Objective To establish a determining method for the content of berberine hydrochloride in Coptis chinensis Franch.and Xianglian pills.Method The sample was extracted and developed,the berberine hydrochloride was determined by thin layer chromatograph-fluorescence of spectrophotometry with Benzen-ethylaceate-isopropanol-methanol-ammonia water(6:3:1.5:1.5:0.5) as the developing system and Ex=365 nm,Em=409 nm.Result The calibration curve was linear in the range of 0.473～ 3.784 ?g,r=0.999 3.The recovery rate in Coptis chinensis Franch.was 99.03%,RSD=1.86%,and in Xianglian pills was 97.10%,RSD=1.09%.Conclusion The method is simple and accurate.

Objective The objective of this study was to observe effects of miR-409-5p on the proliferation and migration of HepG2 cells.Methods HepG2 cells were transfected with miR-409-5p mimics and verified by Real-Time quantitative PCR.After high expression of miR-409-5p,MTT and wound healing assays were used to detect the proliferation and migration ability of HepG2 cells.Results The cell viability of HepG2 cells transfected with miR-409-5p mimics was significantly decreased in comparison with the control group and showed a dose-and time-dependent manner(P<0.05).The migration ability of cells overexpressing miR-409-5p mimics was significantly lower than that in the NC group(P<0.05).There was significant difference between the two groups at treatments for 24 h and 48 h.Conclusion Up-regulated miR-409-5p can inhibit the proliferation and migration of HepG2 cells.

[Objective]To determine the content of Polygonum multiflorum thunb. Polysaccharide and the refined. [Methods] The sulfuric acid-phenol method was used. [Results] ?_ max = 486nm. The linear rang was 10.5～94.5?g?L -1 .And the average recovery was 101.23%(n=5,RSD=3.03%). The content of Polygonum multiflorum thunb. Polysaccharide was 14.28% and the refined was 19.03%. [Conclusion] The method was simple,rapid and reliable.

Objective To establish a detection method of Salmonella in laboratory animals based on a high-throughput sequencing technology, and to apply it in detection of Salmonella in laboratory animals.Methods DNA samples were extracted from mouse feces.Universal primers for 16S rDNA, 23S rDNA, 16S-23S rDNA, 23S-5S rDNA region, gyrB preferred area were designed, respectively.Each primer was tested and analyzed to determine the best amplification conditions and build a database.Forty-two samples of Salmonella were assayed by Illumina high-throughput sequencing technology and evaluated the specificity and stability of this method.Results The species preferred region of Salmonella was gyrB gene region.The primers for gyrB gene were FP5 ’-AACCACCGCAATCAGACCTT3‘ and FP5 ’-AGCCACGAAACCTTCACYA-3’.The primers were optimized and determined, through a high-throughput sequencing, and the sequence analysis detected very small amount of Salmonella in the 42 samples, indicating that this detection method is stable, highly sensitive, and the limit of detection reached to 0-102 CFU.Conclusions We have established a complete detection system for detection of Salmonella in laboratory animals based on a high-throughput sequencing technology, This system can detect trace amounts of Salmonella in laboratory animals, and this detection method is stable and highly sensitive, which can be also used in detection of other kinds of pathogenic microorganism in laboratory animals.

Objective To explore the effect of electroacupuncture intervention on tumor growth and PCNA expression in HepG2 nude mice.Methods Thirty-two nude mice were randomized to electroacupuncture, electrostimulation, model and blank control groups, 8 mice each. A nude mouse model of subcutaneous tumor was made with HepG2. The electroacupuncture group received electroacupuncture at points Zusanli and Sanyinjiao on bilateral lower limbs and the electrostimulation group, direct electrical stimulation of the same points. The tumor volume, the tumor/weight ratio and the tumor growth inhibition rate were observed. PCNA expression in the tumor tissue was determined.Results There was no statistically significant pre-/post-intervention difference in the tumor volume in the electroacupuncture, electrostimulation and model groups (P>0.05). There was no statistically significant post-intervention difference in the tumor volume between the electroacupuncture or electrostimulation group and the model group (P>0.05), but tumor growth tended to slow in the electroacupuncture group. There was no statistically significant pre-/post-intervention difference in the nude mouse weight in every group (P>0.05). There was a statistically significant post-intervention difference in the tumor/weight ratio between the electroacupuncture group and the electrostimulation or model group (P<0.05), but no statistically significant post-intervention difference in the tumor/weight ratio between the electrostimulation and model groups (P>0.05). There was a statistically significantpost-intervention difference in the tumor growth inhibition rate between the electroacupuncture and electrostimulation groups (P<0.05). There was a statistically significant post-intervention difference in the PCNA expression level between the electroacupuncture or electrostimulation group and the model group (P<0.05), no statistically significant post-intervention difference in the PCNA expression level between the electroacupuncture and electrostimulation groups (P>0.05).Conclusion Electroacupuncture can delay HepG2 tumor growth to a certain extent.

Objective:To study the extraction method of total flavonoids in Mosla hangzhowensis Matsuda.Methods:Orthogonal experiment design was used,with the content of the total flavonoids as the index,to study the effect of ethanol concentration,solvent quantity and extraction time on recovery rate of total flavonoids.Results:The best extraction condition was:extract two times;adding 20 fold 65% ethanol and to extract an hour each time.Conclusion:This extraction method is convenient and feasible.

