Abstract
DNA damage response (DDR) pathways have been extensively studied in cancer cell lines and mouse models, but little is known about how DNA damage is recognized by different cell types in nonmalignant, slowly replicating human tissues. Here, we assess, using ex vivo cultures of human prostate tissue, DDR caused by cytotoxic drugs (camptothecin, doxorubicin, etoposide, and cisplatin) and ionizing radiation (IR) in the context of normal tissue architecture. Using specific markers for basal and luminal epithelial cells, we determine and quantify cell compartment-specific damage recognition. IR, doxorubicin, and etoposide induced the phosphorylation of H2A.X on Ser(139) (γH2AX) and DNA damage foci formation. Surprisingly, luminal epithelial cells lack the prominent γH2AX response after IR when compared with basal cells, although ATM phosphorylation on Ser(1981) and 53BP1 foci were clearly detectable in both cell types. The attenuated γH2AX response seems to result from low levels of total H2A.X in the luminal cells. Marked increase in p53, a downstream target of the activated ATM pathway, was detected only in response to camptothecin and doxorubicin. These findings emphasize the diversity of pathways activated by DNA damage in slowly replicating tissues and reveal an unexpected deviation in the prostate luminal compartment that may be relevant in prostate tumorigenesis. Detailed mapping of tissue and cell type differences in DDR will provide an outlook of relevant responses to therapeutic strategies.
Highlights
DNA damage response (DDR) pathways protect genomic integrity and form a barrier against cancer [1]
Expression of p63 and cytokeratins 5 and 14 was detectable in the basal cell compartment, p27 and androgen receptor (AR) were expressed in the luminal cell compartment, and α-smooth muscle actin (α-SMA) was expressed by stromal cells at comparable levels to standard paraffin-embedded prostate sections, AR levels somewhat declined after a week (Fig. 1C; Supplementary Fig. S2A)
We present here an alternative method to assay how different cell types respond to DNA damage in ex vivo cultured human prostate tissues
Summary
DNA damage response (DDR) pathways protect genomic integrity and form a barrier against cancer [1]. The nature and amount of DNA damage, tissue-specific factors, Authors' Affiliations: 1Molecular Cancer Biology Program, Biomedicum Helsinki and Department of Virology, Haartman Institute and 2Molecular Imaging Unit and Computational Systems Biology Laboratory, Institute of Biomedicine and Genome-Scale Biology Program, University of Helsinki; 3Department of Virology and 4Department of Pathology and Haartman Institute, University of Helsinki Hospital Laboratory Diagnostics; 5Department of Clinical Chemistry, University of Helsinki and Helsinki University Central Hospital; 6Department of Urology, Helsinki University Central Hospital, Helsinki, Finland; 7The Sidney Kimmel Comprehensive Cancer, Johns Hopkins University School of Medicine, Baltimore, Maryland; and 8Department of Pathology and Oncology, Karolinska Institutet, Stockholm, Sweden. Phone: 410-502-9748; Fax: 443-502-2821; E-mail: mlaiho1@ jhmi.edu
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