The Protective Effect Of Chlorogenic Acid On Experimental Abdominal Aortic Aneurysms Via Nrf2/HO-1

Objective:With the aging of the population,the events of age-related cardiovascular and cerebrovascular diseases are increasing year by year,threatening the health of middle-aged and elderly people.As a member of the vascular diseases,the incidence of Abdominal Aortic Aneurysm（AAA）is also on the rise.Moreover,because of its hidden signs,it is easy to be ignored.Once obvious signs appear,there is a greater risk of rupture and a high risk of death after rupture.Therefore,early detection and timely treatment are the key factors to save patients with abdominal aortic aneurysm.At present,surgical treatment is generally adopted for aneurysm with a diameter of more than 5.5cm,which mainly includes traditional open vascular replacement and the newly developed endovascular repair for aneurysms in recent years.However,for patients with small aneurysm diameter and not up to the surgical indications,treatment strategies such as controlling blood pressure,making a few small healthy lifestyle changes and regular follow-up are usually adopted.This strategy based solely on"watch and wait"has insufficient benefits for patients,and it is of great clinical significance whether innovative and reasonable drug therapy can be sought to delay the expansion of the aneurysm and provide greater benefits to patients during the follow-up period.In recent years,Chinese medicine has been vigorously developed,and the concept of"medicine and food with the same origin"has gradually entered people’s life circle.Chlorogenic acid（CGA）,as a phenolic acid compound existing in many kinds of fruits and vegetables,tea and Chinese herbal medicine,has obvious anti-inflammatory,antioxidant,anti-tumor and cardiovascular protective effects.Currently,it has been widely studied in a variety of diseases,but there has not been any research related to the treatment of abdominal aortic aneurysm.Therefore,in this study,we investigated whether chlorogenic acid can enhance the antioxidant capacity of the body by up-regulating the expression of Nrf2/HO-1,and then realize the protective effect on experimental abdominal aortic aneurysm.Methods:This study was conducted in vivo and in vitro respectively.First,healthy male SD rats were selected as the research object for the in vivo experiment,and the model of abdominal aortic aneurysm was constructed by the way of Calcium chloride（Ca Cl₂）infiltration,and the treatment group was respectively given intraperitoneal injection of chlorogenic acid（25mg/Kg,50mg/Kg and 100mg/Kg）.The contents of hydrogen peroxide（H₂O₂）,malondialdehyde（MDA）and reduced glutathione（GSH）in serum of each group were determined by related kits.Histological staining was used to observe the rupture of elastic fibers in the middle artery of the aneurysm.The expression of matrix metalloproteinases（MMP-2,MMP-9）and Nrf2 in each group was detected by immunohistochemical method.Western blotting was used to detect the protein levels of MMP-2,MMP-9,SM22α,Nrf2,NQO1 and HO-1 in aneurysm tissues of each group.Vascular smooth muscle cells（VSMC）were treated with H₂O₂（200μM）for 4H to establish an oxidative stress model.The effects of chlorogenic acid（25μM,50μM,and100μM）on the reduction of Reactive oxygen species（ROS）production and activation of Nrf2 pathway were investigated.CCK8 reagent and flow cytometry were used to detect the effects of H₂O₂and CGA treatment on VSMC cell viability.The oxidative stress level of VSMC was detected by flow cytometry and ROS detection kit.The expressions of NQO1,HO-1 and Nrf2 were detected by Western blot.The nuclear translocation of Nrf2was detected by immunofluorescence method.Results:To a certain extent,chlorogenic acid delayed the progression of experimental abdominal aortic aneurysm in rats,slowed the expansion of the aneurysm,and dose-dependent reduction of oxidative stress in experimental abdominal aortic aneurysm rats.At the same time,the rupture and dissolution of elastic fibers in aneurysm tissue were significantly decreased in the chlorogenic acid treatment group,the expression of matrix metalloproteinases（MMP-2,MMP-9）was significantly decreased,and the expression of antioxidant related proteins Nrf2,NQO1,HO-1 and contractile phenotypic marker SM22αwere increased.Similarly,similar results were obtained in vitro experiments.Chlorogenic acid reduced the oxidative stress level of VSMC induced by H₂O₂.Western blotting showed that the expression trend of anti-oxidation-related proteins NQO1 and HO-1 was consistent with that of Nrf2.Immunofluorescence results further confirmed that chlorogenic acid pretreatment promoted the expression of Nrf2and western blot showed that Nrf2 tended to transpose from cytoplasm to nucleus.Conclusions:Chlorogenic acid can eliminate excess reactive oxygen species from the body and decrease expression of matrix metalloproteinases（MMPs）to reduced the dissolution and rupture of elastic fibers in aneurysm tissue.In addition,it can maintain the normal contractile phenotype of vascular smooth muscle.Therefore,it has certain protective effect on the experimental rat model of abdominal aorta induced by calcium chloride infiltration.Chlorogenic acid activated the expression of Nrf2 and nuclear translocation in a dose-dependent manner,and then increased the expression of anti-oxidation-related proteins NQO1 and HO-1.Therefore,chlorogenic acid may play a protective role on experimental abdominal aortic aneurysms to some extent by activating Nrf2/HO-1 expression.