Levo-Tetrahydropalmatine Attenuates Progression of Abdominal Aortic Aneurysm in an Elastase Perfusion Rat Model via Suppression of Matrix Metalloproteinase and Monocyte Chemotactic Protein-1.

BACKGROUND Levo-tetrahydropalmatine (L-THP) is a tetrahydro protoberberine isoquinoline alkaloid obtained from the genera Stephania and Corydalis. In the present research, we evaluated the effects of L-THP on the progression of aortic aneurysms (AAs) in experimental rats induced with perfusion of elastase. MATERIAL AND METHODS Thirty-six Sprague-Dawley rats were divided into sham-operated, control, and L-THP treated groups (n=12 in each group). The rats in the control group and the L-THP group received intra-aortic perfusion of elastase to induce AAs; the sham-operated group received perfusion of saline. The rats in the L-THP group received a dose of 15 mg/kg/day, the control and the sham group received saline treatment. The animals were evaluated for aortic diameters (ADs) and systolic blood pressure (SBP) just before and after the elastase perfusion, and 24 days after perfusion. The extracts of the aortas were evaluated by western blotting and immunohistochemistry. RESULTS In the control group, a significant increase in aortic size was observed (p<0.05) compared to the sham group after 24 days post-perfusion, whereas the L-THP group showed a decrease in diameter compared to the control group (p<0.05). The SBP increased significantly in the control group compared to the sham group. The L-THP group showed reduction in SBP, exhibited decreased expression of metalloproteinase and monocyte chemotactic protein-1, and the tissue samples also exhibited significant decreased levels of iNOS compared to the control group. L-THP treatment prevented loss of vascular smooth muscle cells (VSMCs) of the aortic walls. CONCLUSIONS L-THP inhibited progression of AAs in rats by curbing inflammation, oxidative stress, and conserving VSMCs, suggesting a new therapeutic approach for managing AAs.

Role of myeloperoxidase in abdominal aortic aneurysm formation: mitigation by taurine.

Oxidative stress plays a fundamental role in abdominal aortic aneurysm (AAA) formation. Activated polymorphonuclear leukocytes (or neutrophils) are associated with AAA and express myeloperoxidase (MPO), which promotes inflammation, matrix degradation, and other pathological features of AAA, including enhanced oxidative stress through generation of reactive oxygen species. Both plasma and aortic MPO levels are elevated in patients with AAA, but the role of MPO in AAA pathogenesis has, heretofore, never been investigated. Here, we show that MPO gene deletion attenuates AAA formation in two animal models: ANG II infusion in apolipoprotein E-deficient mice and elastase perfusion in C57BL/6 mice. Oral administration of taurine [1% or 4% (wt/vol) in drinking water], an amino acid known to react rapidly with MPO-generated oxidants like hypochlorous acid, also prevented AAA formation in the ANG II and elastase models as well as the CaCl2 application model of AAA formation while reducing aortic peroxidase activity and aortic protein-bound dityrosine levels, an oxidative cross link formed by MPO. Both MPO gene deletion and taurine supplementation blunted aortic macrophage accumulation, elastin fragmentation, and matrix metalloproteinase activation, key features of AAA pathogenesis. Moreover, MPO gene deletion and taurine administration significantly attenuated the induction of serum amyloid A, which promotes ANG II-induced AAAs. These data implicate MPO in AAA pathogenesis and suggest that studies exploring whether taurine can serve as a potential therapeutic for the prevention or treatment of AAA in patients merit consideration.NEW & NOTEWORTHY Neutrophils are abundant in abdominal aortic aneurysm (AAA), and myeloperoxidase (MPO), prominently expressed in neutrophils, is associated with AAA in humans. This study demonstrates that MPO gene deletion or supplementation with the natural product taurine, which can scavenge MPO-generated oxidants, can prevent AAA formation, suggesting an attractive potential therapeutic strategy for AAA.

Increased galectin-3 levels are associated with abdominal aortic aneurysm progression and inhibition of galectin-3 decreases elastase-induced AAA development.

