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2018 Vol. 48, No. 4 Published: 2018-04-25   235 Changes in the expression of SIRT1 in nonalcoholic fatty liver disease in rat liver and the roles of Curcumin derivatives L6H4 DING Tingting, WU Hangfei, LIU Cheng, TANG Wen, WANG Shengkai, ZHENG Jingyu, ZHU Zaisheng, PENG Tianqing. Objective: To explore the expression changes of SIRT1 in rat liver with high-fat induced nonalcoholic fatty liver disease (NAFLD), and the effects of Curcumin derivatives L6H4 on it. Methods: Twenty-four male SD rats were randomly divided into normal control group (control group), model group (NAFLD group) and L6H4 treatment group (L6H4 group). Rats in the control group were fed with normal-diet while rats in NAFLD group and L6H4 group were fed with high-fat diet. At the same time, rats in L6H4 group were treated with Curcumin derivatives L6H4 from the 9th week till the 20th week of the experiment. At the end of the 20th week, all rats were sacrificed. The serum triglyceride, total cholesterol, low-density lipoprotein cholesterol and high-density lipoprotein cholesterol levels were detected by biochemical method respectively. The pathological changes of liver tissues were observed by HE staining and Masson’s trichrome staining. The total SOD activity was detected by hydroxylamine method while the content of MDA was analyzed by thiobarbituric acid method.The expressions of SIRT1, DRP1 and MFN2 mRNA and protein in liver tissues were detected by real-time PCR and Western blot respectively. Results: Obvious steatosis and fibrosis were observed in NAFLD group under microscope. The pathological changes were significantly reduced in L6H4 group compared with NAFLD group. Compared with control group, the serum triglyceride, total cholesterol, low-density lipoprotein cholesterol levels, the content of MDA and the expression of DRP1 mRNA and protein were increased significantly, while the serum high-density lipoprotein cholesterol level, the total SOD activity and the expressions of SIRT1 and MFN2 mRNA and protein were decreased significantly (P<0.05) in NAFLD group. Compared with NAFLD group, the serum triglyceride, total cholesterol, low-density lipoprotein cholesterol levels, the content of MDA and the expression of DRP1 mRNA and protein were decreased significantly, while the serum high-density lipoprotein cholesterol level, the total SOD activity and the expressions of SIRT1 and MFN2 mRNA and protein were increased significantly (P<0.05) in L6H4 group. Conclusion: Curcumin derivatives L6H4 has a protective effect on NAFLD, and its mechanism may be through increasing the expression of SIRT1, thereby reducing the resistance of hepatocyte to insulin and lipid peroxidation, attenuating oxidative stress damage and inhibiting mitochondrial fission. 2018 Vol. 48 (4): 235-240 [Abstract] ( 643 ) HTML (1 KB)  PDF (1525 KB)  ( 950 ) 241 Construction of the recombinant plasmid of human YAP1 gene and the expression of fusion protein DONG Jinglai, ZHANG Jinsan, GUO Qiang, CHEN Qianjie, GUO Lisha, LI Xiaokun. Objective: To construct two isoforms of YAP1 for further research into their difference. Methods: YAP1-1δ and YAP1-2δ fragments were obtained by using overlapping PCR. And then the two DNA fragments were respectively ligated to the plasmid contained partial YAP1 gene via ClaI and SalI two restriction enzyme cutting sites. The recombinant vectors named pCI2-Flag-YAP1-1δ and pCI2-Flag-YAP1-2δ were identified by restriction enzyme digestion and gene sequencing. After transfection of 293 cells with the two vectors for 48 h, the expression of YAP1 protein was detected by Western blot. Results: The two over-expressing plasmids were successfully constructed and YAP1-1δ and YAP1-2δ proteins were strongly expressed in HEK293 cells. Conclusion: The HEK293 cells could transiently express YAP1-1δ and YAP1-2δ by using the recombinant plasmids pCI2-Flag-YAP1-1δ and pCI2-Flag-YAP1-2δ, which lays a foundation for further study of the different isoforms of YAP1 and its mechanism in the development of tumorigenesis. 