Objective To screen a cell line which stably suppresses DAPK expression and to observe the growth characters.Methods Four pairs of shRNA were designed,synthesized and inserted into the pDsRed1-N1-U6 vector.The recombinant plasmids were purified and transfected into PC12 cell.Meanwhile,a pDsRed1-N1-U6 vector was transfected as control.The cell clones were screened by G418,and the stable PC12 cell line was established.DAPK expression was detected by Western blot.MTT method and Flow Cytometry(FCM) assay were used to assess the growth characters of the cell line.Results The shRNAs were transfected into PC12 cell and the cell clones were successfully screened out.Of the four recombinant plasmids,the F2 was the best interfering shRNA.Beyond our expectation,the F1 recombinant plasmid had an enhanced effect on DAPK expression.Conclusion A stable PC12 cell line with stable inhibition of DAPK expression by was established using siRNA expression vectors.

Objective To analyze the reasons and mechanisms of stereoscopic function abnormality in elder patients after the cataract extraction with intraocular lens implantation. Methods 150 cases with artificial lens implantation were randomly selected.Routine eye examinations were carried out and the associated refractive error and presbyopia were corrected.Bilateral simultaneous visual perception and fusion function were examined.The stereoscopic function was tested using "the stereoscopic function examination diagram"created by Yan Shao-ming.Ninety-seven patients who were found to have unrecover-ed or abnormal stereoscopic function were enrolled for analysis. Results In these 97 cases,53(54.6%)were found to have preexisting eye disorders that could affect visual acuity and binocular single vision before the operation.Macular problem was the most prevalent problem in this group.Twenty-seven(27.8%)patients had complicated with corneal astigmatism,after cataract,paralysing strabismus and diplopia as well as macular edema after the operation.In addition,the contralateral unoperated cataract in 17(17.5%)patients and post-operative anisometropia in 9(9.3%)patients were also the causes of stereoscopic function abnormality.There was no reason could be identified in 8 cases. Conclusions The pre-existing eye disorders before lenses implantation,complications of the operation,contralateral unoperated cataract and anisometropia are all the major factors that affect visual acuity recovery and bilateral stereoscopic function rehabilitation.

Objective To test serum differentially expressed proteins between early-stage (stage IB-ⅢA) and late-stage (stage Ⅳ) lung cancer patients by proteinchip technology and investigate its clinical value. Methods SELDI-TOF-MS and WCX-2 protein chip were used to detect the serum protein of 30 cases of early stage lung cancer patients and 30 cases of late stage lung cancer patients. The data were analyzed by using Biomarker Wizard software. Results There are ten different proteins in the serum between the two groups of lung cancer patients. Four protein markers 7978, 8139, 15 951 and 16 133 are over expressed and seven protein markers 2867, 6885, 8701, 8840, 13 781 and 13 955 are low expressed in the late group. Conclusion SELDI-TOF-MS proteinchip technology is a convenient, sensitive and high-throughput analysis method which can screen several relatively specific protein markers for late stage lung cancer from the serum samples. This selected protein markers can predict metastasis of lung cancer patients.

Animals ; Basilar Artery ; Brain Ischemia ; pathology ; Brain Stem ; blood supply ; Cats ; Medulla Oblongata ; blood supply

OBJECTIVETo observe the expressions of Calbindin(CB) and Parvalbumin (PV), the two calcium-binding protein, in auditory pathway in mice of wild type C57BL/6J and kit⁺/kitW⁻ ²Bao, a kit gene mutant.

METHODSSix mutated kit gene kit⁺/kitW⁻ ²Bao mice and 6 wild type C57BL/6J (B6) mice were anaesthetized i. p. with chloral hydrate. After the mice were fixed by heart perfusion, the brains were removed and coronal sections were cut with a freezing microtome.

RESULTSWe found that wild type mice had significant expressions of PV on ventral cochlear nucleus, anterior part (AVCN), ventral cochlear nucleus, posterior part (PVCN), inferior colliculus (IC) and auditory cortex (AC). CB was expressed in wild type mice on PVCN and nucleus of the trapezoid body (Tz). The mutant of kit gene induced the less expression of PV on PVCN, IC and AC (P < 0.01), but increased the expression of Tz (P < 0.01). CB could not be observed on PVCN in mutant mice, and the expression of AC was increased( P < 0.01).

