As a part of the project for the Chinese Pharmacopoeia (2015 edition), the quality standard of Sophora flavescens root extract was investigated and established. According to the methods described in the Appendix of Chinese Pharmacopoeia (2010 edition), the water and ash inspections were carried out. The marker components trifolirhizin, sophoraflavanone G, oxymatrine and oxysophocarpine in the samples were identified by qualitative TLC. The determination of oxymatrine, matrine, oxysophocarpine and sophocarpine was conducted by HPLC and the total flavonoids were measured by ultraviolet spectrophotometry, using sophoraflavanone G as reference substance. The results indicated the spots on the plate were clear with good resolution and the contents of oxymatrine, matrine, oxysophocarpine and sophocarpine in the 13 batches of the samples were 3.87% - 11.1%, 0.970% - 4.33%, 1.30% - 2.59% and 0.260% - 1.14%, respectively. The total flavoids in the 13 batches of the samples were 3.88% - 7.93%. In the study, the validated methods were reproducible and the established quality standard was feasible, which could be used for the quality control of S. flavescens root extract and related preparations.
Chromatography, High Pressure Liquid ; Flavonoids ; analysis ; Plant Extracts ; analysis ; Plant Roots ; chemistry ; Sophora ; chemistry

This study aims to investigate the serotype distribution of non-polio enterovirus (NPEV) isolated from patients with acute flaccid paralysis (AFP) during 2011-2012 in Hebei Province, China and to analyze the relationship between these viruses and AFP. NPEV strains were isolated from the stool specimens from AFP cases in Hebei using human rhabdomyosarcoma cells (RD) and the mouse cell line expressing the gene for the human cellular receptor for poliovirus (L20B) according to the WHO requirements. The nucleotide sequence of VP1 region was determined, and the serotypes of NPEV were identified by molecular typing. The results showed that among the 82 strains of NPEV isolated from the AFP cases during 2011-2012, 42 isolates (55.3%) were identified as human enterovirus A (HEV-A), which were classified into 4 serotypes, 34 (44.7%) as human enterovirus B (HEV-B), which were classified into 13 serotypes, 2 as adenovirus, and 4 were untyped; human enteroviruses C and D were not found in these cases. Enterovirus A71 (EV-A71) was the main type of HEV-A, accounting for 85.7% of all HEV-A strains. HEV-A, especially EV-A71, was predominant among the NPEV strains isolated from AFP patients during 2011-2012 in Hebei Province.
Acute Disease ; China ; epidemiology ; Enterovirus ; classification ; physiology ; Humans ; Paralysis ; epidemiology ; virology ; Seasons ; Serotyping

Objective To prepare the 99Tcm-survivin mRNA antisense peptide nucleic acid (PNA)and investigate its value as a gene imaging agent in tumor bearing mice and early diagnosis in tumor.Methods Survivin mRNA antisense PNA and mismatch PNA were synthesized.Four amino acids (Gly- (D)Ala-Gly-Gly) and Aba (4-aminobutyric acid) were linked to the 5' end of PNA.Gly- (D)Ala-Gly-Gly served as a chelating moiety for strong chelation of 99Tcm and Aba acted as a spacer to minimize the steric hindrance.PNAs were labeled with 99Tcm by the ligand-exchange method.The labeling efficiency and radiochemical purity were measured by HPLC and ITLC methods.There were five BALB/c nude mice bearing human lung carcinoma ( A549 ) in each of antisense PNA and mismatch PNA groups.Gene imaging of 99Tcm-survivin mRNA antisense and mismatch PNAs were performed at 1,2 and 4 h post the injection,respectively,and the T/NT ratio was measured by the method of ROI.The statistical comparisons of average values were performed with the two-group t-test for independent sample by SPSS 13.0.Results The product kept stable in vitro.The labeling efficiency of 99Tcm-survivin mRNA antisense PNA was (95.48 ±1.92)％ and more than 85％ after the incubation for24 h in serum.The radiochemical purity was ＞ 95％.The labeling efficiency of mismatch PNA was similar to the antisense PNA.99Tcm-survivin mRNA antisense PNA was especially uptaken by tumor lesion,and its accumulation reached the top at 4 h post the injection.T/NT ratios at 1,2,and 4 h were 2.70 ± 0.28,3.44 ± 0.35,4.21 ± 0.63,respectively.In the comparison,the T/NT ratio of 99Tcm-survivin mRNA mismatch PNA at 4 h (3.12 ±0.50) was significantly lower (t =2.918,P =0.019).Conclusions 99Tcm-survivin mRNA antisense PNA has high labeling efficiency,good stability and no need of purification.Its characteristic of especial uptake by tumor lesion provides the potential value in early diagnosis of tumor.

