Downregulation of P38 phosphorylation correlates with low-grade differentiation and proliferation of lung squamous cell carcinoma

Abstract

Background: P38MAPK has been investigated as a tumor-related signaling molecule because of its apparent association with tumorigenesis. This study aimed to investigate P38MAPK expression and its role in lung squamous carcinoma (LSCC).

Methods: The expression of P38MAPK and phosphorylated P38 (P-P38) in LSCC tissues and cells was examined by Western blot, real-time PCR, and immunohistochemistry. The influence of P38MAPK inhibitor SB203580 on the proliferation of LSCC cells was detected by MTT and flow cytometry.

Results: The expression of P-P38 in LSCC tissues and cells was lower than that in cancer-adjacent normal tissues and normal bronchial epithelial cells (P<0.05). In addition, the expression of P-P38 was downregulated in LSCC tissues of poor differentiation, stages III and IV, and with lymph node metastasis compared with the LSCC tissues of well differentiation, stages I and II, and without lymph node metastasis (P<0.05). Moreover, the cell proliferation of LSCC SK-MES-1 cells treated by P38MAPK inhibitor SB203580 significantly increased in a concentration-dependent manner compared with that of SK-MES-1 cells without SB203580 (P<0.05). The inhibition of P38MAPK promoted the transition of the S phase to the G2 phase.

Conclusions: P-P38 was poorly expressed in LSCC tissues and cells. Its low expression was correlated with low-grade differentiation, lymph node metastasis, and advanced stage of LSCC. Inhibition of P38MAPK expression could significantly increase the proliferation of LSCC cells by promoting the transition of the S phase to the G2 phase.

Keywords: LSCC; Lung squamous carcinoma; P-P38; P38MAPK; proliferation.

Figure 1

Expression levels of P38MAPK and P-P38 in LSCC tissues and CANT. A. The amplification product size of P38MAPK gene was 190 bp, the expression of P38MAPK mRNA did not show obvious difference between in LSCC and in CANT (P>0.05); B. The expression of P-P38 in LSCC tissues was significantly lower than that in CANT (P<0.05); C. The expression of P38MAPK mRNA in LSCC tissues was not different from CANT (P>0.05); D. There was no significant difference in the expression of P38MAPK between LSCC tissues and CANT (P>0.05); E. Obviously, the expression of P-P38 in LSCC was lower than that in CANT (P<0.05); LSCC, lung squamous cell carcinoma; CANT, cancer-adjacent normal tissues.

Figure 2

Correlation between the expressions of P38MAPK and P-P38 and clinicopathological features of LSCC. A. Western Blot showed that the expression of P-P38 in well differentiated LSCC cells was significantly higher than that in moderately and poorly differentiated LSCC (P<0.05); B. Western Blot showed that the expression level of P-P38 protein in LSCC tissues of stage I-II was significantly higher than that in stage III-IV (P<0.05); C. IHC analysis showed that P-P38 protein expression in LSCC tissues displayed a significant down-regulation compared with the CANT (P<0.05); D. IHC analysis showed that well-differentiated LSCC tissues showed a higher expression than in moderately and poorly differentiated LSCC (P<0.05); E. IHC analysis showed that increased P-P38 protein was observed in LSCC of stage I-II compared with stage III-IV (P<0.05); F. IHC analysis showed LSCC tissues with lymph node metastasis had a lower expression of P-P38, as compared with those without lymph node metastasis (P<0.05); LSCC, lung squamous cell carcinoma; CANT, cancer-adjacent normal tissues; IHC, immunohistochemistry.

Figure 3

The IHC staining of P38MAPK and P-P38 in LSCC tissues and CANT (IHC×400). A. Low staining of P-P38 in normal lung tissues; B. Low staining of P38MAPK in well differentiated LSCC; C. High staining of P38MAPK in poorly differentiated LSCC; D. Low staining of P-P38 in poorly differentiated LSCC; E. Moderate staining of P-P38 in moderately differentiated LSCC; F. High staining of P-P38 in well differentiated LSCC; LSCC, lung squamous cell carcinoma.

Figure 4

Expression of P38MAPK in LSCC cell line and its role in cell cycle regulation and proliferation of LSCC cells. A, B. The expression of P-P38 in LSCC SK-MES-1 cells was significantly lower than that in normal human bronchial epithelial BEAS-2B cells (P<0.05); C, D. Compared with SK-MES-1 cells without containing P38 specific inhibitor SB203580, the proliferation ability of K-MES-1 cells treated by SB203580 was promoted, and this proliferation capacity was positively correlated with SB203580 concentration (P<0.05). E, F. The down-regulation of P38 protein promoted the transition of S phase to G2 phase of LSCC SK-MES-1 cells, which meant the down-regulation of P38 protein speeded up the proliferation of LSCC SK-MES-1 cells (P<0.05).