MiR-199a attenuates endometrial stromal cell invasiveness through suppression of the IKKβ/NF-κB pathway and reduced interleukin-8 expression
Advanced Access publication on October
MiR-199a attenuates endometrial stromal cell invasiveness through suppression of the IKKb/NF-kB pathway and reduced interleukin-8 expression
Lan Dai 0 1
Liying Gu 0 1
0 Shanghai Key Laboratory of Gynecologic Oncology, Focus Construction Subject of Shanghai Education Department , Shanghai , People's Republic of China
1 Department of Obstetrics and Gynecology, Renji Hospital, Shanghai Jiaotong University School of Medicine , 1630 Dong Fang Rd., Shanghai 200127 , People's Republic of China
MicroRNAs have recently been identified as regulators that modulate target gene expression and are suggested to be involved in the development and progression of endometriosis. This study was undertaken to analyze the expression level of microRNA199a (miR-199a) in paired ovarian endometrioma and eutopic endometrium from women with endometriosis, and to investigate the contribution of miR-199a to the invasive capability of endometrial stromal cells (ESCs). Cell adhesion, migration and Matrigel invasion assays were carried out to measure the invasiveness of ESCs. Bioinformatics prediction, reporter gene assay, PCR, western blotting and ELISA were performed to identify miR-199a targets and related signaling pathways. The results showed that the expression level of miR-199a was lower in the eutopic endometrium from women with endometriosis, and even lower in the paired ovarian endometrioma, compared with the expression in normal controls. Moreover, ectopic expression of miR-199a attenuated ESC adhesion, migration and invasiveness. MiR-199a targeted and inhibited IkappaB kinase beta (IKKb) in ESCs. Accompanied by IKKb reduction, miR-199a suppressed nuclear factor-kappa B (NF-kB) pathway activation and interleukin-8 (IL-8) production in ESCs. All these findings suggest that miR-199a, down-regulated in endometriosis, attenuates the invasive capability of ESCs in vitro partly through IKK/NF-kB pathway suppression and reduced IL-8 expression. In conclusion, miR-199a could be involved in the pathogenesis of endometriosis.
endometriosis / miR-199a / invasion / nuclear factor-kappa B / interleukin-8
Introduction
Endometriosis, characterized by the presence and growth of functional
endometrial-like tissues outside the uterus, is a common, chronic and
estrogen-dependent gynecological disorder associated with pelvic pain
and infertility. An understanding of the pathogenesis of endometriosis
has steadily improved, but the molecular mechanisms mediating
endometriosis are just beginning to be elucidated. In particular, epigenetic
factors have been suggested as a regulatory source promoting
endometriosis development, including the expression and function of
microRNAs (miRNAs)
(Guo, 2009; Qian et al., 2009; Teague et al.,
2010; Hawkins et al., 2011)
.
MiRNAs are small non-coding RNAs of 20 – 22 nucleotides, which
post-transcriptionally regulate gene expression and can control a
† Lan Dai and Liying Gu contributed equally to this work.
broad spectrum of normal and pathological cellular functions
(Djuranovic et al., 2011; Huntzinger and Izaurralde, 2011)
. First
discovered in 2003
(Lagos-Quintana et al., 2003; Lim et al., 2003)
,
microRNA-199a (miR-199a) has been shown to be down-regulated
in several cancerous tissues and to contribute to various malignant
processes, such as tumor invasion, metastasis and angiogenesis
(Shen et al., 2010; Cheung et al., 2011)
. Furthermore, miR-199a can
target IkappaB kinase beta (IKKb) (Chen et al., 2008), a co-factor
required for nuclear factor-kappa B (NF-kB) pathway activation,
which may drive the expression of several genes related to malignant
transformation. Recently, multiple published studies have identified
differentially expressed miRNAs in endometriotic tissues
(Pan et al.,
2007; Toloubeydokhti et al., 2008; Burney et al., 2009; Ohlsson
Teague et al., 2009; Filigheddu et al., 2010; Hawkins et al., 2011;
Ramon et al., 2011)
, and the results of one research study suggest that
miR-199a may decrease in endometriosis
(Pan et al., 2007)
. However,
to the best of our knowledge, the expression level of miR-199a in
endometriosis has not been validated and the role of miR-199a in
the disease remains largely unknown.
Numerous in vitro and in vivo studies have suggested that NF-kB
plays an important role in regulating key cell processes that may
contribute to the initiation and progression of endometriosis, such as cell
adhesion, migration, invasion and angiogenesis
(Gonzalez-Ramos et al.,
2010; Zhang et al., 2010, 2011)
. NF-kB is composed of homo- and
heterodimers of five members of the Rel family including NF-kB1
(p50), NF-kB2 (p52), RelA (p65), RelB and c-Rel (Rel). NF-kB
p50/p65 is generally considered to be the predominant and inducible
form of NF-kB in most cells, and the general term NF-kB traditionally
refers to the p50/p65 heterodimer
(McKay and Cidlowski, 1999;
Karin et al., 2004)
. For this reason, throughout the remainder of this
text, the (...truncated)