Microsatellite instability(MSI)was defined according to the frequency of positive findings in a panel of MSI markers.High frequency MSI(MSI-H)was the phenotype in which repeat sequences were extraordinarily unstable, and was considered to be the bona fide phenotype of DNA mismatch repair defection. However base substitutions in some well studied oncogenes or tumor suppressors were reported to be uncommon in MSI-H tumors. To explore this obvious contradiction, the relationship between MSI and KRAS gene mutations were studied in a panel of 76 human colorectal carcinomas, the whole exon of MLH1 and MSH2 were sequenced for MSI-H tumors. KRAS gene mutation was confirmed by similar frequencies in tumors of different MSI status. Intriguingly, all of the KRAS mutant MSI-H tumors harbored sequence alterations in MLH1gene, which was a key player in DNA mismatch repair system. This implied that in MSI-H tumors carrying MMR mutations, KRAS mutation were frequently and almost exclusively occurred. Furthermore, these MMR mutants were uniformly carrying a unique "modification" + "jumping" type MSI, which was different to MSI-H tumors without MLH1 or MSH2 gene mutations. This study shaded lights on the heterogeneity of MSI-H tumors, and implied the connection between "modification" type MSI and DNA mismatch defection.

The methods to determine the total phenols, total saponins, and marker constituents salidroside, chlorogenic acid and 3, 4-dihydroxy-phenylethyl-β-D-glucopyranoside in the samples of Sargentodoxae Caulis were established to provide the evidence for the improvement and revision of the quality standard of the crude material recorded in the Chinese Pharmacopoeia (2015 Edition). The content of total phenols was determined by ultraviolet spectrophotometry, using gallic acid as a reference substance. The content of total saponins was determined by ultraviolet spectrophotometry, using 3-O-[β-D-xylopyranosyl-(1-2)-O-β-D-glucuronopyranosyl]-28-O-[β-D-glucopyranosyl] asiatic acid as a reference substance. The contents of salidroside, chlorogenic acid and 3,4-dihydroxy-phenylethyl-β-D-glucopyranoside were detected by HPLC. The linear ranges were 1.01-7.04 mg x L(-1) for total phenols, 37.7-201 μg for total saponins, 0.025 8-1.55 μg for salidroside, 0.076 2-5.44 μg for chlorogenic acid, and 0.064 9-3.47 μg for 3,4-dihydroxy-phenylethyl-βP-D-glucopyranoside, respectively. Their average recoveries were 99.12%, 99.11% 105.5%, 99.08%, and 101.6%, respectively. The contents of total phenols and total saponins were 3. 04% -11. 9% and 0. 87% -3. 63%. The contents of salidroside, chlorogenic acid and 3,4-dihydroxy-phenylethyl-β-D-glucopyranoside fluctuated from 0.018% to 0. 572%, from 0.041% to 1.75% and from 0.035% to 1.32%. The established methods were reproducible, and they could be used for the quality control of Sargentodoxae Caulis. The present investigation suggested that total phenols, salidroside, and chlorogenic acid should be recorded in the quality standard of Sargentodoxae Caulis and their contents should not be less than 6.8% for total phenols, 0.040% for salidroside, and 0.21% for chlorogenic acid.
China ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; analysis ; Magnoliopsida ; chemistry ; Phenol ; analysis ; Plant Stems ; chemistry ; Saponins ; analysis ; Triterpenes ; analysis

