Common and distinct mechanisms of different redox‐active carcinogens involved in the transformation of mouse JB6P+ cells

Abstract

We transformed JB6P+ cells with prolonged intermittent low-dose UVB radiation or prolonged exposure to low-dose H2O2 or CdCl2. Stable transformation was confirmed by an anchorage-independence assay. The JB6P+ transformants formed more colonies (∼six folds) in soft agar as compared to their JB6P+ parent cells and were associated with increased intracellular reactive oxygen species (ROS) levels. Activating protein-1 (AP-1) is a family of transcription factors that are rapidly activated by elevated intracellular ROS levels, and their composition is important in the process of cellular transformation and/or tumor progression. To investigate if carcinogenesis induced by distinct carcinogens was via similar molecular mechanisms in these transformants, gel mobility shift and immunoblot analyses were utilized to determine the distinct AP-1 compositions. Compared to parent JB6P+ cells, the gain of JunB and Fra-1 in AP-1 DNA binding complexes was markedly increased in all transformed cells, which might contribute to a more proliferative phenotype, while loss of Fra-2 occurred in JB6P+/H2O2 and JB6P+/Cd cells. Differential AP-1 components in the transformants suggested that their transformations might be mediated by distinct transcription signalings with distinct AP-1 dimer compositions. However, all three transformants exhibited increased activation of pathways involved in cell proliferation (ERK/Fra-1/AP-1 and JNK/c-jun/AP-1) and anti-apoptosis (Bcl-xl). The development of the JB6P+ transformants (JB6P+/UVB; JB6P+/H2O2; JB6P+/Cd) provides a unique tool to study the mechanisms that contribute to different redox-active carcinogens in a single model.

Topically Applied Carvedilol Attenuates Solar Ultraviolet Radiation Induced Skin Carcinogenesis

“…Anchorage-independent growth assay of JB6 P þ cells promoted by EGF or UV EGF-mediated transformation of JB6 P þ cells was conducted in 96-well plates as previously described (29). Low-dose UV irradiation was used to transform the JB6 P þ cells following a modified method (30). In brief, radiant doses of 15 mJ/cm 2 UV was applied to the cells in PBS three times per week for a total of 8 weeks.…”

“…As 4-OHC absorbs UV irradiation, it is expected that 4-OHC will have some degree of UV protection, while carvedilol is hypothesized to provide greater preventive activity against UVinduced effects due to its pharmacological activity. This hypothesis was first tested in the JB6 P þ cell transformation assay using UV as the tumor promoter because low and re-occurring dosages of UV irradiation can lead to malignant transformation of JB6 P þ cells (30). Treating JB6 P þ cells with 15 mJ/cm 2 UV three times a week for 8 weeks then plating the cells in agar with growth media without any promoter or drugs resulted in more colonies than EGF as the promoter (48 AE 4 vs. 35 AE 4, P ¼ 0.002922, respectively; data expressed as median AE median absolute deviation (MAD)).…”

Fos-related antigen-1 (Fra-1) is a regulator of glioma cell malignant phenotype

“…37 The preferential association of Fra-1 and Jun B was shown to be related with malignant transformation of JB6 epithelial cells, with gain of JunB and Fra-1 binding complexes in all transformed cells, which might contribute to a more proliferative phenotype. 38 The heterodimer Fra-1-JunB is able to regulate the expression of multiple genes including the interleukin 2 (IL-2), 39 and cyclin A, 40 which in turn promote neoplastic cell growth and proliferation.…”

Retinol induces morphological alterations and proliferative focus formation through free radical-mediated activation of multiple signaling pathways

“…Although the involvement of MAPKs in cell division control and transformation is well recognized, as mentioned earlier, different members of the MAPK family and related kinases may regulate different path- Many of the cell cycle regulatory effects promoted by different members of the MAPK family have also been observed to be evoked or strongly influenced by free radicals and related species. ERK was demonstrated to be involved in the free radical-mediated promotion of transformation or in the increase to transformation susceptibility [40,41] . The p38 MAPK was observed to be a key contributor for the redox signaling involved in UVA-mediated transformation [42] and was postulated to exert a selective role in promoting free radicalmediated tumorigenesis [43] .…”