays an important role as a mediator of fever and acute phase reaction and IL–10 is the cytokine with the greatest anti-inflammatory power. Although there seems to be some evidence about the possible effect of the immune system on ovarian function and implantation, the role it plays in ART remains unknown. Our aim was to investigate the effect of cytokines in ovarian reserve and response. Study design, size, duration One hundred and fifty-two patients were included in a retrospective study between February 2016 and December 2020. Serum cytokines IL–2, IL–4, IL- 6, IL–8, IL–10, VEGF, IFN↖, TNF α, IL–1 α, IL–1 β, MCP–1 and EGF were measured previously to the ovarian stimulation cycle. Patients with altered karyotype, mutation or premutation in the FMR1 gene or endometriosis or with any other factor that could alter the ovarian reserve or response were excluded from the study. Participants/materials, setting, methods To measure the levels of the different cytokines, a sandwich immunoassay with specific antibodies for the cytokines IL–2, IL–4, IL–6, IL–8, IL–10, VEGF, IFN↖, TNF α, IL -1 α, IL–1 β, MCP–1 and EGF were used. The statistical analysis was performed with R Statistical Software, version 4.0.3 and the Software Statistical Product and Service Solutions, version 20.0 (SPSS, Chicago, IL, EE.UU.). Main results and the role of chance We found that the ratio between IL–6 and IL–10 cytokines is higher in those patients in whom four or fewer oocytes have been recovered after ovarian puncture (2.15 versus 1.55; p = 0.035; Mann-Whitney test). If we establish 0.9 as a cut-off point for the IL–6 / IL–10 ratio, we observed that above this value the risk of having a low response to ovarian stimulation is more than 3 times greater than below this value (22.9% versus 6.0%; p = 0.007; Fischer exact test). There were no statistically significant differences between both groups in terms of age (p = 0.136), dose of gonadotropin administered (p = 0.415) and duration of ovarian stimulation (p = 0.706). In addition, performing hierarchical cluster analysis with the analyzed cytokines and the associated variables to ovarian reserve and response, we observed that the antral follicle count, the total oocytes recovered and the MII recovered are grouped in the same cluster as the cytokines IL–2, IL–4, IL–6, IL–10, IL–1α, IL–1B, IFNγ y TNFα. We determined the number of clusters based on the tree diagram and k-means method. Limitations, reasons for caution The retrospective study design and the sample size could be a limitation. The study was performed in patients with suspected implantation failure. Wider implications of the findings: The ratio between IL–6 and IL–10 could be used as a potential biomarker to predict the ovarian response and provide real expectations regarding the success of IVF cycle. The action of IL–6 could be reduced by blocking its receptor using humanized monoclonal antibodies as Tocilizumab. Trial registration number Not applicable

Detection of interleukin 8 biological activity in synovial fluids from patients with rheumatoid arthritis and production of interleukin 8 mRNA by isolated synovial cells

The presence of neutrophils in the synovial joint of patients with rheumatoid arthritis (RA) is thought to be due to the activity of chemotactic factors released by activated cells in the joint. We have shown in this report, for the first time, the abundance of one such factor, interleukin 8 (IL 8), in the synovial fluid of patients both with RA and other non-RA joint diseases, and the spontaneous production of IL 8 mRNA by RA synovial cells in culture. There was no correlation between the levels of chemotactic activity and IL 8 protein, suggesting that other factors with similar neutrophil chemotactic activity are also present in the synovial fluid exudate. In support of this concept neither the level of chemotactic activity nor IL 8 protein levels correlated with neutrophil or leukocyte infiltration, indicating that the mechanism of migration into the inflammatory environment of the joint is complex. Such migration is likely to be due to a number of chemotactic signals in addition to IL 8, which may either synergize with, or inhibit, the action of IL 8.

Interleukin 1 stimulates platelet activating factor production in cultured human endothelial cells

Monocyte-derived interleukin 1 (IL 1) induced the platelet activating factor (PAF) production in cultured human endothelial cells (HEC). The product was identified as PAF by the chemical properties, the susceptibility to phospholipase A2 and C and to lipase A1, its behaviour in thin layer chromatography and in high pressure liquid chromatography and the recovery of biological activity tested on rabbit platelets. The action of IL 1 was concentration-dependent and took more than 2 hours to became apparent. Most of the PAF produced was cell-associated and only the 25% of the total was released.

Sequential synergistic effect of interleukin 2 and interferon-gamma on the differentiation of a Tac-antigen-positive B cell line.

