Analysis of endoplasmic reticulum stress in placentas of HIV-infected women treated with protease inhibitors.

Combined antiretroviral therapy has proven efficacy in decreasing vertical HIV transmission. However, endoplasmic reticulum stress is a known side effect of HIV protease inhibitors. We investigated endoplasmic reticulum stress in placentas of HIV-infected and uninfected mothers by PCR-based splicing analysis of the specific endoplasmic reticulum stress marker XBP1 in post-delivery placental samples of uninfected mothers and in HIV-infected mothers taking antiretroviral therapy. No elevated XBP1 splicing could be detected in placentas of uninfected mothers and most of the mothers receiving combined anti-retroviral therapy. However, markedly elevated XBP1 splicing was found in the placentas of three individuals on combined antiviral therapy, all receiving lopinavir or atazanavir. In vitro experiments confirmed induction of endoplasmic reticulum stress by lopinavir and atazanavir in trophoblast-derived cell lines. Since endoplasmic reticulum stress occurred in selective patients only, individual differences in susceptibility of HIV-infected mothers to protease inhibitor induced endoplasmic reticulum stress can be postulated.

The inhibin-ßC subunit is down-regulated, while inhibin-ßE is up-regulated by interferon-ß1a in Ishikawa carcinoma cell line.

INTRODUCTION: Inhibins are important regulators of the female reproductive system. Recently, two new inhibin-subunits ßC and ßE have been described, although, their function is still quite unclear. Interestingly, there is an association between interferon and TGF-ß expression. Therefore, the aim of this study was to determine expression changes of inhibin-ßC and -ßE subunits in endometrial Ishikawa carcinoma cell line after stimulation with interferon-ß1a. MATERIALS AND METHODS: The Ishikawa cell line was cultured until confluence was observed (after 2 days). After adding interferon-ß1a (1,000 IE/ml), Ishikawa cells were analyzed for inhibin-ßC and -ßE subunits by RT-PCR. The fibroblast cell line BJ6 served as negative control. Experiments were performed in triplicates. RESULTS: The endometrial adenocarcinoma cell line Ishikawa synthesized the inhibin- ßC and -ßE subunits. The fibroblast cells BJ6 did not demonstrate an inhibin -ßC and -ßE mRNA expression, while inhibin-ßC subunit is down-regulated and inhibin-ßE is up-regulated in Ishikawa carcinoma cell line after stimulation with interferon-ß1a in Ishikawa. DISCUSSION: We demonstrated for the first time a functional relationship between interferon and the novel inhibin-ßC and -ßE subunits. It might be possible that interferon exerts a possible apoptotic function through the ßE-subunit, while, by down-regulating the ßC isoform, cell proliferation is inhibited. However, the precise function of the novel ßC- and ßE-subunits are still not known in human endometrial tissue and a possible association with interferon is still unclear and warrants further research.

Inhibin/activin-betaE subunit is expressed in normal and pathological human placental tissue including chorionic carcinoma cell lines.

BACKGROUND: Inhibins and activins are important regulators of the female reproductive system. Recently, a novel inhibin subunit, named betaE, has been identified and shown to be expressed in several human tissues. However, only limited data on the expression of this novel inhibin-betaE subunit in normal and pathological human placenta as well as and human chorionic carcinoma cell lines exist. MATERIALS AND METHODS: Tissue specimens of normal, preeclamptic and HELLP pregnancies (n = 18) were obtained at the course of an cesarean section. Normal and pathological placental tissues as well as chorionic carcinoma cells (BeWo and JEG) were analyzed by using immunohistochemistry and RT-PCR. RESULTS: Expression of the inhibin betaE subunit could be demonstrated at the protein level by means of immunohistochemical evaluation and at the transcriptional level by betaE-specific RT-PCR analysis. The immunoreactive score for inhibin-betaE did not show any significant differences between normal, preeclamptic and HELLP tissue in extravillous trophoblast and syncytiotrophoblast cells. Expression of inhibin betaE could further be demonstrated for the human chorionic carcinoma cell lines JEG and BeWo. DISCUSSION: We demonstrated that inhibin-betaE is expressed in normal and pathological human placenta tissues. Although the precise role of this novel inhibin subunit for human placenta development is quite unclear, similarities with the well-characterized betaA- and betaB-subunits suggest an involvement in autocrine/paracrine signaling pathways, angiogenesis, decidualization and tissue remodeling under normal as well as malignant conditions. Additionally, the human chorionic carcinoma cell lines JEG and BeWo synthesize this subunit and therefore can be used as a cell culture model for further functional analysis of this subunit in human placental tissue.

Immunohistochemical labeling of the inhibin/activin betaC subunit in normal human placental tissue and chorionic carcinoma cell lines.

Inhibins and activins are important regulators of the female reproductive system. A novel inhibin subunit, named betaC, has been identified and demonstrated to be expressed in several human tissues. We demonstrate here that inhibin betaC is expressed in human placenta. Expression of the inhibin betaC subunit was demonstrated at the protein level by means of immunohistochemical evaluation and at the transcriptional level by an inhibin betaC-specific RT-PCR analysis. Expression of inhibin betaC was detected in the human chorionic carcinoma cell lines JEG and BeWo. Although the precise role of this novel inhibin subunit in human placenta development and homeostasis is unclear, analogies with other inhibin subunits and the strong expression of betaC in normal human trophoblast cells and chorionic carcinoma cells suggest that betaC may be involved in autocrine/paracrine signaling pathways, angiogenesis, decidualization, and tissue remodeling under normal and malignant conditions. Additionally, JEG and BeWo express betaC and, therefore, can be used as a cell culture model for further functional analysis of this subunit in the human placenta.

Inhibin/activin-betaC and -betaE subunits in the Ishikawa human endometrial adenocarcinoma cell line.

BACKGROUND: Inhibins and activins are important regulators of the female reproductive system. Recently, two novel inhibin subunits, named betaC and betaE, have been identified and shown to be expressed in several human tissues. However, only limited data on the expression of these novel inhibin subunits in normal human endometrial tissue and endometrial adenocarcinoma cell lines exist. MATERIALS AND METHODS: Samples of proliferative and secretory human endometrium were obtained from five premenopausal, non-pregnant patients undergoing gynecological surgery for benign diseases. Normal endometrial tissue and Ishikawa endometrial adenocarcinoma cell lines were analyzed by immunohistochemistry, immunofluorescence and RT-PCR. RESULTS: Expression of the inhibin betaC and betaE subunits could be demonstrated at the protein level by means of immunohistochemical evaluation and at the transcriptional level by establishing a betaC- and betaE-specific RT-PCR analysis in normal human endometrial tissue and the parental Ishikawa cell line. Interestingly, in a highly de-differentiated subclone of the Ishikawa cell line lacking estrogen receptor expression, the expression of the inhibin-betaC subunit appeared strongly reduced. DISCUSSION: Here, we show for the first time that the novel inhibin/activin-betaC and -betaE subunits are expressed in normal human endometrium and the estrogen receptor positive human endometrial carcinoma cell line Ishikawa using RT-PCR and immunohistochemical detection methods. Interestingly, the Ishikawa minus cell line (lacking estrogen receptor expression) demonstrated no to minimal expression of the betaC subunit as observed with immunofluorescence and RT-PCR, suggesting a possible hormone- dependency of this subunit in human endometrial cancer cells. Moreover, because the Ishikawa cell line minus is thought to be a more malignant endometrial cell line than its estrogen receptor positive counterpart, inhibin-betaC subunit might be substantially involved in the pathogenesis and malignant transformation in human endometrium.