1.Effect of ferulic acid on expression of microtubule-associated protein light chain 3,Beclin1,Bcl-2 and Bax in spinal cord injury rats
Xuefeng YU ; Xi JIANG ; Guokang WANG ; Jiajia ZHANG ; Yingzheng ZHAO ; Jianchun PAN
Chinese Journal of Pharmacology and Toxicology 2016;30(7):714-722
OBJECTIVE To investigate the effect of ferulic acid(FA)on motor function of rats with a spinal cord injury model(SCI)and its possible effects on expression of microtubule-associated protein light chain 3(LC3),Beclin1,Bcl-2 and Bax. METHODS SD rats were randomly divided into 3 groups:sham group,SCI group,FA(100 mg·kg-1,po)group. Rats were subjected to moderate contusion inju?ries using a vascular clip for 2 min to establish an SCI animal model beforfe they were given BBB scores and inclined plate scoring function test on 6 h,1,3,7 and 14 d after SCI. After the test,rats were sacrificed. Spinal cords were observed by hematoxylin eosin(HE)staining. Furthermore,the expressions of LC3,Beclin1,Bcl-2 and Bax were detected by Western blotting. RESULTS Compared with the sham group,BBB scores and inclined plate function scores significantly decreased in model group. The BBB scores decreased from 21 in sham group to(0.5±0.5)in SCI group,and inclined plate function scores decreased from 70° in sham group to(5.8±2.0)° in SCI group. However,this was reversed by FA treatment. BBB scores and inclined plate function scores increased from 3.0 ± 1.7 to 6.2 ± 3.6(P<0.05)and from (13.3 ± 4.1)° to(26.7 ± 8.7)°(P<0.05)after FA was po given for 7 d,respectively. HE staining showed the gradual emergence of internal spinal cord edema,while the structural changes associated with spinal cord injury could be significantly reversed by administration of FA. Moreover,the expression of LC3-Ⅱ/LC3-Ⅰand Beclin1 was significantly increased in SCI 1 d group(P<0.01),but was decreased in 14 d group when compared with SCI 1 d group(P<0.05). The expression of Bcl-2 was increased in SCI 14 d group,and the ratio of Bcl-2/Bax was increased on 14 d after SCI(P<0.05). Compared with the SCI group,LC3-Ⅱ/LC3-Ⅰand Beclin1 expression was increased in FA-treated 1 d group(P<0.05),Bcl-2 expression was increased in FA-treated 14 d group and the ratio of Bcl-2/Bax was significantly increased on 14 d after SCI(P<0.05). CONCLUSION FA has a therapeutic effect on SCI,which may be related to the impact of neuronal autophagy and apoptosis. Meanwhile,autophagy of SCI may be a process of gradual enhancement followed by weakening,and the anti-apoptosis effect may gradually increase with autophagy.
2.Establishment of a TaqMan probe real-time quantitative PCRfor detection of rat Theiler's-like virus
Xiaoyao CAI ; Xiaofeng WEI ; Wei XIONG ; Yifei CHEN ; Yingzheng LIN ; Quan ZHANG ; Hongjun CHEN
Acta Laboratorium Animalis Scientia Sinica 2017;25(4):433-437
Objective To establish an accurate TaqMan probe real-time quantitative PCR (qPCR)method for detection of Theiler''s-like virus of rats (TLV).Methods Primers and TaqMan probes specific to 3622~3729 nt region were designed according to the whole genomic sequence of TLV representative strain.Using a synthesized plasmid as DNA standard template, the stability, specificity, and sensitivity of the qPCR method were determined.Results In the standard curve, R2 value was 0.99 with a high specificity.The sensitivity of the real-time PCR was less than 10 copies/μL, which was 100 times higher than the ordinary PCR method.No cross reactions appeared to the other rat viruses.Conclusions The TaqMan probe qPCR method established in this study has advantages such as simple to use, high sensitivity and specificity.
