Opposing Effects on Modulation of Angiogenesis by Protein Kinase C and cAMP-Mediated Pathways

Abstract

The role of cAMP-mediated pathway in modulating angiogenesis was investigated. We have shown previously that activators of protein kinase C (PKC) caused a marked increase in angiogenesis, while the specific inhibitor of PKC, Ro 318220 suppressed angiogenesis. Here we show that forskolin, which activates adenylate cyclase and elevates the intracellular levels of cAMP, and the Sp-diastereomer of adenosine cyclic-3’,5’-monophosphothioate (Sp-cAMPS), caused a dose-dependent suppression of collagenous protein biosynthesis and angiogenesis in the chick chorioallantoic membrane system (CAM). The opposite modulation of angiogenesis by activators of PKC and elevated cAMP levels was further confirmed by the suppression of 4β-phorbol-12-myristate-13-acetate (4β-PMA)-stimulated angiogenesis by either forskolin or Sp-cAMPS. On the contrary, the Rp-diastereomer of adenosine cyclic-3’,5’-monophospho-thioate (Rp-cAMPS), which antagonises endogenous cAMP biochemical actions, had no effect on angiogenesis alone and did not suppress 4β-PMA stimulated angiogenesis. However, Rp-cAMPS antagonised the effect of forskolin and Sp-cAMPS on 4β-PMA induced angiogenesis. Similar results were obtained in the human umbilical vein endothelial cell tube formation assay. In this system, the PKC inhibitor, Ro 318220, caused a dose-dependent inhibition and 4β-PMA reversed this effect. Also, forskolin and Sp-cAMPS caused an inhibition in tube formation. These results indicate that increased levels of intracellular cAMP have a negative effect in normal angiogenesis and cause a large reduction of the promotion of angiogenesis resulting from PKC activation.

Vascular Abnormalities in Mice Deficient for the G Protein-Coupled Receptor GPR4 That Functions as a pH Sensor

“…This agrees with previous results showing that cAMP can attenuate angiogenesis in other systems (15,25,53). Collectively, the results from the aortic ring assay and wound healing assay indicate that GPR4 can serve as a sensor for blood vessels to detect pH changes, and this pH-sensing function may be related to vascular growth and other processes.…”

“…In the aortic ring assay, microvessel outgrowth from GPR4-null aortas was less inhibited by low pH. In the cell wound healing assay, acidic pH delayed the HUVEC wound closure and overexpression of GPR4 synergistically inhibited this process, consistent with the production of cAMP and its inhibitory effects on angiogenesis (15,25,53). Our results suggest that GPR4 functions as an acidic pH sensor for vascular cells, which may provide new insights into the pH-sensing mechanisms of blood vessels.…”

“…Previous cell line overexpression studies demonstrate that acidic pH activates GPR4 to induce the production of cyclic AMP (cAMP) (34,44). In this respect, cAMP and its downstream kinase protein kinase A have been shown to inhibit angiogenesis (11,15,24,25,53).…”

Protein kinase C and downstream signaling pathways in a three-dimensional model of phorbol ester-induced angiogenesis

“…By using multiple PKC inhibitors, these findings reveal the importance of PKC for vessel assembly in this threedimensional assay. Moreover, these results are in agreement with previously published studies demonstrating that PKC inhibition prevents angiogenesis in vitro [32] and in vivo [33,34].…”

Chorioallantoic membrane capillary bed: A useful target for studying angiogenesis and anti‐angiogenesis in vivo

“…However, the cellular mechanisms involved are not well defined. Activation of protein kinase c (PKC) was reported to induce CAM angiogenesis between days 9 and 11 (Tsopanoglou et al, 1993), whereas increased levels of intracellular cyclic adenosine monophosphate substantially downregulated this PKC-mediated angiogenic stimulus (Tsopanoglou et al, 1994).…”