Abstract
Angiotensin-converting enzyme (ACE) can hydrolyze many peptides and plays a central role in controlling blood pressure. Moreover, ACE overexpression in monocytes and macrophages increases resistance of mice to tumor growth. ACE is composed of two independent catalytic domains. Here, to investigate the specific role of each domain in tumor resistance, we overexpressed either WT ACE (Tg-ACE mice) or ACE lacking N- or C-domain catalytic activity (Tg-NKO and Tg-CKO mice) in the myeloid cells of mice. Tg-ACE and Tg-NKO mice exhibited strongly suppressed growth of B16-F10 melanoma because of increased ACE expression in macrophages, whereas Tg-CKO mice resisted melanoma no better than WT animals. The effect of ACE overexpression reverted to that of the WT enzyme with an ACE inhibitor but not with an angiotensin II type 1 (AT1) receptor antagonist. ACE C-domain overexpression in macrophages drove them toward a pronounced M1 phenotype upon tumor stimulation, with increased activation of NF-κB and signal transducer and activator of transcription 1 (STAT1) and decreased STAT3 and STAT6 activation. Tumor necrosis factor α (TNFα) is important for M1 activation, and TNFα blockade reverted Tg-NKO macrophages to a WT phenotype. Increased ACE C-domain expression increased the levels of reactive oxygen species (ROS) and of the transcription factor C/EBPβ in macrophages, important stimuli for TNFα expression, and decreased expression of several M2 markers, including interleukin-4Rα. Natural ACE C-domain-specific substrates are not well-described, and we propose that the peptide(s) responsible for the striking ACE-mediated enhancement of myeloid function are substrates/products of the ACE C-domain.
Highlights
Angiotensin-converting enzyme (ACE) can hydrolyze many peptides and plays a central role in controlling blood pressure
We found that ACE overexpression by the macrophages of Tg-ACE and Tg-NKO mice strongly suppressed the growth of melanoma
Founder mice were screened for ACE overexpression in macrophages, and mice homozygous for transgene expression were made by mating heterozygotes
Summary
Mice were made transgenic for one of three different ACE constructs in which ACE cDNA is under the control of the mouse c-fms promoter (Fig. 1A). Alterations in STAT and NF-B pathways play a major role in determining the M1 versus M2 polarization of macrophages [12] To study whether these pathways are affected by ACE up-regulation and, if so, by which ACE domain, macrophages were isolated from WT, Tg-ACE, Tg-NKO, and Tg-CKO mice and conditioned for 3 days with supernatant from B16-F10 cells. To test the role of TNF␣ in ACE-mediated tumor resistance, we examined Tg-NKO macrophage activation by melanoma-conditioned medium in the presence of anti-TNF␣ antibody This significantly suppressed M1 activation of Tg-NKO macrophages, as indicated by a substantial reduction of the pro-inflammatory cytokines IL-6 and IL-12 and an increase of several anti-inflammatory markers including IL-10, CCL17, and CCL20 (Fig. 5, A and B). This may underpin the increased tumor resistance observed in Tg-ACE and Tg-NKO mice
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have