Objective To investigate the interference effect of baicalin on acute brain injury induced by aconitine in rats and its mechanism. Methods A total of 200 Sprague-Dawley(SD)rats were randomly divided into five groups:normal control,baicalin control,aconitine poisoning,baicalin 15 mg/kg intervention and baicalin 30 mg/kg intervention groups(each,n=40). Aconitine(20μg/kg)was given via tail vein in aconitine poisoning group. The rats in the normal control group and baicalin control group were respectively injected with saline 2 mL/kg and baicalin 30 mg/kg via tail vein. The aconitine poisoning rats were given with baicalin at the dose of 15 mg/kg and 30 mg/kg respectively in the low and high dose baicalin intervention groups within 2-3 minutes after injection of aconitine. Rats in all groups in the study were anesthetized and sacrificed at 1,6,12,24 hours after various agents were respectively given in the groups,the rat cerebral cortex samples were collected,the histological changes in normal and baicalin control groups and pathological changes of the aconitine poisoning rats were observed,the levels of glutamate(Glu),aspartate(Asp),γ-aminobutyric acid(GABA),glycine(Gly)were detected and the apoptotic cells were determined at the above time points. Results Compared with the normal control group,the aconitine poisoning group had significantly higher levels of excitatory amino acids Glu and Asp and the number of apoptotic neurons. After exposure to aconitine for 1 hour, the levels of inhibitory amino acids of GABA and Gly were markedly decreased in the rat cortex in the poisoning group compared to the normal control group(both P<0.05),at 6 hours and 12 hours they were significantly increased and after 24 h,they began to decline,but still maintained at relatively high levels. Compared with the aconitine poisoning group, after baicalin intervention for 1 hour,in the 15 mg/kg and 30 mg/kg baicalin intervention groups,the levels of Glu and Asp were markedly decreased〔Glu(μmol/L):309.39±14.59,307.22±23.69 vs. 370.46±40.31,Asp(μmol/L):143.43±8.36,129.12±4.86 vs. 222.97±6.26〕,while the levels of GABA and Gly were increased〔GABA(μmol/L):55.91±4.76,59.61±13.11 vs. 32.05±2.20,Gly(μmol/L):32.33±1.85,33.90±0.66 vs. 21.96±4.75〕,and the number of neuronal apoptosis was obviously decreased(cell/mm2:18.65±4.10,14.80±1.89 vs. 58.15±3.68,both P<0.05). Under microscope and electron microscope,the pathological and ultrastructural changes indicated that the aconitine poisoning group had the most marked cerebral cortex damage at 12 hours after poisoning,while the two baicalin intervention groups showed milder damage than that in aconitine poisoning group. Conclusions The neural toxic effect of aconitine in rats may be related to the imbalance between the neurotransmitter contents of excitatory Glu. Asp and inhibitory GABA,Gly in the cerebral cortex. Baicalin can decrease the contents of excitatory amino acid and elevate the inhibitory amino acid,therefore it may ameliorate the cerebral injury of acute aconitine intoxication in rats.

Objective To investigate the Human papillomavirus(HPV)infection in different age groups of women in Shenyang, and explore its correlation with cervical biopsy diagnosis.Methods 7 311 women aged 13-85 did HPV test and thin-cytologic test (TCT)in the hospital.Some of them had biopsy detection under electronic colposcopy,and the pathological diagnosis was the golden standard for the diagnosis of cervical lesions.SPSS18.0 statistical software was used for all statistical analysis.Results The infection rate of <30 years old women was significantly higher than that of 30 - <40,40 - <50,≥50 years old women (P <0.05).The most prevalent high-risk HPV genotype in Shenyang were subtype 16,52,58,53,33,31 and 18,and the most prevalent low-risk HPV subtypes were 81,11 and 6.The former 4 subtypes of high-risk HPV infection accounted for 67.3% of all high-risk infection.As to the 4 subtypes with higher infection rate,the infection rate of ≥40 years old women was higher than that of <40 years old(χ2 =20.29,P =0.00).The top two low-risk HPV subtypes accounted for 74.8% of the infections.The mean age of the ICC patients were 48.3,which was statistically different from the other groups(P <0.05).Cervical lesions occured mostly in 40-49 years old,which accounted for 37.1% and was higher than the other agees(P <0.01).HPV16 infection rate increased with the severity of cervical lesions.Conclusion HPV DNA genotyping is a necessary methord for cervical cancer screen,an effective com-plement for precancerous lesions diagnosis which was missed in cytology test,and also an indispensable test for CIN treatment and follow-up after operation.

