| Background:adriamycin(ADR)belongs to anthracycline broad-spectrum anticancer drugs,and is one of the most widely used chemotherapeutic drugs with remarkable curative effect.However,ADR can cause cumulative dose-dependent cardiotoxicity and further develop into irreversible myocardial injury,which seriously limits its clinical application.At present,the drugs used to reduce the myocardial injury caused by ADR and other chemotherapeutics in clinical practice mainly include mesterodium,dexradezosin and amphostine etc,but the mechanism of action is unclear,the price is expensive and the safety is low.In addition,many chemical drugs,such as histamine 2 receptor antagonists,can also alleviate the cardiotoxicity of ADR to a certain extent,but the side effects are more obvious and interfere with the chemotherapeutic efficacy of ADR to varying degrees.Therefore,the development of new drugs that can reduce the toxic and side effects of ADR without affecting its anticancer activity has become a focus of clinical research and an urgent problem to be solved.Psoralidin(PSO)is a natural coumarin extracted from the seeds of Psoralea corylifolia L.Previous studies have shown that PSO has anti-inflammatory,antibacterial,antiviral,anti-osteoporosis and other pharmacological activities.Our previous study found that Bakuchiol(BAK),another active component of Psoralea corylifolia L,can significantly reduce ADR-induced myocardial cell injury,suggesting that PSO may have a similar protective effect on ADR-induced myocardial injury.Silent Information Regulator 1(SIRT1)is a nicotinamide adenine dinucleotide(NAD+)-depend ent histone deacetylase.Studies have found that a variety of natural drugs,such as BAK and curcumin,can play a protective role in the heart and other organs by activating SIRT1-related signaling pathways.However,whether SIRT1 is involved in the process of PSO anti-ADR myocardial injury has not been studied up to now.Peroxisome proliferator-activated receptor-γ(PPAR-γ)is a member of the nuclear hormone receptor superfamily and has been reported that it plays a key role in cardiovascular diseases such as hypertension,atherosclerosis and other cardiovascular diseases.SIRT1 may play an important role in lipid metabolism and liver metabolism by regulating PPAR-γ-associated signaling pathways.However,it is not clear whether SIRT1 also plays a role by regulating PPAR-γ signaling pathway in myocardial injury of ADR.Therefore,we will explore whether PSO has a protective effect on ADR myocardial injury in vivo and in vitro through a series of experimental methods,and confirm whether PSO plays a role by regulating the SIRT1/PPAR-γ signaling pathway.Research Methods:1.Animal experiments and related tests: BALB/c mice as the research object,using intraperitoneal injection of ADR method to simulate the myocardial injury,model of chemotherapy drugs were divided into Control group、ADR group(10 mg/kg)、PSO low,medium and high dose(12.5 mg/kg、25 mg/kg、50 mg/kg)+ADR group;Routine blood detector,automatic blood biochemical analyzer,small animal ultrasound detector,histopathological staining,Western blot and other instruments and techniques were used to detect and compare the related indicators of each group,so as to study and clarify the effect of PSO on myocardial injury of ADR in mice.2.Cell experiments and related testing: myocardial cells of HL-1 as the research object,explore ADR damage model,set up the Control group,the ADR group,the different concentration of PSO(10,20,50,100 μM)and PSO(20 μM)+ADR group,using Muse cells analyzer,fluorescence microscope,Western blot,and other instruments and methods for testing and comparison between groups related indicators.The si RNA of SIRT1 was synthesized and PPAR-γ blocker GW9662 was used to further investigate the effect of PSO on ADR induced-myocardial injury,and to prove whether PSO plays an anti-ADR myocardial injury role by regulating the SIRT1/PPAR-γ signaling pathway.Research Results:1.Animal experiment:(1)Compared with ADR(10 mg/kg)group,survival rate of mice in PSO(12.5 mg/kg,25mg/kg,50 mg/kg)group was significantly increased(P<0.05);the survival rate of PSO(25mg/kg)increased most significantly,which was used as the best experimental concentration in the follow-up;(2)Compared with ADR(10 mg/kg)group,PSO(25 mg/kg)group significantly increased body weight(P<0.05);(3)Compared with ADR(10 mg/kg)group,the number of white blood cells(WBC),monocytes(MON)and neutrophils(GRA)in PSO(25 mg/kg)group was significantly increased compared with ADR(10 mg/kg)group(P<0.05);(4)Compared with ADR(10 mg/kg)group,the blood biochemical related indexes:the content of lactate dehydrogenase(LDH)、creatine kinase(CK)、glutamic oxalacetic transaminase(AST)、 albumin(ALB)、blood urea nitrogen(BUN)was significantly decreased(P<0.05);(5)Compared with ADR(10 mg/kg)group,cardiac function related indexes: stroke volume(SV)and cardiac output(CO)were significantly increased(P<0.05);(6)Compared with ADR(10 mg/kg)group,myocardial morphology was improved(HE staining)and fibrosis degree was reduced(Masson and Sirius red staining);(7)Compared with ADR(10 mg/kg)group,oxidative stress level of myocardial tissue was significantly decreased(DHE staining).The above data indicated that PSO could significantly reduce the myocardial injury of ADR in mice;(8)Compared with ADR(10 mg/kg)group,the expression levels of SIRT1,PGC-1α,PPAR-γ,Bcl2,Bax,Nrf2,NQO1,HO-1,SDH5 and COXIV in PSO(25 mg/kg)group were significantly up-regulated.These results suggested that PSO may play a protective role in ADR myocardial injury in mice by activating SIRT1 signaling pathway,which will be further proved in vitro.2.Cell experiment:(1)Compared with the Control group,the viability of HL-1 cardiomyocytes decreased to about 50% after 12 h treatment with 2 μM ADR,and the concentration and treatment time were the optimal injury conditions for the subsequent experiments;(2)Compared with the Control group,there were no significant changes in cell morphology and viability in different concentrations of PSO groups,indicating that PSO had no toxicity on cells within the experimental concentration range;(3)Compared with the Control group,the expression levels of SIRT1,PPAR-γ,PGC-1α,TFAM,UCP-2,NRF1,HO-1,Bcl2,NQO1 and other proteins in different concentrations of PSO groups were significantly up-regulated(P<0.05);Bax was not significantly changed,suggesting that PSO can activate the SIRT1/PPAR-γ signaling pathway in normal cardiomyocytes;(4)Compared with the ADR group,the cell morphology of the PSO administration group was improved,the cell viability was increased,the cell apoptosis rate was decreased,the LDH level of the culture medium was decreased,and the intracellular ROS level was decreased(P<0.05),indicating that PSO had a significant protective effect on ADR injury of HL-1 cardiomyocytes;(5)Compared with ADR group,PSO group significantly upregulated the expression levels of SIRT1,PPAR-γ,PGC-1α,TFAM,UCP-2,NRF1,HO-1,NQO1,Bcl2 and other proteins(P<0.05);which proved that PSO played an anti-ADR effect on HL-1 cardiomyocytes through SIRT1/PPAR-γ pathway;(6)Compared with Control group,the cell viability of SIRT1 si RNA1269 and PPAR-γantagonist GW9662 were significantly decreased((P<0.05),and the expression levels of SIRT1,PPAR-γ,PGC-1α,HO-1,NRF-2,UCP-2,Bcl2 and other proteins were significantly decreased;after SIRT1 si RNA treatment(P<0.05),the results showed that inhibiting the expression of SIRT1 and PPAR-γ gene could reverse the effect of PSO on ADR myocardial injury.Research Conclusions:In conclusion,(1)PSO has a significant protective effect on ADR-induced myocardial injury;(2)PSO plays an anti-ADR myocardial injury role by activating the SIRT1/PPAR-γ related pathway;(3)PSO is expected to be a potential drug for alleviating myocardial injury in ADR... |
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