2018 Vol. 48 (4): 241-244,250 [Abstract] ( 735 ) HTML (1 KB)  PDF (1501 KB)  ( 1201 ) 245 Efficacy evaluation of obesity measurement indicators combined with family history of diabetes in pathogly-cemia identification XU Jing, GU Xuejiang, HONG Yilian, LIN Yi, ZHANG Jiale, YANG Lijuan. Objective: To compare the effectiveness of obesity measurement index in identifying abnormal blood glucose (prediabetes and diabetes) in populations with family history of diabetes (FHD+) and those with non-diabetic family history (FHD-). Methods: A cross-sectional study was carried out in four communities of Wenzhou city. A total of 4 867 people aged above 40 years old were included in the study, among whom, 979 people were FHD+ group and 3 878 people in FHD- group. A correlation analysis was made of body mass index (BMI), waist circumference (WC), waist-hip ratio (WHR), waist-to-height ratio (WHtR) and blood glucose level (fasting plasma glucose, 2-hour postprandial blood glucose and hemoglobin A1c) using Pearson correlation. Areas under the working characteristic curve of four indexes in identifying subjects with abnormal blood glucose were compared and the optimal cut-off value was determined. Results: BMI, WC, WHR, WHtR were all related to blood glucose indexes (r=0.114~0.253, P<0.05). FHD was layered, and the incidence of abnormal blood glucose in FHD+ group was significantly higher than that in FHD- group (P<0.05). The optimal cut-off value of WHtR in screening glucose abnormality in FHD+ group and FHD- group was 0.54 and 0.56 respectively; that of BMI was 23.5 kg/m2 and 24.8 kg/m2 respectively; that of WC was 86.1 cm and 88.2 cm respectively; that of WHR was both 0.89; ROC curve analysis showed that the area under curve of waist-to-height ratio was the largest (both 0.65). Conclusion: The cut-off value of obesity measurement indexes in identifying glucose abnormality in FHD+ group is lower than that in FHD- group. It is suggested that stricter cut-off value should be given for FHD+ population. Of the two groups, WHtR is the best index to identify glucose abnormality. 2018 Vol. 48 (4): 245-250 [Abstract] ( 683 ) HTML (1 KB)  PDF (1367 KB)  ( 940 ) 251 Relationship between the development of spleen deficiency syndrome and the changes of myocardial energy metabolism in rats QU Xiaohu, CHEN Hui, HUANG Ling, LYU Bin, JIN Liqin Objective: To explore the developmental changes of energy metabolism in myocardium of the spleen deficiency from light to heavy dynamic process by replicating the rat models of spleen qi deficiency syndrome and spleen deficiency and failure to control circulating blood syndrome. Methods: The rat models of spleen qi deficiency syndrome and spleen deficiency and failure to control circulating blood syndrome were replicated. The levels of ATP, the activity of citrate synthase (CS), the activity of mitochondrial respiratory chain complex (Complex I, Complex II, Complex III, Complex IV), the mtDNA copy number, the expression of COXIV and Cyt-C and their mRNA were measured in the myocardium of the rats. Results: The ATP content and activities of CS, Complex I, Complex II, Complex III, Complex IV, expression of COXIV protein and its mRNA in rats with spleen qi deficiency syndrome and spleen deficiency and failure to control circulating blood syndrome were significantly lower than those in normal rats (P<0.05). The mtDNA copy number and expression of Cyt-C protein and its mRNA in rats with spleen deficiency and failure to control circulating blood syndrome were significantly higher than those in normal rats (P<0.05). The activities of CS and expression of Cyt-C protein and its mRNA in rats with spleen deficiency and failure to control circulating blood syndrome were significantly higher than those in spleen qi deficiency syndrome rats (P<0.05). Conclusion: In the early stage of spleen qi deficiency syndrome in rats, there was no significant change in energy metabolism of the myocardium. Dysfunction of myocardial tissue energy metabolism in rats began to develop with the prolonged course of deficiency in spleen and the changing from spleen qi deficiency syndrome to spleen deficiency and failure to control circulating blood syndrome. The level of ATP was significantly reduced. Tricarboxylic acid cycle electron transport chain related enzyme activity and the expressions of important proteins and genes also changed, indicatin had been adversely affected in the aggravated process of spleen deficiency.  