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Decreasing the ratio of matriptase/HAI‑1 by downregulation of matriptase as a potential adjuvant therapy in ovarian cancer.

Sun P, Jiang Z, Chen X, Xue L, Mao X, Ruan G, Song Y, Mustea A - Mol Med Rep (2016)

Bottom Line: The migration distance and number of invading HO‑8910PM cells were significantly increased compared with HO‑8910 cells.HO‑8910PM cells exhibited a significantly higher ratio of matriptase/HAI‑1 mRNA levels compared with HO‑8910 cells (0.51 vs. 0.24, ~2.2 fold increase).Similar increases in protein expression levels were also observed in HO‑8910PM cells compared with HO‑8910 cells.

View Article: PubMed Central - PubMed

Affiliation: Institute of Gynecologic Oncology, Department of Gynecology, Fujian Maternity and Children Health Hospital, Teaching Hospital of Fujian Medical University, Fuzhou, Fujian 350001, P.R. China.

ABSTRACT
Tumor invasion and metastasis are complex biological processes. Matriptase and its endogenous inhibitor, hepatocyte growth factor activator inhibitor‑1 (HAI‑1) are involved in invasion and metastasis. To evaluate the ratio of matriptase/HAI‑1 and their potential therapeutic value in ovarian cancer, HO‑8910 human ovarian cancer cells and the homologous high‑metastatic HO‑8910PM cells were used as in vitro cellular models ovarian cancer. The invasive and metastatic abilities, and the expression of matriptase and HAI‑1 in these cells were detected using scratch assays, Transwell chamber assays, reverse transcription‑quantitative polymerase chain reaction, western blotting and fluorescent immunocytochemistry. Following infection with lentivirus‑mediated matriptase‑targeting small interfering RNA (siRNA), cell cycle progression and apoptosis were also analyzed. The migration distance and number of invading HO‑8910PM cells were significantly increased compared with HO‑8910 cells. HO‑8910PM cells exhibited a significantly higher ratio of matriptase/HAI‑1 mRNA levels compared with HO‑8910 cells (0.51 vs. 0.24, ~2.2 fold increase). Compared with HO‑8910 cells, the matriptase mRNA level was increased by ~3.6 fold in HO‑8910PM cells, whereas the HAI‑1 mRNA level was increased by ~1.7 fold. Similar increases in protein expression levels were also observed in HO‑8910PM cells compared with HO‑8910 cells. Migration and invasiveness were positively correlated with matriptase expression level (r=0.994, P<0.01) and the ratio of matriptase/HAI‑1 (r=0.929, P<0.01). Downregulation of matriptase using siRNA resulted in inhibition of the invasive and metastatic abilities of HO‑8910PM cells, cell cycle arrest in the G0/G1 phase and increased apoptosis. The present study demonstrated that ovarian cancer cell metastasis and invasion were more dependent on upregulation of matriptase levels than downregulation of HAI‑1. Matriptase may be a potential adjuvant therapeutic target for inhibiting ovarian cancer invasion and metastasis.

No MeSH data available.