CONCLUSIONCB and PV has differential expression level in auditory pathway. Since mutated kit gene can affect expression of PV on PVCN, IC, Tz and AC, as well as CB on PVCN and AC, it suggests that the mutation of kit gene can affect the advanced function of central nervous system in auditory pathway.

Animals ; Auditory Cortex ; metabolism ; Auditory Pathways ; metabolism ; Calbindins ; metabolism ; Inferior Colliculi ; metabolism ; Mice ; Mice, Inbred C57BL ; Mutation ; Parvalbumins ; metabolism ; Pons ; metabolism ; Proto-Oncogene Proteins c-kit ; genetics

OBJECTIVETo investigate the effect of extracellular regulating kinase (ERK) signaling pathway on the secretion of gamma-aminobutyric acid (GABA) in cultured rat hippocampal neurons induced by stromal cell derived factor-1 (SDF-1).

METHODSThe hippocampal neurons of newborn SD rats were cultured and identified in vitro; the phosphorylation level of ERK1/2 was examined by Western blot; ELISA was used to detect the effect of PD98059, a ERK1/2 specific blocker on GABA secretion of cultured hippocampal neurons and Western blot were adopted to measure the protein expression levels of glutamate decarboxylase (GAD65/67) and gamma aminobutyric acid transporter (GAT); after blocking ERK1/2 signaling pathway with PD98059; RT-PCR was used to detect the mRNA expression levels of GAT-1 and GAD65 after treated with PD98059.

RESULTSThe levels of ERKl/2 phosphorylation were increased significantly by SDF1 acting on hippocampal neurons, and CX-CR4 receptor blocker AMD3100, could inhibit SDF-1 induced ERK1/2 activation; SDF-1 could inhibit the secretion of GABA in cultured hippocampal neurons, and ERK1/2 specific inhibitor PD98059, could partly reverse the inhibition of GABA secretion by SDF-1. The effects of SDF-1 on cultured hippocampal neurons was to decrease the mRNA genesis of glutamic acid decarboxylase GAD65 and GABA transporter GAT-1, besides, ERK inhibitor PD98059 could effectively flip the effect of SDF-1. The results of Western blot showed that SDF-1 could inhibit the protein expression of GAT-1 and GAD65/67 in hippocampal neurons and the inhibition of GAT-1 and GAD65/67 protein expression could be partially restored by ERK1/2 blocker.

CONCLUSIONSDF-1 acts on the CXCR4 of hippocampal neurons in vitro, and inhibits the expression of GAD by activating the ERK1/2 signaling pathway, and this may represent one possible pathway of GABA secretion inhibition.

Animals ; Blotting, Western ; Cells, Cultured ; Chemokine CXCL12 ; pharmacology ; Flavonoids ; pharmacology ; Glutamate Decarboxylase ; metabolism ; Hippocampus ; cytology ; MAP Kinase Signaling System ; Neurons ; metabolism ; Phosphorylation ; RNA, Messenger ; Rats ; Rats, Sprague-Dawley ; Receptors, CXCR4 ; metabolism ; gamma-Aminobutyric Acid ; secretion

Objective To investigate the clinical and imaging features in patients with vertebrobasilar dolichoectasia (VBD) through a comparative study in patients with ischemic stroke with or without VBD.Methods The patients with acute cerebral infarction were divided into either a VBD group or a non-VBD group according to magnetic resonance angiography.The VBD group was further divided into an anterior circulation infarction subgroup and a posterior circulation infarction subgroup.The cardiovascular risk factors,the diameter of basiar artery (BA),bifurcation height,and horizontal displacement were compared in all groups.Results A total of 269 patients with acute cerebral infarction were included,28 had VBD,accounting for 10.41％ of the patients with acute cerebral infarction during the same period.The proportion of male patients (78.6％ vs.66.8％ ;x2 =4.392,P =0.036),age (70.38 ± 10.58 years vs.62.86 ± 12.20 years; t =2.870,P =0.009),and the proportion of hypertension (89.3％ vs.47.7％ ; x2 =17.367,P =0.000) in the VBD group were significantly higher than those in the non-VBD group.The multivariate logistic regression analysis showed that the advanced age (odds ratio [OR] 1.248,95％ confidence interval [CI] 1.137-1.371; P=0.000),hyperglycemia (OR 1.599,95％ CI 1.181-2.164; P =0.002),hypertension (OR 1.251,95％ CI 1.020-1.534; P =0.032) and increased triglyceride level (OR 1.876,95％ CT 1.021-3.445; P =0.043) were the independent risk factors for VBD,while female gender (OR 0.133,95％ CI 0.024-0.735; P =0.021) was the independent protective factor for VBD.Of the 28 cerebral infarction patients with VBD,9 had anterior circulation infarction and 19 had posterior circulation infarction.There were significant differences in BA diameter ([5.40 ± 0.49] cm vs.[6.00 ± 0.77] cm; t =2.046,P =0.041),and the proportions of high score in bifurcation height (x2 =6.768,P =0.037) and horizontal displacement (x2 =5.241,P =0.042) between the 2 groups (all P ＜0.05).The multivafiate logistic regression analysis showed that the BA bifurcation height was an independent risk factor for posterior circulation infarction (OR 1.347,95％ CI 1.069-2.457; P =0.038) in patients with VBD.Conclusions VBD accounted for 10.41％ of the patients with acute cerebral infarction during the same period.Advanced age,hyperglycemia,hypertension and increased triglyceride level were the independent risk factors for VBD.Female gender was the independent protective factors for VBD,and the BA bifurcation height was an independent risk factor for VBD occurring posterior circulation infarction.

