1.Construction of HPV18E7 recombinant plasmid and exploration of its optimization expression condition in Escherichia coli
Renjian HU ; Jiali CAI ; Li LIU ; Manyu TU ; Tao XU ; Cuirong DU ; Jia LUO ; Sen DING
Chongqing Medicine 2013;(30):3647-3649
Objective To construct recombinant plasmids containing HPV18E7 gene ,and explore the optimization condition of its expression in Escherichia coli .Methods The genomic DNA extracted from HeLa cell line which served as a template to the HPV18 E7 gene was amplified using PCR method ;and the amplified product of HPV18E7 gene was connected to the pET-32a(+ ) vector ,which composed the pET-32a(+ )-HPV18E7 recombinant plasmid ;the positive recombinant plasmids were transformed into BL21-DE3-pLysS competent cells and the optimized expression condition was explored in order to obtain a large amount of HPV18E7 oncogenic protein .Results The fragment length of PCR products of HeLa cell genomic DNA was consistent with that of HPV18 E7 gene .In LB medium ,the expression level of the target protein was not high under such conditions as different concentra-tion of IPTG and lactose ,different temperatures and different induction starting amount .Therefore the ZYM-5052 auto-induction medium was tried in this experiment ,and the expression amount of the fusion protein was much higher than that induced with IPTG and lactose .Conclusion The amount of HPV18E7 fusion protein in ZYM-5052 automatic induction medium is much higher than that induced with IPTG and lactose .
2.Endothelin-1 regulates SOCC/TGF-β and involved in atrial fibrosis in rats with atrial fibrillation
Zhuoran JIA ; Manyu DAI ; Shichu LIANG ; Jian WU ; Yangcheng XUE ; Dingxin ZHANG ; Bing SHEN ; Ren ZHAO
Acta Universitatis Medicinalis Anhui 2024;59(3):429-435
Objective To investigate the effect and mechanism of endothelin-1(ET-1)on atrial fibrosis in Atrial fibrillation(AF)rats.Methods Fourteen adult male SD rats were randomly divided into normal control(NC)group and Atrial fibrillation(AF)group.The rat model of Atrial fibrillation was established by injecting 0.1 ml/100g CaCl2-Ach mixture into the tail vein once a day for one week.The control group was injected with the same dose of normal saline.An electrocardiogram of normal or atrial fibrillation was recorded on the first day and the eighth day in each group,and echocardiography was used to monitor atrial size and cardiac function.The fibrosis of atrial was observed using Masson and HE staining.The expression of endothelin-1(ET-1),collagen-I(Col-I),transforming growth factor-β(TGF-β)and the store operated calcium channel(SOCC)protein Orai1,stromal in-teraction molecule 1(STIM1)in atrial tissue were detected by Western blot.HL-1 cells were cultured and treated with gradient concentration of ET-1 for 24 hours.Western blot was used to observe changes in the expression of TGF-β,Orai1 and STIM1 proteins in ET-1/SOCC/TGF-β signaling pathway of HL-1 cells.Small interfering RNA(siRNA)transfection method was used to knock down the expression of Orai1 in HL-1 cells,then the cells were treated with appropriate concentrations of ET-1 for 24 hours,and the expression of TGF-β protein in HL-1 cells was detected by Western blot.Results Compared with the control group,echocardiography showed a significant in-crease in left atrial diameter(LAD)of the heart in atrial fibrillation rats(P<0.05).The HE and Masson staining results showed significant fibrosis in the myocardial tissue of AF group rats(P<0.05),and the Western blot re-sults indicated the expression of ET-1,Orai1,STIM1,TGF-β and COL-Ⅰ in the myocardial tissue of AF group significantly increased compared to the NC group(P<0.05).After ET-1 treatment of HL-1 cells,the protein ex-pression of Orai1,STIM1and TGF-β increased(P<0.05),while knocking down Orai1 in HL-1 cells,ET-1 treat-ment no longer caused the expression of TGF-β a significant upregulation.Conclusion AF caused by atrial fibril-lation results in a significant increase in ET-1 expression in atrial tissue,and ET-1/SOCC/TGF-β signal pathway promotes atrial fibrillation and fibrosis.