3.Improvement of left ventricular myocardial perfusion after acidic fibroblast growth factor delivered by using ultrasound-targeted microbubble destruction in rats with diabetic cardiomyopathy:an preliminary study
Lei ZHENG ; Xinqiao TIAN ; Ming ZHANG ; Yingzheng ZHAO ; Ao RU ; Jianmin LI ; Keke JIN
Chinese Journal of Ultrasonography 2016;(1):69-74
Objective To investigate the protective effects of left ventricular myocardial perfusion after delivery of acidic fibroblast growth factor (aFGF) in rats with diabetic cardiomyopathy (DCM) by using ultrasound‐targeted microbubble destruction ( UTMD ) with real‐time myocardial contrast echocardiography( RT‐MCE) . Methods Among 64 male SD rats ,52 rats were randomly selected and were induced DCM by streptozotocin through intraperitoneal injecting ,the other rats as normal control group . DCM rats were randomly divided into the DCM model group ,aFGF only group ,SonoVue‐aFGF group and the SonoVue‐aFGF+ UTMD group in this study . The aFGF only group rats were injected with aFGF solution through tail vein ,the SonoVue‐aFGF group were injected with SonoVue‐aFGF solution through tail vein and SonoVue‐aFGF+ UTMD group rats were injected with SonoVue‐aFGF solution through tail vein and using UTMD simultaneously . All rats underwent conventional echocardiography and RT‐MCE exams before and 4 weeks after intervention . Left ventricular ejection fraction ( LVEF) and left ventricular fraction shortening( LVFS ) were measured by conventional echocardiography . The plateau intensity ( A ) ,initial slope of the curve (β) and myocardial blood flow ( A ×β) of left ventricular anterior wall at the papillary muscle level were measured in left ventricular short‐axis view by RT‐MCE . Results Before intervention , LVEF and LVFS in the DCM model group ,aFGF only group ,SonoVue‐aFGF group and the SonoVue‐aFGF+UTMD group were significantly lower than in the normal control group ( P <0 .01) .During 4 weeks after intervention ,LVEF and LVFS in the aFGF only group ,SonoVue‐aFGF group and the SonoVue‐aFGF+UTMD group were slightly increased than those in the DCM model group ,but no statistically significant differences were found ( P > 0 .05) ,however A and A × β in the SonoVue‐aFGF+ UTMD group were significantly increased than those in the DCM model group( P < 0 .01) . Compared with the same group before intervention ,A and A ×βin the SonoVue‐aFGF+UTMD group were higher ( P <0 .05) and these in the DCM model group were lower during four weeks after intervention ( P < 0 .05) . Conclusions Acidic fibroblast growth factor delivered by using UTMD can improve the left ventricular myocardial perfusion in diabetic cardiomyopathy rats .
4.Effects of Electroacupuncture at Baihui (DU20) on Learning and Memory and Expression of Brain-derived Neu-rotrophic Factor in APP/PS1 Double-transgenic Mice
Jixiang CHEN ; Yunan WU ; Yaxuan ZHENG ; Peiyuan ZHUO ; Yingzheng ZHANG ; Lidian CHEN
Chinese Journal of Rehabilitation Theory and Practice 2015;21(6):642-647
Objective To explore the effects of electroacupuncture at Baihui (DU20) acupoint on learning and memory and its possible mechanism through the expression of brain-derived neurotrophic factor (BDNF) in APP/PS1 double-transgenic mice. Methods 30 female APP/PS1 double transgenic mice were randomly divided into model group, DU20 group and non-acupoint group, and 10 wild type mice con-sisted of wild group. DU20 group received electroacupuncture at Baihui and the non-acupoint group received electroacupuncture at non-acu-point for 28 days. Learning and memory was tested by Morris water maze. Deposition ofβ-amyloid (Aβ) peptide was determined by immu-nohistochemical staining. The expression of BDNF in cortex was examined by RT-PCR and Western blotting. Results Compared with the model group, DU20 group ameliorated the learning and memory ability of APP/PS1 double-transgenic mice (P<0.05), decreased the deposi-tion of Aβpeptide (P<0.05) and upregulated the gene and protein levels of BDNF (P<0.01). There was no significant difference between the model group and non-acupoint group (P>0.05). Conclusion Electroacupuncture at DU20 acupoint could ameliorate learning and memory in APP/PS1 double-transgenic mice. The mechanism may be related to increase the expression of BDNF and decrease the deposition of Aβ.
