FHL1 Overexpression as A Inhibitor of Lung Cancer Cell Invasion via Increasing RhoGDIß mRNA Expression.

Figures

• 

  Fig.1. FHL1 was downregulated in human lung cancer tissues and cell lines. A, B. Bioinformatics analysis was performed on the FHL1 transcript with 574 lung adenocarcinoma (LUAD) patients from the TCGA database. The FHL1 transcripts were reduced in lung cancer tissues, in comparison with the normal lung tissues. The expression of FHL1 transcript was downregulated in lung cancer patients with an advanced stage. C. FHL1 mRNA expression was detected in 15 paired lung cancer tissues and their adjacent normal tissues. Student’s t test was used to detect the P value, P<0.05. D. Expression of FHL1 in Beas-2B, A549, and H1299 cell lines was determined by western blot assay. β-Actin was used as a protein loading control. E. FHL1 mRNA expression was determined by real-time polymerase chain reaction (PCR), and the asterisk (*) represents a notable decrease from normal Beas-2B cells (P<0.05).

• 

  Fig.2. FHL1 overexpression inhibited the invasion of A549 cell line. A. The Flag-tagged FHL1 plasmid was stably transfected into A549 cell line. B. The invasive ability was determined using the Biocoat™ matrigel® invasion chamber, while the migration ability was detected using the same system without the matrigel (scale bar: 200 µm), C. The invasive ability was normalized to the insert control. The asterisk (*) represents a significant reduction as compared to A549 (Vector) cell lines (P<0.05).

• 

  Fig.3. FHL1 overexpression inhibited the invasion of H1299 cell line. A. The Flag-tagged FHL1 plasmid was stably transfected into the H1299 cell line. B. The invasive ability was determined using the Biocoat™ matrigel® invasion chamber, while the migration ability was detected using the same system without the matrigel (scale bar: 200 µm). C. The invasive ability was normalized to the insert control. The asterisk (*) represents a significant reduction as compared to H1299 (Vector) cell lines (P<0.05).

• 

  Fig.4. FHL1 ectopic expression promoted the protein and mRNA expression of RhoGDIβ. A, B. The Flag-tagged FHL1 plasmid was transfected into A549 and H1299 cell lines stably. The western blot assay was utilized to detect the expression of RhoGDIα and RhoGDIβ protein. β-Actin was used as a protein loading control. C, D. RhoGDIβ mRNA expression was determined by real-time polymerase chain reaction (PCR). The bars indicate the mean ± standard deviation (SD) of 3 independent experiments. The asterisk (*) represents a notable enhancement in comparison with vector control cells (P<0.05).

• 

  Fig.5. RhoGDIβ acted as a FHL1 downstream mediator responsible for the FHL1-inhibited human lung cancer invasion. A. The RhoGDIβ knockdown constructs were transfected into A549 (Flag-FHL1) cell lines stably. B. The invasion abilities of A549 (Flag-FHL1/Nonsense), A549 (Flag-FHL1/shRhoGDIβ#1), and A549 (Flag-FHL1/shRhoGDIβ#2) cell lines were detected (scale bar: 200 µm). C. The bars indicate mean ± SD of 3 independent experiments. Student’s t test was used to detect the P value. The asterisk (*) represents a significant increase as compared to A549 (Flag-FHL1/Nonsense) transfectants (P<0.05).