Xianglin Shi

A critical role of PI-3K/Akt/JNKs pathway in benzo[a]pyrene diol-epoxide (B[a]PDE)-induced AP-1 transactivation in mouse epidermal Cl41 cells

Mouse skin tumorigenicity studies indicate that benzo[a]pyrene-7,8-diol-9,10-epoxide (B[a]PDE) contributes to carcinogenesis as both a tumor initiator and promoter. However, the mechanisms that mediate B[a]PDE tumor promotion effects remain unclear. Our results demonstrated that in mouse epidermal Cl41 cells, B[a]PDE treatment resulted in marked activation of AP-1 and its upstream MAPKs, including ERKs, JNKs and p38K. B[a]PDE exposure also led to activation of phosphotidylinositol 3-kinase (PI-3K), Akt and p70 S6 kinase (p70S6k). B[a]PDE-induced AP-1 transactivation was inhibited by pretreatment of cells with PI-3K inhibitors, wortmannin or Ly294002. In contrast, inhibition of p70S6k with rapamycin did not show any inhibitory effects. An overexpression of dominant-negative mutant of PI-3K, Δp85, impaired B[a]PDE-induced activation of PI-3K, Akt and AP-1 transactivation. Furthermore, an overexpression of dominant-negative Akt mutant, Akt-T308A/S473A, blocked B[a]PDE-induced activation of Akt, AP-1 and JNKs, while it did not affect the activation of p70S6k, ERKs and p38 kinase. These results demonstrated that B[a]PDE was able to induce AP-1 transactivation and this AP-1 induction was specific through PI-3K/Akt/JNKs-dependent and p70S6k-independent pathways. This study also indicated that Akt-T308A/S473A blocks B[a]PDE-induced AP-1 activation specific through impairing JNK pathway. These findings will help us to understand the signal transduction pathways involved in the carcinogenic effects of B[a]PDE.

Downregulation of cyclin D1-CDK4 protein in human embryonic lung fibroblasts (HELF) induced by silica is mediated through the ERK and JNK pathway.

Silica is a factor in the induction of acute injury and chronic pulmonary fibrosis. In 1996, silica was also listed as a human carcinogen by the International Agency for Research on Cancer (IARC). However, the molecular mechanisms involved in its pathologic effects are not well understood. We found that exposure of human embryonic lung fibroblasts (HELF) to crystalline silica for 2 h decreased cyclin D1 and cyclin‐dependent kinase 4 (CDK4) expression levels. Extracellular signal‐regulated protein kinase (ERKs), c‐Jun NH2‐terminal amino kinase (JNKs), and p38 kinase, as well as their downstream transcription factor, AP‐1, had different effects on the regulation of expression levels of cyclin D1 and CDK4 alterations induced by silica.

Different Patterns of Cyclin D1/CDK4-E2F-1/4 Pathways in Human Embryo Lung Fibroblasts Treated by Benzo[a]pyrene at Different Doses

OBJECTIVE To investigate the roles of the cyclin D1/CDK4 and E2F-1/4 pathways and compare their work patterns in cell cycle changes induced by different doses of B[a]P. METHODS Human embryo lung fibroblasts (HELFs) were treated with 2 micromol/L or 100 micromol/L B[a]P which were provided with some characteristics of transformed cells (T-HELFs). Cyclin D1, CDK4 and E2F-1/4 expressions were determined by Western blotting. Flow cytometry was used to detect the distribution of cell cycle. RESULTS After B[a]P treatment, the proportion of the first gap (G1) phase cells decreased. CDK4 and E2F-4 expression did not change significantly. In 2 micromol/L treated cells, a marked overexpression of cyclin D1 and E2F-1 was observed. However, in T-HELFs overexpression was limited to cyclin D1 only, and no overexpression of E2F-1 was observed. The decreases of G1 phase in response to B[a]P treatment were blocked in antisense cyclin D1 and antisense CDK4 transfected HELFs (A-D1 and A-K4) and T-HELFs (T-A-D1 and T-A-K4). After 2 micromol/L B[a]P treatment, overexpression of E2F-1 was attenuated in A-D1, and E2F-4 expression was decreased significantly in A-K4. In T-A-D1 and T-A-K4, E2F-4 expression was increased significantly, compared with T-HELFs. The E2F-1 expression remained unchanged in T-A-D1 and T-A-K4. CONCLUSIONS Cyclin D1/CDK4-E2F-1/4 pathways work in different patterns in response to low dose and high dose B[a]P treatment. In HELFs treated with 2 micromol/L B[a]P, cyclin D1 positively regulates the E2F-1 expression while CDK4 negatively regulates the E2F-4 expression; however, in HELFs treated with 100 micromol/L B[a]P, both cyclin D1 and CDK4 negatively regulate the E2F-4 expression.