Objective To investigate the effect of apigenin-loaded nanoliposomes on myocardial cells apoptosis induced by diabetic cardiomyopathy. Methods The apigenin-loaded nanoliposomes were prepared by film hydration and ultrasonic dispersion technology and their quality inspections were also investigated. Sixty SD rats were randomly divided into normal control group, model control group, blank and apigenin-loaded nano-liposomes group. Rat model of typeⅠdiabetes was induced by single intraperitoneal injection of STZ. After two weeks of STZ injection, the model rats were used for this study. The rats of apigenin loaded nano-liposomes treatment group were treated with apigenin loaded nano-liposomes via caudal vein administration for 12 weeks (three times a week). And the rats of normal control group and model control group were administrated equivalent volume of 0.9% sodium chloride solution. After treatment for 12 weeks, the experimental animals were sacrificed and their hearts were harvested after 0.9% sodium chloride solution perfusion. The myocardial cell apoptosis index was detected by TUNEL staining and the expressions of Bcl-2 and Bax, which were related to the cell apoptosis, were examined by Western blotting analysis. Results TUNEL staining showed the apoptosis index in the rats of diabetes and blank nano-liposomes groups were significant higher than the normal control group (P<0.05). However, there was significant decrease of apoptosis index in the diabetes rats after the treatment of apigenin loaded nano-lipsomes (P<0.05). Moreover, Western blotting analysis confirmed that there was significant lower expression of Bcl-2 and higher expression of Bax in the hearts of model control group and blank nano-liposomes treated rats compared with normal control rats (P<0.01). Compared with the diabetes and blank nano-liposomes group, the apigenin loaded nano-liposomes treated groups showed higher expression of anti-apoptosis protein Bcl-2 (P<0.01) and lower expression of pro-apoptosis protein Bax (P<0.01). Conclusion Apigenin loaded nano-liposomes have reduced the apoptosis of myocardial cells in diabetic cardiomyopathy rats via the Bcl-2/Bax pathway.
Fig.2
Myocardial apoptosis in four groups of rats(n=15) A.normal control group;B.model control group;C.blank nanoliposome group;D.apigenin loaded nanoliposome group.Compared with normal control group, *1P<0.05;compared with apigenin-loaded nanoliposome group, *2P<0.05
Fig.3
Expression of Bcl-2 and Bax in myocardium in four groups of rats(n=15) A.normal control group;B.model control group;C.blank nanoliposome group;D.apigenin-loaded nanoliposome group.Compared with normal control group, *1P<0.01;compared with apigenin-loaded nanoliposome group, *2P<0.01
SHAWJ E,SICREER A,ZIMMETP Z.Global estimates of the prevalence of diabetes for 2010 and 2030[J].,2010,87(1):4-14.
We estimated the number of people worldwide with diabetes for the years 2010 and 2030. Studies from 91 countries were used to calculate age- and sex-specific diabetes prevalences, which were applied to national population estimates, to determine national diabetes prevalences for all 216 countries for 2010 and 2030. Studies were identified using Medline, and contact with all national and regional International Diabetes Federation offices. Studies were included if diabetes prevalence was assessed using a population-based methodology, and was based on World Health Organization or American Diabetes Association diagnostic criteria for at least three separate age-groups within the 20 79 year range. Self-report or registry data were used if blood glucose assessment was not available. The world prevalence of diabetes among adults (aged 20 79 years) will be 6.4%, affecting 285 million adults, in 2010, and will increase to 7.7%, and 439 million adults by 2030. Between 2010 and 2030, there will be a 69% increase in numbers of adults with diabetes in developing countries and a 20% increase in developed countries. These predictions, based on a larger number of studies than previous estimates, indicate a growing burden of diabetes, particularly in developing countries.
CAIL,KANGY J.Cell death and diabetic cardiomyopathy[J].,2003,3(3):219-228.
