Avaliação dos efeitos dos peptídeos ricos em prolina do veneno da serpente Bothrops jararaca Bj-PRO-7a e Bj-PRO-10c na revascularização de membros posteriores isquêmicos

Abstract

Therapeutic angiogenesis has been proposed as an alternative for treating ischemic diseases. Angiotensin converting enzyme inhibitors (ACEi) beyond its antihypertensive effects are also able of enhacing ischemia-induced angiogenesis and improving function of angiogenic stem cells. ACEi were developed based on proline-rich oligopeptides found in the venom of Bothrops jararaca (Bj-PRO) - formerly known as bradykinin potentiating peptides (BPP). However, the antihypertensive effect of the newly described Bj-PRO-7 and -10c is not related to its ability to inhibit ACE or potentiate bradykinin. In this study, we evaluated whether Bj-PRO-7a and -10c, akin to ACEi, would affect hindlimb revascularization after ischemia in a murine model after permanent femoral artery occlusion (FAO). Using a daily treatment schedule for 14 days (started immediately after FAO) with Bj-PRO-7a and Bj-PRO-10c at 71nmol/kg (i.p.), we observed stimulation of blood flow recovery and the increase in capillary density, capillary/myocyte ratio and arteriogenesis in ischemic muscles. Interestingly, we observed that only the Bj-PRO-10c, but not the Bj-PRO-7a, increased levels of pro-angiogenic molecules such as nitric oxide (NO) and VEGF in the ischemic muscle and VEGF in bone marrow, besides has stimulated subpopulations of angiogenic stem cells in the bone marrow and their mobilization into the bloodstream. Supporting a possible role for NO as a pro-angiogenic mechanism of the peptides, the simulated limb blood flow recovery, angiogenesis and arteriogenesis in ischemic muscles of animals treated with Bj-PRO-10c, but not with Bj-PRO-7a, were partly inhibited in the presence of the nitric oxide synthase inhibitor L-NAME. Furthermore, we evaluated the role of the bradykinin B2 receptor (B2R) as another possible mechanism using transgenic animals deficient in the expression of this receptor (B2R-KO). There was no difference in the blood flow recovery, angiogenesis in ischemic muscle of both wild-type (WT) and B2R-KO animals treated with the peptides. Thus, our data suggest that both peptides Bj-PRO-7a and Bj-PRO-10c are able to stimulate revascularization of ischemic hindlimbs independently of the bradykinin B2 receptor and, in the case of Bj-PRO-10c, dependently of NO release. Furthermore, only the peptide Bj-PRO-10c is able to increase the mobilization of bone marrow-derived angiogenic stem cells after FAO.