Twelve compounds were isolated from the venom of Bufo bufo gargarizans. On the basis of their physical and chemical properties and spectral data, their structures were identified as resibufagenin (1), bufotalin (2), desacetylcinobufagin (3), 19-oxodesacetylcinobufotalin (4), cinobufotalin (5), 1beta-hydroxylbufalin (6), 12alpha-hydroxybufalin (7), bufotalinin (8), Hellebrigenin (9), telocinobufagin (10), hellebrigenol (11) and cinobufagin-3-hemisuberate methyl ester (12), respectively. Compounds 7 and 12 are new natural products.
Animals ; Bufanolides ; chemistry ; Bufo bufo ; Medicine, Chinese Traditional ; Molecular Structure ; Venoms ; chemistry

The safety of Chinese patent medicine has become a focus of social. It is necessary to carry out work on post-marketing clinical safety evaluation for Chinese patent medicine. However, there have no criterions to guide the related research, it is urgent to set up a model and method to guide the practice for related research. According to a series of clinical research, we put forward some views, which contained clear and definite the objective and content of clinical safety evaluation, the work flow should be determined, make a list of items for safety evaluation project, and put forward the three level classification of risk control. We set up a model of post-marketing clinical safety evaluation for Chinese patent medicine. Based this model, the list of items can be used for ranking medicine risks, and then take steps for different risks, aims to lower the app:ds:risksrisk level. At last, the medicine can be managed by five steps in sequence. The five steps are, collect risk signal, risk recognition, risk assessment, risk management, and aftereffect assessment. We hope to provide new ideas for the future research.
Clinical Trials as Topic ; Drug-Related Side Effects and Adverse Reactions ; epidemiology ; etiology ; Drugs, Chinese Herbal ; adverse effects ; chemistry ; economics ; therapeutic use ; Herbal Medicine ; economics ; legislation & jurisprudence ; Humans ; Patents as Topic ; Product Surveillance, Postmarketing ; Quality Control

OBJECTIVETo evaluate the changes in programmed death 1 (PD-1) and programmed death ligand 1 (PD-L1) expression on peripheral blood T lymphocytes of patients with chronic hepatitis C (CHC) over the 24 weeks course of antiviral therapy.

METHODSTwenty-four CHC patients administered 24 weeks of combination antiviral therapy with pegylated-interferon-alpha-2a (Peg-IFNa-2a) and ribavirin (RBV) were enrolled for study from the Nanjing Second Hospital between October 2008 and October 2011. Peripheral blood was collected before treatment initiation, at treatment weeks 4, 12 and 24, and post-treatment week 24 (to investigate sustained virologic response (SVR), and used to measure expression of PD-1 and PD-L1 on CD4+ and CD8+ T lymphocytes by flow cytometry, load of serum hepatitis C virus (HCV) RNA by real-time polymerase chain reaction, and level of serum alanine aminotransferase (ALT) by auto-biochemical analyzer. Intergroup differences were analyzed by the two-sample t-test, and the significance of differences between pre- and post-treatment measurements was determined by one-way or two-way repeated measurements analysis of variance tests.

