Cytometric evaluation of intracellular IFN-γ and IL-4 levels in thyroid follicular cells from patients with autoimmune thyroid diseases

<p>Abstract</p> <p>Background</p> <p>In recent few years is underlined that altered balance of pro- and anti-inflammatory cytokines play an important role in the pathogenesis of AITD.</p> <p>The aim of this study was to estimate intracellular INF-γ and IL-4 levels in thyroid-infiltrating lymphocytes and thyrocytes isolated from thyroid tissues in 54 adolescent patients aged 8-21 years, with Graves' disease (GD; n = 18), Hashimoto's thyroiditis (HT; n = 18) and non-toxic multinodular goiter (NTMG; n = 18).</p> <p>Methods</p> <p>Fresh thyroid tissues were taken on culture medium RPMI -1640, it was mechanically prepared. In next step were added cell activators -12- myristate 13- the acetate (PMA) and Ionomycin as well as the inhibitor of transportation of proteins - Breferdin A. They were cultured 24 hours in 50 ml flasks at 37°C in a 5-95% CO2-air water-saturated atmosphere. After that, thyrocytes were identified by mouse mAb directed against human TPO epitope 64 conjugated with rabbit anti-mouse antibodies IgG (Fab')₂labeled by FITC. After incubation at room temperature to each of samples added reagent A fixative the cellular membrane. In next step into the cell suspensions were added reagent B to permeabilization of cellular membrane and specific anti-IL-4-PE or anti-IFN-γ-PE mAbs. Identification of intracellular cytokines in T lymphocytes was performed in the same procedure with application of anti-CD4-PerCP and anti-CD8-PerCP mAbs specific for T lymphocytes. The cells were analyzed in a flow cytometry (Coulter EPICS XL).</p> <p>Results</p> <p>In examined group of patients with GD we observed statistically significant higher mean percentage of cells with phenotype CD4+IL-4 (p < 0.05; p < 0.025), CD8+IL-4 (p < 0.033; p < 0.01) and TFCs-IL-4+ (p < 0.05; p < 0.01) in comparison to patients with HT and NTMG. The analysis of mean percentages of positive TILs and TFCs with intracellular INF-g levels in patients with HT revealed statistically significant increase percentage of CD4+INF-γ (p < 0.04; p < 0.001), CD8+ INF-γ (NS; p < 0.025), TFCs+INF-γ (p < 0.03; p < 0.001) cells in comparison to the percentage of positive cells from patients with GD and NTMG.</p> <p>Conclusions</p> <p>We conclude that human thyrocytes in autoimmune thyroid disorders could be a source of cytokine production and that their activation influences local interaction with T lymphocytes inflowing to the thyroid gland.</p

Increased percentage of T cells with the expression of CD127 and CD132 in hypertrophic adenoid in children with otitis media with effusion

The hypertrophic adenoid may promote chronic suppurative otitis media in children as it fulfills its immune function. The number of lymphocytes in the adenoid and their cooperation in the immune response depend of on their proliferation and migration to the effector sites. Interleukin 7 (IL-7) is essential for the normal development and function lymphocytes. IL-7 plays pivotal role for activation and proliferation of T and B cells. The heterodimeric interleukin-7 receptor (IL-7R) is composed of the IL-7Rα (127) and the common cytokine receptor γc (CD132). The aim of this study was to evaluate the percentage of lymphocytes T (CD4+ and CD8+) with IL-7R (CD127 and CD132) expression in hypertrophic adenoid in children suffering with otitis media with effusion for a duration of 3 months. Adenoid excised due to hypertrophy with or without chronic otitis media with effusion was used as study material. CD4+ CD127+, CD4+132+, CD8+CD127+ and CD8+CD132+ cell subpopulations were identified using monoclonal antibodies and flow cytometry. The percentage of CD4+ and CD8+ T cells with CD127 receptor expression in hypertrophic adenoid of children with otitis media with effusion was statistically significantly higher than in hypertrophic adenoid group. The percentage of CD4+ T cells with CD132 expression in the study group was statistically significantly higher than in the reference group. The percentage of CD8+ T cells with CD132+ expression was not statistically different in both groups. The increased percentage of T lymphocytes with IL-7R expression (CD127 and CD132) in hypertrophic adenoid seems to influence the quantity of lymphocytes and upset the immunological function of tonsils which can influence the course of otitis media with effusion

Select aspects of oogenesis and folliculogenesis

The processes of oogenesis and folliculogenesis begin very early in foetal life and are continued until the end of the reproductive period. Despite the numerous research conducted, the mechanism which initiates the growth of primary follicles has not been fully elucidated. The process of oocyte maturation begins after the recruitment of the primary follicle and continues until ovulation. This involves synthesis and accumulation of an appropriate number and kind of compounds and redistribution of cellular organelles inside the oocyte, which are necessary for the processes of fertilization and early embryonic development. The complexity and multi-functionality of these mechanisms and the factors which take part in the process of oogenesis indicate a necessity to conduct further research concerning this issue. A more thorough understanding of the processes of oogenesis and folliculogenesis will allow in time to develop more effective methods of treatment for a range of gynaecological, oncological, and endocrinological disorders

The effect of functional status of the ovaries on the embryological results of controlled ovarian hyperstimulation

Introduction: Controlled ovarian hyperstimulation is an integral part of infertility treatment. Despite many years of use, some aspects of controlled ovarian stimulation have not yet been clarified, especially the role of the functional status of the ovaries before hormonal stimulation. Aim of the research: To assess the effect of the functional status of the ovaries on the embryological results of controlled ovarian hyperstimulation. Material and methods: The retrospective study included female patients treated for infertility. The patients were divided into two groups depending on the ultrasonographic appearance of the ovaries before controlled ovarian hyperstimulation. Patients with small antral follicles < 6 mm in diameter were selected for group I. Patients with five or more antral follicles ≥ 8 mm in diameter in each ovary were included in group II. Patients from both groups underwent the same treatment process. We performed a detailed analysis of the number, type and quality of the obtained embryos. Results: The number of two- and three-blastomere embryos were comparable in the two groups. There were significantly more four-blastomere embryos in group I than II (p > 0.05). The numbers of A, C, D quality embryos were comparable between the groups (p > 0.05). There were more B quality embryos in group I than II (p > 0.05). The embryo growth rate was significantly faster in group I than II. Conclusions: The results of the present study indicate that the functional status of the ovaries before controlled ovarian hyperstimulation plays a pivotal role in treatment outcome