Matrix Metalloproteinase-2 Proteolyzes the Sarcoplasmic Reticulum Calcium ATPase (SERCA), thus Resulting in Morphofunctional Vascular Alterations in Arterial Hypertension

Abstract

Introduction: Arterial hypertension is characterized by matrix metalloproteinase (MMP)-2 increased activity, which proteolyzes extra- and intracellular proteins, thus contributing to cellular migration and proliferation and resulting in vascular remodeling and dysfunction. MMP-2 also proteolyzes the sarcoplasmic reticulum calcium ATPase (SERCA) in myocardial ischemia and reperfusion injury, thus impairing cardiac contractile function. SERCA activity can be also reduced in the arteries of hypertensive animals. The hypothesis is hypertension-induced increased MMP-2 activity contributes to proteolyze aortic SERCA, thus resulting in vascular remodeling and dysfunction. Method: Male Sprague-Dawley rats were submitted to two kidney-one clip (2K-1C) or Sham surgery and treated with water or doxycycline (Doxy, 30 mg/kg/day). Systolic blood pressure (SBP) was assessed basally and daily from 1st to 7th day post-surgery by tail-cuff plethysmography. Then, aortas were collected for gelatin and in situ zymography, western blot to SERCA, immunofluorescence to calcium and Ki-67, morphological analysis by hematoxilin and eosin stain, vascular reactivity to phenylephrine and acetylcholine and ATPase (SERCA) activity assay. Statistical analysis was done by two-way ANOVA followed by Tukey post-test. The Ethics Committee for Animal Research of the Ribeirao Preto Medical School approved all protocols (122/2019). Results: SBP increased in 2K-1C rats vs. Sham groups (n=9; p<0.05) and Doxy did not reduce it. 2K-1C aortas presented increased MMP-2 activity (n=5-8; p<0.05) and it was reduced by Doxy (n=5-8; p<0.05). In addition, SERCA proteolysis was increased in the 2K-1C rats (n=4-8; p<0.05) and Doxy prevented it. The SERCA activity, in its turn, was reduced in the 2K-1C animals (n=5-8; p<0.05) and Doxy prevented it (n=5-8; p<0.05). Cytosolic calcium trended to increase in 2K-1C aorta (n=3; p=0.08) and Doxy decreased it (n=3; p<0.05). As calcium accumulation results in VSMC proliferation and vascular remodeling, cross sectional area (CSA), M/L ratio and the immunofluorescence of Ki-67 were increased in aortas of hypertensive rats (n=5-6; p<0.05) and Doxy decreased cell proliferation (n=5; p<0.05). Arterial contraction in response to phenylephrine was also increased in the 2K-1C rats (n=5-6; p<0.05 PD2), while relaxation in response to acetylcholine was impaired (n=4-6; p<0.05 PD2; p=0.06 Emax.), and treatment with Doxy seems to prevent this effect (n=4-6; p=0.06 Emax). Conclusions: SERCA levels and activity seem to be reduced by hypertension-induced increased MMP-2 activity in aortas, thus contributing to structural and functional vascular alterations. These effects were ameliorated in the presence of Doxy, an MMP inhibitor. CAPES, CNPq and FAPESP This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.