Purpose To explore the relationship between the mutations of epidermal growth factor receptor ( EGFR) and KRAS genes and clinicopathological characteristics in patients with non-small cell lung cancers (NSCLC). Methods Clinical samples from 431 NSCLC patients were obtained for EGFR and KRAS gene analysis. PCR based direct DNA sequencing was used to investigate mutations in exon 18-21 of EGFR gene and codon 12 and 13 of exon 2 of KRAS gene. Results The overall EGFR mutation rate of primary NSCLC was 53. 6% (231/431) in this study cohort and eight cases showed double EGFR mutations. Mutation rates in female and male were 65. 2% (122/187) and 46. 9% (98/209), respectively. The mutation rate was higher in patients with non-smokers and adeno-carcinoma and adenosquamous carcinoma subtypes than in their counterparts (P<0. 05), with the percentage of 57. 2% (124/216), 60. 3% (199/330), 42. 9% (6/14), respectively. In squamous cell carcinomas and other subtypes, EGFR mutation rates were 11. 6% (5/43) and 11. 1% (1/9), respectively. The EGFR mutation types included exon 18 point mutations (4. 0%, 9/227), exon 19 deletion mutations (4. 5%, 101/227), exon 20 insert or point mutations (9. 7%, 22/227) and exon 21 point mutations (41. 4%, 94/227). Activating mutations of KRAS gene were detected in 7. 8%(31/396) of NSCLC. Twenty-eight patients showed codon 12 mutations ( G>T, G>A, G>C) , and three patients had codon 13 mutations ( G>A, G>T) . Most of these mutations were G to T transversion (64. 5%, 20/31). Conclusion Polymerase chain reaction-direct sequencing is a reliable and effective method for the detection of the EGFR and KRAS gene mutation in NSCLC patients. The mutation rate of EGFR is higher in Chinese patients, especial-ly in non-smoking female patients with adenocarcinoma.

Objective To investigate the potential clinical factors associated with the prognosis and relapse of adult onset Still's disease(AOSD).Methods The factors possibly influencing the prognosis and relapse of AOSD were analyzed by logistic regression and COX regression in the cohort study.Ninety-six con- secutive inpatients of AOSD diagnosed based on Yamaguchi criteria in the hospital from March 1996 to September 2004 were included in the study.Results Nine cases(9.4%)were lost during the follow-up. Eleven patients(12.6%)were diagnosed as other diseases(5 with other rheumatic diseases,4 with tumor and 2 with infections)in the 87 follow-up cases.In 76 cases,3 patients(3.9%)died and 33 patients(43.4%) got remission over one year after treatment.Splenomegaly(OR=3.14,95%CI=1.01～9.74)and treated with methotrexate(OR=0.22,95%CI=0.07～0.67)were associated with the prognosis from the logistic regression analysis of the 76 cases.The serum ferritin(RR=I.05,95%CI=1.01～1.08)and treated with methotrexate (RR=0.13,95%CI=0.02～0.76)were associated with relapse from the COX regression analysis of the 61 remis- sion cases.Conclusion We need to be very cautious in the follow-up of AOSD patients because some of them may change to other diseases.Methotrexate may be an importent therapy of AOSD not only in improve- ment the prognosis but also in reduction of relapse.

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.

Through protein-protein BLAST of homologous sequences in different species in NCBI database and preliminary simulating molecular docking and molecular dynamics by computer software discovery studio 3.1, three amino acids R25K26K27 of natural human parathyroid hormone (1-34) with Q25E26L27 were mutated and the biological activity of the mutant peptide was evaluated. Result showed that: root mean superposition deviation RMSD value between PTH (1-34)-(RKK-QEL) and PTH (1-34) peptide main chain was 2.509 3, indicating that the differences between the two main chain structural conformation was relatively small; the interaction energy between PTH (1-34)-(RKK-QEL) and its receptor protein PTH1R had been enhanced by 7.5% compared to nature PTH (1-34), from -554.083 kcal x mol(-1) to -599.253 kcal x mol(-1); the number of hydrogen bonds was increased from 32 to 38; PTH (1-34)-(RKK-QEL) can significantly stimulate the RANKL gene expression (P < 0.01) while inhibiting the OPG gene expression (P < 0.01) in UAMS-32P cells; in the co-culture system of UAMS-32P cells and mouse primary femur bone marrow cells, PTH (1-34)-(RKK-QEL) stimulated the formation of osteoclasts (P < 0.01) and had a higher biological activity than PTH (1-34) standard reagents.

OBJECTIVETrying to find out the mechanism of microstructure influencing bacterial adhesion, we prepared different microstructures to compare the adsorptive behavior of graphite powder and adhesive behavior of oral microbe.

METHODSWe used polydimethylsiloxane (PDMS) to copy 23 microstructures of hydroxyapatite (HA) chip, and cultured them with different sizes graphite powder and oral microbes respectively, to observe and compare their behavior on microstructures.

