We prepared protoplasts from Salvia miltiorrhiza Bunge suspension culture cells. Then, the protoplasts' vitality and functions were tested by fluorescein diacetate staining method and Fluo-3/AM flourescent probe. The optimal condition of protoplast isolation was Cellulase R-10 1.5%, Pectinase Y-23 0.3%, Macerozyme R-10 0.5%, 40 r/min 12 h, 600 r/min 5 min, and the protoplasts yield was 1.1x10(6) cells/g FW, the vitality was more than 95% by using fluorescein diacetate staining method. It has been confirmed that calcium fluorescent probe Fluo-3/AM can be successfully loaded into protoplasts.
Aniline Compounds ; chemistry ; Cell Culture Techniques ; Cellulase ; chemistry ; Fluorescent Dyes ; chemistry ; Protoplasts ; chemistry ; Salvia miltiorrhiza ; growth & development ; Xanthenes ; chemistry

Objective To investigate the efficacy of TACE combined with radiofrequency ablation (RFA) in treatment of advanced hepatocellular carcinoma.Methods A total of 72 patients with advanced hepatocellular carcinomas were analyzed retrospectively,including 35 patients underwent TACE combined with RFA (combined group),37 patients underwent single TACE (control group).After the operation,the short-term effect,the changes of liver function,serum alpha-fetoprotein (AFP) level,the complication and the long-term survival rates of the two groups were compared.Results The total effective rate of the combined group (29/35,82.86％) was significantly higher than that of control group (20/37,54.05％;P=0.009).In the combined group,the AFP reduced to (102.19±32.13)μg/L,and the control group reduced to (218.46±49.87)μg/L,which had statistical difference (P＜0.001).The survival rates of 1-year,2-year and 3-year in the combined group were 82.86 ％,54.29 ％,34.29 ％ with a median survival time of 25 months;while in the control group those were 54.05％,32.43％,13.51％ with a median survival time of 16 months;there were statistically significant differences in the survival rate between two groups (P=0.009).After treatment,the hepatic functions of both group had a transient change,and 2 weeks after the operation,there was no significant difference between the two groups (all P＞0.05).Conclusion The combination of TACE and RFA is an effective method for the treatment of advanced hepatocellular carcinoma.

Cerebrospinal Fluid Otorrhea ; surgery ; Craniotomy ; Encephalocele ; Humans ; Mastoid ; surgery

Linalool, as a major volatile compound, is widely distributed in natural plant essential oil. In addition, it can also be artificially synthesized. Linalool is used frequently as an important ingredient of perfumes and household detergents. It is still employed in food flavor and industries. Besides, linalool has some positive effect on healthcare. Many studies have showed that linalool exhibited a variety of pharmacological activities, including analgesic, anxiolytic, sedative, anti-inflammatory, anti-tumor and anti-bacterial effects. Therefore, linalool will be a promising agent for clinical application. This article reviews the pharmacological effects and formulation studies of linalool so as to provide a theoretical basis for its further development and utilization.
Animals ; Anti-Anxiety Agents ; chemistry ; pharmacology ; Anti-Inflammatory Agents ; chemistry ; pharmacology ; Chemistry, Pharmaceutical ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Humans ; Hypnotics and Sedatives ; chemistry ; pharmacology ; Monoterpenes ; chemistry ; pharmacology

Objective To compare two different dumping cooking methods (boiling vs.frying) in their effect on postprandial glucose level in diabetic patients using continuous glucose monitoring (CGM).Methods 10 type 2 diabetes mellitus (T2DM) in-patients in the Department of Endocrinology of Peking Union Medical College Hospital between February and May 2011 were enrolled,whose fasting and preprandial glucose levels were controlled with a insulin pump.On day 2 and day 4 in the study period,the patients were given fried dumplings and boiled dumplings for lunch respectively,with the same nutrient contents.The starch digestibility of these two kinds of dumplings were compared using in vitro resistant starch digestion,measuring the concentrations of rapidly digestible starch,slowly digestible starch,and resistant starch.CGM was used to record blood glucose changes,in order to evaluate glycemic effect of these two dumpling cooking methods on postprandial glucose levels at 9 time points (0,15,30,60,90,120,150,180,and 240 minutes),peak blood glucose,and area under the curve (AUC) in 4 time periods (0-60,63-120,123-180,and 183-240 minutes).Results The percentage of rapidly digestible starch was remarkably lower in fried dumplings than in boiled dumplings (30.8％ vs.77.0％),but the content of slowly digestible starch in fried dumplings was higher than that in boiled ones (63.7％ vs.20.7％),and the content of resistant starch in both dumplings were similarly low (1.9％ and 2.3％).The average time to the peak glucose value was shorter in fried dumplings compared with boiled dumplings [(93 ± 53) minutes vs.(156 ± 61) minutes,P =0.02],but the average glucose levels at all the 9 time points and the AUC in all the 4 time periods were not significantly different (all P ＞ 0.05).Conclusions Compared with fried dumplings,boiled dumplings show faster starch digestion,but long time to the peak postprandial glucose level.Fried dumplings may raise the glucose level faster than boiled dumplings do in T2DM patients.