3.Study on the relationship between adenoid and tonsil hypertrophy and obesity in children
Manyu JIA ; Shizhen ZOU ; Jinrang LI
Chinese Journal of Otorhinolaryngology Head and Neck Surgery 2020;55(8):760-763
Objective:To study the relationship between adenoid hypertrophy, tonsillar hypertrophy and overweight or even obesity in children.Methods:A total of 799 children aged 2 to 12 years with tonsillar and adenoid hypertrophy from January 2015 to December 2019 in the Department of Otolaryngology Head Neck Surgery, the Sixth Medical Center of Chinese PLA General Hospital were selected. The body mass index (BMI) was calculated according to the height and weight measured routinely at the time of admission. The difference of BMI between children and normal children of the same age, and the correlation between adenoid, tonsil hypertrophy and obesity were compared. Chi-square test was used to compare the surgical children′s BMI of different genders with normal children of the same age, and Spearman rank correlation was used to analyze the correlation between BMI and adenoid and tonsil hypertrophy.Results:The Spearman correlation coefficient between tonsil hypertrophy and BMI was 0.078, P=0.077, the Spearman correlation coefficient between adenoid hypertrophy and BMI was -0.058( P=0.100). χ 2 test showed that the proportion of overweight and obesity in school-age children (7~12 years old) was significantly higher than that in preschool children (2~6 years old), and the difference was statistically significant (χ 2male=22.386, Pmale<0.001, χ 2female=4.478, Pfemale<0.001). Conclusion:There is no obvious correlation between adenoid hypertrophy, tonsil hypertrophy and overweight or obesity in children, but the probability of children overweight or obesity increases with age, and the proportion of obesity in children aged 7-12 years is higher.
4.Comparative study of left atrial matrix ablation and posterior wall linear isolation ablation strategies in elderly patients with atrial fibrillation
Hui Xiao ; Haoyang Lu ; Manyu Dai ; Yangcheng Xue ; Zhuoran Jia ; Zhuoran Jia ; Jie Guo ; Ren Zhao
Acta Universitatis Medicinalis Anhui 2022;57(11):1844-1848
Objective :
In the present study,the changes of left atrial matrix characteristics and the effect of cathe- ter ablation on cardiac function in elderly patients with persistent atrial fibrillation were observed.Furthermore,the application of two catheter ablation methods including circular pulmonary vein isolation combined with left atrial ma- trix modification or left atrial posterior wall isolation were compared by researchers in order to explore effectiveness and safety of the two methods in elderly persistent atrial fibrillation patients.
Methods :
A total of 86 elderly pa- tients with persistent atrial fibrillation were selected and divided into two groups using randomization principle ; the pulmonary vein isolation combined with left atrial matrix modified catheter ablation method was named matrix modi- fied group (43 cases) ; the pulmonary vein isolation combined with left atrial posterior wall isolation catheter abla- tion method was named posterior wall isolation group (43 cases) .During ablation left atrial electro matrix mapping were analyzed by multipole catheter for all patients.After ablation all patients were follow-up in 12 months by car- diac color doppler ultrasound examination,ECG and Holter.The incidence of arrhythmia ,serious adverse event and complications were compared between two groups. Successful rate of single ablation and blank period recur- rence after ablation were observed in 12 months follow-up time.
Results :
Through electrogram mapping of left atri- um low-voltage areas or scar areas were founded in 72 cases among the 86 cases after ablation,After a 12-month follow-up,the incidence of arrhythmia in the blank period between the two groups was 41.9% ( matrix modified group) and 23. 3% (posterior wall isolation group) ,independently,P<0. 01,successful rates of single ablation in the two groups were 83. 7% (matrix modified group) ,60. 5% (posterior wall isolation group) ,independently, P <0. 01 .No adverse event occurred in the two groups.After AF ablation LVEF increased,LVEDD and LVAD significantly decreased in all of the cases.There was no significant difference between the two groups in patients ’ characters ablation time and complications rate.
Conclusion
Circular pulmonary vein isolation combined with left atrial matrix modification is as safe as left atrial posterior wall isolation,and it is a simplified,personalized,and more effective ablation strategy in elderly patients with persistent atrial fibrillation.