Related in: MedlinePlus

Downregulation of matriptase expression by RNA interference. (A) Microscopic examination of the infection efficiencies of different lentiviral-mediated SI constructs targeting matriptase in HO-8910PM cells (magnification, ×100). There was no signal observed in HO-8910PM-PBS. The infection efficiencies exceeded 80% at a MOI of 80. (B) Relative matriptase mRNA expression levels in HO-8910PM cells infected with different lentiviral particles at high MOI. Matriptase SI2 achieved the highest inhibition of matriptase mRNA expression, and little inhibition was observed in HO-8910PM-NC cells. The SI inhibited matriptase expression but not HAI-1 expression. The ratio of matriptase/HAI-1 was significantly reduced in the HO-8910PM cells treated with matriptase SI2. (C) Relative matriptase protein expression levels in HO-8910PM cells transfected with different lentiviral particles at high MOI. Protein quantification was normalized using β-actin. The highest silencing efficiency was achieved in HO-8910PM-SI2 cells using matriptase SI2. *P>0.05 vs. HO-8910PM; #P<0.05 vs. HO-8910PM. MOI, multiplicity of infection; HAI-1, hepatocyte growth factor activator inhibitor-1; NC, negative control; SI, small interfering RNA.
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f3-mmr-14-02-1465: Downregulation of matriptase expression by RNA interference. (A) Microscopic examination of the infection efficiencies of different lentiviral-mediated SI constructs targeting matriptase in HO-8910PM cells (magnification, ×100). There was no signal observed in HO-8910PM-PBS. The infection efficiencies exceeded 80% at a MOI of 80. (B) Relative matriptase mRNA expression levels in HO-8910PM cells infected with different lentiviral particles at high MOI. Matriptase SI2 achieved the highest inhibition of matriptase mRNA expression, and little inhibition was observed in HO-8910PM-NC cells. The SI inhibited matriptase expression but not HAI-1 expression. The ratio of matriptase/HAI-1 was significantly reduced in the HO-8910PM cells treated with matriptase SI2. (C) Relative matriptase protein expression levels in HO-8910PM cells transfected with different lentiviral particles at high MOI. Protein quantification was normalized using β-actin. The highest silencing efficiency was achieved in HO-8910PM-SI2 cells using matriptase SI2. *P>0.05 vs. HO-8910PM; #P<0.05 vs. HO-8910PM. MOI, multiplicity of infection; HAI-1, hepatocyte growth factor activator inhibitor-1; NC, negative control; SI, small interfering RNA.

Mentions: Three lentivirus-mediated siRNAs targeting matriptase were constructed and used to infect HO-8910PM cells at high and low MOIs (Fig. 3A). Compared with NC, matriptase mRNA levels in HO-8910PM cells were decreased by 19.63, 89.72 and 32.43% by Ma-siRNA-1, Ma-siRNA-2, and Ma-siRNA-3, respectively (Fig. 3B). Additionally, western blot analysis demonstrated that Ma-siRNA-2 significantly reduced the matriptase protein expression level compared with NC (P<0.01; Fig. 3C). The siRNA inhibited matriptase expression but did not affect HAI-1 levels. The HO-8910PM cells infected with Ma-siRNA-2, which induced the greatest inhibition of matriptase levels, were termed HO-8910PM-SI2 and selected for further analysis.


Decreasing the ratio of matriptase/HAI‑1 by downregulation of matriptase as a potential adjuvant therapy in ovarian cancer.

Sun P, Jiang Z, Chen X, Xue L, Mao X, Ruan G, Song Y, Mustea A - Mol Med Rep (2016)

Downregulation of matriptase expression by RNA interference. (A) Microscopic examination of the infection efficiencies of different lentiviral-mediated SI constructs targeting matriptase in HO-8910PM cells (magnification, ×100). There was no signal observed in HO-8910PM-PBS. The infection efficiencies exceeded 80% at a MOI of 80. (B) Relative matriptase mRNA expression levels in HO-8910PM cells infected with different lentiviral particles at high MOI. Matriptase SI2 achieved the highest inhibition of matriptase mRNA expression, and little inhibition was observed in HO-8910PM-NC cells. The SI inhibited matriptase expression but not HAI-1 expression. The ratio of matriptase/HAI-1 was significantly reduced in the HO-8910PM cells treated with matriptase SI2. (C) Relative matriptase protein expression levels in HO-8910PM cells transfected with different lentiviral particles at high MOI. Protein quantification was normalized using β-actin. The highest silencing efficiency was achieved in HO-8910PM-SI2 cells using matriptase SI2. *P>0.05 vs. HO-8910PM; #P<0.05 vs. HO-8910PM. MOI, multiplicity of infection; HAI-1, hepatocyte growth factor activator inhibitor-1; NC, negative control; SI, small interfering RNA.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4940087&req=5

