Cytokine microenvironments in human first trimester decidua are dependent on trophoblast cells  Ulrike von Rango, PhD, Irmgard Classen-Linke, PhD, Gabie Raven, MD, Frans Bocken, MD, Henning M Beier, MD, PhD  Fertility and Sterility  Volume 79, Issue 5, Pages 1176-1186 (May 2003) DOI: 10.1016/S0015-0282(02)04829-X

FIGURE 1 Cytokine fibroblasts messenger (m)RNA expression in first-trimester epithelial cells, stromal cells (fibroblasts), and leukocytes. (A) Cytocentrifugation of epithelial cells and leukocytes. The various cell populations were identified by immunohistochemistry. (1) Epithelial cells identified by staining for cytokeratin. (2) Stromal cells (fibroblasts) identified by staining for vimentin. (3) Staining for leukocyte common antigen, which confirmed the identity of the leukocyte population. The stromal cell population contained cytokeratin-positive epithelial (including trophoblast) cells (4) and CD45-positive leukocytes (5). Original magnification, ×150. (B) Typical reverse transcription polymerase chain reaction (RT-PCR) signals for interleukin (IL)-2, IL-4, interferon (IFN)-γ, and IL-1β. Interleukin-2, IL-4, and IFN-γ are expressed predominantly in decidual leukocytes, whereas IL-1β was found in the leukocyte and fibroblast fraction. Values are means (±SEM, indicated by the bars) corrected for minimal differences in the amount of complementary DNA by internal control Cyt-1A. (C) Typical cycle test to define the linear amplification range of IL-1β, using fibroblasts (a) and epithelial cells (b) to obtain the optimal number of cycles for RT-PCR. In this case, 27 cycles were used. von Rango. Cytokine microenvironments in human decidua. Fertil Steril 2003. Fertility and Sterility 2003 79, 1176-1186DOI: (10.1016/S0015-0282(02)04829-X)

FIGURE 2 Cytokine messenger (m)RNA and protein expression in decidua basalis and decidua parietalis. (A) Immunostaining for cytokeratin (epithelial cell marker), which clearly differentiated decidua basalis (presence of trophoblast confirmed by staining of extravillous trophoblast cells) from decidua parietalis (no invading trophoblast cells; only glandular epithelial cells are positive). Representative reverse transcription polymerase chain reaction (RT-PCR) signals are given for each cytokine. In each pair of samples (n = 6), the OD value of the RT-PCR signal for the decidua basalis was expressed in relation to that of the decidua parietalis (defined as 1.0). Values for mRNA expression in the decidua basalis are means (± SEM). Interferon (IFN)γ mRNA expression was significantly higher in decidua basalis than decidua parietalis (P=.0426; paired t-test). Interleukin (IL)-4 mRNA expression was nonsignificantly higher in decidua parietalis than decidua basalis. (B) Enzyme-linked immunosorbent assay of IFN-γ content of decidua basalis and decidua parietalis (7 pairs). Resulting data were processed as described in case of RT-PCR. Values are means (±SEM) in the decidua basalis. The significantly higher IFN-γ protein expression in decidua basalis compared with decidua parietalis confirmed the RNA data (P=.0324; paired t-test). von Rango. Cytokine microenvironments in human decidua. Fertil Steril 2003. Fertility and Sterility 2003 79, 1176-1186DOI: (10.1016/S0015-0282(02)04829-X)

FIGURE 3 Cytokine protein content in decidual tissue homogenate, aspirated fluid, serum of pregnant women, and serum from nonpregnant women. Values are expressed as means (±SEM, indicated by the bars). Cytokines were found predominantly in decidual tissue homogenate and did not appear in serum of pregnant or nonpregnant women. The P value (Student two-tailed t-test) indicated differences between decidual tissue and aspirated fluid. von Rango. Cytokine microenvironments in human decidua. Fertil Steril 2003. Fertility and Sterility 2003 79, 1176-1186DOI: (10.1016/S0015-0282(02)04829-X)