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The Role Of CoQ10-lip In Renal IRI And The Evaluation Of Ultrasound RGD Targeted Microbubble

Posted on:2019-03-31Degree:DoctorType:Dissertation
Country:ChinaCandidate:X Q WangFull Text:PDF
GTID:1364330590966387Subject:Internal medicine Kidney disease
Abstract/Summary:PDF Full Text Request
ObjectiveTo explore the protective role of coenzyme Q10 loaded liposomes in renal ischemia reperfusion injury and mechanism.To approach the feasibility on evaluation of ischemia reperfusion injury in rat with RGD peptides targeted nano microbubble and contrast-enhanced ultrasound.Methods1 The protective role of Co Q10-lip in renal ischemia reperfusion injury and mechanism.(1)Preparation the Co Q10-lip with thin-film-hydration method(2)Male rat right kidney was removed,left kidney was subjected to IRI.The rats were randomly divided into four groups: N group(Sham group),B group(IRI group),C group(Co Q10 group),G group(Co Q10-lip group).C group and G group were respectively injected through the tail vein by Co Q10 solution or Co Q10-lip.After 24 hours,the rats were sacrificed.(3)The renal function and kidney morphology were analyzed.The level of superoxide dismutase(SOD),malondialdehyde(MDA),myeloperoxtidase(MPO)and tumor necrosis factor-?(TNF-?)in renal tissue were assayed by ELISA.The expression of Bax,Bcl-2,caspase-3,and NF-?B were detected by Western blot analysis.The apoptotic cells of the renal tissue were detected using a TUNEL apoptosis assay kit.Ultrasound and MRI were performed before the operation and 24 hours after the operation.2 The evaluation of ischemia reperfusion injury with RGD peptides targeted nano microbubble and contrast-enhanced ultrasound(1)Preparation the RGD peptides targeted nano microbubble and general lipid microbubbles with thin-film-hydration method(2)The rats of IRI were randomly divided into two groups: RGD peptides targeted microbubble group and general lipid microbubble group.After 1 min of intravenous bolus injection of two contrast agents,the ultrasound targeted microbubble destruction(UTMD)treatment immediately followed.The db values were measured respectively before and after UTMD,and comparation the variation of the db values.The db values are namely the number of microbubble.(3)The expression of ?v?3 in renal tissue were detected by Immunohistochemistry staining.3 Statistical analysis: The data are presented as the means±SD and were analyzed by SPSS 20.0 software.The P-value of less than 0.05 was considered statistically significant between different groups.Results1 The protective role of Co Q10-lip in renal ischemia reperfusion injury and mechanism.(1)In the glomeruli,there appeared no morphological change of four groups.There were severe architectural disruptions in tubules of group B,C and G,including tubular dilatation and brush border loss and cast formation at the corticomedullary junction,.whereas IRI rats with Co Q10 treatment demonstrated significantly less tubular damage.Moreover,the histopathology damage scores in group G were significantly lower than those in Group C.(2)IRI caused kidney dysfunction in group B with a peak Scr(125.83±64.39 VS 26±2.45?mol/L),P=0.01;Urea(26.32±13.87 VS 3.75±0.76 mmol/L),P=0.01.There were significant lower SCr and Urea levels in group G rats than group B,(41.17±7.19 VS 125.83±64.39?mol/L),P=0.02;(10.1±2.37 VS 26.32±13.87 mmol/L),P=0.04.There were also lower SCr and Urea levels in group C rats than group B,but there were no statistically significant between group C and group B.Scr(65.5±15.29 VS 125.83±64.39?mol/L)P=0.06;Urea(19.93±2.69 VS 26.32±13.87 mmol/L)P=0.23.(3)The SOD levels were significantly reduced in group B compared with those in group N.(408.83±6.46 VS 559.43±15.81 ng/ml),P<0.001.Co Q10-lip-treated rats had significantly higher SOD levels compared with untreated rats following IRI.454.73±13.86 VS408.83±6.46 ng/ml),P<0.001.Co Q10 solution has a similar role,(424.77±5.78 VS 408.83±6.46 ng/ml),P=0.01.Moreover,there were statistically significant between group G and group C,P<0.001.The MDA levels were significantly increased in group B compared with those in group N.(1225.27±51.95 VS 726.62±19.75 nmol/L),P<0.001.Co Q10-lip-treated rats had significantly lower MDA levels compared with untreated rats following IRI.(869.50±50.02 VS1225.27±51.95 nmol/L),P<0.001.Co Q10 solution has a similar role,(1043.59±42.35 VS 1225.27±51.95 nmol/L),P<0.001.Moreover,there were statistically significant between group G and group C,P<0.001.The MPO levels were significantly increased in group B compared with those in group N.(30.52±0.88 VS 19.24±0.86 ng/ml),P<0.001.Co Q10-lip-treated rats had significantly lower MPO levels compared with untreated rats following IRI.