IL-10-producing B cells play important role in the pathogenesis of recurrent pregnancy loss.

Interleukin (IL)-10-producing B cells are recently known for their regulatory function in several disorders. However, the possible role of these cells remains unclear in recurrent pregnancy loss (RPL) pathogenesis. Total B cells from 24 RPL patients with cellular immune abnormalities, as well as that of 25 normal pregnant women were cultured and stimulated by Toll Like Receptor (TLR) agonists (CpG oligodeoxynucleotides (ODN) and imiquimod). Then, the frequency of IL-10+ CD19+ B cells was found out using flow cytometry. Enzyme-linked immunosorbent assay (ELISA) was used to assess the levels of IL-10 in supernatant medium and serum of stimulated B cells, as well as those of several serum autoantibodies. Real-Time PCR method was carried out for determining the IL-10 expression level and specific genes transcripts. RPL patients indicated a lower proportion of IL-10+ CD19+ B cells, and reduced levels of IL-10 in both serum and supernatant of the culture medium of the stimulated B cells. According to the results, total IgG levels was greater in serum of RPL patients in comparison with healthy pregnant women. Similarly, the percentage of these cells was negatively correlated with serum total IgG levels and the number of miscarriages. The expression levels of the mRNA of programmed death-ligand 1 (PD-L1) and IL-10 were lower in RPL patients, while those of x binding protein 1 (XBP-1), interferon regulatory factor 4 (IRF4), and B lymphocyte-induced maturation protein 1 (BLIMP1) were significantly increased. These observations indicated that the reduction in the population of peripheral blood IL-10-synthesizing B cells may prompt RPL pathogenesis, suggesting suppressive effects of these cells on autoantibody production and successful pregnancy outcomes.

The role of IL-10-producing B cells in repeated implantation failure patients with cellular immune abnormalities.

There are a few data of the role of B cells in RIF pathogenesis. Accordingly, the objective of the current study was to determine the role of IL-10-producing B cells in RIF. Twenty-three RIF women with cellular immune abnormalities and 25 normal controls were enrolled in this experiment. Isolated naïve B cells from peripheral blood of the subjects were cultured in vitro, divided into two parts and activated by CpG ODN and imiquimod as TLR agonists. Afterwards, the number of CD19+ IL-10+ B cells was evaluated by flow cytometry and their related IL-10 cytokine level was assessed by ELISA. The mRNA expression levels of related genes in just CPG stimulated B cell population were also analyzed using real-time PCR. RIF patients exhibited a decreased level of the cells (P = 0.014, P = 0.023, respectively) and IL-10 cytokine (P = 0.009, P = 0.045, respectively) in both CPG and imiquimod stimulated B cell groups. IL-10 serum level was also lower in these patients (P = 0.0014). Additionally, we found a negative relationship between the frequency of these cells with the number of failed ET and total IgG titers in RIF patients. The mRNA levels of IL-10-producing B cells related genes (IL-10 and PD-L1) was also significantly lower in RIF women, whereas the expression of plasma cells-associated transcriptional factors (BLIMP1, IRF4, and XBP1) was higher. Summing up the obtained results, we concluded that peripheral blood IL-10-producing B cells down-regulation might result in RIF pathogenesis. It is further suggested that these cells can suppress autoantibody generation and contribute to a successful implantation.

Oxidative stress, inflammatory settings, and microRNA regulation in the recurrent implantation failure patients with metabolic syndrome.

PROBLEM: Increased oxidative stress (OS) and inflammatory factors in metabolic syndrome (MS) patients are considered as risk factors for recurrent implantation failure (RIF). The aim of this study was to investigate OS markers, inflammatory factors, related microRNAs (miRNA) expression, and cytokine and transcription factors RNA expression. METHOD OF STUDY: We evaluated the frequency of helper T (Th) 17 and regulatory T (Treg) cells in recurrent implantation failure (RIF) women with or without MS. miRNA expression, an inflammatory cytokine, and transcription factors were measured by real-time PCR. The level of interleukin (IL)-1ß, IL-6, IL-17, tumour necrosis factor-alpha (TNF-alpha) and chemokine (C-C motif) ligand 2 (CCL-2), and C-X-C motif chemokine ligand 8 (CXCL-8) were measured by enzyme-linked immunosorbent assay (ELISA). OS markers were evaluated by spectrophotometric assay. Th17 and Treg cell frequencies were determined by flow cytometry. RESULTS: The expression of AP1, NF-κB, FOXP3, miRNA-21; serum or plasma level of OS markers (ie, nitric oxide, total oxidant status, and myeloperoxidase); serum level of inflammatory factors (ie, IL1-ß, IL-6, IL-17, TNF-alpha, CXCL-8, and CCL-2); and frequency of Th17 cells were increased in RIF-MS patients in comparison with RIF women without MS (RIF-NMS) and control group. The expression of miRNA-223 and 146a, antioxidant enzymes, namely superoxide dismutase (SOD) and catalase (CAT), and frequency of Treg also declined in RIF-MS patients. CONCLUSION: Overall, our findings suggest that MS in RIF patients causes increased inflammatory factors and OS, which in turn leads to implantation failure.

