Anna Chelmonska-Soyta

CD40, CD80, and CD86 Costimulatory Molecules are Differentially Expressed on Murine Splenic Antigen‐presenting Cells During the Pre‐implantation Period of Pregnancy, and they Modulate Regulatory T‐cell Abundance, Peripheral Cytokine Response, and Pregnancy Outcome

The object of the study was to investigate the costimulatory phenotype of spleen antigen‐presenting cells (APCs) in mice after mating and the influence of costimulatory signal blocking on pregnancy outcome, cytokine production, and Treg cell concentration.

Matrix metalloproteinase (MMP)-2 and MMP-9 expression in tumor infiltrating CD3 lymphocytes from women with endometrial cancer.

Objective In this study, we hypothesized that not only endothelial malignant cells but also lymphocytes infiltrating tumor epithelium, in patients with endometrial cancer, could be an important source of the gelatinases (matrix metalloproteinase [MMP]-2 and MMP-9) extensive production, which in turn, may facilitate tumor cells infiltration and progression due to the extracellular matrix degradation. Materials and Methods First, we isolated lymphocytes from the endometrial carcinoma samples taken from 41 patients who were operated on and from healthy endometrial tissue taken of the same patients after histological verification. Then, we detected the level of CD3-positive cells in endometrial tissues by flow cytometry. Simultaneously, we studied the messenger RNA expression of MMP-2 and MMP-9 in the isolated cells from malignant and unchanged endometrial tissues. Using immunohistochemistry, we compared the protein expression of MMP-2, MMP-9, and CD3 in the studied samples. Results We showed the enhanced abundance of CD3 lymphocytes both by flow cytometry and immunohistochemistry in the samples from malignant tissues. The expression of MMP-9 in the endometrial carcinoma was increased significantly at the protein level but not at the messenger RNA level. We could not observe any differences concerning MMP-2 expression in both methods of detection. Conclusions CD-3 lymphocytes significantly infiltrate endometrial cancer tissue, but they do not seem to be the source of enhanced metalloproteinases 2 and 9 expression in the tumor environment. Still, owing to the immunohistochemistry staining, we could show the significant increase of MMP-9 protein in the very close vicinity of tumor-infiltrating CD3 lymphocytes. Could it be the result of CD3 lymphocyte action, or is it just the imperfection of the detecting method we used? This remains unclear. Further studies explaining the role of tumor infiltrating lymphocytes in mediating the endometrial cancer milieu are needed.

The influence of mating on estrogen receptor alpha protein level in spleen and uterine macrophages in female mice.

Macrophages are antigen-presenting cells that have a key role in the regulation of immune phenomena and are responsible for the recognition of paternal antigens during pregnancy. The aim of this study was to investigate changes in the estrogen receptor alpha (ERalpha) protein level in splenic and uterine mature (F4/80(+)MHC II(+)) and immature (F4/80(+)MHC II(-) ) macrophages in female mice during time corresponding to the preimplantation period. C57BL/6J females in estrus were mated with Balb/c male mice or were mechanically stimulated through the vagina to achieve pseudopregnancy. Uterine and spleen cells were isolated on days 0.5 and 3.5 after mating or after uterine cervix stimulation. ERalpha content in macrophages was measured by flow cytometry and expressed as mean fluorescence intensity (MFI). The ERalpha level in splenic macrophages on 0.5 day after mating was higher than that in splenic macrophages of pseudopregnant mice on 0.5 day after stimulation. The ERalpha level was also higher in mature than in immature macrophages present in both the spleen and uterus, especially in mated mice. In the spleen, a correlation was found between the percentage of mature macrophages and ERalpha level in these cells. In conclusion, the elevated alpha level observed shortly after mating in splenic but not in uterine macrophages indicates an early systemic response to male antigens.

CD80 and CD86 Costimulatory Molecules Differentially Regulate OT-II CD4+ T Lymphocyte Proliferation and Cytokine Response in Cocultures with Antigen-Presenting Cells Derived from Pregnant and Pseudopregnant Mice

Immune phenomena during the preimplantation period of pregnancy are poorly understood. The aim of our study was to assess the capacity for antigen presentation of splenic antigen-presenting cells (APCs) derived from pregnant and pseudopregnant mice in in vitro conditions. Therefore, sorted CD11c+ dendritic cells and macrophages F4/80+ and CD11b+ presenting ovalbumin (OVA) were cocultured with CD4+ T cells derived from OT-II mices (C57BL6/J-Tg(TcraTcrb)1100Mjb/J) spleen. After 132 hours of cell culture, proliferation of lymphocytes (ELISA-BrdU), activation of these cells (flow cytometry), cytokine profile (ELISA), and influence of costimulatory molecules blocking on these parameters were measured. We did not detect any differences in regulation of Th1/Th2 cytokine balance. CD86 seems to be the main costimulatory molecule involved in the proliferation response but CD80 is the main costimulatory molecule influencing cytokine secretion in pregnant mice. In conclusion, this study showed that CD80 and CD86 costimulatory molecules regulate OT-II CD4+ T lymphocyte proliferation and cytokine response in cocultures with antigen-presenting cells derived from pregnant and pseudopregnant mice. The implications of these changes still remain unclear.

