Prevention By Sulforaphane Of Diabetic Cardiomyopathy Is Most Likely Mediated By Nrf2up-Regulation And Activation

Diabetic cardiomyopathy refers to myocardial disease occur in diabetes, independent ofhypertension, coronary heart disease, valvular heart disease and other heart disease. Diabeticcardiomyopathy as one of microvascular diseases is the most common complications ofdiabetes，and also a main cause of the mortality for diabetic patients. Accumulating evidencealready indicates that to prevent the development and progression of cardiomyopathy in thepatients with diabetes could not be done by controlling glucose level or blood pressure,lowing lipid level, and blocking the renin-angiotensin system. As three main metabolicabnormalities, hyperglycemia, hyperlipidemia and inflammation all stimulate the generationof reactive oxygen or nitrogen species (ROS or RNS). Extra generation of these species iscausative of the development of diabetic complications, including cardiomyopathy.Accordingly, antioxidant prevention or therapy of diabetic complications has been attractive,but to date, there was no any antioxidant that was found to be efficiently applied in clinics.Therefore, activation of tissue’s endogenous antioxidant components has been proposed as anattractive strategy.The transcription factor Nuclear transcription factor erythroid2p45-related factor2(Nrf2) as one member of the cap“n”collar family is a master regulator of cellulardetoxification responses and redox status. Upon exposure of cells to oxidative stress orelectrophilic compounds, Nrf2translocates into the nucleus to bind to antioxidant-responsiveelements (ARE) in the genes encoding antioxidant enzymes, increasing their expression toplay a role of detoxification, antioxidant, and anti-inflammatory. Accordingly, the antioxidantdefense system is the main protection mechanism. Cells using antioxidants balancing outoxidant and electrophilic reagent. Direct antioxidant is redox active, life is short, and they inthe process of consumption；moreover indirectly antioxidants may not necessarily be redox active, but playing antioxidant effect through the raise all kinds of cell protection complex andprotein. Indirectly antioxidants have relatively long half-life, in playing antioxidation processare not consumed, and also can catalyze all kinds of chemical detoxification reaction,participate in some direct antioxidant’s regeneration.Sulforaphane (SFN) is an organosulfur compound that exhibits anticancer andantidiabetic properties in experimental models, which obtained from cruciferous vegetablessuch as broccoli. SFN as a kind of indirect antioxidant, can activate transcription factor Nrf2signal pathway, induction of antioxidant enzyme produce, which will play to toxicity,antioxidant and anti-inflammatory effect. But at present, the SFN research mainly focus onthe chemical prevention action, and about SFN and cardiovascular disease, the researchersreported less.This study was to investigate whether SFN as one of Nrf2activator can protect diabeticcardiomyopathy. For induction of the type1diabetes, mice were injected intraperitoneally withmultiple low-dose streptozocin（MLD-STZ），at50mg/kg body weight daily for5consecutivedays, while age-matched control mice were received multiple injections of the same volume ofsodium citrate buffer. Five days after the last injection of STZ, take the mouse tail venousblood monitoring non-fasting blood glucose. Blood glucose levels≥250mg/dl were defined asdiabetic. After the establishment of the type1diabetes model, hyperglycemic and age-matchedcontrol mice were treated with or without SFN at0.5mg/kg every day, in five days of eachweek for3months, and then stopped SFN injection and kept until6months. In the wholeexperiment process, monitoring group mice weight and non-fasting blood glucose change. At3and6months of diabetes, blood pressure and cardiac function were assessed. Cardiac fibrosis,inflammation, and oxidative damage were also assessed by Western blot, real-time PCR, andhistopathological methods. The results showed that from2weeks to24weeks, there’s nosignificant difference of each group body weight. After MLD-STZ injection, in with orwithout SFN injection of diabetes group, blood glucose levels increased significantly, butthere’s no significant difference of non-fasting blood glucose between the diabetes group andSFN treated diabetes group. Though small dose SFN is not significant hypoglycemic action,SFN significantly prevented diabetes-induced high blood pressure and cardiac dysfunction(Mainly displays in the interventricular septum and left ventricular posterior wall thickening increase, while cardiac ejection fraction decrease), at both3and6months. And alsosignificantly prevented diabetes induced cardiac pathological changes including cardiachypertrophy along with myocytes showing strong eosinophilic cytoplasmic staining oracidophilic degeneration. Diabetes of these pathological changes resulted in obviousmyocardial structure disorder and increased interstitial collagen fiber (Performance in Sirius-red staining collagen content increased, CTGF and TGF-β1expression enhancement). Thesepathological changes also accompanied with significant increases in cardiac oxidative damage,shown by increased3-NT and4-HNE accumulation, and inflammation (TNF-α and PAI-1expression increase), observed in diabetic hearts, were nearly completely prevented inSFN-treated diabetic hearts. SFN up-regulated Nrf2expression and function, reflected byincreased Nrf2nuclear accumulation and phosphorylation as well as mRNA and proteinexpression of Nrf2downstream antioxidants.In order to further explore the mechanism of SFN preventing diabetic cardiomyopathy,we conducted system in vitro experiment. The in vitro experiment of culture H9c2myocardialcells, high glucose cultivate myocardial cell, which induced the fibrosis and inflammationfactor expression (Performance in CTGF and PAI-1express enhancement). And through themRNA and protein level detection, SFN raised Nrf2and downstream target gene expressionand function, which fully protect the high glucose induced inflammation and fibrosis reaction.Silencing Nrf2gene with its siRNA abolished the prevention by SFN of high glucose-inducedfibrotic response.This study is the first system to research the correlation between Nrf2agonist SFN anddiabetic cardiomyopathy. These results suggest that diabetes induced cardiac dysfunction,structure derangement and remodeling, and oxidative damage can be prevented by SFN viaup-regulation of Nrf2expression and function.