Prolactin stimulates cell migration and invasion by human trophoblast in vitro.

INTRODUCTION: Prolactin (PRL) is present in endometrium at the time of embryo implantation and throughout pregnancy. Extrapituitary PRL acts as a cytokine in cells expressing PRL receptor (PRLR). So far no specific function has been demonstrated for PRL in the trophoblast of early pregnancy. METHODS: PRLR in placental tissue and trophoblast cells was shown here immunochemically. The possibility that PRL could influence trophoblast cell migration and invasion was investigated in vitro using isolated cytotrophoblast of the first trimester of pregnancy placental tissue and HTR-8/SVneo cell line. Wound healing cell migration test was performed on HTR-8/SVneo cells, and both cell types were used in Matrigel invasion test. RESULTS: PRLR is expressed by extravillous cytotrophoblast of the cell column and the placental bed, as well as in isolated cytotrophoblast (CT) and HTR-8/SVneo cells. PRL (at 100 and 1000 ng/ml) stimulated HTR-8/SVneo cell migration and cell invasion in both cell types, which could be blocked by anti-PRLR. Integrins α1 and α5, and galectin-1 (gal-1) were variably increased in PRL treated CT and HTR-8/SVneo cells. DISCUSSION: To our knowledge this is the first study demonstrating that PRL stimulates trophoblast invasiveness through PRLR, which is accompanied by increased integrins and gal-1, not excluding change in other potential mediators. This finding further supports relevance of PRLR for invasive trophoblast. CONCLUSION: This report supports a possibility that PRL may have a role in trophoblast invasion in vivo.

Effects of conventional versus biocompatible peritoneal dialysis solutions on peritoneal and systemic inflammation, malnutrition and atherosclerosis in CAPD patients.

BACKGROUND: Chronic inflammation, malnutrition and atherosclerosis (MIA syndrome) are important predictors of high mortality in continuous ambulatory peritoneal dialysis (CAPD) patients. We aimed to evaluate the effects of PD solutions (standard vs. biocompatible) on some parameters of MIA syndrome in patients undergoing CAPD. METHODS: 42 stable patients who were on CAPD at least 2.5 years participated in this cross-sectional study. Patients who had severe anemia (Hb < 10 g/l), immunomodulatory therapy, peritonitis or any inflammatory conditions for at least 3 months before the analysis, malignant disease and acute exacerbation of heart failure, were excluded. 21 (50%) patients were treated with standard PD solutions (CAPDP-1), while the remaining 21 (50% of patients) were treated with biocompatible PD solutions (neutral solutions with lower level of glucose degradation products and lower concentration of calcium, CAPDP-2). All patients underwent echocardiography and B-mode ultrasonography of common carotid arteries together with assessments of nutrition status and parameters of systemic and local inflammation. RESULTS: There were no significant differences between the groups concerning age, gender, underlying disease, residual renal function, peritoneal transport characteristics, comorbidity or therapy applied. Patients from group CAPDP-2 had a significantly lower serum level of hs-CRP (3.7 ± 2.6 mg/l vs. 6.3 ± 4.5 mg/l; p = 0.023) and significantly better nutritional status confirmed by mid-arm circumference (p = 0.015), mid-arm muscle circumference (p = 0.002) and subjective global assessment (14.28% of patients in CAPDP-2 vs. 71% of patients in CAPDP-1 were malnourished; p = 0.000). Group CAPD-2 had less frequent left ventricular hypertrophy (p = 0.039), thinner intima-media thickness (p = 0.005), smaller carotid narrowing (p = 0.000) and fewer calcified plaques of common carotide arteries (p = 0.003). No significant difference between the CAPDP groups was observed in serum and effluent levels of inflammatory cytokines (IL-1, IL-6 and TNF-α) and CA-125 effluent level. Logistic regression analysis did not confirm that biocompatibility of PD solutions was an independent predictor of any parameter of MIA syndrome. CONCLUSIONS: According to the present study and logistic regression analysis, the effect of biocompatible CAPD solutions on parameters of malnutrition, inflammation and atherosclerosis have to be confirmed by well-designed and controlled studies in a higher number of patients.

Immunomodulatory activity of IL-27 in human periapical lesions.

IL-27, a cytokine with pro-inflammatory and anti-inflammatory properties, is a new member of the IL-6/IL-12 family, whose function in periapical lesions is unknown. We hypothesized that the production of IL-27 and its effect depend upon the type of immune/inflammatory response and clinical presentation of periapical lesions. We tested this hypothesis by studying the expression and function of IL-27 in human periapical lesions, both in situ and in culture. Immunohistochemistry demonstrated the strongest expression of IL-27 by endothelial cells and mononuclear phagocytes. Its production by periapical lesion mononuclear cells (PL-MNC), especially in symptomatic lesions, was significantly higher compared with that in peripheral blood MNC and correlated with the frequency of CD14(+) and CD3(+) cells. Exogenous IL-27 stimulated Th1 and down-regulated Th17 cytokine production by PL-MNC from symptomatic lesions, but down-regulated Th1 and Th2 responses in asymptomatic lesions. These findings suggest that IL-27 is an immunomodulatory cytokine in periapical lesions, with complex biological effects.

Regulatory T-cells in periapical lesions.

