AIM:To determine four bioactive components in Tongda Granule ( Radix Paeoniae alba,Flos Carthami,Radix Scutellariale,etc. ),namely hydroxy safflower yellow A,peoniflorin,ferulic acid,and baicalin. METHODS:The isolation was performed on an Alltech Apollo C18 (250 mm ? 4. 6 mm,5 ?m) with the mobile phase consisting of methanol-0. 2% (w)phosphoric acid for gradient elution. The flow rate was 1. 0 mL/min and the column temperature was set at 30 ℃. RESULTS:This method was successfully applied to determining four bio-active components in Tongda Granule. The standard curves were linear over the ranges of 5. 20 -104 mg/L for hydroxy safflower yellow A(r =0. 999 1),25. 6 -512 mg/L for peoniflorin (r =0. 999 6),7. 72 -154. 4 mg/L for ferulic acid (r =0. 999 5),32. 5 -650 mg/L for baicalin (r = 0. 999 5),respectively. The average recoveries were 101. 1% ,99. 6% ,98. 9% and 101. 9% ,RSD were 2. 3% ,1. 8% ,2. 6% and 1. 3% ,respectively. CONCLUSION:The method is simple and can be used as a quality control method for Tongda Granule.

We described the first results of a quantitative ultra performance liquid chromatographytandem mass spectrometry method for a novel antimicrobial peptide （phylloseptin, PSN-1）. Chromatographic separation was accomplished on a Waters bridged ethyl hybrid （BEH） C18 （50mm× 2.1 mm, 1.7 μm） column with acetonitrile-water （25：75, v/v） as isocratic mobile phase. Mass spectrometry detection was performed in the positive electrospray ionization mode and by monitoring of the transitions at m/z 679.6/120, 509.6/120 （PSN-1） and m/z 340.7/165 （Thymopentin, IS）. Protein precipitation was investigated and the recovery was satisfactory （above 82%）. The method was shown to be reproducible and reliable with intra-day precision below 5.3%, inter-day precision below 14.2%, and linear range from 0.02 to 2 lag/mL with r〉0.994. The method was successfully applied to a pharmacokinetic study of PSN-1 in rats after intravenous administration.

We described the first results of a quantitative ultra performance liquid chromatographytandem mass spectrometry method for a novel antimicrobial peptide (phylloseptin,PSN-1).Chromatographic separation was accomplished on a Waters bridged ethyl hybrid (BEH) C18 (50 mm × 2.1 mm,1.7 μm) column with acetonitrile-water (25∶75,v/v) as isocratic mobile phase.Mass spectrometry detection was performed in the positive electrospray ionization mode and by monitoring of the transitions at m/z 679.6/120,509.6/120 (PSN-1) and m/z 340.7/165 (Thymopentin,IS).Protein precipitation was investigated and the recovery was satisfactory (above 82％).The method was shown to be reproducible and reliable with intra-day precision below 5.3％,inter-day precision below 14.2％,and linear range from 0.02 to 2 μg/mL with r＞0.994.The method was successfully applied to a pharmacokinetic study of PSN-1 in rats after intravenous administration.

An HPLC method for the determination of 18alpha-glycyrrhetinic acid and 18beta-glycyrrhetinic acid in rat plasma was established, which was used subsequently to determine the pharmacokinetic profiles of both epimers of glycyrrhetinic acid in rats. alpha-glycyrrhetinic acid, beta-glycyrrhetinic acid, and a mixture of alpha-glycyrrhetinic and beta-glycyrrhetinic acids were administered to rats via gastric infusion. Blood samples were collected at different time intervals and extracted by liquid-liquid extraction. Separation was achieved by using a Kromasil C18 column (150 mm x 4.6 mm, 5 microm) with the mobile phase composed of acetonitrile--4 mmol x L(-1) ammonium acetate solution (46 : 54, v/v) at a flow rate of 1.0 mL x min(-1), and the detection wavelength was set at 250 nm. The pharmacokinetic parameters were calculated using the software DAS 2.0. In a combined administration, the main pharmacokinetic parameters of beta-glycyrrhetinic acid are significantly different from that of alpha-glycyrrhetinic acid (P < 0.05), while no significant difference was obtained when administrated individually. Compared to the single administration, significant differences (P < 0.05) on the values of AUC(0-t) and AUC(0-infinity) of beta-glycyrrhetinic acid were observed when this chemical was administrated together with alpha-glycyrrhetinic acid. In contrast, the pharmacokinetic parameters of alpha-glycyrrhetinic acid were not affected even under the co-administration. Here, a sensitive, specific, rapid and reproducible HPLC method was developed for the pharmacokinetic studies of alpha-glycyrrhetinic acid and beta-glycyrrhetinic acid in rat plasma.

