Abstract 176: IL6 and CXCL1 induce senescent phenotype of cancer-associated fibroblast via autocrine loops in oral squamous cell carcinoma
Authors: Kim Eun Kyoung , Moon Sook , Kim Do Kyeong , Kim Jin , Bae Jung Yoon 
Published: May 2015

Background & Objective

Tumor-stroma interaction plays a key role in tumor development and progression. One of the prominent components of tumor stroma is cancer-associated fibroblasts (CAFs). Although CAFs have been well known to contribute to tumorigenesis, the characteristics of CAFs are still poorly understood. For this reason, we characterized CAFs and investigated a mechanism underlying the transformation of NOFs into CAFs in oral squamous cell carcinoma (OSCC).

Materials & Methods

For this study, three primary cultured NOFs and three primary cultured CAFs were used, respectively. Expression of proliferating cell nuclear antigen (PCNA) and senescence-associated β-galactosidase (SA-β-Gal) enzyme activity were compared between NOFs and CAFs. Population doubling levels and expression of activated fibroblast marker were measured in NOFs and CAFs. The expression levels of senescence-related markers were detected by reverse transcription polymerase chain reaction (RT-PCR) and western blotting. In search of mechanism that triggers senescence in CAFs, cytokine antibody array was employed in NOFs co-cultured with OSCC cells, and the results were confirmed by real-time PCR.

Results

In comparison between NOFs and CAFs, α-SMA, a marker of activated fibroblasts, showed no difference in expression pattern. CAFs showed higher SA-β-Gal enzyme activity and lower PCNA expression than NOFs at the same passage. In addition, CAFs exhibited lower population doubling level than NOFs, indicating that CAFs had senescent phenotype. NOFs co-cultured with OSCC cells showed higher SA-β-Gal enzyme activity, p16 and p21 expression compared with mono-cultured NOFs, whereas NOFs co-cultured with human epidermal keratinocytes (HEK) showed no SA-β-Gal enzyme activity, indicating that the induction of senescence in CAFs was not merely an artifact of co-culture system but was triggered specifically by the co-cultured cancer cells. Cytokine antibody array revealed that co-culture conditions induced cytokine secretion from CAFs. In particular, IL6 and CXCL1 showed the highest secretion level, and mRNA expression levels corresponded with the results from cytokine antibody array. Upon treating NOFs with IL6 and CXCL1, higher SA-β-Gal enzyme activity was detected in NOFs compared with non-treated NOFs, indicating that IL6 and CXCL1 were capable of inducing senescence in NOFs.

Conclusion

From these results, we propose that the senescent phenotype of CAFs might be elicited by cytokines such as IL6 and CXCL1, which are secreted from CAFs in an autocrine manner. Additional studies are in progress to identify specific factors to induce cytokine secretion in a carcinoma milieu.

Acknowledgments

This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (2009-0094027).

Citation Format: Eun Kyoung Kim, Sook Moon, Do Kyeong Kim, Jin Kim, Jung Yoon Bae. IL6 and CXCL1 induce senescent phenotype of cancer-associated fibroblast via autocrine loops in oral squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 176. doi:10.1158/1538-7445.AM2014-176