Expression of renin–angiotensin system (RAS) components in endometrial cancer

Figures

• 

  Figure 1. Tissue renin–angiotensin system cascade. Prorenin is activated by the (pro)renin receptor ((P)RR) and possibly by proteolysis, to cleave angiotensin (Ang) I from angiotensinogen (AGT). Angiotensin-converting enzyme (ACE) then converts Ang I to the biologically active Ang II. Ang II can bind to angiotensin II type 1 receptor (AGTR1) to promote proliferation, angiogenesis, fibrosis, migration and invasion through stimulation of growth factors and intracellular signalling pathways. Furthermore, angiotensin (Ang) II binds to angiotensin II type 2 receptor (AGTR2) and antagonises AGTR1 activation. Ang I can also be further converted by angiotensin-converting enzyme 2 (ACE2) to Ang(1–7). Ang(1–7) acts upon its receptor Mas. This results in antagonism of Ang II/AGTR1 stimulation, thus inhibiting proliferation, angiogenesis, fibrosis, migration and invasion.

• 

  Figure 2. Prorenin and (P)RR mRNA and protein expression in endometrial cancer and matched adjacent non-cancerous endometrium. (A) There was no difference in REN mRNA expression between tumour and adjacent non-cancerous endometrium. (B) ATP6AP2 mRNA expression was greater in tumour (*P = 0.023) than that in adjacent endometrium. Scale bars = 500 μm. Images in the bottom panels were taken at a higher magnification, scale bars = 200 μm. Prorenin immunostaining was intense in both glandular (g) and stromal (s) tissue of both the adjacent non-cancerous endometrium (C) and tumour (E and G). ATP6AP2, although intensely expressed in tumour glandular epithelium (F and H), was localised to the perivascular space (closed head arrow) and endothelium (open head arrow) in the adjacent normal endometrium (E). Immunostaining was not seen in the negative controls containing no primary antibody (inserts, tumour tissues).

• 

  Figure 3. CTSD mRNA expression in tumour and matched adjacent non-cancerous endometrium. CTSD tended to be higher in tumour tissue when compared with matched adjacent non-cancerous endometrium (*P = 0.058). CTSD has the potential to activate prorenin in a low pH milieu (12).

• 

  Figure 4. Expression of RAS mRNA levels in matched tumour and adjacent non-cancerous endometrium. AGT mRNA (A) was similar between tumour and adjacent non-cancerous endometria. ACE1 (B), ACE2 (C) and AGTR1 (D) mRNAs were all increased in tumour tissue compared with adjacent non-cancerous endometria (*P = 0.004, 0.046 and 0.008, respectively).

• 

  Figure 5. Localisation of RAS protein levels in matched tumour and adjacent non-cancerous endometrium. Scale bars = 500 μm. Images in the right hand panel were taken at a higher magnification, scale bars = 200 μm. Immunostaining of AGT, ACE1, ACE2, AGTR1, AGTR2 and MASR was present in the stroma (s) of both tumour and adjacent non-cancerous endometria and was localised to the perivascular space (closed head arrow) and endothelium (open head arrow). AGT, ACE1, ACE2, AGTR1, AGTR2 and MASR staining was most intense in the glandular epithelium (g) of the tumour. Immunostaining was not seen in the negative controls containing no primary antibody (inserts, tumour tissues).

• 

  Figure 6. Expression of downstream targets of the RAS in matched tumour and adjacent non-cancerous endometrium. (A) VEGFA and (D) SERPINE1 mRNA levels were the same in tumour and matched adjacent non-cancerous endometrium. (B) PIK3R1 tended to be higher in tumour tissue (P = 0.053). (C) TGFB1 mRNA was significantly increased in tumour tissue when compared with matched adjacent non-cancerous endometrium (*P = 0.001).

• 

  Figure 7. Tissue renin–angiotensin systems in endometrial cancer. In tumour tissue, upregulation of cathepsin D and (P)RR allows for greater activation of prorenin, potentially leading to increased enzymatic cleavage of Ang I from AGT. Additionally, increased ACE1 abundance may lead to increased production of the biologically active Ang II. Together, upregulation of the pro-angiogenic/pro-proliferative components may lead to increased activation of both (P)RR- and AGTR1-mediated intracellular signalling cascades to stimulate the production of TGFB1 and PI3KR1 and promote tumourigenesis. The role of increased ACE2 and presence of MAS1 and AGTR2 may enhance tumour development by stimulating vascularisation; however, this is unknown.