Objective To evaluate the association between 8 single nucleotide polymorphisms (SNPs)of vascular endothelial growth factor (VEGF ) gene and the risk of Alzheimer’s disease (AD ) in Shaanxi Han population.Methods We examined the potential association between AD and 8 SNPs of VEGF gene using the MassARRAY system.The participants enrolled in this study included 214 patients with AD and 249 healthy controls from Shaanxi Han population.SPSS16.0 and Haploview 4.2 were employed to analyze differences in genotypes, alleles and haplotypes between the two groups.Results The results showed that rs3025039 (3’UTR)were significantly associated with AD (P<0.05).Greater frequency of rs3025039 T allele (P=0.008,OR=1.527,95%CI=1.116-2.088)was found in AD subjects.Furthermore,strong linkage disequilibrium (LD)was observed in 2 locks (block1:rs699947-rs1570360-rs2010963;block 2:rs3024997-rs3024998-rs3025006)(D’>0.9).There were no significant haplotypes in block 1 and block 2 (P=0.034)found between the patients and controls.Conclusion These findings point to the role for VEGF gene polymorphisms (rs3025039)in AD of a Shaanxi Han population. Individuals with T allele of rs3025039 may be at a higher risk for AD.

ABSTRACT:Objective To study the effect of curcumin on the learning and memory ability in a rat model of Alzheimer’s disease (AD).Methods AD rat model was prepared using intraventricular injection of Aβ1-42. Curcumin was acutely (single injection before the behavioral tests)or chronically (injected for 6 consecutive days) injected intraperitoneally at doses of 50,100 or 300 mg/kg.Brain-derived neurotrophic factor (BDNF)protein (1 μg/side)or BDNF shRNA (2×10 5 units/side)was infused into the hippocampus.The behavioral changes in Y-maze,open field test and Morris water maze and the expression of BDNF in the hippocampus were analyzed. Results Acute treatment with curcumin had no significant effects on the spontaneous alteration,locomotor activity or water maze latency of AD rats.AD rats treated chronically with curcumin (300 mg/kg ) showed significant elevation in the spontaneous alternation (P <0.000 1)in Y-maze and memory ability in the water maze test (P <0.05 )compared with those in the saline group.Chronic treatment with 100 and 300 mg/kg of curcumin induced an increased level of BDNF in the hippocampus as compared with the saline controls (P <0.05 and <0.000 1). Intrahippocampal injection of BDNF significantly decreased the escape latency of AD rats in the water maze (F 4,2 9 5=5.813,P <0.01 ).Rats chronically injected with curcumin combined with shBDNF showed no difference in the swimming time in Ⅱ quadrant as compared with saline controls (P =0.657).However,rats in 100 mg/kg curcumin group,BDNF group and sham group had significantly increased swimming time than the saline controls (P <0.05, P <0.05 and P <0.000 1,respectively).Conclusion Curcumin may activate the downstream signaling pathways by upregulating the expression of BDNF and ultimately contribute to the improvement of learning and memory in AD rats.

Objective:To study the adverse reactions (ADRs) of recombinant human erythropoietin (rHuEPO) to provide refer-ence for clinical rational and safe medication. Methods:ADRs induced by rHuEPO reported at home and abroad were collected and analyzed in respects of age,gender,original illness, occurrence time, clinical manifestations and the results. Results: After the re-trieval,there were 149 cases of rHuEPO-induced ADRs with the damage of cardio vascular system, hematologic system, skin and its appendents accounting for 43.4%,20.8% and 12.7%,respectively. The top three main clinical manifestations of rHuEPO drug reac-tions were hypertension,pure red-cell aplastic anemia (PRCA) and hyperkalemia. The occurrence time should be paid particular at-tention in 5-12 weeks after the administration (43.0%). Conclusion:Physicians should be aware of rHuEPO-induced ADRs (espe-cially the occurrence time),pay attention to patients' medication education and avoid serious adverse reactions.