Objective To investigate the relationship between the expression of eonnexin 43 (Cx43) and clinicopathologieal characteristics of gastric cancer, and to study the role of Cx43 in peritoneal metastasis of gastric cancer. Methods Thirty-two patients who had gastric cancer and with peritoneal metastasis had been admitted to Southwest Hospital from January 2000 to December 2008. Gastric cancer tissues, adjacent tissues and metastatic peritoneal tissues were obtained postoperatively, and the expression of Cx43 was detected by immunohistochemistry. The relationship between Cx43 expression and clinicopathological characteristics of gastric cancer was analyzed. All data were analyzed via Spearman rank correlation coefficient, Fisher exact probability and chi-square test. Results The expression of Cx43 was mainly detected in the cell membrane and cytoplasm. The positive expres-sion rates of Cx43 in gastric cancer tissues, adjacent tissues and metastatic peritoneal tissues were 34% (11/32), 100% (32/32) and 94% (30/32), respectively. There were significant differences in the Cx43 expression between gastric cancer tissues and adjacent tissues (X~2=28.350, P < 0.01), and between gastric cancer tissues and metastatic peritoneal tissues (X~2 = 21.989, P < 0.01). The expression of Cx43 did not correlate with age and sex of patients (r = -0.030, - 0.169, P > 0.05), but with tumor differentiation, histological type and lymph node metastasis (r = 0.750, 0.642, - 0.357, P < 0.05). Conclusions There is a decreased expression of Cx43 in gastric cancer tissues and a up-regulated expression of Cx43 in metastatic peritoneal tissues. Cx43 may play a positive role in the peritoneal metastasis.

Objectives To investigate the expression and diagnostic value of 14-3-3 protein in brains of patients with sporadic Creutzfeldt-Jakob disease(sCJD).Methods 14-3-3 protein was immunohistochemically analyzed in tissue from the frontal lobe of 5 patients with sCJD and 4 non-CJD eases Using 14-3-3 ?and ?antibodies with reference to the results of KB,GFAP and PrP detection.Results The expressions of 14-3-3 protein in five brains of sCJD were more obviously,mostly in gray matters and astrocytes in three cases.The concentration was related to PrP deposition type,but not related to prion protein genotype.Except few expression of 14-3-3 protein in neurous of two cases of acute contusion,there were no expression in the other two cases in control group.Conclusions The expression of 14-3-3 protein in brain is useful to pathological diagnosis of CJD.

OBJECTIVETo assess the regulating effects of interleukin-1 (IL-1) on gene expression of cartilage specificity gene Sox9 and type II collagen mRNA in the human intervertebral discs.

METHODSRT-PCR were used to investigate the effects of IL-1 on gene expression of Sox9 and type II collagen mRNA in intervertebral discs cells cultures of embryo.

RESULTSThe Sox9 and type II collagen mRNA in intervertebral discs were decreased progressively along with the addition concentrations of IL-1 than the controls. And the mRNA of Sox9 and type II collagen also markedly decreased with the time of culture.

CONCLUSIONSIL-1 could cause dose-dependent and time-dependent inhibition effects on Sox9 and type II collagen gene expression in human intervertebral discs.

Cells, Cultured ; Collagen Type II ; genetics ; Dose-Response Relationship, Drug ; Gene Expression ; drug effects ; High Mobility Group Proteins ; genetics ; Humans ; Interleukin-1 ; pharmacology ; Intervertebral Disc ; cytology ; drug effects ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; SOX9 Transcription Factor ; Time Factors ; Transcription Factors ; genetics

To investigate the chemical constituents of the whole plants of Bidens bipinnata, the separation and purification of constituents were performed by chromatography on macroporous resin, silica gel, MCI and Sephadex LH-20. Their structures were elucidated by spectroscopic data as quercetin (1), quercetin-3-0-alpha-L-rhamnoside (2), keampferol-3-O-beta-D-glucopyranoside (3), keampferol-3-O-alpha-L-rhamnoside (4), 3', 5-dyhydroxy-3, 6, 4'-trimethoxyl -7-O-beta-D-glucopyranoside flavonoid (5), 7, 8, 3', 4'-tetraflavanone(6), (2S)- and (2R)-isookanin-7-O-beta-D- glucopyranoside (7a/7b), (2S)- and (2R)-3'-methoxy-isookanin-8-O-beta-D-glucopyranoside (8a/8b), 6, 7, 3', 4'-tetrahydroxyaurone(9), maritimetin (10), esculetin (11), 3-O-caffeoyl-2-methyl-d-erythrono-1, 4-lactone (12), (7S, 8R) balanophonin-4-O-beta-D-glucopyranoside (13), eugenyl-O-beta-apiofuranosyl-( 1"-6') -O-beta-glucopyranoside (14), and (+)-syringaresinol-4'-O-beta-D-glucopyranoside (15). Compounds 8, 13, 14, and 15 were isolated from this genus for the first time. Compounds 1 and 6 were potent inhibitors against HSC-T6 cells in vitro and compounds 1, 2, 6, and 7 were capable of decreasing the inflammatory cytokine production of macrophage cells in vitro.
Bidens ; chemistry ; Drugs, Chinese Herbal ; chemistry ; Molecular Structure ; Spectrometry, Mass, Electrospray Ionization

