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. 2020 Apr 16;18(1):31.
doi: 10.1186/s12958-020-00590-3.

hsa-miR-100-5p, an overexpressed miRNA in human ovarian endometriotic stromal cells, promotes invasion through attenuation of SMARCD1 expression

Kanetoshi Takebayashi  1 , Kaei Nasu  2   3 , Mamiko Okamoto  1 , Yoko Aoyagi  1 , Tomoko Hirakawa  1 , Hisashi Narahara  1
Affiliations

hsa-miR-100-5p, an overexpressed miRNA in human ovarian endometriotic stromal cells, promotes invasion through attenuation of SMARCD1 expression

Kanetoshi Takebayashi et al. Reprod Biol Endocrinol. .

Abstract

Background: A number of microRNAs are aberrantly expressed in endometriosis and are involved in its pathogenesis. Our previous study demonstrated that has-miR-100-5p expression is enhanced in human endometriotic cyst stromal cells (ECSCs). The present study aimed to elucidate the roles of has-miR-100-5p in the pathogenesis of endometriosis.

Methods: Normal endometrial stromal cells (NESCs) were isolated from normal eutopic endometrium without endometriosis. Using hsa-miR-100-5p-transfected NESCs, we evaluated the effect of hsa-miR-100-5p on the invasiveness of these cells by Transwell invasion assay and in-vitro wound repair assay. We also investigated the downstream signal pathways of hsa-miR-100-5p by microarray analysis and Ingenuity pathways analysis.

Results: hsa-miR-100-5p transfection enhanced the invasion and motility of NESCs. After hsa-miR-100-5p transfection, mRNA expression of SWItch/sucrose non-fermentable-related matrix-associated actin-dependent regulator of chromatin subfamily D member 1 (SMARCD1) was significantly attenuated. Whereas, the expression of matrix metallopeptidase 1 (MMP1) mRNA and active MMP1 protein levels was upregulated.

Conclusion: We found that SMARCD1/MMP-1 is a downstream pathway of hsa-miR-100-5p. hsa-miR-100-5p transfection enhanced the motility of NESCs by inhibiting SMARCD1 expression and MMP1 activation. These findings suggest that enhanced hsa-miR-100-5p expression in endometriosis is involved in promoting the acquisition of endometriosis-specific characteristics during endometriosis development. Our present findings on the roles of hsa-miR-100-5p may thus contribute to understand the epigenetic mechanisms involved in the pathogenesis of endometriosis.

Keywords: Endometriosis; Hsa-miR-100-5p; Invasion; Matrix metallopeptidase 1; SMARCD1.

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Conflict of interest statement

There are no conflicts of interest to declare.

Figures

Fig. 1
Fig. 1
Downstream signaling pathway of hsa-miR-100-5p in NESCs. A gene expression microarray and pathway analyses of hsa-miR-100-5p-transfected NESCs revealed that hsa-miR-100-5p upregulated the motility of NESCs by direct inhibition of SMARCD1 expression followed by MMP1 activation. SMARCD1, SWItch/sucrose non-fermentable-related matrix-associated actin-dependent regulator of chromatin subfamily D member 1; MMP1, matrix metallopeptidase 1; NESCs, normal endometrial stromal cells
Fig. 2
Fig. 2
hsa-miR-100-5p expression in NESCs and ECSCs. The relative hsa-miR-100-5p levels in ECSCs (n = 6) were significantly higher than those in the NESCs (n = 6). *p < 0.0005 vs. NESCs (Student’s t-test). Data are shown as the mean ± SD. ECSCs, endometriotic cyst stromal cells; NESCs, normal endometrial stromal cells
Fig. 3
Fig. 3
Effects of hsa-miR-100-5p transfection on the downstream target molecule expression in NESCs. (a) hsa-miR-100-5p expression after precursor miRNA transfection. Note that the vertical axis is expressed as a logarithmic scale. (b) SMARCD1 mRNA expression. (c) MMP1 mRNA expression. (d) Active MMP1 protein expression. *p < 0.05, **p < 0.005, #p < 0.0005 vs. controls (Student’s t-test). MMP1, matrix metallopeptidase 1; NESCs, normal endometrial stromal cells; SMARCD1, SWItch/sucrose non-fermentable-related matrix-associated actin-dependent regulator of chromatin subfamily D member 1
Fig. 4
Fig. 4
Effects of SMARCD1 siRNA transfection on the MMP1 mRNA expression in NESCs. (a) SMARCD1 mRNA expression after SMARCD1 siRNA transfection. (b) MMP1 mRNA expression. *p < 0.05, **p < 0.005 vs. controls (Student’s t-test). MMP1, matrix metallopeptidase 1; NESCs, normal endometrial stromal cells; SMARCD1, SWItch/sucrose non-fermentable-related matrix-associated actin-dependent regulator of chromatin subfamily D member 1
Fig. 5
Fig. 5
Effects of hsa-miR-100-5p transfection on the motility of NESCs. (a) Results of transwell invasion assay. (b) Representative photographs of transwell invasion assay. (c) Results of in-vitro wound repair assay. (d) Representative photographs of in vitro wound repair assay. *p < 0.05, **p < 0.0005 vs. controls (Student’s t-test). NESCs, normal endometrial stromal cells
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