ObjectiveTo investigate the value of 99Tcm labeled survivin mRNA antisense peptide nucleic acid (PNA) as an imaging agent in the specific diagnosis for carcinoma.MethodsSurvivin mRNA antisense PNA was labeled directly with 99Tcm by the ligand-exchange method.Twenty nude mice with lung carcinoma A549 xenografts were randomly divided into 4 groups.Three groups were used for biodistribution study and one group was used for imaging study.Other twenty mice infected by staphylococcus aureus underwent the same procedure.The biodistribution and imaging of 99Tcm-survivin mRNA antisense PNA was studied at 1,2 and 4 h respectively after the intravenous injection in nude mice bearing lung carcinoma A549 xenografts or inflammation models.SPSS 13.0 was used in the study and all data were analyzed by t test.ResultsBiodistribution results showed that the highest radioactivity was found in the liver,and then in the kidney.Four hours after the administration of the imaging agent,the radioactivity ratios of target-tonon target (T/NT,tumor or inflamumatory lesions to the contralateral regions) in tumor model group were significantly higher than those in inflammation model group ( 3.69 ± 1.13 vs 2.03 ± 0.47,t =3.01,P =0.02 ).Tumors were clearly visible in the tumor model groups at 0.5 h and still clearly seen at 4 h after the injection of antisense PNA.On the contrary,inflammatory lesions could not be seen clearly.Conclusion 99Tcm labeled survivin mRNA antisense PNA can be used to distinguish tumor from inflammation and it may provide a new feasible method for specific tumor diagnosis.

OBJECTIVETo investigate the mechanism of DDP-resistance in gastric cancer cell line SGC7901/DDP mediated by phosphate(p)-JNK.

METHODSThe p-JNK expression was blocked by the JNK inhibitor SP600125. The drug sensitivity was detected by MTT. Cell apoptosis rate was measured by flow cytometry. The expression of p-JNK and P-glycoprotein (P-gp) was examined by Western blot. The expression of both proteins were detected in a tissue microarray containing 168 spots of cancer tissue and 27 spots of normal gastric tissue by SP immunohistochemistry. Pearson method was used to analyze the correlation between p-JNK and P-gp.

RESULTSThe drug sensitivity and cell apoptosis rate significantly increased (P<0.01), and the protein expression levels of p-JNK and P-glycoprotein were down-regulated after the inhibition of p-JNK by SP600125 in both SGC7901(p-JNK: 1.17+/-0.03 vs 0.38+/-0.071, P-gp: 0.21+/-0.01 vs 0.06+/-0.01) and SGC7901/DDP (p-JNK: 2.56+/-0.14 vs 1.02+/-0.12, P-gp: 0.77+/-0.05 vs 0.52+/-0.06 )cells(all P<0.01). The protein expression rates of p-JNK and P-glycoprotein were 45.8% and 51.8% respectively in gastric cancer tissue, which were significantly higher than those in normal gastric tissue 7.4% and 18.5% (P<0.01). The correlation of protein expression of p-JNK and P-gp was positive (P<0.01).

CONCLUSIONJNK anti-apoptosis pathway with the regulation of P-gp expression plays an important role in the DDP-resistance of gastric cancer, which may be a novel target for reversing multidrug resistance.

ATP-Binding Cassette, Sub-Family B, Member 1 ; metabolism ; Apoptosis ; drug effects ; Cell Line, Tumor ; Cisplatin ; metabolism ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Gene Expression Regulation, Neoplastic ; Humans ; JNK Mitogen-Activated Protein Kinases ; metabolism ; Signal Transduction ; Stomach Neoplasms ; drug therapy ; metabolism

OBJECTIVETo evaluate the effect of JAK/STAT signaling pathway activation on the transdifferentiation and secretion of transforming growth factor-beta1 (TGF-beta1) induced by high glucose in renal proximal tubular epithelial cells.