Objective To investigate the role of mitochondrial cytochrome c on hepatocyte apoptosis in non-alcoholic fatty liver disease in rabbits and its pathogenesis.Methods Forty Japanese white rabbits were randomly assigned to control group and model group.The model group was divided into three subgroups: 4-week, 6-week, and 8-week groups, with 10 rabbits in each group.The model groups were subcutaneously injected with peanut oil (1.2 mL/kg), twice a week for 4 weeks, 6 weeks or 8 weeks.The rabbits of all groups were killed at the right time.Serum samples were collected to detect the serum biochemical index levels.Liver tissue samples were taken for pathological observation using HE staining.The hepatocyte apoptosis index ( AI ) was measured by flow cytometry, and mitochondrial permeability transition pore ( MPTP) was evaluated by ultraviolet spectrophotometry.Immunohistochemistry was employed to detect the hepatic expressions of Bcl-2, Bax, CYT C and caspase-3.Western blot was performed to detect the changes of CYT C and caspase-3 protein expressions.Results The model groups showed hepatic injury and high level of TC, TG, CRP, IL-6 and TNF-αbeginning from 4 weeks.With the NAFLD process, the hepatocyte apoptosis index was significantly increased at 4-8 weeks and the MPTP was gradually increased.In the model group, hepatic Bcl-2, Bax, CYT C and caspase-3 expressions were increased steadily with the time passing.Conclusions NAFLD-induced liver damage is associated with apoptosis, and the mitochondrial cytochrome c-mediated apoptotic pathway plays a role in the occurrence of NAFLD.

Non-alcoholic fatty liver disease (NAFLD) has become one of the major metabolic diseases.In view of the defects of traditional animal models, this study was the first to establish the NAFLD model of Mongolian gerbil (Meriones unguiculatus) with simple feed formula which is similar to human (from simple fatty liver to steatohepatitis, fibrosis,Liver cirrhosis).This study discussed the mechanism of rapid fatty liver deposition in Mongolian gerbil, revealed its molecular mechanism,main regulatory target and network function of fatty liver susceptibility.We provide a new animal model of NAFLD with relatively clear background and less time-consuming for clinical treatment and new drug development.The theoretical and practical basis for the breeding of inbred strain NAFLD gerbil was established.

Objective To establish a method for determination and optimize the extraction process of the content of plantamajoside in plantain.Methods Plantamajoside content was determined by HPLC.The effects of ethanol concentration, ethanol amount and extraction time on the extraction of plantamajoside from plantain were studied by orthogonal design.Results The calibration curve was linear (r=0.999 6) over the range of 12.52-125.10μg/ml.The average recovery was 98.57％ (RSD=1.45％).The optimum extraction process was as follows:60％ethanol, 10times volumes, extracted 2times, 1heach time.Conclusion The established method was simple, accurate and reproducible for determination of the content of plantamajoside in plantain.The optimal extraction process was stable and feasible.

Objective To explore the efficacy and safety of anterior cervical discectomy and fusion assisted with microscope.Methods Thirty-seven patients with cervical spondylosis were included to be retrospectively ana lyzed,including 21 males and 16 females.All these patients had accepted anterior cervical discectomy and fusion (ACDF) assisted with microscope from October,2015 to February,2018,and they were aged from 22 to 77 years old (51.5±6.2 years on the average).In these patients,30 cases were operated on single segment,6 cases were operated on double segments,and 1 case was operated on 3 segments.Among all the patients,15 patients of which (40.54％) had cervical spondylotic myelopathy and 22 patients of which (59.46％) sufferered from cervical spondylotic radicu lopathy.All the operations were performed with a conventional transverse anterior cervical incisions,an intervertebral distractor was placed.The decompression was completed under the microscope,and the fixation was performed under direct vision.Moreover,the operative time,intraoperative blood loss and surgery-related complications were recorded.Follow-up was carried out at different times,including 7 days,1 month,3 months,6 months and every year after operation.Japanese Orthopaedic Association (JOA) score was used to calculate the rate of improvement in neurological function,which can evaluate the clinical efficacy.And cervical dysfunction index (NDI) was used to assess cervical function.Results All patients in this group underwent successful decompression under the microscope.The operation time was 90-160 min,with an average of (110.67±36.42) min;The intraoperative blood loss was 20-110 ml,with an average of (36.00±29.11) ml.All patients were followed-up for 12-31 months,with an average of (15.2±4.7) months.The JOA score improved from 8.37±3.26 preoperatively to 15.96 ± 1.50 at the last follow-up,and its difference had signifi cance in statistics (t=8.592,P=0.000).Neurological function improvement rate could be graded:excellent in 31 cases and good in 6 cases,the excellent and good rate was 100％;NDI was reduced from 19.01 ± 6.47 preoperatively to 5.81 ± 2.58 at the last follow-up,with statistical significant difference (t=5.127,P=0.000).During the follow-up,1 screw was found loosened and slightly withdrawn in 1 female patient at 3 months after operation,of whom had not obvious discomfort.The patient was continuously observed and there was no screw withdrawal again.Moreover,there were no complications such as cerebrospinal fluid leakage,hoarseness and difficulty in swallowing.Conclusion Microscope-assisted ACDF can provide safe and adequate decompression without significantly extending the operation time,which is satisfactory in clinical results.Even in some cases of 1 or 2 segments of intervertebral disc nucleus prolapse,it is possible to avoid a more traumatic ACDF.And it is worthy of clinical promotion.