Metabolic syndrome mediates proinflammatory responses of inflammatory cells in preeclampsia.

PROBLEM: To investigate whether metabolic syndrome (MetS) is associated with exacerbation of inflammatory responses in preeclamptic (PE) patients, the dynamic changes of Th17 and Treg cells, monocytes, cytokines, and transcription pattern of inflammasome-related genes were analyzed in 35 women with PE suffering from MetS in comparison to 38 PE women without MetS and healthy pregnant women. METHOD OF STUDY: Expression of inflammasome-related genes, cytokines, and also TLR4 was measured using real-time PCR. Serum and medium supernatant cytokines levels of PBMCs and serum levels of HMGB1 and Caspase-1 were also evaluated by ELISA. Monocytes, Th17, and Treg cells frequency were also determined by flow cytometry. RESULT: PE women with MetS exhibited increased percentage of non-classical and intermediate monocytes and Th17 cells (P = 0.025). Furthermore, decreased Treg cells frequency was also observed in PE women with MetS compared to PE women (P = 0.019). The mRNA expression of inflammasome-related genes (Caspase-1, NLRP3, HMGB1), TLR4, IL-1ß, IL-6, IL-17, IL-18, and TNF-α was significantly higher in PE patients with MetS than that of the healthy pregnant individuals (P < 0.0001) and PE patients (P < 0.0001). Serum levels of TGF-ß and TNF-α in PE patients with MetS were increased compared to other two groups, while IL-10 levels were significantly reduced. A significant sFlt (P = 0.016), Caspase-1 (P = 0.012), HMGB1 (P = 0.016) upregulation, and VEGF (P = 0.023) downregulation were also observed in the serum of PE women having MetS compared to PE women. CONCLUSION: MetS is closely related to the exacerbation of inflammatory reactions in PE. This study indicates that, in order to diminish the systemic features of PE, prior to conceive and start a pregnancy, MetS should be severely considered and managed.

NK cell frequency and cytotoxicity in correlation to pregnancy outcome and response to IVIG therapy among women with recurrent pregnancy loss.

BACKGROUND: Recurrent miscarriage (RM) has a multifactorial etiology mainly due to chromosomal abnormalities and immunological factors. Treating RM has remained to be a challenging issue and the role of intravenous immunoglobulin (IVIG) in treating RM is still controversial. MATERIALS AND METHODS: This study aimed to evaluate the changes in natural killer (NK) cells' frequency and cytotoxicity in patients with RM who received the IVIG therapy. A total of 78 women with a history of three or more recurrent miscarriages were included and their peripheral blood was drawn in case of positive pregnancy test. On the same date, 400 mg/kg of IVIG was administrated intravenously in 38 women and it continued every four weeks through weeks 30-32 of gestation. The remaining 40 patients with RM were included to be the untreated control group. Then, the effects of IVIG on NK cell frequency, cytotoxic activity, and the expression of inhibitory and activating receptors in the patients with RM, pre and posttreatment were assessed. RESULTS: NK cells percentage and cytotoxicity were significantly reduced in the IVIG-treated patients after 32 weeks of gestation (p < 0.0001). Expression levels of inhibitory receptors was increased, however, the expression levels of activating receptors were significantly decreased after the IVIG therapy. Pregnancy outcome after the treatment was significantly higher (86.8%) in the IVIG-treated patients than controls (45%; p = 0.0006). CONCLUSION: Our results suggested that women with RM may benefit from IVIG as a therapeutic approach and the frequency and functional status of peripheral NK cells may serve as a valuable predictive factor of therapy response.