METHODSHuman kidney cells (HKC) were cultured and then divided into four groups: low glucose (LG) group, high glucose (HG) group, high mannitol (LG + M) group, and HG + AG490 group. Immunoprecipitation and Western blot analysis were used to determine the expression of tryosine phosphorylated Janus kinase 2 ( p-JAK2). The protein expressions of STAT1, STAT3, p-STAT1, and p-STAT3 and the expressions of alpha-SMA and E-Cadherin were observed by Western blot. The contents of TGF-B1, fibronectin and type I collagen in the supernatants of the cultured HKC were detected by enzyme-linked immunosorbent assay (ELISA). The expression of TGF-beta1 mRNA was measured by reverse transcription and polymerase chain reaction (RT-PCR).

RESULTSCompared with LG group, the expressions of JAK2, p-STAT1, p-STAT3, and TGF-beta1, mRNA were significantly increased in HG group from 6 to 72 hours. Meanwhile, the contents of TGF-beta1 and collagen I in the supernatants and the expression of alpha-SMA increased and the expression of E-Cadherin decreased. The expressions of JAK2, p-STAT1, p-STAT3, and TGF-beta mRNA as well as the levels of TGF-beta1 and collagen I in the supernatant s in HG + AG490 group were significantly lower than in the HG group. The expressions of alpha-SMA and E-Cadherin were also decreased in HG + AG490 group.

CONCLUSIONActivation of JAK/STAT signaling pathway may be involved in the high glucose-induced transdifferentiation and overproduction of TGF-beta1, and ECM proteins in HKCs.

Cell Line ; Cell Transdifferentiation ; Epithelial Cells ; cytology ; metabolism ; Glucose ; metabolism ; pharmacology ; Humans ; Janus Kinases ; physiology ; Kidney Tubules, Proximal ; cytology ; metabolism ; STAT Transcription Factors ; physiology ; Signal Transduction ; Transforming Growth Factor beta1 ; biosynthesis ; secretion ; Urothelium ; cytology ; metabolism

OBJECTIVETo study the technical feasibility of QAMS to determine different structural types of ingredients in Zhizi Jinhua pill, a Chinese patent medicine.

METHODEmodin was chosen as the internal reference substance. The relative correction factors (RCF) of berberine, baicalin, wogonoside, baicalein, aloe-emodin, wogonin, rhein, chrysophanol and physcion to emodin were calculated with the peak areas determined at 254 nm. The ruggedness of these RCFs was validated. The contents of emodin in the samples of Zhizi Jinhua pill were determined by using the external standard method, and the contents of the other nine ingredients were calculated by their RCFs. The contents of these ten ingredients in all samples were also determined by the external standard method.

RESULTNo significant differences were observed between the quantitative results of the two methods. The obtained RCFs were credible.

CONCLUSIONThe established QAMS method can be used for quantitative determination of different structural types of multi-compounds in Zhizi Jinhua pill. It can provide reference for the quality control of Chinese patent medicine.

Chromatography, High Pressure Liquid ; methods ; Dosage Forms ; Drugs, Chinese Herbal ; chemistry ; Quality Control

OBJECTIVETo explore the effects of aptamer-siRNA nucleic acid compound on growth and apoptosis in myeloid leukemia cell line K562.

METHODSthe changes of cellular morphology and structure were observed by using fluorescence microscope, laser confocal microscope, JEM-4000EX transmission electron microscopy; MTT assay were performed to evaluate the sensibility of K562 cells to aptamer-siRNA compound, the apoptosis was detected by DNA gel electro-phoresis.

RESULTSThe remarkably changes of morphology and structure of K562 cells treated with 200 µmol/L aptamer-siRNA were observed under fluorescence microscopy and electromicroscopy. As compared with control, the aptamer-siRNA compound showed more inhibitory effect on K562 cells and there was significant difference (P<0.05). The MTT assay showed that the IC50 value of aptamer-siRNA compound for K562 cells was 150 µmol/L. According to agarose gel electrophoresis observation, when the aptamer-siRNA compound showed effect on K562 cells, the typical DNA lader could be observed.

CONCLUSIONThe aptamer-siRNA compound can significantly induce K562 cell apoptosis, and provide reference for gene therapy of patients with chronic myelocytic lenkemia.

Apoptosis ; Cell Proliferation ; Humans ; K562 Cells ; Leukemia, Myeloid ; RNA, Small Interfering