BACKGROUNDArthritogenic T lymphocytes with common T cell receptor (TCR) Vbeta clonotypes, infiltrating in the articulars of rheumatoid arthritis (RA) patients, play a central role in the pathogenesis of RA. TCR Vbeta5.2 and TCR Vbeta8.2 are the main pathogenic T cell clonotypes in the course of collagen-induced arthritis (CIA) progression in Lewis rats. To investigate a TCR-based immunotherapy for RA, we constructed recombinant DNA vaccines encoding TCR Vbeta5.2 and TCR Vbeta8.2, and evaluated the inhibitive effects of the two vaccines on CIA rats.

METHODSGenes encoding TCR Vbeta5.2 and TCR Vbeta8.2 were amplified by RT-PCR from spleen lymphocytes of Lewis rats and cloned into the eukaryotic expression vector pTargeT. The expression of vaccines was confirmed by RT-PCR and immunohistochemistry. The inhibitive effects of the vaccines on articulars of CIA rats were assessed with arthritis index evaluation and histology. Interferon gamma (IFN-gamma) and interleukin (IL)-4 production by spleen lymphocytes were tested with enzyme-linked immunospot assay (ELISPOT) technique, the changes in peripheral CD4(+) and CD8(+) lymphocyte populations were tested by flow cytometry, and the level of anti-CII antibody in serum was assayed by enzyme-linked immunosorbent assay (ELISA).

RESULTSRecombinant DNA vaccines pTargeT-TCR Vbeta5.2 and pTargeT-pTCR Vbeta8.2 were successfully constructed. Both vaccines inhibited CIA, which alleviated the arthritis index score (P < 0.05), decreased the level of IFN-gamma (P < 0.05), and reduced the ratio of CD4(+)/CD8(+) lymphocytes (P < 0.05) and the anti-CII antibody in serum (P < 0.05). In addition, the histological change in DNA-vaccinated rats was less serious than CIA rats. Compared to pTCR Vbeta 8.2 and pTCR Vbeta 5.2 groups, the group that was injected with a combination of the two vaccines showed stronger inhibitive effects on CIA than either individual vaccine.

CONCLUSIONThe recombinant plasmids pTargeT-TCR Vbeta5.2 and pTargeT-TCR Vbeta8.2 have obvious inhibatory effects on CIA rats and better effects could be achieved when the vaccines were used in combination.

Animals ; Arthritis, Experimental ; metabolism ; prevention & control ; CD4-Positive T-Lymphocytes ; drug effects ; CD8-Positive T-Lymphocytes ; drug effects ; Enzyme-Linked Immunosorbent Assay ; Female ; Flow Cytometry ; Immunohistochemistry ; Interferon-gamma ; metabolism ; Interleukin-4 ; metabolism ; Muscles ; drug effects ; metabolism ; Peptide Fragments ; antagonists & inhibitors ; Rats ; Rats, Inbred Lew ; Receptors, Antigen, T-Cell, alpha-beta ; antagonists & inhibitors ; Reverse Transcriptase Polymerase Chain Reaction ; Vaccines, DNA ; pharmacology

Background and Objective: Elevated expression of matrix metalloproteinases (MMPs) has been found in multiple carcinoma tissues.MMP-26 is highly expressed in prostate and breast cancer tissues,and promotes the invasion of human prostate cancer cells not only through the cleavage of fibronectin and type Ⅳ collagen but also by the activation of pro-MMP-9,a powerful gelatinase. This study was to present a comprehensive protein expression profile of MMP-26 in multiple human cancer tissues. Methods: The protein expression pattern of MMP-26 was examined using immunohistochemistry and multiple-tissue microarray. MMP-26 mRNA expression in coronary artery smooth muscle cells was detected by reverse transcription-polymerase chain reaction(RT-PCR). Results: The expression of MMP-26 in breast,colon,lung, brain, head and neck, prostate cancer, and melanoma tissues was significantly elevated when compared with parallel normal tissues (P＜0.05), while not significantly elevated in kidney cancer,ovarian cancer,and non-Hodgkin's lymphoma (P＞0.05).MMP-26 was also detected to express in gastric,rectal,thyroid, esophageal,and pancreatic cancers.MMP-26 protein was expressed in smooth muscle cells of the prostate and associated blood vessels. MMP-26 mRNA was also detected to express in human coronary artery smooth muscle cells. Conclusions: MMP-26 expression may be associated with multiple human carcinomas,and it may serve as a molecular marker for the early diagnosis of these carcinomas.MMP-26 may also contribute to smooth muscle function in the human prostate and cardiovascular system.