RESULTSThe adsorption of 30-50 microm powder on different microstructures was insignificant, while 10-20 microm powder had maximum adsorption on 10 microm and 20 microm microstructures. Saccharomyces albicans was most likely to adhere to 5 microm microstructures which was equivalent to its own size. However, microstructures had little effect on adhesion of Streptococcus mutans which grew in a chain.

CONCLUSIONThe size of microstructure was the most effective factor that affects the adsorption of non-living powder, and it also had the same effect on the microbial adhesion; but some special bacteria, such as Streptococcus mutans which grew in a chain, was not affected by the sizes or shapes of microstructures.

Adsorption ; Bacteria ; Bacterial Adhesion ; Durapatite ; Graphite ; Mouth ; microbiology ; Streptococcus mutans

Objective To explore the role of white matter injuries in the schizophrenia induced by the NMDA re-ceptor antagonist. Methods Adult male C57BL/6J mice (8 week old) were equally divided into four groups. One group was sub-chronically treated with saline solution, and the other three groups were intraperitoneally treated with MK-801 at dose of 0.025 mg/mL (M1), 0.050 mg/mL (M2) and 0.100 mg/mL (M3) in a volume 10 ml per kilogram body weight. All ani-mals were tested using Morris water maze at the 9th-15th day and using the Hole Board exploration as well as Rota Rod performance tests on the 16th day. The myelin basic protein (MBP) and the ultrastructure of the myelin sheaths in the cor-pus callosum were then examined using immunohistochemical methods, transmission electron microscope technique and stereological methods. Results The repeated sub-chronic MK-801 treatment did not induce impairment of spatial learning and memory in Morris water maze. The MK-801 treatment at 0.25 mg/kg and 1.00 mg/kg but not at 0.50 mg/kg resulted in less exploration to a new environment. The myelin staining with anti-MBP antibody was less intense in all three schizo-phrenic groups when compared to saline control group (P<0.01). Furthermore, MK-801 treatment caused pathological al-terations of the myelin sheaths including segmental demyelination of myelinated fibers and splitting of myelin sheath lamel- lae in schizophrenic groups. The ratio of the injured myelinated nerve fibers in the corpus callosum of MK-801 treated mice [M3 group, (22.42 ± 4.24)%] was significantly higher when compared to the control mice [(3.84 ± 1.35)%,P<0.01)]. Conclusions The present study demonstrated the white matter damages, mainly low MBP expression and segmental demye-lization in the corpus callosum in the mice sub-chronic treated with MK-801, indicating that the white matter changes might be involved in the schizophrenia induced by NMDA antagonist.

OBJECTIVETo study the levels of CD4+CD25+CD127- and CD3+CD4-CD8- regulatory T (Treg) cells in peripheral blood of children with idiopathic thrombocytopenic purpura (ITP).

METHODSThe flow cytometry was used to detect the expression of CD4+CD25+CD127- and CD3+CD4-CD8- Treg cells in peripheral blood of 33 children with ITP and 21 healthy children.

RESULTSThe expression levels of CD4+CD25+CD127-［(2.7±1.7)% vs (4.8±1.6)%; P<0.01］and CD3+CD4-CD8-［(5.2±3.1)% vs (8.1±3.5)%; P<0.01］Treg cells in children with ITP were significantly lower than in the controls.

CONCLUSIONSThe expression levels of CD4+CD25+CD127- and CD3+CD4-CD8- Treg cells decrease in children with ITP, suggesting that CD4+CD25+CD127- and CD3+CD4-CD8- Treg cells might play a role in the pathogenesis of ITP.

Adolescent ; CD3 Complex ; analysis ; CD4 Antigens ; analysis ; Child ; Child, Preschool ; Female ; Flow Cytometry ; Humans ; Infant ; Interleukin-2 Receptor alpha Subunit ; analysis ; Interleukin-7 Receptor alpha Subunit ; analysis ; Male ; Purpura, Thrombocytopenic, Idiopathic ; etiology ; immunology ; T-Lymphocytes, Regulatory ; immunology

Purpose To investigate the positive rate and concordance rate of BRAF mutation in papillary thyroid carcinoma detected by real-time PCR method and Sanger sequencing. Methods 312 papillary thyroid carcinomas patients were enrolled in this study. Real-time PCR method and Sanger sequencing were performed to detect BRAF gene mutations. The frequency of BRAF mutation and the concordance of two methods were analyzed. Results BRAF mutation was detected in 65. 4% (204/312) and 63. 8% (199/312) of 312 papillary thyroid carcinoma samples by using real-time PCR method and Sanger sequencing, respectively. There was no significant correlation between BRAF gene mutations and patients’ gender. There was significant correlation between BRAF gene mutations and patients’ age. The overall concordance between real-time PCR method and Sanger sequencing for BRAF mutation detection was 98. 4%. Conclusion Real-time PCR method provides an effective method in BRAF gene mutation detection.