Abdominal aortic aneurysm (AAA) evolution is unpredictable and no specific treatment exists for AAA, except surgery to prevent aortic rupture. Galectin-3 has been previously associated with CVD, but its potential role in AAA has not been addressed. Galectin-3 levels were increased in the plasma of AAA patients (n=225) compared with the control group (n=100). In addition, galectin-3 concentrations were associated with the need for surgical repair, independently of potential confounding factors. Galectin-3 mRNA and protein expression were increased in human AAA samples compared with healthy aortas. Experimental AAA in mice was induced via aortic elastase perfusion. Mice were treated intravenously with the galectin-3 inhibitor modified citrus pectin (MCP, 10 mg/kg, every other day) or saline. Similar to humans, galectin-3 serum and aortic mRNA levels were also increased in elastase-induced AAA mice compared with control mice. Mice treated with MCP showed decreased aortic dilation, as well as elastin degradation, vascular smooth muscle cell (VSMC) loss, and macrophage content at day 14 postelastase perfusion compared with control mice. The underlying mechanism(s) of the protective effect of MCP was associated with a decrease in galectin-3 and cytokine (mainly CCL5) mRNA and protein expression. Interestingly, galectin-3 induced CCL5 expression by a mechanism involving STAT3 activation in VSMC. Accordingly, MCP treatment decreased STAT3 phosphorylation in elastase-induced AAA. In conclusion, increased galectin-3 levels are associated with AAA progression, while galectin-3 inhibition decreased experimental AAA development. Our data suggest the potential role of galectin-3 as a therapeutic target in AAA.

Dipeptidyl peptidase-4 inhibitor decreases abdominal aortic aneurysm formation through GLP-1-dependent monocytic activity in mice.

Abdominal aortic aneurysm (AAA) is a life-threatening situation affecting almost 10% of elders. There has been no effective medication for AAA other than surgical intervention. Dipeptidyl peptidase-4 (DPP-4) inhibitors have been shown to have a protective effect on cardiovascular disease. Whether DPP-4 inhibitors may be beneficial in the treatment of AAA is unclear. We investigated the effects of DPP-4 inhibitor sitagliptin on the angiotensin II (Ang II)-infused AAA formation in apoE-deficient (apoE-/-) mice. Mice with induced AAA were treated with placebo or 2.5, 5 or 10 mg/kg/day sitagliptin. Ang II-infused apoE-/- mice exhibited a 55.6% incidence of AAA formation, but treatment with sitagliptin decreased AAA formation. Specifically, administered sitagliptin in Ang II-infused mice exhibited decreased expansion of the suprarenal aorta, reduced elastin lamina degradation of the aorta, and diminished vascular inflammation by macrophage infiltration. Treatment with sitagliptin decreased gelatinolytic activity and apoptotic cells in aorta tissues. Sitaglipitn, additionally, was associated with increased levels of plasma active glucagon-like peptide-1 (GLP-1). In vitro studies, GLP-1 decreased reactive oxygen species (ROS) production, cell migration, and MMP-2 as well as MMP-9 activity in Ang II-stimulated monocytic cells. The results conclude that oral administration of sitagliptin can prevent abdominal aortic aneurysm formation in Ang II-infused apoE-/-mice, at least in part, by increasing of GLP-1 activity, decreasing MMP-2 and MMP-9 production from macrophage infiltration. The results indicate that sitagliptin may have therapeutic potential in preventing the development of AAA.

Effect of hyperbaric oxygen treatment in a rat model of abdominal aortic aneurysm.

Matrix metalloproteinase (MMP) plays a pivotal role in the pathophysiology of abdominal aortic aneurysms (AAA). Experimental studies have demonstrated that hyperbaric oxygen (HBO2) therapy decreases MMP levels in different tissues. However, the effect of HBO2 therapy on AAA has yet to be investigated. This study aimed to examine the effects of HBO2 on MMPs in an experimental AAA model. The model was implemented with CaCl2 in 12-week-old male Wistar albino rats. The rats were randomized into four groups: Group I: received NaCl (n = 6) (Sham group); Group II: received NaCl and were treated with HBO2 (n = 6); Group III: received CaCl2 (n = 6); and Group IV: received CaCl2 and were treated with HBO2 (n = 6). HBO2 therapy was applied for five of seven days over a period of six weeks. Although in the CaCl2 groups, aortic diameters were significantly higher than the NaCl groups (p < 0.05), there was no difference between pre- and post-HBO2 in the CaCl2 groups (p > 0.05). In the CaCl2 group, the MMP-2 and MMP-9 levels were significantly higher than those in the NaCl group (p < 0.05). HBO2 therapy had no statistically significant effect on the MMP-2 and MMP-9 levels in Groups III and IV. However, it was observed that both levels clearly decreased in Group IV. In conclusion, the study suggested that HBO2 may have favorable effects on MMP levels.

Orally administered dipeptidyl peptidase-4 inhibitor (alogliptin) prevents abdominal aortic aneurysm formation through an antioxidant effect in rats.

