1.Role of endoplasmic reticulum stress in ketamine-induced apoptosis in rat neurons: an in vitro experiment
Huan CHEN ; Di WEN ; Chunling MA ; Guanjie XU ; Qiongmei GUO ; Changhao ZHOU
Chinese Journal of Anesthesiology 2016;36(10):1225-1228
Objective To evaluate the role of endoplasmic reticulum stress in ketamine-induced apoptosis in rat neurons.Methods Rat adrenal pheochromocytoma cell line (PC12 cells) was seeded in the culture dishes 100 mm in diameter (10 ml/dish) or in 6-well plates (2 ml/well) at a density of 5 × 105 cells/ml.PC12 cells were divided into 4 groups (n =6 each) using a random number table:control group (group C);ketamine group (group K);endoplasmic reticulum stress inhibitor salubrinal group (group S);ketamine + salubrinal group (group K+S).In group C,the cells were cultured in the plain culture medium.In group K,1.5 mmol/L ketamine was added.In group S,30 mmol/L salubrinal was added.In group K + S,1.5 mmol/L ketamine and 30 mmol/L salubrinal were added.At 24 h of incubation,the cell morphology was observed under light microscope,the expression of Bip and caspase-12 in PC12 cells was detected by Western blot,and the cell apoptosis was measured by flow cytometry.The apoptosis rate was calculated.Results Compared with group C,the expression of Bip and caspase-12 was significantly upregulated,and the apoptosis rate was increased in K and K + S groups (P < 0.05),and no significant change was found in the parameters mentioned above in group S (P> 0.05).Compared with group K,the expression of Bip and caspase-12 was significantly down-regulated,and the apoptosis rate was decreased in group K+S (P<0.05).The degree of damage to PC12 cells was more serious in group K than in group C..The degree of damage to PC12 cells in group K+S was significantly mnilder than that in group K and more serious than that in group C.Conclusion The mechanism by which ketamine induces neuronal apoptosis is related to the enhancement of endoplasmic reticulum stress in rats.
2.Simultaneous Determination of the Contents of 5 Indicator Components in Yanyan Tablets by QAMS
Changhao MA ; Huaiwei LI ; Guiping TONG ; Haichun MA ; Siteng FENG
China Pharmacy 2019;30(16):2232-2236
OBJECTIVE: To establish QAMS method for content determination of paeoniflorin, rutin, oroxin B, baicalin and cinnamates in Yanyan tablets. METHODS: HPLC method was adopted. The determination was performed on Hypersil GOLD-C18 column (250 mm×4.6 mm,5 μm) with mobile phase consisted of methanol-0.35% phosphoric acid solution (gradient elution) at flow rate of 1 mL/min. The detection wavelengths were set at 280 nm (rutin, oroxin B, baicalin, cinnamates) and 230 nm (paeoniflorin). The column temperature was 30 ℃, and sample size was 10 μL. Using paeoniflorin as internal reference, relative correction factors (RCF) of rutin, oroxin B, baicalin and cinnamates were established. Effects of different chromatogram system, chromatogram column, mobile phase proportion, flow rate and column temperature on relative correction factors were investigated; the chromatographic peaks of the components were located according to the relative retention time. The content of paeoniflorin as internal reference was determined by external standard method, and the other four components were determined by QAMS, and then compared with the results of external standard method. RESULTS: The separation degree of each component to be measured was greater than 1.5. The linear range was 3.97-119.22 μg/mL for paeoniflorin,1.96-58.68 μg/mL for rutin,2.39-71.64 μg/mL for oroxin B, 1.92-57.51 μg/mL for baicalin, 0.54-16.24 μg/mL for cinnamates(r≥0.999 7), respectively. RSDs of precision, reproducibility and stability tests were all lower than 2%. Average recoveries were 97.20%-98.07%(RSD<3%,n=6). RCFs of rutin, oroxin B, baicalin and cinnamates were 0.554 6,1.815 6,2.489 3 and 5.423 2, using paeoniflorin as internal reference. RSDs of RCF and relative retention time were all lower than 5% under different chromatogram conditions. Absolut relative error of four components (except for internal reference) in 10 batches of Yanyan tablets sampled by QAMS and external standard method were all less than 1%. The results of the two methods were identical. CONCLUSIONS: The established method is accurate, rapid, efficient and inexpensive, and it can be used for simultaneous determination of 5 indicator components in Yanyan tablet.
3.Effect of microRNA-1290 on invasion and metastasis of pancreatic cancer
Zili CHEN ; Yifei MA ; Yaozhen PAN ; Chao YU ; Huajian GU ; Changhao ZHU ; Chengyi SUN
Chinese Journal of Hepatobiliary Surgery 2019;25(6):457-461
Objective To investigate the expression of microRNA-1290 in pancreatic cancer and its role in invasion and metastasis of pancreatic cancer.Methods The expression of microRNA-1290 in pancreatic cancer tissue microarray and pancreatic cancer cell lines (AsPC-1,BxPC-3,Capan-2,Panc-1,and MIA PaCa-2) were detected by immunohistochemistry and QT-PCR.The pancreatic cancer cell lines Panc-1 and MIA PaCa-2 in logarithmic growth phase were treated with microRNA-1290 inhibitor,and the invasion and metastasis ability of pancreatic cancer cells were detected by Transwell and wound healing asssay.Western Blot was used to detect the expression of invasion and metastasis-associated proteins cyclooxygenase 2 (COX-2) and matrix metalloproteinase 2(MMP-2) in pancreatic cancer cell lines.Results (1) The expression of microRNA-1290 in pancreatic cancer tissues was significantly higher than that in normal pancreatic tissues and adjacent tissues (P < 0.05).(2) Compared with pancreatic normal epithelial cells (HPDE),the expression of microRNA-1290 was significantly higher in different pancreatic cancer cell lines (P < 0.05).The expression level of MicroRNA-1290 in Panc-1 and MIAPaCa-2 pancreatic cancer cells was significantly higher than that in other pancreatic cancer cell lines (P < 0.05).(3) The number of invasive and metastatic cells was significantly decreased after treatment with microRNA-1290 inhibitor (P <0.05).(4) The expression of MMP-2 and COX-2 were decreased in Panc-1 and MIAPaCa-2 pancreatic cancer cells treated with MicroRNA-1290 inhibitor.Conclusion The expression of MMP-2 and COX-2 may be involved in the invasion and metastasis of pancreatic cancer cell by regulating the expression of microRNA-1290 in pancreatic cancer.