5.Effect of leptin on expression of lipoic acid synthase in the liver and kidney of Leprdb/dbmice
Qiang PENG ; Yingzheng ZHAO ; Tingting YAN ; Xiaonan ZHAI ; Xuxu ZHANG ; Xianwen YI ; Hexi ZHANG ; Guangcui XU
Acta Laboratorium Animalis Scientia Sinica 2018;26(2):145-149
Objective To study the expression of lipoic acid synthase(LIAS)in the liver and kidney of Leprdb/db mice with deficient leptin receptor. Methods Eight 10-week old male Leprdb/ +mice and Leprdb/dbmice were included in this study. The body weight of rats in the two groups was measured. Fasting blood glucose(FPG)was measured with blood glucose test strips for all mice after fasting for 8 hours. Blood samples were obtained from the abdominal aorta and the animals were sacrificed. The liver and kidney were weighed. The right lobe of liver and the left kidney samples were fixed in 4% paraformaldehyde for pathological examination. Serum samples were separated and the sereum contents of CHO, TG,HDL and LDL were detected. The mitochondria of liver and kidney tissues were extracted with a mitochondrial isolation kit, and the protein was extracted. The expression of LIAS protein was detected by western blot. Results Histopathological observation showed that the liver and kidney tissues of Leprdb/ +mice have intact and clear structure. But the liver tissue of Leprdb/dbmice showed fatty degeneration, the kidney tissue showed glomerular hypertrophy, basement membrane thickening, mesangial area widened, including mesangial cells and mesangial matrix increased. The GLU,CHO,TG,LDL and AST of Leprdb/dbmice were significantly increased compared with those of Leprdb/ +mice(P<0.05). Compared with Leprdb/ +mice,the LIAS protein expression was significantly increased in the liver and kidney mitochondria of Leprdb/dbmice(P<0.05). Conclusions There is impaired glucose and lipid metabolism in the Leprdb/dbmice which has defect leptin receptor,and the expression of LIAS protein in liver and kidney of the Leprdb/dbmice is higher than that of Leprdb/ +mice.
6.Genotyping of the offsprings of Leprdb/ +mice by TaqMan probe fluorescence quantitative PCR
Yingzheng ZHAO ; Qiang PENG ; Tingting YAN ; Xuxu ZHANG ; Xiaonan ZHAI ; Weidong WU ; Xianwen YI ; Guangcui XU
Acta Laboratorium Animalis Scientia Sinica 2018;26(2):207-210
Objective To establish an efficient method of genotyping for Leprdb/ +mouse offsprings by TaqMan probe quantitative fluorescence PCR. Methods Genome DNA was extracted from tails of 228 Leprdb/ +mouse offsprings. PCR primers and TaqMan probes were designed according to the mutation sites of Lepr gene(rs1801133). Real time PCR assay was applied and SNP loci were typed with SDS software. The genotyping of 2-month old Leprdb/dbmice was validated by the phenotype and Hardy-Weinberg equilibrium test was performed. Results 228 samples were detected by the established TaqMan fluorescence quantitative PCR assay. 64 mice were of GG genotype, with a genotype frequency of 0.1929. 123 mice were of GT genotype, with a genotype frequency of 0.5395. 41 mice were of TT genotype, with a genotype frequency of 0.2807. Compared with the phenotype typing,the sensitivity of the TaqMan fluorescence quantitative PCR was 97.56% and the specificity was 99.47%. Conclusions TaqMan probe quantitative fluorescence PCR assay is a simple and efficient method,and can be used to detect the genotype of Leprdb/ +mouse offsprings.
7. Therapeutic effect of combined use of FGF1-loaded nano-liposomes and ultrasound-targeted microbubble destruction technique on treating rats with experimental diabetic cardiomyopathy
Ming ZHANG ; Yingzheng ZHAO ; Weicheng MA ; Jinlong XU ; Jingling WANG ; Mengjia CHEN ; Lu YU ; Yuanna CHEN
Chinese Journal of Cardiology 2017;45(5):427-433
Objective:
The therapeutic effect of acid fibroblast growth factor 1(FGF1) on rats with diabetic cardiomyopathy (DCM) was evaluated by using nano-liposomes combined with ultrasound-targeted microbubble destruction technique (UTMD).
Methods:
The FGF1-loaded nano-liposomes were prepared by water-in-water emulsion method combined with lyophilization technique.TypeⅠdiabetes model was induced by intraperitoneal injection of streptozotocin (STZ, 70 mg/kg) in 60 male SD rats.Sixteen weeks later, diabetic rats were randomly divided into: placebo group (saline treatment), FGF1 group, FGF1-loaded nano-liposomes group, and FGF1-loaded nano-liposomes plus UTMD group (