OBJECTIVE: Dipeptidyl peptidase-4 (DPP-4) inhibitor, a novel antidiabetic drug, has a cardioprotective effect on ischemia-reperfusion injury through an antioxidant effect. However, the effect of DPP-4 inhibitor on aneurysm formation has not been investigated. We aimed to test the hypothesis that the DPP-4 inhibitor, alogliptin, attenuates vascular oxidative stress and thus inhibits abdominal aortic aneurysm (AAA) formation. METHODS: AAAs were created with intraluminal elastase and extraluminal calcium chloride in 36 male rats. Rats were divided into three groups: a low dose of alogliptin group (group LD; 1 mg/kg/d), a high-dose group (group HD; 3 mg/kg/d), and a control group (group C, water). Alogliptin was administered by gastric gavage once daily beginning 3 days before surgery. On day 7 after aneurysm preparation, reactive oxygen species (ROS) expression was semiquantified by dihydroethidium staining, and the oxidation product of DNA produced by ROS, 8-hydroxydeoxyguanosine (8-OHdG), was measured by immunohistochemical staining. Blood glucose concentrations were measured. Hematoxylin and eosin and elastica Van Gieson stainings were performed on day 28, and the AAA dilatation ratio was calculated. RESULTS: On day 7 (six in each group), dihydroethidium staining of the aneurysm wall showed a reduced level of ROS expression (4.6 ± 0.6 in group C, 2.7 ± 0.3 in group LD, and 1.7 ± 0.5 in group HD; P < .0001) and showed fewer 8-OHdG-positive cells in alogliptin-treated samples (138.1 ± 7.4 cells in group C, 102.5 ± 4.5 cells in group LD, and 66.1 ± 4.5 cells in group HD; P < .0001) The treatment significantly reduced messenger RNA expression of matrix metalloproteinases (MMPs) in aneurysm walls (relative expression: MMP-2: 2.1 ± 0.4 in group C, 1.3 ± 0.3 in group LD, and 0.9 ± 0.2 in group HD; P < .001; MMP-9: 2.0 ± 0.5 in group C, 0.3 ± 0.3 in group LD, and 0.3 ± 0.2 in group HD; P < .001). On day 28 (six in each group), the aortic wall in groups LD and HD was less dilated (dilatation ratio: 199.2% ± 11.8% in group C, 159.6% ± 2.8% in group LD, and 147.1% ± 1.9% in group HD; P < .02 group C vs HD) and had higher elastin content than in group C. The difference in blood glucose levels among the three groups was not significant. CONCLUSIONS: The DPP-4 inhibitor, alogliptin, attenuates aneurysm formation and expansion dose-dependently in a rat AAA model via an antioxidative action.

Doxycycline therapy for abdominal aneurysm: Improved proteolytic balance through reduced neutrophil content.

BACKGROUND: Matrix metalloproteinase-9 (MMP-9) is thought to play a central role in abdominal aortic aneurysm (AAA) initiation. Doxycycline, a tetracycline analogue, has direct MMP-9-inhibiting properties in vitro, and it effectively suppresses AAA development in rodents. Observed inhibition of AAA progression, and contradictory findings in human studies evaluating the effect of doxycycline therapy on aortic wall MMP-9, suggest that the effects of doxycycline extend beyond MMP-9 inhibition and that the effect may be dose-dependent. METHODS: This clinical trial evaluated the effect of 2 weeks of low- (50 mg/d), medium- (100 mg/d), or high-dose (300 mg/d) doxycycline vs no medication in four groups of 15 patients undergoing elective AAA repair. The effect of doxycycline treatment on MMP and cysteine proteases, and their respective inhibitors, was evaluated by quantitative polymerase chain reaction, Western blot analysis, immunocapture protease activity assays, and immunohistochemistry. RESULTS: Doxycycline was well tolerated and no participants dropped out. Doxycycline treatment reduced aortic wall MMP-3 and MMP-25 messenger RNA expression (P < .045 and P < .014, respectively), selectively suppressed neutrophil collagenase and gelatinase (MMP-8 and MMP-9) protein levels (P < .013 and <.004, respectively), and increased protein levels of the protease inhibitors tissue inhibitor of metalloproteinase 1 and cystatin C (P < .029). As for the apparent selective effect on neutrophil-associated proteases, we sought for a reducing effect on aortic wall neutrophil content that was indeed confirmed by immunohistochemical analysis that revealed a 75% reduction in aneurysm wall neutrophil content (P < .001). CONCLUSIONS: Independent of its dose, short-term preoperative doxycycline therapy improves the proteolytic balance in AAA, presumably through an effect on aortic wall neutrophil content. This study provides a rationale for doxycycline treatment in patients with an AAA as well as in other (vascular) conditions involving neutrophil influx such as Kawasaki disease and Behçet disease.

