Doxycycline ameliorates ischemic and border-zone remodeling and endothelial dysfunction after myocardial infarction in rats.

BACKGROUND: Although matrix metalloproteinase (MMP) activity increases, endothelial function decreases after myocardial infarction (MI). The antibiotic doxycycline inhibits MMP activity in vitro. The role of doxycycline-mediated MMP inhibition in endothelial function is unclear. HYPOTHESIS: Doxycycline ameliorates endothelial dysfunction, in part, by inhibiting MMP activity. METHODS: We subjected Sprague-Dawley male rats to MI by ligating the left anterior descending arteries. We subjected another group of rats to sham surgery. We administered doxycycline in drinking water (0.67 mg/ml) to both groups 2 days before surgery: the sham group underwent sham surgery and received doxycycline therapy, and the MI group underwent MI and received doxycycline therapy (n = 6 in each group). After 4 weeks, we anesthetized rats and prepared left ventricular rings from infarcted-ischemic (I), non-infarcted near-infarcted (NI), and sham surgery hearts with and without doxycycline treatment. RESULTS: The MMP-2 activity increased significantly in I and NI hearts, and we observed a selective increase in MMP-9 activity only in I hearts, when compared with other groups (p < 0.05), measured by zymography. Cardiac inhibitor of metalloproteinase decreased only in I hearts (p < 0.05 vs other groups), measured by Western analysis, and doxycycline treatment reversed this decrease. Contractile response of rings to acetylcholine was attenuated in the I group, suggesting nitric oxide-mediated dysfunction, and was reversed by doxycycline. The response to nitroprusside was attenuated in I hearts and ameliorated by doxycycline, suggesting cardiomyocyte dysfunction. Bradykinin induced relaxation in rings from sham surgery hearts and from NI hearts, but induced paradoxic contraction in rings from I hearts. Treatment with doxycycline reversed the paradoxic contraction. CONCLUSION: Results suggest a protective action of doxycycline in the ischemic heart, possibly because of additional pharmacologic actions such as metalloproteinase inhibition.

Peroxisome proliferator ameliorates endocardial endothelial and muscarinic dysfunction in spontaneously hypertensive rats.

Spontaneously hypertensive rats (SHR) develop hypertension (HT) at the age of 2-6 weeks. Endocardial endothelial (EE) dysfunction, autonomic suppression, left ventricle hypertrophy (LVH), and fibrosis are hallmarks of HT. The mechanism of EE dysfunction, LVH, and fibrosis in SHR is largely unknown. It is known, however, that the levels of peroxisome proliferator-activated receptor gamma (PPARgamma) are negatively correlated with EE function, LVH, and HT. PPARgamma ameliorates EE dysfunction and LVH, in part, by increasing endothelial nitric oxide (eNO) and muscarinic activity. Male SHR and normotensive Wistar rats (NWR) at 1 week of age were administered 8 micro g/ml ciglitazone (CZ), a PPARgamma agonist, in drinking water. The rats were grouped as follows: NWR, NWR+CZ, SHR, SHR+CZ, at 2 and 6 weeks (n = 6 in each group). The levels of PPARgamma were low in the nuclear extracts of the left ventricle (LV) in SHR, but increased in CZ-treated rats, measured by western analysis. The contractile response to norepinephrine in cardiac rings prepared from the above groups of rats, measured in tissue myobath and normalized by tissue weight, demonstrated no difference in the maximum response to norepinephrine in any group. However, the EC(50) was significantly lower in SHR at 2 weeks (SHR2wk) than in any other groups, and CZ normalized this decrease. The response to acetylcholine demonstrated no difference in EC(50); however, the maximum response was attenuated in SHR2wk, and substantially increased in SHR6wk as compared with age-matched NWR, suggesting that early in HT eNO dysfunction in SHR2wk leads to depression of autonomic muscarinic cholinergic receptor in SHR6wk. The PPARgamma agonist ameliorated both the early eNO dysfunction and late autonomic suppression in HT. The response to nitroprusside demonstrated no change in EC(50); however, the maximum response was attenuated only in SHR6wk. There were significant fibrosis, LVH, and increased LV pressure in SHR6wk compared with any other group, and CZ regressed LVH and LV pressure. Results suggest that early in HT, except for eNO dysfunction, other contractile responses are preserved; however, at 6 weeks there is significant nonendothelial cell dysfunction, and treatment with CZ reverses this nonendothelial dysfunction as well.

Mechanism of matrix accumulation and glomerulosclerosis in spontaneously hypertensive rats.

BACKGROUND: Renal interstitial fibrosis and thickening of the glomerular basement membrane are associated with hypertension. However, the mechanism of matrix accumulation is unclear. Spontaneously hypertensive rats (SHR) develop hypertension at between 2 and 6 weeks of age. METHODS: To test the hypothesis that increased matrix metalloproteinase (MMP) and tissue inhibitor of metalloproteinase (TIMP) contribute to the pathomechanisms of hypertensive nephropathy, the cortex and medulla of male SHR at 2 and 6 weeks were analyzed for MMP-2, MMP-7, and MMP-9 by gelatin and elastin gel zymography. The levels of TIMP-4 were measured by western blot analysis. The bands in blots were scanned and normalized with actin. To localize MMP-2 and TIMP-4 in situ, immuno-labeling was performed. To determine proteinuresis, urinary protein was measured by Bio-Rad dye binding assay. The mean arterial pressure (mmHg) was measured in Inactin-anesthetized rats by a PE-50 catheter in the femoral artery. Age-sex matched normotensive Wistar rats (NWR) were used as controls and grouped: (1). SHR, 2 weeks; (2). SHR, 6 weeks; (3). NWR, 2 weeks; and (4). NWR, 6 weeks (n = 6 in each group). RESULTS: Levels of cortex MMP-2 and MMP-9 were increased in 6 week SHR as compared with NWR. In the medulla, MMP-9 and MMP-7 were increased, but there was no change in MMP-2. The levels of cortex TIMP-4 tended to increase but insignificantly. In contrast, there were significant increases in the levels of TIMP-4 in the medulla of 6 week SHR as compared with 2 week SHR or NWR. In addition, there were substantial elastinolytic activity in the cortex of 6 week SHR. The in situ labeling suggested no TIMP-4 in the glomeruli. There was substantial TIMP-4 in the epithelial layer of tubules. The levels of fibrotic collagen were significantly higher in both the glomeruli and tubular interstitium. Urinary protein excretion was increased significantly in 6 week SHR when compared with other groups. The mean arterial pressure was 1.6-fold higher in 6 week SHR than in controls. CONCLUSION: These results suggest that increased MMP-2 and MMP-9 activity contributes to glomerular injury and hypertensive remodeling. The increased levels of TIMP-4 in the medulla may inhibit the collagenolytic activity of MMP but is unable to inhibit the elastinolytic activity. An important role of MMP-2, MMP-9, and TIMP-4 in hypertensive remodeling of the cortex and medulla in the SHR is demonstrated.