Reduction of estrogen production by interleukin-6 in a human granulosa tumor cell line may have implications for endometriosis-associated infertility

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Reduction of estrogen production by interleukin-6 in a human granulosa tumor cell line may have implications for endometriosis-associated infertility Imari Deura, M.D., Tasuku Harada, M.D., Fuminori Taniguchi, M.D., Tomio Iwabe, M.D., Masao Izawa, Ph.D., Naoki Terakawa, M.D. Fertility and Sterility Volume 83, Issue 4, Pages 1086-1092 (April 2005) DOI: 10.1016/j.fertnstert.2004.12.014 Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

FIGURE 1 Expression of IL-6, IL-6R, and gp130 in KGN cells and granulose lutein cells (GLCs). The polymerase chain reaction (PCR) products for IL-6 (173 bp), IL-6R (263 bp), gp130 (423 bp), and internal control G3PDH (450 bp) are indicated. Specificity of each PCR product was confirmed by Southern blot analysis. Fertility and Sterility 2005 83, 1086-1092DOI: (10.1016/j.fertnstert.2004.12.014) Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

FIGURE 2 Effect of IL-6 on E2 production in KGN cells. KGN cells were primed with 500 mIU/mL hMG for 48 hours after the addition of 10−5 M of T (test: substrate for E2) in the presence or absence of 0.1–10 ng/mL of IL-6. The concentrations of E2 in the medium were measured by EIA. The data are the mean with SE of three different experiments. Asterisks show significant differences. *P<.01. Fertility and Sterility 2005 83, 1086-1092DOI: (10.1016/j.fertnstert.2004.12.014) Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

FIGURE 3 Effect of IL-6 on aromatase activity in KGN cells. KGN cells were cultured for 48 hours with IL-6 (0.1–10 ng/mL), IL-6 (10 ng/mL) plus U0126 (10 μM), or IL-6 plus anti-IL-6 antibody (A-IL-6) (50 ng/mL) in the presence of hMG (500 mIU/mL). The data are the mean with SE of three different experiments. Asterisks show significant differences. *P<.01. Fertility and Sterility 2005 83, 1086-1092DOI: (10.1016/j.fertnstert.2004.12.014) Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

FIGURE 4 Effect of IL-6 on the level of mRNA for aromatase in KGN cells. Subconfluent cells were cultured for 48 hours with IL-6 (10 ng/mL) or IL-6 (10 ng/mL) plus U0126 (10 μM) in the presence of hMG (500 mIU/mL). Reverse transcriptase-polymerase chain reaction and Southern blot analysis for aromatase mRNA are shown. Fertility and Sterility 2005 83, 1086-1092DOI: (10.1016/j.fertnstert.2004.12.014) Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

FIGURE 5 The time course of ERK1/2 phosphorylation induced by IL-6 in KGN cells. Interleukin-6 (10 ng/mL) was added to the medium for 0–90 minutes as indicated. Phosphorylated and total ERK1/2 (44 and 42 kD) were detected by Western blot analysis. Fertility and Sterility 2005 83, 1086-1092DOI: (10.1016/j.fertnstert.2004.12.014) Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions

FIGURE 6 Effect of IL-6 on phosphorylation of ERK1/2 in KGN cells by Western blot analysis. KGN cells were pretreated for 30 minutes with U0126 (10 μM). Next, IL-6 (10 ng/mL) was treated after 10 minutes. Phospho-ERK1/2 (P-ERK1/2) and total ERK1/2 (ERK1/2) were indicated. Relative densitometric units of the bands are shown in the lower panel, with the density of the control band set arbitrarily at 1.0. Each bar represents the means with SE. ND: not detectable. Significant differences are shown by asterisks: **P<.05. Fertility and Sterility 2005 83, 1086-1092DOI: (10.1016/j.fertnstert.2004.12.014) Copyright © 2005 American Society for Reproductive Medicine Terms and Conditions