RESULTSAt treatment week 4, 19 of the CHC patients were HCV RNA-negative. Among those patients the PD-1 expression on both T lymphocyte subsets showed a significant decrease from pre-treatment to post-treatment week 24 (CD4+: 18.6 +/- 6.1% vs. 10.3 +/- 7.7%, F = 12.406, P = 0.002; CD8+: 16.6 +/- 13.8% vs. 9.4 +/- 4.6%, F = 4.955, P = 0.039). However, the CD8+ lymphocyte subset showed significant increase in PD-L1 expression during treatment (pre-treatment: 17.5 +/- 13.7% vs. treatment week 4: 25.9 +/- 11.1%, F = 9.063, P less than 0.01; 12: 29.6 +/- 15.1%, F = 8.365, P less than 0.01; 24: 32.0 +/- 15.7%, F = 9.736, P less than 0.01). Among the five CHC patients showing HCV RNA-positivity at treatment week 4 there was only a significant difference observed in the increased expression of PD-L1 on CD8+ lymphocyte subset from pre-treatment to treatment week 24 (17.4 +/- 16.7% vs. 39.2 +/- 15.6%, F = 10.292, P = 0.033). Twenty of the CHC patients achieved SVR. among whom the PD-1 expression was significantly decreased during treatment on the CD4+ lymphocyte subset (pre-treatment: 20.2 +/- 7.5% vs. treatment week 4: 14.4 +/- 7.5%, F = 6.133, P less than 0.05; 12: 14.0 +/- 6.9%, F = 5.541, P less than 0.05; 24: 10.7 +/- 7.6%, F = 14.780, P less than 0.05) and on the CD8+ lymphocyte subset (pre-treatment: 16.8 +/- 13.4% vs. treatment week 12: 10.2 +/- 4.6%, F = 4.964, P less than 0.05; 24: 10.1 +/- 4.9%, F = 4.613, P less than 0.05). Additionally, the PD-L1 expression was significantly increased during treatment on the CD8+ lymphocyte subset (pre-treatment: 19.0 +/- 14.5% vs. treatment week 12: 30.8 +/- 16.6%, F = 6.442, P = 0.020; 24: 35.2 +/- 16.5%, F = 12.349, P = 0.002). Among the four CHC patients who relapsed there were no significant differences observed in the expressions of PD-1 or PD-L1 on the CD4+ or CD8+ T lymphocytes.

CONCLUSIONThe standard Peg-IFNa-2a + RBV combination antiviral therapy reduces PD-1 expression on CD4+ and CD8+ T lymphocytes and increases PD-L1 expression on CD8+ T lymphocytes in peripheral blood. The clinical outcome of CHC patients may be related to the antiviral therapy-induced changes in expressions of PD-1 and PD-L1 on T lymphocytes.

Antiviral Agents ; therapeutic use ; Hepatitis C, Chronic ; drug therapy ; Humans ; Interferon-alpha ; therapeutic use ; Real-Time Polymerase Chain Reaction ; Ribavirin ; therapeutic use

Objective To investigate the dynamics and development trends of drinking water type of endemic fluorosis after water improvement in Xinbaerhuyouqi of Hulunbeir city, Inner Mongolia and to provide a scientific evidence for the development of countermeasures. Methods We mainly selected Adunchulusumu and Kerlunsumu in Xinbaerhuyouqi of Hulunbeir city as the two monitoring points after water improvement in 2000 -2009. Of these, 1 sample of centralized water supply source water and 3 samples of tap water and 5 samples of noncentralized water supply source water according to water well locations of east, west, south, north and center were collected and the levels of water fluoride were tested; the prevalence of dental fluorosis of school children aged 8 to 12 were examined; from 2002 onwards, the urine samples of 30 children aged 8 to 12(five age groups, six urine samples for each age group) were collected, and all urine samples were collected in the case of less than 30, and urine fluoride was tested. Dental fluorosis was diagnosed using Dean method; water fluoride was tested using fluoride ion selective electrode(WS/T 106-1999); urinary fluoride was tested by determination of fluoride in urine using ion-selective electrode(WS/T 89-1996). Results In 2000 - 2009, the mean levels of fluorine in drinking water in Adunchulusumu and Kerlunsumu were 1.79 - 4.35 mg/L and 1.38 - 3.18 mg/L, respectively; the detection rate of dental fluorosis of children aged 8 to 12 were 45.24％(19/42) - 89.78％(123/137) and 40.00％ (28/70) - 74.47％ (70/94), respectively; the median urinary fluoride of them were 2.30 - 4.15 mg/L and 2.73 - 4.55 mg/L, respectively. ConclusionsThe detection rate of children's dental fluorosis remains high in Xinbaerhuyouqi during the past 10 years after changing water. The endemic fluorosis remains a serious disease. Effective prevention and control measures must be taken to control the occurrence of fluorosis in the future.

Objective To study the clinical application anatomy of interventricular foramen and offer a base for operation.Methods Interventricular foramens were observed in 15 adult cadaveric brainThirty- two patients of obstructive hydrocephalus were operated to observe the structure of interventricular foramen un- der neuroendoscope.Results Interventricular foramen was a poriform structure which consists of fornixan- terior pole of thalamencephalon and choroid plexus and was a oval shape in most of themThe plane of the fo- ramen was a included angle with the median sagittal planeThe septal veinthalamostriate veinthalamen- cephal and even the floor of third ventricle could be observed clearly in endoscope.At the same timewe found the foramen had a significant change in obstructive hydrocephalus.Conclusion The interventricular foramen has a simple relatively structure but a variation on size and shape especially in obstructive hydroceph- alusA clearly comprehension of it's structure and adjacent is a base to microsurgery and endoscopic surgery on the foramen.