Myocardial cell death is a key element in the pathogenesis and progression of various etiological cardiomyopathies such as ischemia-reperfusion, toxic exposure, and various chronic diseases including myocardial infarction, atherosclerosis, and endothelial dysfunction. Myocardial cell death is also observed in the hearts of diabetic patients and antimal models; however, its importance in the development of diabetic cardiomyopathy is not completely understood. The goal of this review is to summarize our current understanding of the characteristics of diabetes-induced myocardial cell death. In the search of themechanisms by which diabetes induces myocardial cell death, multiple cell death pathways have been proposed. Reactive oxygen and nitrogen species accumulation plays a critical role in the cell death by antioxidants or inhibitors for apoptosis-specific signaling pathways results in a significant prevention of diabetic cardiotoxicity, suggesting that cell death in diabetic subjects plays an important role in the development of diabetic cardiomyopathy.
ZHANGJ,LIUD,HUANGY,et al.Biopharmaceutics class-ification and intestinal absorption study of apigenin[J].,2012,436(1/2):311-317.
The aim of the study was to characterize the biopharmaceutics classification system (BCS) category of apigenin (AP) using intrinsic dissolution rate (IDR) and rat intestinal permeability, and to investigate the intestinal absorption mechanism of AP in rats. In the present investigation, equilibrium solubility and intrinsic dissolution rate (IDR) of AP were estimated in phosphate buffers. Effective intestinal permeability (Peff) of AP was determined using single-pass intestinal perfusion (SPIP) technique in four segments (duodenum, jejunum, ileum and colon) of rat intestine at three concentrations (10, 50 and 100μg/ml). The aqueous solubility of AP in tested phosphate buffers was very poor with maximum solubility of 2.16μg/ml at pH 7.5. The IDR of AP was very low with a value of 0.006mg/min/cm2. The minimum and maximum Peffs determined by SPIP were 0.198×10614 and 0.713×10614cm/s at jejunum and duodenum site, respectively. In addition, the concentration-dependent permeability behavior was observed in the duodenum and jejunum, which suggested that AP was transported by both passive and active carrier-mediated saturable mechanism in these two intestinal segments. However, the observed concentration-independent permeability behavior in ileum and colon indicated primarily passive transport mechanism of absorption of AP in the last two intestinal segments. AP was classified as class II drug of the BCS due to its low solubility and high intestinal permeability. AP could be well absorbed in the whole intestine with the main absorption site at duodenum. The absorption of AP in four intestinal segments exhibited different transport mechanisms.
JORAHOLMENM W,BASNETP,ACHARYAG,et al.PEGylated liposomes for topical vaginal therapy improve delivery of interferon alpha[J].,2017,113(1):132-139.
Recent studies regarding mucosal drug delivery indicate that nanosystems with surface-available polyethylene glycol (PEG) are able to penetrate mucus barrier, assure closer contact with the epithelium, and improve drug delivery to vagina. In the present work, we developed the mucus-penetrating PEGylated liposomes containing interferon alpha-2b (IFN α-2b), destined to provide localized therapy for human papilloma virus (HPV) vaginal infections. The PEGylated liposomes were of a mean size of 18102±02802nm, bearing a negative zeta potential of – 1302mV and an entrapment efficiency of 8102±0210%. In vitro release experiments on model membrane showed a nearly non-existent IFN α-2b release from both the control and liposomally-associated IFN α-2b. However, the ex vivo penetration studies performed on the vaginal tissue obtained from pregnant sheep, showed the clear elevated IFN α-2b penetration from PEGylated liposomes as compared to the control. Furthermore, mucin studies confirmed the absence of interaction between the PEG-modified liposomes and mucin, confirming their ability to penetrate mucus and reach the deeper epithelium. The system holds a promise in improving topical delivery of IFN α-2b through enhanced efficacy of local anti-viral therapy.
JINX,YANGQ,ZHANGY.Synergistic apoptotic effects of apigenin TPGS liposomes and tyroservatide:implications for effective treatment of lung cancer[J].,2017,12(5):109-118.