OBJECTIVETo evaluate the molecular mechanism and prognostication of bcl-2 protein expression in different subgroups of diffuse large B-cell lymphoma(DLBCL) in Guangxi Zhuang Autonomous Region, China.

METHODSImmunohistochemical stains for CD10, bcl-6, MUM-1, bcl-2 and NF-κB were performed in 214 cases of DLBCL. The Hans immunologic classification was applied to classify DLBCL into GCB and non-GCB subgroups. Using a dual-probe fluorescence in-situ hybridization (FISH) assay, IgH/bcl-2 gene translocation and bcl-2 amplification were analyzed.

RESULTSIn 214 cases of DLBCL, 30.8% (66/214) of cases were GCB and 69.2% (148/214) were non-GCB. Twenty-seven point three percent (18/66) of GCB subgroups and 59.5% (88/148) of non-GCB subgroups had bcl-2 protein expression, with a significant difference (P < 0.01). IgH/bcl-2 translocation was positive in 3.7% (8/214) of cases, even majority of them (6/8) was found in GCB subgroup, while represented only 9.1% of GCB case. There was a significant difference (P = 0.02) in bcl-2 gene amplification between GCB (27/66, 40.9%) and non-GCB subgroup (86/148, 58.1%). Among non-GCB cases, the expression of bcl-2 was correlated with that of NF-κB expression and bcl-2 gene amplification (r = 0.216 and 0.219, respectively, P < 0.05). No similar correlation was observed in GCB cases. The overall survival time of bcl-2-positive patients (31.4 ± 3.8) months was shorter than that of bcl-2-negative patients (40.2 ± 4.2) months. In conjunction with immunophenotypes and clinical stages, the bcl-2 positive patients had a 1.89 times higher risk than that of the bcl-2 negative patients.

CONCLUSIONSMajority of the cases were prognostically unfavorable non-GCB subgroups among DLBCL, which were characterized by high frequency of bcl-2 gene amplification and low frequency of IgH/bcl-2 translocation. The anti-apoptotic gene bcl-2 was frequently expressed in non-GCB subgroups and closely related to the gene amplification and NF-κB activation. bcl-2 positive patients had more short overall survival times, would face significant higher risk of death, these results suggested that bcl-2 could be a prognostic marker independent to clinical staging and immunophenotyping.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Antineoplastic Combined Chemotherapy Protocols ; therapeutic use ; Child ; Child, Preschool ; Cyclophosphamide ; therapeutic use ; Doxorubicin ; therapeutic use ; Female ; Gene Amplification ; Gene Expression Regulation, Neoplastic ; Genes, bcl-2 ; Humans ; Immunoglobulin Heavy Chains ; genetics ; In Situ Hybridization, Fluorescence ; Lymphoma, Large B-Cell, Diffuse ; classification ; drug therapy ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; NF-kappa B ; metabolism ; Prednisone ; therapeutic use ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; Survival Rate ; Translocation, Genetic ; Vincristine ; therapeutic use ; Young Adult

OBJECTIVETo investigate the effect of mito chondrial K(ATP) channels (mitoK(ATP)) inhibitor 5-hydroxydecanoate(5-HD) on chronic hypoxic pulmonary artery hypertension (CHPAH) rats and its underlying mechanisms.

METHODSForty-eight male SD rats were equally divided into 4 groups randomly (n=12): normal group, hypoxia group, hypoxia + 5-HD group, hypoxia + Diazoxide group. Except the first group, the other three groups were put into hypoxic [O2 (10.0% +/- 0.3%] and nonrmobaric chamber for four weeks to establish chronic hypoxic model and received different interference. When the interference completed, right heart catheter was used to detect the mean pulmonary arterial pressure (mPAP) of each rat and PKC-alpha mRNA expression in pulmonary arteries was detected by reverse transcription-polymerase chain reaction (RT-PCR) and protein expression by Western blot.

RESULTS(mPAP was much higher in hypoxia group than that in normal group (P < 0.01) while in hypoxia + 5-HD group and hypoxia + diazoxide were decreased significantly compared to hypoxia group (P < 0.01). (2) The protein and mRNA levels of PKC-alpha in the hypoxic group were higher than those in normal group (P < 0.05).

CONCLUSION5-HD plays a protective role on CHPAH. The mechanism of its effect may be attributed to inhibiting MitoK(ATP).

Animals ; Decanoic Acids ; pharmacology ; Hydroxy Acids ; pharmacology ; Hypertension, Pulmonary ; etiology ; metabolism ; physiopathology ; Hypoxia ; complications ; physiopathology ; Male ; Muscle, Smooth, Vascular ; metabolism ; Potassium Channel Blockers ; pharmacology ; Potassium Channels ; drug effects ; Protein Kinase C-alpha ; genetics ; metabolism ; Pulmonary Artery ; metabolism ; Rats ; Rats, Sprague-Dawley