It was consistent with the theory of traditional Chinese medicine in that five viscera will not be nourished and gradually become weak in the case of spleen qi deficiency. 2018 Vol. 48 (4): 251-256 [Abstract] ( 694 ) HTML (1 KB)  PDF (1961 KB)  ( 1006 ) 257 Application of the STOP-Bang questionnaire in screening obstructive sleep apnea hypopnea syndrome WU Wan, HUANG Chenyao, DONG Li, LIU Lingjie, ZENG Haihuan, XIE Yupeng. Objective: To assess the reliability and validity and value of the STOP-Bang questionnaire in the screening of obstructive sleep apnea (OSAHS) in sleep clinic. Methods: Polysomnography (PSG) and STOP-Bang questionnaire survey was simultaneously done in 649 subjects who were consulted at the sleep clinic. Indicators including reliability, validity, sensitivity and specificity of STOP-Bang questionnaire were calculated and evaluated by a comparison of the data obtained. Influencing factors were analyzed by Logistic regression. Results: The Cronbach’s alpha value of the STOP-Bang questionnaire was 0.732. The results of factor analysis showed that the common values of common factors 1, 2 and 3 were 1.789, 1.556 and 1.167 respectively, with the total factor being 56.40%. Based on the diagnosis criteria via PSG as AHI ≥5, ≥15, or ≥30, the sensitivity and specificity via STOP questionnaire were 84.0%, 68.8%; 88.8%, 53.1%; 90.3%, 43.6% respectively. Logistic analysis shows: loud snoring, neck circumference over 40 cm and aged 50 plus combined with hypertension are the risk factors of OSAHS. Conclusion: The STOP-Bang questionnaire has good reliability and validity and can be a useful tool to screen OSAHS in sleep clinic with relatively high sensitivity and specificity. However, it should be further studied in general population. Loud snoring, neck circumference over 40 cm and aged 50 plus combined with hypertension are the risk factors of OSAHS. 2018 Vol. 48 (4): 257-261 [Abstract] ( 1126 ) HTML (1 KB)  PDF (1249 KB)  ( 999 ) 262 The role of long non-coding RNA FENDRR in cervical cancer LAN Xiaoxiao, ZHOU Zhiyang, XU Xinxin, WU Xueqing Objective: To investigate the effects of long non-coding RNA FENDRR in cervical cancer. Methods: Cervical cancer cells were transfected with siRNA that specifically targeting FENDRR. QRT-PCR was used to verify the transfection effect of siRNA. Proliferation and motility capacities of cervical cancer cells were demonstrated by CCK-8 and Transwell assays, respectively. Tumorigenesis in nude mice was performed to identify the proliferation ability of cervical cancer cell in vivo. Results: With siRNA transfection, the expression levels of FENDRR in SiHa and HeLa were significantly reduced from (1.013±0.120), (1.001±0.025) to (0.226± 0.024), (0.099±0.010) (P<0.01). After transfecting with siRNA targeting FENDRR, the capacity of proliferation was significantly enhanced in cervical cancer cells (P<0.01), as well as migration and invasion abilities (P<0.01). The tumorigenesis in nude mice was obviously improved by interfering FENDRR expression in HeLa. Compared with the control group, the tumor volume increased from (853.2±59.4) mm3 to (1 155.0±51.5) mm3 (P<0.01), the tumor weight increased from (0.544±0.053) g to (0.814±0.061) g (P<0.05). Conclusion: Knockdown the expression of FENDRR in cervical cancer cells significantly enhanced their proliferation, migration as well as invasion abilities in vitro, and improved their tumorogenesis in vivo, which indicated that FENDRR may play an important inhibitory role in the progression of cervical cancer. 