Objective:To investigate the changes of circulating tumor DNA (ctDNA), circulating B cell-specific Moloney leukemia virus insertion site 1 mRNA (Bmi-1 mRNA) and microRNA-21 (miR-21) before and after treatment with epidermal growth factor receptor (EGFR) monoclonal antibody in advanced colorectal cancer, and analyze their association with treatment response.Methods:The clinical data of 98 patients with advanced colorectal cancer from March 2019 to March 2021 in Yantai Yuhuangding Hospital were retrospectively analyzed. After treatment with cetuximab, complete remission was in 4 cases, partial remission in 26 cases, stable disease in 39 cases, and progressive disease in 29 cases. The patients with complete remission and partial remission were classified as remission group (30 cases), the stable disease and progressive disease were classified as non-remission group (68 cases). Before treatment and after 2 cycles of treatment, the plasma level of ctDNA was detected by high-throughput sequencing; the levels of Bmi-1mRNA and miR-21 were detected by real-time fluorescence quantitative polymerase chain reaction. Spearman correlation was used to analyze the relationship between ctDNA, Bmi-1mRNA, miR-21 and treatment responsiveness after 2 cycles of treatment; multivariate Logistic regression was used to analyze the independent risk factors affecting treatment responsiveness; receiver operating characteristic (ROC) curve was drawn to evaluate the value of ctDNA, Bmi-1mRNA and miR-21 in predicting remission after 2 cycles of treatment.Results:There were no significant differences in ctDNA, Bmi-1mRNA and miR-21 before treatment between 2 groups ( P>0.05); the ctDNA, Bmi-1mRNA and miR-21 after 2 cycles of treatment in remission group were significantly lower than those in non-remission group: (10.03 ± 3.32) μg/L vs. (15.33 ± 5.14) μg/L, 0.13 ± 0.04 vs. 0.19 ± 0.05 and 0.81 ± 0.26 vs. 1.08 ± 0.24, and there were statistical differences ( P<0.01). Spearman correlation analysis result showed that ctDNA, Bmi-1mRNA and miR-21 were negatively correlated with treatment response ( r = -0.500, -0.506 and -0.531; P<0.01). Multivariate Logistic regression analysis result showed that, after controlling for the number of distant metastatic organs and clinical stage, ctDNA, Bmi-1mRNA and miR-21 were still independent risk factors for treatment response in patients with advanced colorectal cancer ( OR = 3.342, 2.725 and 1.838; 95% CI 3.116 to 3.584, 2.647 to 2.805 and 1.768 to 1.911; P<0.01). ROC curve analysis result showed that the area under the curve (AUC) of ctDNA, Bmi-1mRNA combined with miR-21 after 2 cycles of treatment to predict the treatment response was the largest with 0.922. Conclusions:The changes of ctDNA, Bmi-1mRNA and miR-21 in patients with advanced colorectal cancer before and after treatment with EGFR monoclonal antibody are related to the treatment response. Combined detection is helpful for screening patients sensitive to EGFR-targeted therapy, and can provide reference for new targets of molecular intervention.