Objective To detect the size distribution and Zeta potential of LHRH-MPG△NLS/CDK-siRNA nanoparticles,to observe the effect of different solvents on the nanoparticle size,and to investigate the inhibitory effect of nanoparticles on HepG2 cell growth.Methods LHRH-MPG △NLS and CDK2-siRNA were mixed by continuous stirring to form nanoparticles at different N/P ratios (10/1,20/1 and 40/1).The size distribution and Zeta potential of LHRH-MPG△NLS/CDK2-siRNA nanoparticles were detected by dynamic light scattering,and the stability of the nanoparticles in normal saline,10％ glucose and pure water was discussed.Finally,the inhibitory effect of the nanoparticles on HepG2 cells was determined by CCK8 kit.Results The mean size of the nanoparticles was within 200 nm,and the Zeta potentials were (70±5) mV (N/P=10/1),(120±5) mV (N/P=20/1) and (130±5) mV (N/P=40/1),respectively.The size of the nanoparticles in normal saline was significantly increased,which demonstrated that strong electrolytes had a great impact on the nanoparticles size.When nanoparticle concentration was 200 nmol/L,LHRH-MPG△NLS/CDK2-siRNA nanoparticles (N/P=10/1) showed significantly inhibitory effect on HepG2 cell growth.Conclusions The mean size of the LHRH-MPG△NLS/CDK2-siRNA nanoparticles was within 200 nm,which was ideal for cellular uptake.The Zeta potential of nanoparticles revealed that nanoparticles could be stable in aqueous solution,while strong electrolytes would affect nanoparticle size.When nanoparticle concentration was 200 nmol/L,LHRH-MPG△NLS/CDK2-siRNA nanoparticles (N/P=10/1) showed significantly inhibitory effect on HepG2 cell growth.

Objective This study explored the expression of polyclonal protein SUZ12 in patients with intrahepatic cholangiocarcinoma(ICC),and its role in predicting the survival and treatment of ICC patients.Methods The expression of SUZ12 and p16INK4a was detected by immunohistochemical assay in 207 liver tissue samples including ICC patients,BilIN-1,-2,-3 and non-tumor-like cholangiocarcinoma.The expression of these proteins was assessed to be related to the pathological characteristics of the ICC patients receiving chemotherapy and the outcome of survival as well as the subsequent chemotherapy response.Results The expression level of SUZ12 was gradually increased from non-neoplastic bile duct tissue to BilIN-1,-2,-3 and ICC.The expression of p16INK4a protein was expressed in non-neoplastic-like cholangiocarcinoma,but it decreased gradually in BilIN-1,-2,-3 and ICC tissues.SUZ12 expression was associated with undifferentiated ICC,lymph node metastasis and advanced cancer.Kaplan-Meier curve analysis showed that ICC patients with high expression of SUZ12 had a significant reduction in overall survival and disease-free survival in comparison with ICC patients with the low expression of SUZ12.SUZ12 expression was significantly associated with overall survival of patients receiving adjuvant gemcitabine-based chemotherapy(AGC).Conclusion SUZ12 expression is able to predict the overall survival and disease-free survival of ICC patients with adjuvant gemcitabine-based chemotherapy.

OBJECTIVETo detect protein expression of ERK(1), ERK(2), JNK(1), p38 and MEK(1), MEK(2) in human hepatocellular carcinoma and adjacent non-neoplastic liver.

METHODSIn 16 surgically resected hepatocellular carcinoma and para-carcinoma tissues, Western blotting was used to detect expression of ERK(1), ERK(2), JNK(1), p38 and MEK(1), MEK(2).

RESULTSIn all cases, ERK(1), ERK(2), p38 expression in hepatocellular carcinoma was significantly higher than that in para-carcinoma: integral optic density (IOD) of ERK(1) was 300 +/- 98 in carcinoma and 98 +/- 48 in para-carcinoma tissues (t = 2.519, P < 0.01); IOD of ERK(2) was 587 +/- 83 in carcinoma and 232 +/- 96 in para-carcinoma tissues (t = 2.745, P < 0.01); IOD of p38 was 270 +/- 85 in carcinoma and 107 +/- 88 in para-carcinoma tissues (t = 2.491, P < 0.01). JNK(1) expression in hepatocellular carcinoma was significantly lower than that in para-carcinoma; IOD of JNK(1) was 111 +/- 93 in carcinoma and 292 +/- 109 in para-carcinoma tissues (t = 2.473, P < 0.01). Protein levels of MEK(1) and MEK(2) in carcinoma were significantly higher than in para-carcinoma. IOD of MEK(1) was 1 418 +/- 244 in carcinoma and 806 +/- 90 in para-carcinoma tissues (t = 2.546, P < 0.01). IOD of MEK(2) was 1 041 +/- 122 in carcinoma and 468 +/- 40 in para-carcinoma tissues (t = 2.861, P < 0.01).

