Yonsei Med J.  2018 Sep;59(7):879-886. 10.3349/ymj.2018.59.7.879.

Effects of Oncostatin M on Invasion of Primary Trophoblasts under Normoxia and Hypoxia Conditions

• Wie JH
• Ko HS
• Choi SK
• Park IY
• Kim A
• Kim HS
• Shin JC

• KMID: 2428915
• DOI: http://doi.org/10.3349/ymj.2018.59.7.879

Abstract

PURPOSE
To investigate the effect of oncostatin M (OSM) on protein expression levels and enzymatic activities of matrix metalloprotainase (MMP)-2 and MMP-9 in primary trophoblasts and the invasiveness thereof under normoxia and hypoxia conditions.
MATERIALS AND METHODS
Protein expression levels and enzymatic activities of MMP-2 and MMP-9 in primary trophoblasts under normoxia and hypoxia conditions were examined by Western blot and zymography, respectively. Effects of exogenous OSM on the in vitro invasion activity of trophoblasts according to oxygen concentration were also determined. Signal transducer and activator of transcription 3 (STAT3) siRNA was used to determine whether STAT3 activation in primary trophoblasts was involved in the effect of OSM.
RESULTS
OSM enhanced protein expression levels and enzymatic activities of MMP-2 and MMP-9 in term trophoblasts under hypoxia condition, compared to normoxia control (p < 0.05). OSM-induced MMP-2 and MMP-9 enzymatic activities were significantly suppressed by STAT3 siRNA silencing under normoxia and hypoxia conditions (p < 0.05). Hypoxia alone or OSM alone did not significantly increase the invasiveness of term trophoblasts. However, the invasion activity of term trophoblasts was significantly increased by OSM under hypoxia, compared to that without OSM treatment under normoxia.
CONCLUSION
OSM might be involved in the invasiveness of extravillous trophoblasts under hypoxia conditions via increasing MMP-2 and MMP-9 enzymatic activities through STAT3 signaling. Increased MMP-9 activity by OSM seems to be more important in primary trophoblasts.

Keyword

Oncostatin M; trophoblast; signal transducer and activator of transcription 3; hypoxia

MeSH Terms

Anoxia*
Blotting, Western
In Vitro Techniques
Oncostatin M*
Oxygen
RNA, Small Interfering
STAT3 Transcription Factor
Trophoblasts*
Oncostatin M
Oxygen
RNA, Small Interfering
STAT3 Transcription Factor

Figure

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  Fig. 1 Primary term trophoblasts after isolation of extravillous trophoblasts (HLA-G immunohistochemistry stain, ×200).

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  Fig. 2 Effect of OSM (20 ng/mL) treatment for 12 and 24 hours on MMP-2 and MMP-9 protein expressions in term trophoblasts (A) under normoxia and hypoxia. A quantitative measure of activities was determined by Western blot. Data shown are means±standard error of three independent experiments (B, C). The results are presented as the percent increase or decrease of protein expression with normalization to GAPDH. Significance versus untreated cells (control). *p<0.05. OSM, oncostatin M; MMP, matrix metalloproteinase.

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  Fig. 3 Effect of OSM (20 ng/mL) treatment for 12 and 24 hours on MMP-2 and MMP-9 secretion in conditioned medium (A). A quantitative measure of activity was determined by gelatine zymography. Results are means±standard error of three experiments (B, C). *p<0.05. OSM, oncostatin M; MMP, matrix metalloproteinase.

• 

  Fig. 4 Effects of OSM on protein expression of total and phosphorylated STAT3 (A). The expression was quantified and expressed as mean±SEM (B), *p<0.05, **p<0.001. Effect of siRNA on MMP-2 and MMP-9 secretion in conditioned medium (C). A quantitative measure of activity was determined by gelatine zymography. The expression was quantified and expressed as mean±SEM (D), black *p<0.05 (compared without OSM control at 0 hr), red *p<0.05 (compared with OSM over the same time line). Effects of OSM on protein expression total and phosphorylated STAT3 after transfection of STAT3 SiRNA (E). STAT3 and pSTAT3 expression after STAT3 SiRNA transfection was quantified and expressed as mean±SEM (F), black *p<0.05 (compared with normoxic or hypoxic control without OSM over the same time), red *p<0.05 (compared with normoxic or hypoxic controls with OSM over the same time). The vertical axis represent the relative value when the value of normoxia control is set to 1.0. OSM, oncostatin M; MMP, matrix metalloproteinase; STAT, signal transducer and activator of transcription 3.

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  Fig. 5 ECMatrix invasion by term primary trophoblasts (×200) after treatment with OSM (20 ng/mL) for 24 and 24 hours under normoxia (NC) and hypoxia (HC) (A). Results are presented as means±standard error of three experiments (B). *p<0.05. OCM, oncostatin M.

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