Interleukin-18 system messenger RNA and protein expression in human endometrium during the menstrual cycle  Hong-Yuan Huang, M.D., She-Hung Chan, B.Sc., Hsing-Tse Yu, M.D., Hsin-Shih Wang, M.D., Ph.D., Chyong-Huey Lai, M.D., Yung-Kuei Soong, M.D., M.Phil.  Fertility and Sterility  Volume 86, Issue 4, Pages 905-913 (October 2006) DOI: 10.1016/j.fertnstert.2006.02.122 Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions

FIGURE 1 A representative blot showed a complete IL-18 system, including IL-18, IL-18R, proIL-18, and IL-18 BP mRNA in human endometrium. M = DNA ladder. Huang. IL-18 system expression in human endometrium. Fertil Steril 2006. Fertility and Sterility 2006 86, 905-913DOI: (10.1016/j.fertnstert.2006.02.122) Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions

FIGURE 2 Immunohistochemistry study of the IL-18 system in human endometrium. (A) The presence of tissue macrophages in human endometrium was demonstrated by the immunohistochemistry. The inset photo was a negative control omitting the primary antibody. Human anti-CD68 was used as the primary antibody. A brown reaction product indicated a positive reaction for antibody staining (200×). Representative studies showed by immunohistochemistry that immunoreactive IL-18 (B), IL-18 BP (C), and IL-18R (D) were identified in the human endometrium samples using horseradish peroxidase staining technique. As a positive control, IL-18 was studied in human placenta samples (E). The inset photos were negative controls omitting the primary antibodies for matched tissues and targets. Significantly strong immunostaining of IL-18 and IL-18BP and weak staining of IL-18R at the protein level were also observed in human endometrium. All experiments were performed a minimum of three times with similar results. Huang. IL-18 system expression in human endometrium. Fertil Steril 2006. Fertility and Sterility 2006 86, 905-913DOI: (10.1016/j.fertnstert.2006.02.122) Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions

FIGURE 3 A representative experiment showing the steady-state amount of human endometrial IL-18 mRNA determined by QC-PCR in ectopic pregnancy. The standard curve was obtained for IL-18 mRNA by plotting the logarithmically transformed ratios of the densities of target cDNA to competitive cDNA against the log amount of initially added target cDNA in each PCR. Interleukin-18 (A) and IL-18R (B) mRNA expression were both decreased in human endometrium in secretory phase compared with proliferative phase. On the other hand, IL-18BP (C) mRNA expression increased in secretory phase compared with proliferative phase. Furthermore, proIL-18 mRNA (D) expression remained constant in secretory phase compared with proliferative phase. Values are mean ± SE of measurements on three representatives blots. M = DNA ladder. Huang. IL-18 system expression in human endometrium. Fertil Steril 2006. Fertility and Sterility 2006 86, 905-913DOI: (10.1016/j.fertnstert.2006.02.122) Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions

FIGURE 4 Pearson correlation of IL-18 system mRNA expression in human endometrium. There is a positive correlation of IL-18 and IL-18R in human endometrium in proliferative phase but not in secretory phase endometrium (A and B). (C) The ratio of IL-18BP to IL-18 mRNA levels in human endometrium showed a significantly higher ratio in secretory phase compared with proliferative phase (P=.041). Values are mean ± SE of measurements on three representatives blots. *P<.05, Mann-Whitney-Wilcoxon analysis. Huang. IL-18 system expression in human endometrium. Fertil Steril 2006. Fertility and Sterility 2006 86, 905-913DOI: (10.1016/j.fertnstert.2006.02.122) Copyright © 2006 American Society for Reproductive Medicine Terms and Conditions