(21.48±0.97 VS 30.52±0.88 ng/ml),P=0.01.Co Q10 solution has a similar role,(27.04±1.17 VS 30.52±0.88 ng/ml),P<0.001.Moreover,there were statistically significant between group G and group C,P<0.001.The TNF-? levels were significantly increased in group B compared with those in group N.(249.74±8.31 VS 134.78±8.74 pg/ml),P<0.001.Co Q10-lip-treated rats had significantly lower TNF-? levels compared with untreated rats following IRI.(181.32±7.56 VS 249.74±8.31 pg/ml),P<0.001.Co Q10 solution has a similar role,(217.52±9.99 VS 249.74±8.31 pg/ml),P<0.001.Moreover,there were statistically significant between group G and group C,P<0.001.(4)After reperfusion for 24 hours,the expression of pro-apoptotic protein Bax and Caspases-3 was increased,while the expression of anti-apoptotic protein Bcl-2 was decreased.Co Q10-lip and Co Q10 solution both inhibited the increase of Bax and Caspases-3.Moreover,the role of Co Q10 was more obvious than Co Q10 solution.Co Q10-lip and Co Q10 solution both inhibited the decrease of Bcl-2.Moreover,the role of Co Q10 was more obvious than Co Q10 solution.The expression of NF-?B were increased after IRI.Co Q10-lip and Co Q10 solution both inhibited the increase of NF-?B.(5)TUNEL staining of the renal sections was then performed to visualize DNA fragmentation in situ.There were little TUNEL-positive cells in group N.In group B,TUNEL-positive cells were densely distributed.Treatment with Co Q10-lip or Co Q10 solution significantly reduced amount of TUNEL-positive cells.(6)Ultrasound examination showed after reperfusion for 24 hours the PSV,RI value in group B,C,G increased than those in group N,and the differences were statistically significant.But there was no statistical difference between the group B,C,G.Renal cortex TIC curve analysis in group N was rising rapidly at peak,and also reduced quickly.In another groups,all curve up and down slowly,especially in the fall.The AUC,TTP,Grad in group G were improved than those in group B and C,but there was no statistically significant difference.(7)MRI showed beside group N,there were statistically significant difference of SSI on OSOM value between preoperative and postoperative in group B,C,G.(P<0.05).But there were no statistically significant difference of SSI values between the three groups.(P>0.05).There were statistically significant difference of kcl values between preoperative and postoperative in group B,C.But treatment of Co Q10-lip was significantly delayed the progression IRI.There were no statistically significant difference of kcl values between preoperative and postoperative in group G.2 The evaluation of ischemia reperfusion injury with RGD peptides targeted nano microbubble and contrast-enhanced ultrasound(1)After ultrasound targeted microbubble destruction(UTMD),the intensity of contrast agent perfusion images changed with RGD peptides targeted nano microbubble,and the intensity was decreased.The changes of db values after UTMD were statistically significant difference between RGD peptides targeted nano microbubble group and general lipid microbubble group.(P=0.009).Therefore RGD peptides targeted nano microbubble has targeted ultrasound imaging for renal ischemia-reperfusion injury.(2)After ischemia-reperfusion injury,there were high?v?3 integrin expression in renal tissue,and lower expression in normal renal.Conclusion1 After renal IRI,renal tubular necrosis obviously,SOD concentration of renal tissues decreased,MDA,MPO and TNF alpha concentration increased,apoptosis increased,inflammatory cytokines expression increased;Co Q10-lip can delayed the decreased of SOD,inhibit the increased of MDA,MPO and TNF alpha and alleviate renal IRI.Co Q10-lip can reduce renal ischemia-reperfusion injury by inhibiting oxidative stress and apoptosis,relieving inflammation and other mechanisms and its effect is superior to Co Q10 solution.2 MRI can show sensitively renal IRI change.After the IRI,T2 WI showed in renal tissues,especially the outer stripe of the outer medulla(OSOM)the signal intensity increased.Time of contrast medium from renal cortex to medulla was prolonged,and kidney removal ability is reduced,Co Q10-lip can alleviate the change.3 RGD peptides as interaction between integrin and its ligand recognition sites,can explain the renal IRI high expression of integrin ?v?3,mediating interactions between cells and extracellular matrix,target recognition binding sites,with RGD targeted nano bubble contrast-enhanced ultrasound can more effectively evaluate rat renal IRI.
Keywords/Search Tags:ischemia reperfusion injury, CoenzymeQ10, liposome, oxidative stress, targeted ultrasound microbubbles, RGD peptide
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