Author ORCID Identifier

https://orcid.org/0000-0002-8878-3841

Year of Publication

2019

Degree Name

Doctor of Philosophy (PhD)

Document Type

Doctoral Dissertation

College

Medicine

Department

Pharmacology and Nutritional Sciences

First Advisor

Dr. Xianglin Shi

Abstract

Hexavalent chromium, Cr(VI), is an established human carcinogen that is a worldwide environmental health concern. It is well understood that reactive oxygen species, genomic instability, and DNA damage repair deficiency are important contributors to Cr(VI)-induced carcinogenesis. After decades of research some cancer hallmarks remain understudied for the mechanism of Cr(VI) carcinogenesis. Dysregulated cellular energetics have been established as a hallmark of cancer. Energy pathways that become dysregulated in cancer include mitochondrial respiration, lipogenesis, pentose phosphate pathway, one carbon metabolism, and increased anaerobic glycolysis in the presence of oxygen or ‘Warburg effect’.

To investigate metabolism changes in Cr(VI) carcinogenesis, we exposed human lung epithelial cells (BEAS-2B cells) to Cr(VI) for six months and isolated a colony from soft agar. To confirm the results in the BEAS-2B cells, we used two other sets of Cr(VI)-transformed cells, human lung epithelial cells (BEP2D cells) and human lung fibroblasts (WTHBF-6 cells).

We found increased lipogenesis related protein expressions including: ATP citrate lyase (ACLY), acetyl-CoA carboxylase 1 (ACC1), and fatty acid synthase (FASN) in Cr(VI)-transformed cells as compared to passage-matched control cells. We also observed increased palmitic acid levels, confirming that Cr(VI)-transformed cells were making more lipids. Cr(VI)-transformed BEAS-2B cells had decreased colony formation in soft agar and decreased cell growth when treated with a FASN inhibitor (C75). ACLY, ACC1, and FASN protein expressions were also increased in chromate-induced lung tumors in human tissue samples.

We also observed that Cr(VI)-transformed human lung cells (BEAS-2B, BEP2D, and WTHBF-6 cells) had no major changes in their mitochondrial respiration as measured by the Seahorse Analyzer when compared to their passage-matched control cells. Conversely, xenograft tumor-derived cells had mitochondrial respiratory dysfunction.

Interestingly, we also found that Cr(VI)-transformed human lung cells (BEAS-2B, BEP2D, and WTHBF-6 cells) had no major changes in their glycolytic function as measured by the Seahorse Analyzer when compared to their passage-matched control cells. Similarly, these cells did not have changes in glycolytic enzymes or extracellular L-lactate levels. Moreover, xenograft tumor-derived cells showed no changes in glycolytic endpoints or L-lactate levels. This indicates these cells did not undergo the ‘Warburg effect’.

These data demonstrate that increased lipogenesis is important to Cr(VI)-induced lung carcinogenesis and are consistent with the cancer literature which reports that increased lipogenesis proteins occur during carcinogenesis. Additionally, our results indicate mitochondrial respiratory dysfunction is likely a result of the tumor microenvironment and a later step during Cr(VI) carcinogenesis. Lastly, we observed the ‘Warburg effect’ is not required for Cr(VI)-induced carcinogenesis in vitro. However, it remains to be shown if the ‘Warburg effect’ is still a consequence or contributing factor for tumorigenesis. Future studies are needed to investigate other metabolic pathways in Cr(VI)-induced carcinogenesis. In conclusion, some metabolism pathways are important to Cr(VI)-induced carcinogenesis, while others appear not to be.

Digital Object Identifier (DOI)

https://doi.org/10.13023/etd.2019.039

Funding Information

This research was supported by National Institutes of Health [(R01ES021771, R01ES020870, R01ES025515, R01ES28984, R01ES21771, and P30ES026529) and the Redox Metabolism Shared Resource Facility of the University of Kentucky Markey Cancer Center (P30CA177558).

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