A randomized, placebo-controlled trial of doxycycline after endoluminal aneurysm repair.

BACKGROUND: The late durability of endovascular aneurysm repair (EVAR) has been limited by progressive aortic degeneration believed to be mediated by matrix metalloproteases (MMP). The goal of this study was to evaluate the effect of a MMP inhibitor, doxycycline, on EVAR. METHODS: Patients undergoing EVAR were randomized to doxycycline (100 mg twice daily) or placebo for 6 months following the procedure. Clinical data, blood samples, and computed tomography (CT) scans were obtained preoperatively, postoperatively (blood only), and at 1- and 6-month follow-up. Forty-four subjects were analyzed based on intention-to-treat. RESULTS: Plasma MMP-9 decreased significantly below baseline in the doxycycline (N = 20) treated patients at 6 months (-16.4% +/- 20.7%, P < .05) while there was a nonsignificant increase in the placebo (N = 24) group (128.1% +/- 73.5%). This was primarily related to changes between 1 and 6 months. In patients with endoleaks at 6 months, plasma MMP-9 increased in 83% of the placebo treated patients, but in only 14% of the doxycycline treated group (P < .03). Among endoleak-free patients with AneuRx or Excluder endografts, doxycycline treatment resulted in greater decreases in maximum aortic diameter than placebo treatment (-13.3% +/- 3.3% vs -3.8% +/- 3.0%, P < .05). Furthermore, doxycycline treatment significantly reduced the aortic neck dilatation at 6 months in Excluder treated patients. CONCLUSION: There is evidence of persistent MMP release representing ongoing aortic degradation after endografting which can be inhibited by doxycycline therapy. In analyses based on the endograft used, treatment with doxycycline also demonstrated evidence of increased aortic dimensional stability, a surrogate marker for long-term success of EVAR. Although encouraging, these results require confirmation in larger patient populations. Doxycycline should undergo more thorough evaluation as a potential adjuvant treatment to improve the results of EVAR, particularly in certain subgroups.

Preoperative treatment with doxycycline reduces aortic wall expression and activation of matrix metalloproteinases in patients with abdominal aortic aneurysms.

PURPOSE: Matrix metalloproteinases (MMPs) are considered to play a central role in the pathogenesis of abdominal aortic aneurysms (AAAs). Doxycycline (Dox) has direct MMP-inhibiting properties in vitro, and it effectively suppresses the development of elastase-induced AAAs in rodents. The purpose of this study was to determine if treatment with Dox suppresses MMPs within human aneurysm tissue and to elucidate the molecular mechanisms underlying this effect. METHODS: Aneurysm tissues were obtained from 15 patients with an AAA, eight of whom had been treated with Dox before surgery (100 mg orally twice a day for 7 days). Protein extracts were examined by means of gelatin zymography and immunoblot analysis, and RNA was examined by means of reverse transcription-polymerase chain reaction (RT-PCR). The effects of Dox on MMP production were further examined in human THP-1 mononuclear phagocytes in vitro. RESULTS: No detectable difference was found between groups by using substrate zymography as a means of assessing total MMP activity, but Dox treatment was associated with a slight (24.4%) reduction in the activated fraction of 72-kDa gelatinase (MMP-2; P <.05). In contrast, a 2.5-fold reduction in the amount of extractable 92-kDa gelatinase (MMP-9) protein in Dox-treated patients was revealed by means of immunoblot analysis (P <.05). Also, a 5.5-fold (81.9%) reduction in MMP-9 messenger RNA (mRNA) in Dox-treated patients was demonstrated by means of quantitative competitive RT-PCR (mean +/- SE, mol MMP-9/mol beta-actin: 1.3 +/- 0.5 vs 7.2 +/- 3.1; P <.04). There was no significant difference between groups in the relative expression of MMP-2 protein or mRNA. In cultured THP-1 monocytes stimulated with phorbol ester, the expression of MMP-9 protein and mRNA were both decreased after exposure to relevant concentrations of Dox in vitro. CONCLUSION: In addition to its recognized effects as a direct MMP antagonist, Dox may influence connective tissue degradation within human aneurysm tissue by reducing monocyte/macrophage expression of MMP-9 mRNA and by suppressing the post-translational processing (activation) of proMMP-2. Through this complementary combination of mechanisms, treatment with Dox may be a particularly effective strategy for achieving MMP inhibition in patients with an AAA.