OBJECTIVETo investigate the effect of farnesoid-X-receptor (FXR) antagonist Z-guggulsterone in an in vivo high-fat fed apolipoprotein E knockout (ApoE(-/-)) mice model of myocardial ischemia/reperfusion (I/R).

METHODSMale ApoE(-/-) mice were randomly divided into three groups: standard ApoE(-/-) group (fed with standard mouse diet for 12 weeks before myocardial I/R procedure, n = 18), high-fat ApoE(-/-) group (fed with high-fat mouse diet for 12 weeks before myocardial I/R procedure, n = 22), and high-fat ApoE(-/-) + FXR antagonist group(fed with high-fat mouse diet for 12 weeks and received FXR antagonist Z-Guggulsterone 30 minutes before myocardial I/R procedure, n = 17). The expression of FXR was detected by real-time quantitative-PCR. Myocardial infarct size was determined by Evans blue/TTC double staining methods. Myocardial apoptosis was determined by in situ TUNEL technique. Markers of the mitochondrial-mediated apoptotic pathway (cytochrome c release, caspase-9 activity, and BAX and BCL-2 levels), endoplasmic reticulum stress apoptotic pathway (caspase-12 activity and CHOP level), and death receptor apoptotic pathway (caspase-8 activity, and Fas and FasL levels) were also measured.

RESULTFXR expression (3.7-fold higher, P < 0.01), myocardial infarct size [(62.1 ± 7.0)% vs. (33.8 ± 5.8)%, P < 0.01] and myocardial apoptosis index[ (36.8 ± 5.7)% vs. (17.2 ± 3.8)%, P < 0.01]were all significantly higher in high-fat ApoE(-/-) group than those in standard ApoE(-/-) group. Compared with high-fat ApoE(-/-) group, myocardial infarct size [(24.4 ± 4.7)% vs. (62.1 ± 7.0)%, P < 0.01] and myocardial apoptosis index [(13.8 ± 2.7)% vs. (36.8 ± 5.7)%, P < 0.01] were significantly reduced in high-fat ApoE(-/-) + FXR antagonist group. Moreover, levels of mitochondrial-mediated apoptotic pathway markers (cytochrome c release, caspase-9 activity, and BAX/BCL-2 levels) and endoplasmic reticulum stress apoptotic pathway markers (caspase-12 activity and CHOP level) were significantly lower in high-fat ApoE(-/-) + FXR antagonist group than those in high-fat ApoE(-/-) group (all P < 0.01). Levels of death receptor apoptotic pathway markers (caspase-8 activity, and Fas and FasL levels) were similar between high-fat ApoE(-/-) group and high-fat ApoE(-/-) + FXR antagonist group.

CONCLUSIONFXR antagonist alleviates myocardial reperfusion injury in cholesterol-fed ApoE(-/-) mice via inhibition of the mitochondrial-mediated and endoplasmic-reticulum stress pathway.

Animals ; Apolipoproteins E ; genetics ; Apoptosis ; drug effects ; Caspase 9 ; metabolism ; Cholesterol, Dietary ; administration & dosage ; Cytochromes c ; metabolism ; Disease Models, Animal ; Endoplasmic Reticulum Stress ; Male ; Mice ; Mice, Knockout ; Myocardial Reperfusion Injury ; metabolism ; pathology ; prevention & control ; Pregnenediones ; pharmacology ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Receptors, Cytoplasmic and Nuclear ; antagonists & inhibitors ; metabolism ; bcl-2-Associated X Protein ; metabolism

OBJECTIVETo study the effect of cigarette smoke on the sexual function of male rats.

