To establish a HepG2 cell line ,which stably expressing CYP3A29 of Bama miniature pig in order to evaluate the drug metabolic characteristics of this isoenzyme,the gene for this system was obtained through total RNA extraction and RT-PCR assay.the gene was subcloned into plasmid PMD18-T,designated as pMD-CYP3A29.This gene was then amplified by PCR, and cloned into eukaryotic expression vector pcDNA3.1(+),and the recombinant plasmid was designated as pcDNA-CYP3A29. Sequencing was used to confirmed the correctness of the gene of this gene.the expressed gene was then transfected into HepG2 cells by lipid-media transfection and the transformants were screened by G418 for 10 generations ;the expression of CYP3A29 was identified by RT-PCR、West-blot and the metabolic activity of the transformant HepG2-CYP3A29 was verified by nifedipine oxidation.In comparison with HepG2, the transformant HepG2-CYP3A29 showed remarkable oxidative activity.It is apparent that the cell line stably expressing CYP3A29 isoenzyme was successfully established, and it may be used for the metabolic study of related drugs.

Aim To study the protective effects of Xinnaoshenkang (XNSK) against focal cerebral injury caused by ischmia-reperfusion in rats and its mechanism. Methods The focal brain ischmia-reperfusion model in rats was made through by using an intraluminal monofilament to occlude the middle cerebral artery for 1.5 h and then reperfusing for 24h.Spectrophotometric assay was used to measure the contents of malondial-dehyde (MDA), lactic acid (LA),superoxide dismutase(SOD), reactive oxygen species(ROS), nitricoxide synthase(NOS) and several ATPase in cerebral cortex homogenates from rats. The effects on platelet aggregation were also observed. Results Compared with model and positive control groups,88,175,350 mg?kg-1 XNSK groups were found having significant inhibition of cerebral infarction,MDA,LA,ROS,NOS,platelet aggregation and significant increase of the activity of SOD,ATPase. Conclusion XNSK has protective effects against focal cerebral injury caused by ischmia-reperfusion in rats.

Objective:To examine the relationship between sarcopenia and DNA methylation in the promoter of the growth differentiation factor 15(GDF15)gene in elderly individuals.Methods:This cross-sectional study collected data from 865 elderly individuals aged 65 years and above who underwent physical examination at the Yuetang Town Community Medical Center in Yangzhou City between May and September 2020.The verification set included 431 males and 434 females with an age range of 65-100 years and a mean age of(76.0±5.9)years.The diagnosis of sarcopenia was based on the consensus diagnostic criteria of the Asian Sarcopenia Working Group in 2019.The study included 295 cases in the non-sarcopenia group and 470 cases in the sarcopenia group.The study selected 50 non-sarcopenia patients and 50 age-gender matched sarcopenia patients as the explore set for DNA methylation sequencing of the GDF15 gene.The sequencing results were then verified using the methylation-specific polymerase chain(PCR)method in the verification center.Additionally, serum GDF15 levels were detected using the enzyme-linked immunosoradsorption method.The study analyzed the correlation between GDF15 methylation levels, serum GDF15 levels, and sarcopenia.Results:The study found that individuals with sarcopenia had lower levels of body mass index(BMI), appendicular skeletal muscle mass(ASMI), grip strength, and gait speed in both the discovery and validation sets compared to those without sarcopenia( P<0.05). Additionally, the DNA methylation of GDF15 was found to be lower in the sarcopenic group compared to the non-sarcopenic group[93.7%(79.6%, 98.0%) vs.97.7%(95.3%, 99.0%), Z=-9.294, P<0.01]. The results of the correlation analysis indicate a positive relationship between the methylation level and appendicular skeletal muscle mass( r=0.206, P<0.01), grip strength( r=0.297, P<0.01), and gait speed( r=0.383, P<0.01). Conversely, there was a negative correlation between the methylation level and serum GDF15 level( r=-0.249, P<0.05). The study conducted ROC analysis to determine the predictive ability of GDF15 methylation for sarcopenia found that the area under the curve was 0.700 with a cut-off score of 92.7%.Furthermore, binary regression analysis revealed that low GDF15 methylation( OR=1.136, 95% CI: 1.098~1.175, P<0.01)was linked to a higher risk of sarcopenia, even after adjusting for age, sex, and BMI.The serum GDF15 level was higher in the sarcopenic group compared to the non-sarcopenic group[(0.665±0.432)pg/L vs.(0.465±0.211)pg/L( t=-2.452, P<0.05)]. Additionally, it was observed that the methylation of GDF15 was negatively correlated with the serum GDF15 level( r=-0.249, P<0.05). Conclusions:A low level of GDF15 methylation has been found to be linked with a higher risk of sarcopenia, suggesting that measuring GDF15 methylation could potentially serve as a biomarker for diagnosing sarcopenia.