To develop an alternative treatment for lung cancer, a combination of two potent chemotherapeutic agents liposomal apigenin and tyroservatide was developed. The therapeutic potential of this combination was investigated using A549 cells. Apigenin and tocopherol derivative-containing D-alpha-tocopheryl polyethylene glycol 1000 succinate (TPGS) liposomes might improve the delivery of apigenin to tumor cells, both in vitro and in vivo. Importantly, compared to either agent alone, the combination of apigenin TPGS liposomes and tyroservatide exhibited superior cytotoxicity, induced stronger G2 arrest, and suppressed A549 cancer cell invasion at a lower dose. The proapoptotic synergistic effects were also observed in A549 cells using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling, flow cytometry, and Western blot analysis. More importantly, in vivo results showed that the combination of apigenin TPGS liposomes and tyroservatide exhibited tumor-growth inhibitory effects in A549 cell-bearing mice. In conclusion, our study showed that this combination therapy could serve as a promising synergistic therapeutic approach to improve outcomes in patients with lung cancer.
ZHAOY Z,ZHANGM,WONGH L,et al.Prevent diabetic cardiomyopathy in diabetic rats by combined therapy of aFGF-loaded nanoparticles and ultrasound-targeted microbubble destruction technique[J].,2016,223(1):11-21.
Abstract Diabetic cardiomyopathy is related directly to hyperglycemia. Cell death such as apoptosis plays a critical role in cardiac pathogenesis. Whether hyperglycemia induces myocardial apoptosis, leading to diabetic cardiomyopathy, remains unclear. We tested the hypothesis that apoptotic cell death occurs in the diabetic myocardium through mitochondrial cytochrome c-mediated caspase-3 activation pathway. Diabetic mice produced by streptozotocin and H9c2 cardiac myoblast cells exposed to high levels of glucose were used. In the hearts of diabetic mice, apoptotic cell death occurred as detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay. Correspondingly, caspase-3 activation as determined by enzymatic assay and mitochondrial cytochrome c release detected by Western blotting analysis were observed. Supplementation of insulin inhibited diabetes-induced myocardial apoptosis as well as suppressed hyperglycemia. To explore whether apoptosis in diabetic hearts is related directly to hyperglycemia, we exposed cardiac myoblast H9c2 cells to high levels of glucose (22 and 33 mmol/l) in cultures. Apoptotic cell death was detected by TUNEL assay and DAPI nuclear staining. Caspase-3 activation with a concomitant mitochondrial cytochrome c release was also observed. Apoptosis or activation of caspase-3 was not observed in the cultures exposed to the same concentrations of mannitol. Inhibition of caspase-3 with a specific inhibitor, Ac-DEVD-cmk, suppressed apoptosis induced by high levels of glucose. In addition, reactive oxygen species (ROS) generation was detected in the cells exposed to high levels of glucose. These results suggest that hyperglycemia directly induces apoptotic cell death in the myocardium in vivo. Hyperglycemia-induced myocardial apoptosis is mediated, at least in part, by activation of the cytochrome c-activated caspase-3 pathway, which may be triggered by ROS derived from high levels of glucose.
ZOUM H,XIEZ.Regulation of interplay between auto-phagy and apoptosis in the diabetic heart:new role of AMPK[J].,2013,9(4):624-625.
Diabetes induces cardiomyocyte apoptosis and suppresses cardiac autophagy, indicating that the interplay between autophagy and apoptotic cell death pathways is important in the pathogenesis of diabetic cardiomyopathy. The potential mechanism, however, remains unknown. We recently reported that diabetes depresses AMP-activated protein kinase (AMPK) activity, inhibits MAPK8/JNK1-BCL2 signaling, and promotes the interaction between BECN1 and BCL2. Concomitantly, diabetes induces cardiomyocyte apoptosis and suppresses cardiac autophagy. Activation of AMPK directly phosphorylates MAPK8, which mediates BCL2 phosphorylation and subsequent BECN1-BCL2 dissociation, leading to restoration of cardiac autophagy, protection against cardiac apoptosis, and ultimately improvement in cardiac structure and function. We conclude that dissociation of BCL2 from BECN1 through activation of MAPK8-BCL2 signaling may be an important mechanism by which AMPK activation restores autophagy, protects against cardiac apoptosis, and prevents diabetic cardiomyopathy.
LEFORTE C,BLAYJ.Apigenin and its impact on gastro-intestinal cancers[J].,2013,57(1):126-144.
