Differential Expression of PAI-RBP1, C1orf142, and COTL1 in Non–Small Cell Lung Cancer Cell Lines with Different Tumor Metastatic Potential

Abstract

Human non-small cell lung cancer (NSCLC) is one of the most common malignancies in the modern world. Its recurrence is mainly due to its ability to invade and metastasize. However, the precise mechanism for tumor development and metastasis is still not fully understood. To shed light on the development of lung cancer, the human giant cell lung carcinoma cell lines 95D with high metastatic potential and 95C with low metastatic potential were selected in this study. The 2 cell lines originated from the same parental cell and share a similar genetic background. In the current study, we identified 3 differentially expressed proteins in 95C and 95D cell lines, namely, PAI-RBP1, C1orf142, and COTL1, by using 2-dimensional electrophoresis proteomics analysis. We found that PAI-RBP1 and C1orf142 expression levels were higher in 95D than in 95C cells, whereas COTL1 expression level was lower in 95D when compared to 95C cells. We also confirmed these results by reverse transcription-polymerase chain reaction and immunoblotting analyses. The messenger RNA and protein levels of PAI-RBP1 and C1orf142 were much higher in 95D than in 95C cells, and COTL1 expression level was lower in 95D than in 95C cells. The PAI-RBP1 expression was assessed by immunohistochemistry in 70 NSCLC and 7 normal lung tissue samples from patients. PAI-RBP1 expression level was higher in tumor tissues (positive staining in 87.1% of cases [61/70]) than in normal tissues (positive staining in 14.3% of cases [1/7]). In conclusion, by studying protein expression in NSCLC cell lines with high and low metastasis as well as in human lung cancer tissues, we have identified 3 proteins, namely, PAI-RBP1, C1orf142, and COTL1, which were differentially expressed in NSCLC cell lines with different metastatic potential. In addition, we also found that PAI-RBP1 might contribute to NSCLC development.

Methylated +58CpG site decreases DCN mRNA expression and enhances TGF-β/Smad signaling in NSCLC cells with high metastatic potential

“…1A), the phenotype differences between the two cell lines are consistent with those reported previously (29,30). Thus, we examined the expression of DCN mRNA in low-(95C) and high-metastatic (95D) human NSCLC cell lines using qRT-PCR analysis, and found that DCN mRNA expression in 95D cells was lower by far than that in 95C cells (P<0.01) (Fig.…”

“…Decreased expression of DCN was found to be significantly associated with lymphatic metastasis of lung cancer patients (23). In the present study, we focused on low-(95C) and high-metastatic (95D) NSCLC cells, which were more genetically similar to each other and could be an ideal model for cancer metastasis (29,30), and found that DCN mRNA was significantly lower in 95D cells compared with that in 95C cells, supporting the idea that DCN was involved in metastasis of malignant tumors, including lung cancer. Additionally, demethylating drug 5-Aza restored DCN mRNA expression in 95D cells, suggesting the involvement of DNA methylation in DCN expression.…”

“…Low-(95C) and highmetastatic (95D) human NSCLC cell lines were purchased from Shanghai Institute of Biochemistry and Cell Biology and cultured in RPMI-1640 (Gibco-Invitrogen) with 5% fetal bovine serum (FBS, Gibco-Invitrogen) and incubated at 37˚C in 5% CO 2 . 95C and 95D cell lines share a similar genetic background and have different tumor metastatic potential (29,30). Cell treatment with DNA demethylating agent, 5-aza-2'-deoxycitidine (5-Aza) (Sigma-Aldrich), was performed according to the protocol described previously by us (25) with some modifications.…”

Retrospective Proteomic Screening of 100 Breast Cancer Tissues

“…Recently coactosin expression levels have been associated with the metastatic potential of lung cancer cell lines [135]. In our samples it showed an average expression level of 13.6x with respect to the nontumoral tissues.…”

RNF111/Arkadia is regulated by DNA methylation and affects TGF-β/Smad signaling associated invasion in NSCLC cells

“…95C and 95D cell lines share a similar genetic background [29][30]. 5-aza-2 -deoxycitidine (5-Aza) (Sigma-Aldrich, St. Louis, MO, USA) was used as demethylating agent to treat cells.…”