METHODSBased on Ozyurt's smoking model, we equally divided 30 male adult Sprague-Dawley rats into a control and a smoking group, and exposed the latter to cigarette smoke for 60 days. A week before the end of the experiment, we added 5 female rats to each group and observed their mating through 24-hour video surveillance. Sixty days later, all the rats were killed for the determination of the level of testosterone (T) in the plasma and the activity of nitric oxide synthase (NOS) in the corpus cavernosum, and Masson-dyeing image analysis of the penile tissue.

RESULTSCompared with the controls, the rats in the smoking group showed a significant reduction in the times of mating, the level of plasma T (P < 0.05) and the activity of NOS in the penile cavernous tissue (P < 0.05), but a slight increase in the collagen fibers and obvious changes in the blood sinuses.

CONCLUSIONCigarette smoke seriously affected penile erection in the experimental rats. The decrease in plasma T, NOS activity and the area of smooth muscle may be an important mechanism underlying their erectile dysfunction.

Animals ; Erectile Dysfunction ; Male ; Muscle, Smooth ; metabolism ; Nitric Oxide Synthase ; metabolism ; Penile Erection ; Penis ; metabolism ; Rats ; Rats, Sprague-Dawley ; Smoke ; adverse effects ; Testosterone ; blood ; Tobacco ; adverse effects

OBJECTIVETo detect over-expression of AChR-gamma mRNA in rhabdomyosarcoma tissues by duplex RT-PCR and discuss its potential in diagnosis of rhabdomyosarcoma.

METHODSDuplex RT-PCR was applied to the simultaneous detection of AChR-alpha and gamma subunit messenger RNA in 17 cases of rhabdomyosarcoma (9 ERMS, 6 ARMS, 2 PRMS). 20 cases of non-rhabdomyosarcomous small round cell tumors (6 poorly differentiated synovial sarcomas, 6 ES/PNET, 6 lymphomas, 2 neuroblastomas) and three normal muscle samples were also detected for AChR-alpha and gamma mRNA by the same method.

RESULTSAChR-alpha and AChR-gamma mRNA were expressed in all the cases of rhabdomyosarcoma. The rate of quantity in both transcripts was AChR-gamma/AChR-alpha >or= 1, but the rate for three normal muscle samples was < 1. Cases of non-rhabdomyosarcomous small round cell tumors were all negative for AChR-gamma.

CONCLUSIONAChR-gamma mRNA expression detected by molecular genetic methods is useful in diagnosis and differential diagnosis of rhabdomyosarcoma.

Diagnosis, Differential ; Humans ; Protein Subunits ; RNA, Messenger ; analysis ; Receptors, Cholinergic ; genetics ; Receptors, Nicotinic ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Rhabdomyosarcoma ; diagnosis

ObjectiveTo investigate the effect of drinking brick tea with low-fluoride level on prevention of tea type fluorosis.MethodsHandahangacha,Hadayinggegacha,Dalainuoyi town,in Keshiketengqi Inner Mongolia endemic fluorosis area were selected as test points,and brick tea with fluoride [(204.5 ± 10.2),(308.2 ±15.4)mg/kg] was given for 12 months.Dental fluorosis,clinical skeletal fluorosis,and X-ray diagnosis of skeletalfluorosis [according to “Endemic Skeletal Fluorosis Diagnostic Criteria” (WS 192-2008)] of adults 20 to 70 years of age were examined and level of fluoride before and after the prevention trial,in brick tea,drinking water,milk tea and urine were tested (fluoride ion selective electrode method),and fluoride intake through tea was calculated.ResultsDetection rate of adult dental fluorosis in Handahangacha was 68.89％ (62/90),clinical detection of skeletal fluorosis was 55.32％ (52/94),and X-ray detection of skeletal fluorosis was 65.17％ (58/89); adult dental fluorosis detection rate in Hadayinggegacha was 54.84％(51/93),clinical detection of skeletal fluorosis was 65.69％(67/102),and X-ray detection rate of skeletal fluorosis was 61.36％ (54/88).Brick tea fluoride was (831.4 ±138.9),(864.3 ± 134.6)mg/kg before the prevention trial in Handahangacha and Hadayinggegacha,respectively,drinking water fluoride content was (0.27 ± 0.05),(0.54 ± 0.24)mg/L,fluoride content of milk tea was (216 ± 1.12),(2.82 ± 1.38)mg/L,adult urine fluoride content was (2.78 ± 1.57),(2.96 ± 1.80)mg/L,and fluoride intake through milk tea was (8.12 ± 5.84),(6.42 ± 5.04)mg/d,respectively; after the prevention trial the fluoride content of brick tea was (204.5 ± 10.2),(308.2 ± 15.4)mg/kg,fluoride content of drinking water (0.34 ± 0.11),(0.62 ± 0.30)mg/L,fluoride content of milk tea(0.97 ± 0.33),(1.83 ± 0.66)mg/L,fluoride content in urine(1.29 ± 0.55),( 1.47 ±0.62)mg/L,fluoride intake through milk tea (3.45 ± 2.05),(3.71 ± 2.07)mg/d,respectively; in Handahan and Hadayinggegacha after the prevention trial the fluoride in brick tea,milk tea,urine fluoride,and fluoride intake through milk tea was significantly lower than that before the trial (t =14.30,12.97 ;6.46,3.95;6.69,5.72;6.27,3.57,all P ＜ 0.01 ).Fluoride intake in Handahangacha through milk tea was within the state heath standard limits( ＜ 3.5mg/d).ConclusionDrinking low-fluoride brick tea can prevent drinking brick tea type fluorosis,the preventive effect is especially more reliable with low fluoride brick tea (204.5 ± 10.2)mg/kg.