Apigenin (4 ,5,7-trihydroxyflavone, 5,7-dihydroxy-2-(4-hydroxyphenyl)-4H-1-benzopyran-4-one) is a flavonoid found in many fruits, vegetables, and herbs, the most abundant sources being the leafy herb parsley and dried flowers of chamomile. Present in dietary sources as a glycoside, it is cleaved in the gastrointestinal lumen to be absorbed and distributed as apigenin itself. For this reason, the epithelium of the gastrointestinal tract is exposed to higher concentrations of apigenin than tissues at other locations. This would also be true for epithelial cancers of the gastrointestinal tract. We consider the evidence for actions of apigenin that might hinder the ability of gastrointestinal cancers to progress and spread. Apigenin has been shown to inhibit cell growth, sensitize cancer cells to elimination by apoptosis, and hinder the development of blood vessels to serve the growing tumor. It also has actions that alter the relationship of the cancer cells with their microenvironment. Apigenin is able to reduce cancer cell glucose uptake, inhibit remodeling of the extracellular matrix, inhibit cell adhesion molecules that participate in cancer progression, and oppose chemokine signaling pathways that direct the course of metastasis into other locations. As such, apigenin may provide some additional benefit beyond existing drugs in slowing the emergence of metastatic disease.
JUNGU J,CHOY Y,CHOIM S.Apigenin ameliorates dyslipidemia,hepatic steatosis and insulin resistance by modulating metabolic and transcriptional profiles in the liver of high-fat diet-induced obese mice[J].,2016,8(5):E305.
Severalin vitroandin vivostudies have reported the anti-inflammatory, anti-diabetic and anti-obesity effects of the flavonoid apigenin. However, the long-term supplementary effects of low-dose apigenin on obesity are unclear. Therefore, we investigated the protective effects of apigenin against obesity and related metabolic disturbances by exploring the metabolic and transcriptional responses in high-fat diet (HFD)-induced obese mice. C57BL/6J mice were fed an HFD or apigenin (0.005%,w/w)-supplemented HFD for 16 weeks. In HFD-fed mice, apigenin lowered plasma levels of free fatty acid, total cholesterol, apolipoprotein B and hepatic dysfunction markers and ameliorated hepatic steatosis and hepatomegaly, without altering food intake and adiposity. These effects were partly attributed to upregulated expression of genes regulating fatty acid oxidation, tricarboxylic acid cycle, oxidative phosphorylation, electron transport chain and cholesterol homeostasis, downregulated expression of lipolytic and lipogenic genes and decreased activities of enzymes responsible for triglyceride and cholesterol ester synthesis in the liver. Moreover, apigenin lowered plasma levels of pro-inflammatory mediators and fasting blood glucose. The anti-hyperglycemic effect of apigenin appeared to be related to decreased insulin resistance, hyperinsulinemia and hepatic gluconeogenic enzymes activities. Thus, apigenin can ameliorate HFD-induced comorbidities via metabolic and transcriptional modulations in the liver.
HUJ,LIZ,XUL T,et al.Protective effect of apigenin on ischemia/reperfusion injury of the isolated rat heart[J].,2015,15(3):241-249.
Abstract Apigenin (Api), a mainly bioactive component of Apium graveolens L. var. dulce DC. (a traditional Chinese medicinal herb), possesses a wide range of biological activities, including antioxidant effects. It also has been shown to associate with lower prevalence of cardiovascular diseases, but its mechanisms of action remain unclear. The aim of the present study is to investigate the role of Api in isolated rat heart model of ischemia/reperfusion (I/R). Langendorff-perfused isolated rat hearts were used in our study. Api was added to the perfusate before ischemia and during reperfusion in the isolated pulsed rat heart exposed to 30-min ischemia followed by 50-min reperfusion. The treatment with Api conferred a cardioprotective effect, and the treated hearts demonstrated an improved ischemic cardiac functional recovery, a decreased myocardial infarct size, a reduced activities of creatine kinase isoenzyme and lactate dehydrogenase in the coronary flow, a reduced number of apoptotic cardiomyocytes, a reduced activity of caspase-3, up-regulation of the anti-apoptotic protein Bcl-2 and down-regulation of the pro-apoptotic protein Bax. In addition, Api inhibited the phosphorylation of p38 MAPKS during I/R. In conclusion, these observations provide preliminary evidence that Api can protect cardiomyocytes from I-/R-induced injury, at least partially, through the inhibition of p38 MAPKS signaling pathway.