f3-mmr-14-02-1465: Downregulation of matriptase expression by RNA interference. (A) Microscopic examination of the infection efficiencies of different lentiviral-mediated SI constructs targeting matriptase in HO-8910PM cells (magnification, ×100). There was no signal observed in HO-8910PM-PBS. The infection efficiencies exceeded 80% at a MOI of 80. (B) Relative matriptase mRNA expression levels in HO-8910PM cells infected with different lentiviral particles at high MOI. Matriptase SI2 achieved the highest inhibition of matriptase mRNA expression, and little inhibition was observed in HO-8910PM-NC cells. The SI inhibited matriptase expression but not HAI-1 expression. The ratio of matriptase/HAI-1 was significantly reduced in the HO-8910PM cells treated with matriptase SI2. (C) Relative matriptase protein expression levels in HO-8910PM cells transfected with different lentiviral particles at high MOI. Protein quantification was normalized using β-actin. The highest silencing efficiency was achieved in HO-8910PM-SI2 cells using matriptase SI2. *P>0.05 vs. HO-8910PM; #P<0.05 vs. HO-8910PM. MOI, multiplicity of infection; HAI-1, hepatocyte growth factor activator inhibitor-1; NC, negative control; SI, small interfering RNA.
Mentions: Three lentivirus-mediated siRNAs targeting matriptase were constructed and used to infect HO-8910PM cells at high and low MOIs (Fig. 3A). Compared with NC, matriptase mRNA levels in HO-8910PM cells were decreased by 19.63, 89.72 and 32.43% by Ma-siRNA-1, Ma-siRNA-2, and Ma-siRNA-3, respectively (Fig. 3B). Additionally, western blot analysis demonstrated that Ma-siRNA-2 significantly reduced the matriptase protein expression level compared with NC (P<0.01; Fig. 3C). The siRNA inhibited matriptase expression but did not affect HAI-1 levels. The HO-8910PM cells infected with Ma-siRNA-2, which induced the greatest inhibition of matriptase levels, were termed HO-8910PM-SI2 and selected for further analysis.

Bottom Line: The migration distance and number of invading HO‑8910PM cells were significantly increased compared with HO‑8910 cells.HO‑8910PM cells exhibited a significantly higher ratio of matriptase/HAI‑1 mRNA levels compared with HO‑8910 cells (0.51 vs. 0.24, ~2.2 fold increase).Similar increases in protein expression levels were also observed in HO‑8910PM cells compared with HO‑8910 cells.

View Article: PubMed Central - PubMed

Affiliation: Institute of Gynecologic Oncology, Department of Gynecology, Fujian Maternity and Children Health Hospital, Teaching Hospital of Fujian Medical University, Fuzhou, Fujian 350001, P.R. China.

ABSTRACT
Tumor invasion and metastasis are complex biological processes. Matriptase and its endogenous inhibitor, hepatocyte growth factor activator inhibitor‑1 (HAI‑1) are involved in invasion and metastasis. To evaluate the ratio of matriptase/HAI‑1 and their potential therapeutic value in ovarian cancer, HO‑8910 human ovarian cancer cells and the homologous high‑metastatic HO‑8910PM cells were used as in vitro cellular models ovarian cancer. The invasive and metastatic abilities, and the expression of matriptase and HAI‑1 in these cells were detected using scratch assays, Transwell chamber assays, reverse transcription‑quantitative polymerase chain reaction, western blotting and fluorescent immunocytochemistry. Following infection with lentivirus‑mediated matriptase‑targeting small interfering RNA (siRNA), cell cycle progression and apoptosis were also analyzed. The migration distance and number of invading HO‑8910PM cells were significantly increased compared with HO‑8910 cells. HO‑8910PM cells exhibited a significantly higher ratio of matriptase/HAI‑1 mRNA levels compared with HO‑8910 cells (0.51 vs. 0.24, ~2.2 fold increase). Compared with HO‑8910 cells, the matriptase mRNA level was increased by ~3.6 fold in HO‑8910PM cells, whereas the HAI‑1 mRNA level was increased by ~1.7 fold. Similar increases in protein expression levels were also observed in HO‑8910PM cells compared with HO‑8910 cells. Migration and invasiveness were positively correlated with matriptase expression level (r=0.994, P<0.01) and the ratio of matriptase/HAI‑1 (r=0.929, P<0.01). Downregulation of matriptase using siRNA resulted in inhibition of the invasive and metastatic abilities of HO‑8910PM cells, cell cycle arrest in the G0/G1 phase and increased apoptosis. The present study demonstrated that ovarian cancer cell metastasis and invasion were more dependent on upregulation of matriptase levels than downregulation of HAI‑1. Matriptase may be a potential adjuvant therapeutic target for inhibiting ovarian cancer invasion and metastasis.

No MeSH data available.


Related in: MedlinePlus