Objective:To investigate the effect of insular involvement on the outcomes of patients with acute anterior circulation ischemic stroke.Methods:Patients with acute anterior circulation ischemic stroke admitted to the Department of Neurology, Nanjing Jinling Hospital, Affiliated Hospital of Medical School, Nanjing University from January 2015 to December 2020 were retrospectively included. Demographic data, vascular risk factors, clinical and laboratory data, as well as treatment and outcomes were collected. Firstly, the correlation between the insular involvement and the outcomes was investigated, and then the bootstrap method was used to clarify the mediating role of infarct volume between the insular involvement and the poor outcomes.Results:A total of 450 patients with acute anterior circulation ischemic stroke were enrolled, among whom 79 cases (17.6%) had insular involvement and 41 (9.1%) had left insular involvement. There were 111 (24.7%) with poor outcomes, including 5 (1.1%) died. Compared to the non-insular involvement group, the insular involvement group had a higher proportion of patients with atrial fibrillation, shorter onset to door time, higher neutrophil-to-lymphocyte ratio (NLR), higher National Institutes of Health Stroke Scale (NIHSS) score at admission, larger infarct volume, and higher proportion of patients with poor outcomes (all P<0.05). In addition, patients with left insular involvement were younger than those with right insular involvement, had a higher baseline NIHSS score, a lower proportion of patients with minor stroke (NIHSS score ≤8), and had a longer onset to door time (all P<0.05). Compared to the good outcome group, the poor outcome group was older, with a higher proportion of female patients, higher systolic blood pressure, blood glucose, NLR, and NIHSS scores at admission, larger infarct volume, and a higher proportion of patients with insular involvement (all P<0.05). Mediation analysis suggested that the mediating effect of infarct volume between the insular involvement and the poor outcomes was significant (95% confidence interval 0.033-0.230; P=0.008). Conclusions:insular involvement in patients with acute anterior circulation ischemic stroke is associated with the poor outcomes, and this association may be mediated by infarct volume. Patients with left insular involvement may have more severe symptoms than those with right insular involvement, but there is no significant difference in the outcomes.

Objective:To investigate the immunophenotypic and molecular biological characteristics of patients with elevated serum alpha-fetoprotein (AFP) and enteroblastic differentiated gastric adenocarcinoma (GAED).Methods:The clinicopathological data of 13 patients with elevated serum AFP and GAED admitted to Shanxi Cancer Hospital from 2018 to 2020 were collected. Immunohistochemistry (IHC) and next-generation sequencing (NGS) were used to analyze the immune markers and molecular biological characteristics of the pathological tissues of the patients. Kaplan-Meier method and log rank test were used for survival analysis.Results:Among the 13 patients with GAED, 12 were male and 1 was female, aged 41-70 years, with a median age of 64 years. The lesions were mainly located in the gastric antrum (5 cases) and gastric body (4 cases). IHC results showed that the tumor embryonic protein (AFP, SALL4, GPC3), intestinal epithelial differentiation protein (CDX-2, CD10), and some original intestinal epithelial phenotype markers (OCT3/4, Claudin6) were expressed in the tumor tissues. Combined application of multiple markers can reduce the rate of missed diagnosis. Among the 13 patients, 12 had at least one mutation (1 mutation: 1 case, 2-5 mutations: 3 cases, 6-15 mutations: 8 cases), and 1 case was not detected. The gene with the highest mutation frequency was TP53 (10 cases), and other mutant genes included EPHB1 (3 cases), ATRX (2 cases), EPHA5 (2 cases), GATA3 (2 cases), LRP1B (2 cases) and MAP2K4 (2 cases) were also detected. Three of the 13 patients had structural variations, which were C14orf177- GNAS, AIM1- FGFR3, and EPHA6- ROS1 gene rearrangements. All 13 patients had copy number variation, and 11 patients had copy number variation of more than 2 genes. The common amplification genes were IRS2 (5 cases), PTEN (5 cases), GNAS (4 cases), CCNE1 (3 cases), CEBPA (3 cases), PCK1 (3 cases) and ERBB2 (2 cases). The common deletion genes were SOX2 (5 cases) and MYC (5 cases). Among the 13 patients, 4 died, and 2 of the dead patients had liver metastasis. There were 4 patients with disease-free survival and 5 patients with disease progression, including 3 cases of abdominal metastasis and 2 cases of liver metastasis. The 3-year survival rate of patients was 65.9 %, and the 3-year progression-free survival rate was 30.7 %. Gene LRP1B point mutation was associated with poor prognosis ( P＜0.001). There was no significant improvement in the prognosis of patients treated with immunotherapy compared with those treated with chemotherapy alone ( P=0.595), but the prognosis of patients treated with postoperative chemotherapy or postoperative chemotherapy plus immunotherapy was better than that of patients treated with surgery alone ( P＜0.05). Conclusions:Elevated serum AFP with GAED is a highly invasive tumor with unique molecular characteristics, often accompanied by multiple molecular events. TP53 mutation is the most common type of gene mutation. In addition, some cases are accompanied by HER2 amplification and gene rearrangement.