2018 Vol. 48 (4): 262-266 [Abstract] ( 785 ) HTML (1 KB)  PDF (1480 KB)  ( 854 ) 267 The effects of salvianolic acid A on mitochondrial autophagy and cell injury in human pulmonary microvascular endothelial cells induced by lipopolysaccharide LI Weiwen, LAN Xiu, LI Yuan, SUN Lei, LYU Zhuqing. Objective: To investigate the effect of salvianolic acid A (Sal A) on mitochondrial autophagy and cell injury in human pulmonary microvascular endothelial cells (HPMVECs) injury induced by lipopolysaccharide (LPS). Methods: LPS 10 μg/mL was used to establish cell injury in vitro. The cells were divided into four groups: control group, Sal A group, LPS group, LPS+Sal A group. MTT was used to analyze cell proliferation. The reactive oxygen species (ROS) content and mitochondrial membrane potential were detected by flow cytometry. Western blot was used to analyze the expression of microtubules associated protein light chain 3 (LC3)-II/I, Beclin1, PTEN mediated putative kinase protein 1 (PINK1) and Parkin. Results: LPS decreased the cell viability in time-dependent manner. The viability in LPS+Sal A group was higher than the LPS group, but lower than the control group (P<0.05). Compared with the control group (100%), the ROS content (283.96%±10.88%) increased and the level of membrane (38.25%±5.69%) potential decreased in LPS group (P<0.05). Compared with LPS group, ROS content (138.76%±11.82%) decreased, and the mitochondrial membrane potential (76.56%±6.22%) increased in LPS+Sal A group (P<0.05). The expression of LC-3II/I, Beclin1, PINK1 and Parkin in LPS group was higher than the control group. The expression of LC-3II/I, Beclin1, PINK1 and Parkin in LPS+Sal A group was lower than LPS group. Conclusion: Sal A alleviates the proliferation inhibition by LPS, which may be related with decreased intracellular ROS content, stabilization of mitochondrial membrane potential and mitochondrial autophagy. 2018 Vol. 48 (4): 267-270 [Abstract] ( 809 ) HTML (1 KB)  PDF (1333 KB)  ( 869 ) 271 The role of expression level of protamines and sperm chromatin packaging quality in oligozoospermia HONG Dan, LOU Zhefeng, ZHENG Jiujia, HUANG Xuefeng, JIN Longjin Objective: To investigate the correlation between mRNA transcription level of protamines (PRM1 and PRM2) in oligozoospermia. Methods: Sperm samples were recruited following WHO criteria and further collected from oligozoospermia patients (28 cases), teratozoospermia (16 cases) and healthy volunteers (15 cases) respectively. The mRNA transcriptions of PRM1, PRM2 and TRF2 in sperm were detected by RT-qPCR. The quantification of sperm chromatin packaging quality was performed by CMA3 staining. Results: Compared with control group, mRNA transcription levels of PRM1 and PRM2 in oligozoospermia group were significantly decreased 4 folds and 5 folds respectively. However, there’s no significant difference in mRNA transcription of PRM1 and PRM2 between teratozoospermia and control groups, or TRF2 between oligozoospermia and control groups, or TRF2 between teratozoospermia and control groups. Elvated CMA3 positive cells were only observed in oligozoospermia samples, compared with teratozoospermia and control samples. Conclusion: PRM1 and PRM2 are critical for maintaining high level of sperm counts. Decreased expression of PRM1 and PRM2 may cause sperm chromatin packaging dysfunction, which can further suppress spermiogenesis. 2018 Vol. 48 (4): 271-274 [Abstract] ( 664 ) HTML (1 KB)  PDF (1282 KB)  ( 1210 ) 275 The prevalence and risk factors of dyslipidemia among young adults based on an analysis of physical examination reports CHEN Keyang, HU Beilei, CHEN Songfang, JIANG Minghua, CHEN Ke, WANG Xiaotong Objective: To explore the status and risk factors of dyslipidemia among young adults of Wenzhou reported by the health examination in order to provide intervention measures. Methods: The individuals aged 18-45 years were recruited through multi-stage random sampling from the Medical Examination Center of the 2nd Affiliated Hospital of Wenzhou Medical University. The survey included questionnaire, physical examination and laboratory detections of triglyceride (TG), total cholesterol (TC), high density lipoprotein cholesterol (HDL-C), low density lipoprotein cholesterol (LDL-C). Results: A total of 8 497 young adults were investigated, including 4 295 males and 4 202 females. The prevalence of dyslipidemia was 34.11%, with 51% in male and 16.7% in female; The average age was (34.07±7.11) years old in male and (34.52±7.08) years old in female. The level of serum TG, TC, HDL-C and LDL-C was (1.69±0.07) mmol/L, (4.66±0.18) mmol/L, (1.21±0.05) mmol/L, (2.69±0.11) mmol/L. Logistic regression analysis showed that the risk factors included age (OR=1.032), smoking (OR=2.016), drinking (OR=1.334), overweight (OR=1.228), occupation (OR=1.058). Conclusion: The prevalence of dyslipidemia among young adults in Wenzhou is high. It suggests that targeted prevention and control measures are needed to lower the prevalence of dyslipidemia. 2018 Vol. 48 (4): 275-279 [Abstract] ( 639 ) HTML (1 KB)  PDF (1244 KB)  ( 1035 ) 280 The effect of estrogen on human papilloma virus 16 E6 gene transfecting HaCat cell line LU Xiaosheng, CHEN Jing, ZHAO Junzhao, LYU Jieqiang Objective: To determine the influence of 17-β-estradiol (E2) on human papilloma virus 16 E6 gene transfecting HaCat cell line and the possible mechanism. Methods: The HaCat cell line was incubated with E2 of varies concentration (10-5, 10-6, 10-7, 10-8, 10-9, 0 mol/L) .The cell number was counted for 7 days to draw a growth curve. The cell viability was determined by MTT method. Flow cytometry was used to detect cell cycle distribution and BrdU to determine the time duration of each cell cycle phase. Then, the E2 treated cells was transfected by the plasmid containing HPV E6. The expression of E6 mRNA was determined by quantitative realtime-PCR. Results: Compared with the control, at the concentration between 10-7-10-5 mol/L, E2 was observed to increase the number of HaCat cells, reduce the doubling time of cell number and increase the cell viability and E6 mRNA expression after transfection (P<0.05). Conclusion: E2 increased the cell growth and viability, enhanced the transfection efficiency of E6 on HaCat cells. The mechanism may be related to the acceleration of cell growth and cell viability. 2018 Vol. 48 (4): 280-283 [Abstract] ( 650 ) HTML (1 KB)  PDF (1323 KB)  ( 915 ) 284 Low tube voltage and low dose contrast agent combined with normal saline in 64 row CT pulmonary angiography: a preliminary study HU Qiaoling, CHEN Xingcan. Objective: To explore the applied value of low tube voltage and low dose contrast agent combined with normal saline in 64 row CT pulmonary angiography. Methods: An experimental study was carried out in 60 patients suspected of pulmonary artery disease by clinic and required CTPA, who were randomly divided as control group and experimental group, with 30 cases in each group. All patients accepted pulmonary artery CTPA by 64 row CT. In control group, tube voltage was 120 kVp, contrast dose was 60 mL, while in experimental group, tube voltage was 80 kVp and contrast dose was 40 mL combined with 20 mL of normal saline, the other parameters being all the same. Independent t-test was used to compare the CT values of pulmonary artery and pulmonary vein, image noise, signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR), volume CT dose index (CTDIvol), dose length product (DLP), effective dose (ED). Chi square test was used to compare the subjective quality between two groups of images. Results: Among 60 patients, 54 completed the examination successfully (28 cases in control group and 26 cases in experimental group), CT values of pulmonary artery, image noise, SNR, CNR of experimental group were higher than those of control group (P<0.05), while CT values of pulmonary vein, CTDIvol, DLP, ED were lower than those of control group (P<0.05), and there was no significant difference in subjective image quality between two groups (P>0.05). Conclusion: Low tube voltage and low dose contrast agent combined with normal saline in 64 row CT pulmonary angiography can produce satisfactory image