BACKGROUND: We have recently identified cancer-stem cell (CSC) subpopulations in metastatic head and neck cutaneous squamous cell carcinoma (mHNcSCC). This study investigated the expression of components of the renin-angiotensin system (RAS): pro-renin receptor (PRR), angiotensin converting enzyme (ACE), angiotensin II receptor 1 (ATIIR1) and angiotensin II receptor 2 (ATIIR2) in relation to these CSC subpopulations in mHNcSCC.
METHODS: 3’3 Diaminobenzidine (DAB) immunohistochemical (IHC) staining was performed on mHNcSCC tissue samples from 15 patients to investigate the expression of the components of the RAS: PRR, ACE, ATIIR1 and ATIIR2. Immunofluorescence (IF) IHC dual staining of three of the mHNcSCC tissue samples for these components of the RAS, with embryonic stem cell markers c-MYC or SOX2, was performed to localise these proteins to the CSCs. Western blotting (WB) and RT-qPCR were performed on six mHNcSCC tissue samples and three mHNcSCC-derived primary cell lines, to confirm protein and mRNA expression of the components of the RAS, respectively.
RESULTS: DAB IHC staining demonstrated expression of PRR, ACE, ATIIR1 and ATIIR2 in all 15 mHNcSCC tissue samples. IF IHC staining demonstrated expression of PRR by the c-MYC+ cells within the tumor nests (TNs) and the peritumuoral stroma (PTS). ACE was localised to SOX2+ cells on the endothelium of the microvessels. ATIIR1 and ATIIR2 were present on c-MYC+ cells within the TNs and the PTS. WB confirmed protein expression of PRR, ACE and ATIIR1 in the mHNcSCC tissue samples. RT-qPCR confirmed transcript expression of PRR, ACE and ATIIR1 in mHNcSCC tissue samples and mHNcSCC-derived primary cell lines.
CONCLUSION: Cancer stem cell subpopulations within the TNs, PTS and endothelium of mHNcSCC express components of the RAS. These primitive cells may be a novel therapeutic target by manipulation of the RAS, using commonly available medications that inhibit the RAS.