Objective:To investigate the application value of Clavien-Dindo classification in evaluation of postoperative short-term complications of Da Vinci robotic-assisted or laparoscopic-assisted total gastrectomy with D 2 lymphadenectomy. Methods:The retrospective cohort study was conducted. The clinicopathological data of 262 patients with gastric cancer who were admitted to the 940th Hospital of Joint Logistic Support Force of Chinese People′s Liberation Army from January 2016 to January 2019 were collected. There were 214 males and 48 females, aged (58±11) years, with a range from 17 to 81 years. Of 262 patients, 120 cases undergoing Da Vinci robotic-assisted total gastrectomy + D 2 lymphadenectomy + Roux-en-Y anastomosis were divided into robotic group, and 142 cases undergoing laparoscopic-assisted total gastrectomy + D 2 lymphadenectomy + Roux-en-Y anastomosis were divided into laparoscopic group. Observation indicators: (1) intraoperative and postoperative situations; (2) postoperative pathological examination; (3)complications; (4) stratified analysis; (5) follow-up. Follow-up using outpatient examination and telephone interview was performed to detect complications, tumor recurrence and survival of patients within postoperative 2 months. The follow-up was up to May 2019. Measurement data with normal distribution were represented as Mean± SD, and comparison between groups was analyzed using the t test. Measurement data with skewed distribution were represented as M (range), and comparison between groups was analyzed using the Mann-Whitney U test. Count data were described as absolute numbers or percentages, and comparison between groups was analyzed using the chi-square test or Fisher exact probability. Comparison of ranked data between groups was analyzed using the rank sum test. Results:(1) Intraoperative and postoperative situations: cases undergoing conversion to open surgery, the operation time, volume of intraoperative blood loss, the number of lymph node dissected, time to first flatus, time to initial fluid diet intake, duration of postoperative hospital stay of the robotic group were 1, (243±42)minutes, 100 mL(range, 100-150 mL), 38±15, (2.8±1.0)days, 3 days(range, 3-4 days), 11 days(range, 9-13 days), respectively. The above indicators of the laparoscopic group were 2, (244±38)minutes, 100 mL(range, 100-150 mL), 34±14, (3.2±1.0)days, 4 days(range, 3-5 days), 10 days(range, 9-13 days), respectively. There were significant differences in the number of lymph node dissected, time to first flatus, time to initial fluid diet intake between the two groups ( t=2.068, -3.030, Z=-3.370, P<0.05), and there was no significant difference in cases undergoing conversion to open surgery, the operation time, volume of intraoperative blood loss, duration of postoperative hospital stay between the two groups ( χ2=0.000, t=-0.158, Z=-1.824, -0.088, P>0.05). (2) Postoperative pathological examination: cases with well differentiated tumor, moderately differentiated tumor, poorly differentiated tumor, signet ring cell carcinoma or other types of tumor, cases in stage T1b, T2, T3 or T4a (pT staging), cases in stage N0, N1, N2, N3a or N3b (pN staging), cases in stage ⅠB, ⅡA, ⅡB, ⅢA, ⅢB or ⅢC (pTNM staging) of the robotic group were 6, 50, 55, 9, 10, 22, 63, 25, 42, 19, 19, 24, 16, 17, 22, 23, 20, 23, 15, respectively. The above indicators of the laparoscopic group were 4, 42, 84, 12, 6, 18, 81, 37, 39, 27, 32, 19, 25, 13, 19, 28, 39, 16, 27, respectively. There was no significant difference in the above indicators between the two groups ( Z=-1.880, -1.827, -0.140, -1.460, P>0.05). (3) Complications: cases with