17β-Estradiol and Interferon Tau Interact in the Regulation of the Immune Response in a Model of Experimental Autoimmune Orchitis

Immunoregulatory activity of type I interferons (IFNs) and estrogen is convergent in some cases of autoimmune disorders. The aim of our study was to determine whether a potent interaction of IFN and estradiol (E2) has an influence on immune response and estrogen receptor alpha (ER-?) expression in antigen-presenting cells in a model of experimental autoimmune orchitis (EAO). C3H/He/W male mice were immunized with testicular germ cells (TGCs) and orally treated with interferon tau (IFN-?), E2, or both simultaneously. The delayed-type hypersensitivity reaction was intensified after the administration of either IFN-? or E2, but their co-administration had no effect. IFN-? treatment increased immunoglobulin G2a (IgG2a) and decreased IgG1 levels of TGC-specific antibodies, whereas E2 abolished the effects of the used cytokine. The total splenic cellularity and the number of spleen CD11c+MHC II+ and F4/80+MHC II? cells were increased after IFN-? treatment, whereas E2 antagonized this effect. After IFN-? administration the level of ER-? was significantly higher in F4/80+MHC II? cells, whereas E2 had no effect. However, the administration of E2 significantly reduced the ER-? level in F4/80+MHC II+ and CD11c+MHC II+ cells in comparison with the IFN-??treated groups. In the EAO model, the type I IFN and E2 cooperated at the general and cellular levels of immune response, but E2 treatment usually abolished the effects exerted by the cytokine.

Effects of tamoxifen on estrogen receptor-α level in immune cells and humoral specific response after immunization of C3H/He male mice with syngeneic testicular germ cells (TGC)

Estrogens and estrogen receptors (ERs) are potent regulators of the immune response. Disruption of ERα or modulation of its function by selective ligands during experimental autoimmune conditions changes the course of disease by influencing specific humoral and cellular responses. However, it is not known whether fluctuation in the ERα level and the variable accessibility to its ligands in immune cells influence the development of specific immune responses against auto-antigens. This study was designed to evaluate the expression level of ERα in splenic immune cells and the specific humoral immune response in male C3H/He/W mice immunized with syngeneic testicular germ cells (TGC) in the presence of tamoxifen. Levels of ERα protein in immune cell subpopulations of immunized mice (assessed by flow cytometry) increased in MHCII+CD86+, MHCII+CD86− , F4/80+MHCII+, immature macrophages (F4/80+/MHCII− ), and CD3+CD4+ T cells. Addition of tamoxifen decreased the level of ERα in MHCII+CD86+, MHCII+CD86− , F4/80+MHCII+, immature macrophages (F4/80+/MHCII− ), and the CD19+CD3− cell subpopulation of immunized mice. Therefore, immunization with syngeneic antigen and tamoxifen treatment evoked cell-type specific changes in the level of ERα. Irrespective of tamoxifen treatment the humoral response in immunized animals toward TGCs was similar, suggesting that modulation of the level of ERα in immune cells is not directly related to specific auto-antibody production.

Differentiated all-trans retinoic acid response of naive CD4+CD25– cells isolated from rats with collagen-induced arthritis and healthy ones under in vitro conditions

Aim o the study To compare the potential of CD4+CD25– cells, isolated from both healthy rats and rats with CIA (Collagen-Induced Arthritis), for differentiation into regulatory T cells in the presence of all-trans retinoic acid in order to learn more about the activation mechanisms and therapeutic potential of regulatory T cells. Material and methods Sorted CD4+CD25– cells were cultured in vitro with/without ATRA, and then the frequency of regulatory T cells and their ability to secrete IL-10 by CD4+ FOXP3+ cells was examined. Gene expression of the foxp3, rarα, rarβ, rxrβ, and ppar β/δ and protein expression of the Rarα, Rarβ, and Rxrβ in cells after stimulation with ATRA were also investigated. Results CD4+CD25– cells isolated from healthy animals or from animals with CIA are characterised by different potential of the differentiation into CD4+CD25+ FOXP3+ cells. Retinoic acid receptor Rxrβ is present in the CD4+CD25– cells isolated from rats with CIA. Conclusions We showed that although ATRA did not increase the frequency of Treg in culture, it significantly increased expression of rarβ and rxrβ only in lymphocytes taken from diseased animals and foxp3 expression only in healthy animals. Moreover, after ATRA stimulation, the frequency of Treg-produced IL-10 tended to be lower in diseased animals than in the healthy group. The results imply that the potential of naïve cell CD4 lymphocytes to differentiate into Tregs and their putative suppressive function is dependent on the donor’s health status.