OBJECTIVETo explore the genetic variations of HLA-Cw and 5 KIR2D loci in 2 Chinese Han populations residing at Southern and Northern mainland China, respectively, and to investigate the HLA-Cw polymorphism of a Mongolian Chinese population.

METHODSHLA-Cw genotyping was performed in a total of 293 healthy individuals including 1 Southern Han population living in Hunan Province (n=112), 1 Northern Han population (n=98) and 1 Mongolian Chinese population(n=83) in the Inner Mongolia Autonomous Region, using polymerase chain reaction-sequence specific primer(PCR-SSP) technique. Dimorphism at residue 80 of domain in the HLA-Cw molecule was examined by an additional set of PCR-SSP reactions. PCR-SSP was also used to detect the presence or absence of inhibitory KIR2DL1/2DL2/2DL3 loci and activating KIR2DS1/2DS2 loci for the 2 Han populations.

RESULTSThe main findings were: (1) Very significant frequency difference in the HLA-Cw alleles and dimorphism at codon 80 was detected between Hunan Han and Northern Han population, and between Hunan Han and Mongolian population (P < 0.001),while there was no such difference between the 2 Northern Chinese populations (P> 0.05); (2) There was no significant difference in frequencies of either the 5 individual KIR2D genes or the genotype distributions between the 2 Han populations (P> 0.05); (3) Asn(80)ls/Asn(80), 2DL1+/2DL2-/2DL3+/2DS1-/2DS2- predominated in both Han populations (45/112, 29/98), followed by Asn(80)/Asn(80), 2DL1+/2DL2-/2DL3+/2DS1+/2DS2- (18/112,16/98) and Asn(80)/Lys(80), 2DL1+/2DL2-/2DL3+/2DS1-/2DS2-(11/112,17/98). Among the 12 types of HLA-Cw codon 80 and KIR2D combinations, only Lys(80)/Lys(80), 2DL1+/2DL2-/2DL3+/2DS1-/2DS2- showed marginally significant frequency difference between the 2 Han populations(1/112 vs 8/98; Fisheros P was 0.0134).

CONCLUSIONOur study provided the polymorphism data of HLA-Cw gene for 3 Chinese populations with different geographic and/or ethnic background, we further analyzed the distribution of 5 KIR2D receptor genes in 2 Han populations. Our data suggest that in spite of HLA-Cw heterogeneity, remarkable similarities may exist between the Southern and Northern Chinese Han populations at the combinational level of HLA-Cw and KIR2D, which are characterized by preponderant inhibitory signal pathways.

Asian Continental Ancestry Group ; Genetic Variation ; genetics ; HLA-C Antigens ; genetics ; Humans ; Polymerase Chain Reaction ; Polymorphism, Single Nucleotide ; genetics ; Receptors, KIR ; genetics ; Receptors, KIR2DL1 ; genetics ; Receptors, KIR2DL2 ; genetics ; Receptors, KIR2DL3 ; genetics