ZHAOY Z,ZHANGM,TIANX Q,et al.Using basic fibroblast growth factor nanoliposome combined with ultrasound-introduced technology to early intervene the diabetic cardiomyopathy[J].,2016,11(4):675-686.
Basic fibroblast growth factor (bFGF)-loaded liposome (bFGF-lip) combined with ultrasound-targeted microbubble destruction (UTMD) technique was investigated to prevent diabetic cardiomyopathy (DCM). Cardiac function and myocardial ultrastructure were assessed. Terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) staining, immunohistochemistry staining, and Western blot assay were used to investigate the signal pathway underlying the expression of bFGF in DCM treatment. From Mason staining and TUNEL staining, bFGF-lip + UTMD group showed significant differences from the diabetes group and other groups treated with bFGF or bFGF-lip. The diabetes group showed similar results (myocardial capillary density, collagen volume fraction, and cardiac myocyte apoptosis index) to other bFGF treatment groups. Indexes from transthoracic echocardiography and hemodynamic evaluation also proved the same conclusion. These results confirmed that the abnormalities including diastolic dysfunctions, myocardial fibrosis, and metabolic disturbances could be suppressed by the different extents of twice-weekly bFGF treatments for 12 consecutive weeks (free bFGF or bFGF-lip +/- UTMD), with the strongest improvements observed in the bFGF-lip + UTMD group. The group combining bFGF-lip with UTMD demonstrated the highest level of bFGF expression among all the groups. The bFGF activated the PI3K/AKT signal pathway, causing the reduction of myocardial cell apoptosis and increase of microvascular density. This strategy using bFGF-lip and UTMD is a potential strategy in early intervention of DCM in diabetes.
XIEZ,KOYAMAT,SUZUKIJ,et al.Coronary reperfusion following ischemia:different expression of bcl-2 and bax proteins,and cardiomyocyte apoptosis[J].,2001,42(6):759-770.
The aim of this work was to examine factors that could be involved in the occurrence of apoptosis in rat hearts subjected to coronary occlusion followed by reperfusion. To this end, we studied the expression of the pro- and anti-apoptotic factors, bax and bcl-2, respectively, in reperfused ischemic hearts and in hearts injected with bFGF or saline. In anesthetized rats the left coronary artery was occluded for 45 min, the anesthesia withdrawn and the occlusion removed to allow reperfusion; in sham-operated rats the occlusion was omitted. After 4 hours the rats were decapitated and the heart excised. Sections from the left ventricle were stained with anti-bcl-2-antibody and anti-bax-antibody using the TUNEL method which detects apoptosis. Fragmentation of DNA isolated from reperfused ventricles was examined by agarose electrophoresis. In reperfused hearts no bcl-2 staining was observed in the discrete area in which many cardiomyocyte nuclei were stained by the TUNEL method; outside this area staining for bcl-2 was more marked than in sham-operated rats. Sections from reperfused hearts were stained for bax protein over a wide area including the apoptotic region; sham-operated hearts showed little reaction. Staining for bcl-2 was demonstrable in some nuclei in hearts from saline-injected rats; the numbers were unaffected by i. v. bFGF. Ischemia/reperfusion increases the overall expression of both bcl-2 and bax proteins, but bcl-2 is lost from the reperfused area as indicated by TUNEL staining. Accordingly, the ratio of bcl-2 to bax was reduced in the reperfused area, indicating a pro-apoptotic trend. The marked increase in bcl-2 outside the reperfused area could be a mechanism with which to salvage surviving cardiomyocytes.
Prevent diabetic cardiomyopathy in diabetic rats by combined therapy of aFGF-loaded nanoparticles and ultrasound-targeted microbubble destruction technique
Apigenin ameliorates dyslipidemia,hepatic steatosis and insulin resistance by modulating metabolic and transcriptional profiles in the liver of high-fat diet-induced obese mice