The presence of Tac-antigen (Tac-Ag) on human B lymphocytes and its functional significance with regard to the ability of interleukin 2 (IL 2) to modulate B cell differentiation is currently an area of high interest. An Epstein-Barr virus-transformed B cell line (CB) that secretes IgG was 30 to 40% Tac-Ag+ and was used as a model for examining the role of Tac-Ag and IL 2 in B cell differentiation. Recombinant IL 2 alone was found to have a modest but significant effect on CB in enhancing IgG secretion, increasing the plaque-forming cell response from 637 to 1734 at high concentrations (1000 U/ml IL 2) and to 888 at lower concentrations (100 U/ml). In contrast, recombinant interferon-gamma (IFN-gamma) alone had no effect on the differentiation of CB. However, both factors together showed marked synergy in increasing the number of plaque-forming cells to over 3000 by using only 10 U/ml of IFN-gamma and 100 U/ml of IL 2. These two factors were shown to act sequentially in that IL 2 was needed initially, while IFN-gamma was required for the next differentiation step into IgG-secreting cells. The effect of IL 2 on stimulating differentiation was blocked by anti-Tac, indicating that the action of IL 2 is mediated through its Tac-Ag receptor. CB cells were also sorted into Tac+ and Tac- populations and were cultured separately. In 2 wk, both populations reverted to the pattern of the original cell line. Moreover, cell cycle analysis when using double staining procedures indicated that Tac-Ag on the cell surface of CB appears and disappears according to the stage of the cell cycle, and that Tac is most strongly expressed in the S and G2 + M phases. Thus, the present study suggests that certain B cells are capable of responding to sequential stimulation by IL 2 and IFN-gamma with terminal differentiation into Ig-secreting cells, and that the amount of Tac-Ag expression is cell cycle dependent.

Interleukin 1 increases the binding of human B and T lymphocytes to endothelial cell monolayers.

Lymphocyte binding to specialized high-endothelial venules (HEV) in lymph nodes and Peyer's patches is the first step in normal lymphocyte emigration and recirculation. The development and maintenance of HEV in these lymphoid organs are thought to be immunologically controlled. Because postcapillary venules in chronic inflammatory tissue often resemble the HEV of lymphoid tissue and may also be a site of lymphocyte emigration, examination of the effects of immunologic and inflammatory mediators on endothelial cells (EC) may provide important information about the physiology of both normal lymphocyte recirculation and chronic inflammation. It is reported here that treatment of human umbilical vein EC monolayers in vitro with affinity-purified human interleukin 1 (IL 1) markedly enhances the binding of both B and T lymphocytes. Increased binding was observed within 1 h of treatment of EC with as little as 0.04 U/ml IL 1. This effect of IL 1 was EC-specific, because pretreatment of T cells or human skin fibroblasts with IL 1 did not increase the binding of lymphocytes. Stimulation of binding required active EC metabolism because incubation of EC with IL 1 at 4 degrees C, or prior fixation of EC, prevented enhanced binding. The action of IL 1 was not associated with EC damage. The secretion of IL 1 by macrophages and perhaps other cells in inflammatory lesions may exert a positive feedback signal on EC to enhance further emigration of lymphocytes into the inflammatory focus.

Characterization of the requirements for human T cell mitogenesis by using suboptimal concentrations of phytohemagglutinin.

To characterize the requirements for T cell proliferation, we have studied the response of purified populations of human T cells to varying concentrations of the mitogen phytohemagglutinin (PHA). Concentrations of PHA which induce optimal proliferative responses induce increases in cytosolic free calcium ([Ca2+]i), expression of interleukin 2 (IL 2) receptors, and production of IL 2. As the concentration of PHA is decreased, each of these processes decreases in parallel. At suboptimal concentrations of PHA, the addition of exogenous IL 2 reconstitutes both the proliferative response and the expression of the IL 2 receptor, as measured by immunofluorescence with antibodies directed against the TAC/IL 2 receptor molecule, but without reconstituting the increase in [Ca2+]i. Therefore, the concentration dependence of responses to PHA appears to be secondary to an absence of IL 2 production due to a failure to induce an increase in [Ca2+]i. The addition of the calcium ionophores A23187 and ionomycin or of accessory cells to low concentrations of PHA induces increases in [Ca2+]i and subsequent proliferative responses, suggesting that the two events are linked. The proliferative response can be inhibited by antibodies directed towards IL 2 or the IL 2 receptor, indicating that the proliferative response was at least partially dependent on the production and action of IL 2. This suggests that, although increases in [Ca2+]i are an integral event in the induction of proliferation by PHA, the increase in [Ca2+]i is required for the production but not the action of IL 2. In addition, low concentrations of PHA deliver an additional signal to cells, independent of an increase in [Ca2+]i, which induces IL 2 receptor expression and allows a proliferative response in the presence of exogenous IL 2.