quality and remarkably decrease the radiation dosage and contrast dose. 2018 Vol. 48 (4): 284-288 [Abstract] ( 722 ) HTML (1 KB)  PDF (1338 KB)  ( 1038 ) 289 Evaluation of left ventricular systolic function in severe aortic stenosis using layer-specific stain LIN Jie, YAN Zhihan Objective: To quantitatively evaluate left ventricular (LV) systolic function in severe aortic stenosis (AS) with normal left ventricular ejection fraction (LVEF) using layer-specific strain (LSS). Methods: Fifty-seven patients with severe AS (including 36 cases with LV wall thickness and 21 cases with normal wall thickness, all patients had LVEF≥55%) and 30 controls from January 2012 to September 2017 in Huangyan Hospital of Wenzhou Medical University were included in this study. The global endocardial (Endo), mid-wall (Mid), epicardial (Epi) longitudinal peak strain (GLS) and circumferential peak stain (GCS) were analyzed. The repeatability of GLS and GCS was evaluated in 20 cases randomly selected from the investigated subjects as the AS and controls. Results: Endo-GLS, Mid-GLS, Epi-GLS in AS with and without wall thickness were significantly reduced compared with the controls (P<0.05); and Endo-GLS, Mid-GLS, Epi-GLS in AS with wall thickness were statistically lower than that with LV normal wall thickness (P<0.05). There was no significant difference in Endo-GCS, Mid-GCS, Epi-GCS between the above three groups (P>0.05). Conclusion: With the increase of LV wall thickness, Endo-GLS, Mid-GLS, Epi-GLS gradually decrease in the AS patients. Endo-GLS, Mid-GLS, Epi-GLS can be used to identify subtle contractile dysfunction triggered by increased LV wall thickness in severe AS with normal LVEF. 2018 Vol. 48 (4): 289-293 [Abstract] ( 707 ) HTML (1 KB)  PDF (1430 KB)  ( 898 ) 294 The application of supplementary extensive surgery for accidental invasive cervical cancer YU Jiangtao, ZHENG Feiyun, ZHAO Hongqin Objective: To explore the clinical pathological features, perioperative complications and prognosis of accidental invasive cervical cancer treated by supplementary extensive surgery. Methods: Retrospective analysis of 6 patients with accidental invasive cervical cancer who accepted supplementary extensive surgery in the First Affiliated Hospital of Wenzhou Medical University from February 2016 to April 2017 and their clinical and pathological factors and prognostic factors were analyzed. The correlation between surgical factors were analyzed by Spearman correlation analysis. Results: The average age of these 6 patients was (50.7±7.5) years. Hysterectomy was carried out in 2 cases of CIN III, 1 case of uterus leiomyoma, 1 case of adenomyosis and 2 cases of cervical cancer in stage Ia1. Five of them had squamous carcinoma and 1 case of adenocarcinoma. There was 1 case in stage Ia1, 2 cases in stage Ia2, 3 cases in stage Ib1, in which patient in stage Ia1 had vascular infiltration. The interval time between the two operations was (23.8±13.9) d. The operative time was (176.0±77.9) min. Intraoperative bleeding was 150 (60, 800) mL. The number of pelvic lymph nodes was 24.5±6.6. The duration of indwelling catheter was (14.3±0.5) d. The time of hospitalization after supplementary extensive surgical was (10.0± 2.4) d. Bladder injury occurred in 1 patient. One patient had vascular infiltration and pelvic lymph node metastasis which was treated by postoperative adjuvant chemo-radiotherapy. One case of cervical adenocarcinoma with 4/5 muscular infiltration after initial operation, the postoperative chemotherapy was given based on platinum. There was significant correlation (r=0.840, P=0.036) between the interval time of the two operations and the time for indwelling catheter after supplementary extensive surgery. Conclusion: Supplementary extensive surgery is difficult but its perioperative complication rate is low and therefore it is feasible to treat the accidental invasive cervical cancer. 2018 Vol. 48 (4): 294-297 [Abstract] ( 698 ) HTML (1 KB)  PDF (1241 KB)  ( 1005 )

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