CD4(+)CD25(hi)Foxp3(+) regulatory T-cells (Tregs) are of crucial importance in regulating the immune response, including the control of any defense against infection. Their presence in periapical lesions has not been demonstrated, as yet. We hypothesized that Tregs infiltrate periapical lesions, where they inhibit T-cell proliferation. The aim of this study was to characterize Tregs in periapical lesions by confocal microscopy, flow cytometry, and functional assays. We showed that CD4(+)CD25(hi)Foxp3(+) cells in periapical lesions expressed IL-10 and TGF-beta. Their frequency was significantly higher than in peripheral blood and correlated with the levels of TGF-beta and IL-10 in culture supernatants of periapical lesion mononuclear cells. Tregs inhibited the proliferation of responder T-cells in vitro, at least in part, by stimulating the production of IL-10. These findings suggest that CD4(+)CD25(hi)Foxp3(+) cells in periapical lesions may play regulatory roles in controlling local immune/inflammatory processes.

Characterization of antigen-presenting cells in human apical periodontitis lesions by flow cytometry and immunocytochemistry.

AIM: To analyse phenotypic characteristics of antigen-presenting cells (APC), isolated from human periapical lesions by flow cytometry and immunocytochemistry. METHODOLOGY: Sixteen periapical lesions were digested for 15 min with 0.05% collagenase. Mononuclear cells, separated from other inflammatory cells by density centrifugation, were processed for flow cytometry and/or immunocytochemistry. Single and double immunostainings were performed using monoclonal antibodies specific for human CD45, CD3, CD19, CD14, HLA-DR, CD1a, CD83 and CD123. RESULTS: Antigen-presenting cells (HLA-DR(+) cells) represented 32.9 +/- 17.8% of total mononuclear cells. Amongst them, B cells (HLA-DR(+) CD19(+)) were the predominant APC population, followed by activated macrophages (HLA-DR(+) CD14(+)), dendritic cells (DC) (HLA-DR(+) CD14(-) CD19(-) CD3(-)) and activated T cells (HLA-DR(+) CD3(+)). Based on the predominance of T cells (CD3(+)) or B cells and plasma cells (CD19(+) and CD19(lo), respectively) amongst mononuclear cell infiltrates, lesions were divided into T- and B-types. The percentage of DC in T-type lesions (27.1 +/- 6.8% of total HLA-DR(+) cells) was higher, compared with B-type lesions (10.3 +/- 5.2%) (P < 0.01). Within the DC population, the percentages of CD1a (Langerhans cell type) and CD123 (probably plasmacytoid DC type) did not differ significantly between the groups (P > 0.05). However, the percentage of mature DC (CD83(+)) was significantly higher in T-type periapical lesions (P < 0.05). CONCLUSIONS: Flow cytometry and immunocytochemistry are suitable methods for phenotypic analysis of APC after their isolation from human periapical lesions. APC, that were phenotypically heterogeneous, constituted a significant component of infiltrating cells. Lesions with the predominance of T cells were characterized by a higher proportion of mature DC (HLA-DR(+)CD83(+) cells) than lesions with predominance of B cells/plasma cells.

A nucleoside analogue, 7-thia-8-oxoguanosine stimulates proliferation of thymocytes in vitro.

7-thia-8-oxoguanosine (immunosine) is a nucleoside analogue with immunoenhancing activity. In this work, its effects on proliferation of thymocytes in vitro were studied. It was found that immunosine stimulated proliferation of thymocytes both of mice and rats. The stimulatory effect depended on antigen presenting cells (APC), since thymocytes depleted of accessory cells did not proliferate to immunosine. In addition, pretreatment of APC with immunosine for 24 h significantly increased proliferation of thymocytes. Immunosine stimulated interleukin 2 (IL-2) production and the expression of activation markers (CD25 and CD71). The upregulation of CD25 (alpha subunit of IL-2R) was detected both on thymocytes and thymic dendritic cells. Proliferation of thymocytes in the presence of immunosine was predominantly mediated by IL-2 since blocking IL-2Ralpha by specific monoclonal antibodies inhibited cell proliferation by 65-85%.

Proliferation of spleen cells in culture stimulated by 7-thia-8-oxoguanosine: evidence that both B- and T-cells are the targets of its action.

7-thia-8-oxoguanosine (immunosine) is a nucleoside analog showing efficient antiviral activity in rodent models as a consequence of enhancement of the immune response. However, little is known about the mechanisms of its action. In this work the effect of immunosine on proliferation of mouse and rat splenocytes in culture was studied. It was found that the compound stimulated proliferation of lymphocytes in a dose-dependent manner without any additional stimuli. The effect is predominantly mediated by interleukin-2 (IL-2) as judged by increased IL-2 production, upregulation of IL-2 receptor alpha (IL-2R alpha) expression and by significant inhibition (60-75%) of cell proliferation by anti-IL-2R alpha monoclonal antibodies (mAbs). Immunosine also stimulated proliferation both of T- and B-cells purified by immunomagnetic sorting. The response of B-cells was much higher than that of T-cells. The stimulatory effect of immunosine on both lymphocyte subpopulations was further increased by the addition of enriched splenic antigen-presenting cells or purified dendritic cells. Proliferation of purified T-cells to immunosine was also significantly potentiated by an anti-alpha beta T-cell receptor mAb (R 73). All these data suggest that T-, B- and accessory cells in splenic cultures are the targets of immunosine action.