Temperature sensitivity of interleukin-dependent murine T cell proliferation: Q2 mapping of the responses of peanut agglutinin-negative thymocytes.

Recent studies in our laboratory have shown that IL 1-dependent mitogenic activity is present in apparently homogeneous preparations of rabbit endogenous pyrogen. This finding suggested that the mitogenic and pyrogenic activities of this molecule might serve a common goal. Initial studies on the temperature dependence of interleukin-dependent thymocyte mitogenesis suggested that high temperature sensitivity was associated with the action of IL 1 but not that of IL 2. The present study has used an expanded range of temperatures and refined tissue culture conditions to further examine the relative temperature dependence of thymocyte mitogenesis due to IL 1 or IL 2. Both the pI 5 and pI 7 species of rabbit IL 1 evoke highly temperature-sensitive responses from mouse thymocytes in the presence of a suboptimal dose of PHA and from peanut agglutinin-negative (PNA-) thymocytes in the absence of PHA. IL 2 also evokes a highly temperature-sensitive response from unseparated thymocytes in the presence of PHA. However, in the absence of PHA, vigorous responses by either unseparated or PNA- thymocytes to IL 2 alone lack strong temperature sensitivity. The temperature-dependent responses of both unseparated and PNA- thymocytes to either IL 1 or IL 2 have been analyzed by Q2 mapping, a determination of the temperature intervals most sensitive to temperature changes. By using this mode of analysis, we have found that IL 1 and IL 2 generate distinct Q2 maps, and that PHA transforms the shape of the IL 2-derived Q2 map but not that of IL 1. The possible significance of the temperature sensitivity of IL 1- and IL 2-driven reactions is discussed with respect to the biological functions of inflammation and fever.

Interleukin 3 augments the murine primary cytolytic T lymphocyte response to allogeneic tumor cells.

The primary anti-H-2k allospecific cytolytic T lymphocyte (CTL) response by BALB/c (H-2d) spleen cells in vitro to x-irradiated RDM4 (H-2k) tumor cells is weak. This response has been shown to be augmented by CTL helper factor (CHF), a factor present in supernatants of spleen cells cultured with Sendai virus (SC-CM). Conditioned medium from WEHI-3 cells (WEHI-CM) also contains activity that augments the BALB/c anti- RDM4 CTL response. Attempts to separate the CHF activity from interleukin 3 (IL 3), also present in WEHI-CM, were unsuccessful. Purified IL 3 was then tested, and was found to increase the BALB/c anti- RDM4 CTL response by five- to 10-fold. IL 3 is apparently the only material in WEHI-CM that is active in this assay. The response is apparently a classical CTL response because: 1) the effector cells are sensitive to monoclonal anti-Thy-1.2 antibody plus C; 2) the response is dependent on antigen stimulation, and it peaks on day 5 or 6 of culture; and 3) the effector cells are specific for H-2k targets. IL 3 must be added very early during the in vitro culture period for maximal augmentation of the response, consistent with possible action of IL 3 as a differentiation factor.

Inhibition of mitogen-induced lymphocyte proliferation by ouabain: interference with interleukin 2 production and interleukin 2 action.

Since the discovery of IL 2 it has been possible to dissect the mitogenic activation of T lymphocytes into two steps: first, the production of IL 2; and second, the response of IL 2-dependent T cell blasts to IL 2. We investigated the influence of ouabain, a known inhibitor of T cell activation, on the production of and the response to IL 2 in mouse T lymphocytes. Ouabain dose-dependently inhibited DNA synthesis in Con A activated C57BL/6 spleen cells; inhibition below background levels occurred at a concentration of 5 X 10(-4) mol/liter. The action of IL 2 was assayed on a mouse T cell line, named M 2, derived from C57BL/6 spleen cells and maintained in continuous IL 2-dependent culture for more than 21 mo. Ouabain dose-dependently inhibited the response of these cells to mouse IL 2 with optimal inhibition at 10(-3) mol/liter ouabain. The production of IL 2 by Con A-stimulated C57BL/6 spleen cells was also totally inhibited by 10(-3) mol/liter ouabain; 10(-4) mol/liter ouabain inhibited IL 2 production partially, and 10(-5) mol/liter and 10(-6) mol/liter ouabain had no inhibitory effect. The ouabain-mediated inhibition of IL 2 production and of IL 2 action was not due to a nonspecific toxicity, because the preincubation of spleen cells with 10(-3) mol/liter ouabain for 24 hr did not impair their ability to produce IL 2, nor was the response of M 2 cells to exogenously added IL 2 abrogated by the preincubation of the cells with 10(-3) mol/liter ouabain. The data suggest that ouabain acts on two steps during the induction of T cell activation.