Washed with PBS, and resuspended in 500 l of PBS with 0.2 Triton X-100, 10 mM EDTA, 100 g/ml RNase A, and 50 g/ml propidium iodide (PI) at room temperature for 30 min.Statistical analysisFisher Exact test were used to assess the correlation between RNPC1 and clinicopathologic parameters. For all the tests p purchase SNDX-275 values < 0.05 was considered statistically significant.ResultsRNPC1 was lower expressed in human breast cancer cellsThe data were analyzed using the SPSS 12.0 software (SPSS, Chicago, IL, USA). All experiments in this study were repeated in triplicate, unless otherwise specified. Student t-test was used to analyze the statistical significance of the differences between groups. 2 test andRNPC1 expression in five breast cancer cell lines and two breast epithelial cell lines were quantified by qRTPCR and Western blot (Figure 1A, p < 0.05). Among the seven cell lines analyzed, RNPC1 was found lower expression in breast cancer cells compared to normal mammary breast epithelial MCF-10A and 184A1 cells. Among breast cancer cells, MCF-7, BT474, ZR-75 cells expressed relatively higher levels of RNPC1, and low expression or barely detectable levels were found inFigure 1 RNPC1 expressive in breast cancer cell lines and tissues. (A) qRT-PCR and Western blot analysis of RNPC1 expression in breast cancer cell lines and normal breast cell lines MCF-10A and 184A1. The two normal breast cell lines showed higher expression of RNPC1 than other cell lines (p < 0.05). The fold change of RNPC1a is shown below each lane. The intensity of the bands was determined using densitometric analysis. (B) RNPC1 mRNA expression in 121 pairs of breast cancer and adjacent tissue. 26 cases showed for example. (C) Average expression level of RNPC1 mRNA in 121 pairs of human breast cancer tissues and adjacent normal breast tissues. Adjacent breast tissues had higher expression of RNPC1, where the breast cancer tissues showed the lower level of expression (p < 0.01). (D) RNPC1 protein expression in 121 pairs of breast cancer and adjacent tissues. The 25 kDa band/GAPDH ratio was markedly lower in tumors compared to adjacent normal tissues. 12 cases showed for example. The fold change of RNPC1a is shown below each lane. The intensity of the bands was determined using densitometric analysis. (E) A scatter plot of RNPC1 protein expression in the same cancer PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/25447644 tissue, adjacent tissue (p < 0.05). Data were means of two separate experiments mean ?SEM, *p < 0.05, **p < 0.01.Xue et al. BMC Cancer 2014, 14:322 http://www.biomedcentral.com/1471-2407/14/Page 5 ofTable 1 The association between RNPC1 mRNA expression and clinicopathologic features of breast cancerClinicopathologic parameters Age (years) 55 >55 Tumor Size (cm) 2 >2 TNM stage I II + III Lymph node metastasis 3 >3 Grade I II III unclear ER status Negative Positive unclear PR status Negative Positive Her2 status Negative Positive suspect Ki67 15 >15 CK5/6 Negative Positive P53 Negative Positive 42 52 26 42 16 10 0.042 46 15 35 11 11 4 0.830 43 52 30 40 13 12 0.430 81 32 8 56 19 7 25 13 1 0.298 58 63 42 40 16 23 0.294 40 70 11 22 51 9 18 19 2 0.097 7 67 30 17 2 50 18 12 5 17 12 5 0.066 96 25 69 13 27 12 0.058 24 97 11 71 13 26 0.010 38 83 24 58 14 25 0.463 68 53 42 40 26 13 0.109 Number of case RNPC1 Low expression RNPC1 High expression p-valueTable 1 The association between RNPC1 mRNA expression and clinicopathologic features of breast cancer (Continued)PCNA Negative + ++ +++ Histology ductal special 106 15 71 11 35 4 0.6.