CONCLUSIONSERK(1), ERK(2), MEK(1) and MEK(2) in the signal transduction pathway for cell proliferation are significantly overexpressed and the expression of JNK(1) is lower in hepatocellular carcinoma. Their unbalance is one of the important reasons for the over growth and infinite proliferation of the hepatocellular carcinoma cell. The p38 and JNK(1) may be activated by different pathway.

Adult ; Aged ; Carcinoma, Hepatocellular ; enzymology ; Enzyme Activation ; Female ; Humans ; JNK Mitogen-Activated Protein Kinases ; Liver Neoplasms ; enzymology ; MAP Kinase Kinase 1 ; Male ; Middle Aged ; Mitogen-Activated Protein Kinase Kinases ; analysis ; Mitogen-Activated Protein Kinases ; metabolism ; Protein-Serine-Threonine Kinases ; analysis

Objective:To investigate the impact of aging on the correlation between the intestinal microorganism Akkermansia and obesity, and to analyze the features of the correlation in the elderly population. Methods:This was a cross-sectional study.A total of 6896 cases were collected from the Guangdong intestinal microbiome in 2018, aged 18-94 years old, including 3806 females, 1641 cases with abdominal obesity(23.7%)and 707 cases with systemic obesity(10.3%). The 16S rRNA sequencing data were from individuals of Cantonese descent.The abundance of Akkermansia was calculated after data cleaning, clustering and annotation.The type of abdominal obesity or systemic obesity was diagnosed based on the standards of the Working Group on Obesity in China(2002). According to the five quintiles of the abundance of Akkermansia, subjects were divided into Q1~Q5(Q1-Q4: n=1379, Q5: n=1380). Logistic regression was used to study the relationship between Akkermansia and obesity after adjusting for common confoundors such as gender.Subjects were subgrouped into two types of age groups: the <65 group(n=5467)and the ≥65 group(n=1519); the <70 group(n=6136)and the ≥70 group(n=850). Age windows were used to analyze changes in characteristics of this relationship with increasing age. Results:There were significant differences in age and gender among different Akkermansia groups( t/ χ2=3.51, -5.03, P<0.01). Logistic regression analysis showed that after adjusting for two main confounding factors, age and gender, the risk of systemic obesity and abdominal obesity gradually decreased from Q2 to Q5 group, compared with Q1 group( P<0.001). The correlation between Akkermansia and obesity decreased with age.The protective effect of Akkermansia on obesity was weaker in the ≥65 and ≥70 groups, respectively, than in the <65 and <70 groups. Conclusions:Akkermansia is a protective factor for obesity, but the protective effect is affected by aging and weakened in the elderly.

Objective To find out the importance of human bocavirus (HBoV) as an infectious agent for population in Beijing, China. Seroprevalence study was conducted by using expressed recombinant major capsid VP2 protein as an antigen.Methods Serum specimens collected from infants and children who visited the Children's Hospital Affiliated to the Capital Institute of Pediatrics for health check-up and adults visiting the Xuanwu Hospital, Beijing for diseases other than respiratory infections from April 1996 to March 1997 were used for the investigation. The major capsid protein VP2 from HBoV was expressed in E. coli strain BL21 (DE3) with the transformed PET30b vector inserted with full-length VP2 gene of HBoV and the specific antigenicity of this expressed protein was validated by previous study. Western blotting was used to detect specific IgG antibody against HBoV in collected serum specimens diluted to 1:200. Mock expressed protein was E. coli cells strain BL21 (DE3) with the transformed PET30b vector without insert. Anti-His monoclonal antibody and rabbit anti-HBoV VP2 polypeptides hyper-immune serum were used as positive control for antibody detection.Results Out of 677 serum specimens tested, 400 (59.1% ) were positive for HBoV by Western blotting. About 45.3% (34/75) of the newborns under 1 month of age had anti-HBoV antibodies, and antibody positive rates were decreased in age groups of 1 and 2 months (41.4% and 31.3%, respectively) then increased in the following ages from 6 months to 7 years old ( from 45.6% to 69.7% ). The antibody positive rates were maintained at a relatively constant level ( about 70% ) in the age groups from 7 years to 40 years of age and became lower ( 61.8% - 62. 8% ) in those over 50 years.Conclusions The high seroprevalence of antibody against recombinant HBoV VP2 protein and early age antibody acquisition indicate that HBoV has been circulating in population of Beijing, China as early as in 1996 and most of children had been exposed to HBoV by the age of 7 years. Infants under the age of 6 months were susceptible to this virus.