This study aimed to establish a method for the detection and identification of H7N9 avian influenza viruses based on the NA gene by pyrosequencing. According to the published NA gene sequences of the avian influenza A (H7N9) virus, a 15-nt deletion was found in the NA gene of H7N9 avian influenza viruses. The 15-nt deletion of the NA gene was targeted as the molecular marker for the rapid detection and identification of H7N9 avian influenza viruses by pyrosequencing. Three H7N9 avian influenza virus isolates underwent pyrosequencing using the same assay, and were proven to have the same 15-nt deletion. Pyrosequencing technology based on the NA gene molecular marker can be used to identify H7N9 avian influenza viruses.
Animals ; Base Sequence ; Birds ; Chickens ; High-Throughput Nucleotide Sequencing ; methods ; Influenza A Virus, H7N9 Subtype ; classification ; enzymology ; isolation & purification ; Influenza in Birds ; virology ; Molecular Sequence Data ; Neuraminidase ; genetics ; Phylogeny ; Poultry Diseases ; virology ; Viral Proteins ; genetics

BACKGROUNDNumerous studies have shown that reducing the level of tumor necrosis factor-alpha (TNFα) through the use of anti-TNF antibodies or soluble TNF receptor is a safe and efficacious treatment to inflammatory diseases such as rheumatoid arthritis. Therefore, novel approaches to achieve this outcome are desired. The aim of this study was to investigate the characterization of a small molecule inhibitor, Y316, which blocks TNF mRNA upregulation and TNF production by lipopolysaccharides (LPS) stimulated monocytes.

METHODSPeripheral blood mononuclear cells (PBMC) from healthy volunteers were plated in 24-well plates and stimulated with LPS (1 µg/ml), phorbol-12-myristate-13-acetate (PMA) (100 ng/ml), zymosan (10 µg/ml) and Tsst (100 ng/ml). Supernatants were collected after 4-hour culture at 37°C, and quantitative determination of TNFα, interleukin-1β (IL-1β), IL-6, IL-8, IL-10 and IL-2 production in the supernatants was performed by colorimetric enzyme-linked immunosorbent assay (ELISA). Total RNA of PBMC was isolated and cytokine mRNA quantitation was performed by using a RNA level measuring kit (R & D Systems). PBMC were pretreated with Y316 (10 µmol/L, 1 µmol/L, 0.1 µmol/L, 0.01 µol/L and 0.001 µmol/L) or dimethyl sulfoxide at 37°C for 10 minutes, and then stimulated with LPS or PMA, protein concentrations of p44.42, IKBα, P38 and Jun NH2-terminal kinase were determined by Western blotting. Cyclic adenosine-3',5'-monophosphate (cAMP) of PBMC was measured by enzyme immunoassay kit (Amersham Pharmacia Biotech).

RESULTSY316 blocked TNF production and inhibited the upregulation of TNF mRNA levels in response to LPS, and also prevented the production of IL-1 and IL-6. In contrast, Y316 augmented the production of IL-10 in LPS-stimulated monocytes. Y316 failed to prevent the production of IL-2 and TNF in antigen-stimulated T cells, suggesting that its effects may be cell-type specific. Y316 prevented the phosphorylation and activation of the MAPK, ERK, and therefore appeared to mediate its effects on TNF by acting at an early point in the signaling cascade induced in response to LPS. There was no effect of Y316 on cAMP levels either alone or in the presence of LPS.

CONCLUSIONSY316 appears to be a small molecule inhibiting TNF production, which may act via a novel mechanism. Identification of the target of Y316 may lead to the development of alternative strategies for achieving selective cytokine inhibition.

Anti-Inflammatory Agents ; pharmacology ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Humans ; Interleukin-1 ; antagonists & inhibitors ; biosynthesis ; Interleukin-6 ; antagonists & inhibitors ; biosynthesis ; Lipopolysaccharides ; pharmacology ; Monocytes ; drug effects ; immunology ; Phosphorylation ; Tumor Necrosis Factor-alpha ; antagonists & inhibitors ; biosynthesis