complication classified as grade Ⅰ, grade Ⅱ, grade Ⅲa, grade Ⅲb, grade Ⅳa, grade Ⅳb of Clavien-Dindo classification, cases with death, cases with overall complications, cases with severe complications of the robotic group were 9, 6, 3, 2, 2, 0, 0, 22, 7, respectively. The above indicators of the laparoscopic group were 12, 15, 9, 6, 3, 1, 1, 47, 20, respectively. There were significant differences in cases with overall complications, cases with severe complications between the two groups ( χ2=7.309, 4.790, P<0.05), and there was no significant difference in cases with complication classified as grade Ⅰ, grade Ⅱ, grade Ⅲa, grade Ⅲb, grade Ⅳa, grade Ⅳb of Clavien-Dindo classification, cases with death between the two groups ( χ2=0.080, 2.730, 1.042, 0.704, 0.000, 0.000, 0.000, P>0.05). (4) Stratified analysis: of the patients with overall complications in robotic group, cases of male or female, cases aged ≥65 years or <65 years, cases with body mass index (BMI) ≥24 kg/m 2 or <24 kg/m 2, cases with tumor diameter ≥5 cm or <5 cm, cases with or without abdominal surgery, cases with tumor located at upper stomach or middle stomach, cases in Ⅰ-Ⅱ grade or Ⅲ grade of American Society of Anesthesiologists (ASA) classification, cases with well differentiated tumor or undifferentiated tumor, cases in stage Ⅰ-Ⅱ or stage Ⅲ (pTNM staging), cases with operation time ≥250 minutes or <250 minutes, cases with volume of intraoperative blood loss ≥150 mL or <150 mL, cases with the number of lymph node dissected ≥25 or <25 were 15, 7, 14, 8, 11, 11, 16, 6, 4, 18, 19, 3, 15, 7, 7, 15, 8, 14, 12, 10, 12, 10, 14, 8, respectively. The above indicators of patients with overall complications in the laparoscopic group were 33, 14, 17, 30, 16, 31, 36, 11, 11, 36, 27, 20, 31, 16, 13, 34, 14, 33, 24, 23, respectively. Of the patients with overall complication, there were significant differences in cases of male, cases aged ≥65 years or <65 years, cases with BMI<24 kg/m 2, cases with tumor diameter≥5 cm, cases without abdominal surgery, cases with tumor located at middle stomach, cases in Ⅰ-Ⅱ grade or Ⅲ grade of ASA classification, cases with well differentiated tumor, cases in stage Ⅲ (pTNM staging), cases with operation time ≥250 minutes, cases with volume of intraoperative blood loss <150 mL, cases with the number of lymph node dissected ≥25 between the two groups ( χ2=6.683, 4.207, 6.761, 7.438, 4.297, 6.325, 9.433, 3.970, 4.850, 4.911, 3.952, 3.915, 6.865, 4.128, P<0.05) and there was no significant difference in cases of female, cases with BMI≥24 kg/m 2, cases with tumor diameter <5 cm, cases with abdominal surgery, cases with tumor located at upper stomach, cases with undifferentiated tumor, cases in stage Ⅰ-Ⅱ (pTNM staging), cases with operation time < 250 minutes, cases with volume of intraoperative blood loss ≥150 mL, cases with the number of lymph node dissected <25 between the two groups ( χ2=0.277, 1.052, 1.996, 1.552, 2.172, 2.594, 2.244, 3.771, 1.627, 3.223, P>0.05). (5) Follow-up: 262 patients were followed up postoperatively for 2 months. During the follow-up, no patient was diagnosed with tumor recurrence, and one patient in the laparoscopic group died of severe infection. Conclusions:The Clavien-Dindo classification can be used in evaluating postoperative short-term complications of Da Vinci robotic-assisted or laparoscopic-assisted total gastrectomy with D 2 lymphadenectomy. Compared with laparoscopic-assisted total gastrectomy with D 2 lymphadenectomy, Da Vinci robotic-assisted total gastrectomy with D 2 lymphadenectomy has the advantages of minimally invasiveness, low incidence of overall and severe complication.