BACKGROUND: Color Doppler flow imaging can exam the early myocardial disorder in type 2 diabetic patients. What does exercise tolerance test work for the examination of such disorder in combination with color Doppler flow imaging (CDFI) ?OBJECTIVE: To analyze in comparison the early evaluation on reduced diastolic function in left ventricle in patients with type 2 diabetes between the examinations of exercise tolerance test combined with color Doppler flow image and simple color Doppler flow imaging.DESIGN: Cases-controlled comparison and self-comparison.SETTING: Department of electrodiagnosis and department of Endocrinology in a municipal hospital.PARTICIPANTS: Thirty-six cases of inpatients with type 2 diabetes were selected from Department of Endocrinology of Shenyang Red Cross Hospital from March to December in 2004, of which, 25 cases were males and 11cases females. The diabetic patients included had no cardiac vascular complications and participated in the study in volunteer. Thirty-two patients who received annual routine health check at the same period were selected as the control, of which, 20 cases were males and 12 cases females.METHODS: Metronics treadmill exercise test equipment was used for exercise tolerance in two groups. Before exercise(at quiescent state) and after exercise tolerance, Vivid 4 CDFI was used to determine flow velocity at E and A peak values respectively, ratio between flow velocities at E peak value and A peak value as well as isovolumtric relaxation time (IRT).MAIN OUTCOME MEASURES: Comparison of cardiac functional indexes before and after exercise tolerance in two groups.RESULTS: Thirty-four diabetic patients accomplished exercise tolerance test, of which, 1 case presented frequent ventricular extrasystole, another one presented precordial pain and stopped the test. In the control, all of 32 cases ratio between flow rates of E peak value and A peak value was remarkably lower than that in the control (0. 90 ± 0. 25, 1.40 ± 0.30, P ＜ 0.05 ); IRT was remarkably longer than that in the control [ (112. 07 ± 20. 16),imental group, the ratio between flow rates of E peak value and A peak value was remarkably lower than that in the control (0.62 ±0. 12, 1.28 ±0.87, P＜ 0.01 ); IRT was remarkably longer than that in the control[ (138. 10± 19.21), (97.37±9.61) ms, P ＜0.01].CONCLUSION: CDFI can supervise and evaluate at early stage the cardiac functional changes in type 2 diabetic patients. Due to the induction of exercise tolerance, the combination of exercise tolerance test and CDFI provides more accurate, objective and valuable conclusions at early stage.

Objective:To investigate the relationship between base level of Cyfra21-1,SCCA,TK1 and lung cancer prognosis.Methods: A nested case-control study was conducted.721 lung cancer cases who had no distant metastasis were recruited baseline population from January 2010 to January 2013.About 2 years follow-up,364 cases of death or brain (or multiple) metastasis were identified as case group, and the other 357 cases were included in the control group.The level of serum Cyfra21-1,SCCA,TK1 was detected.The relationship between base level of Cyfra21-1,SCCA,TK1 and lung cancer prognosis were analyzed.Results: The age in the case group was (59.3±10.1),and the control group was (59.0±9.9).There were obvious differences in body mass index,smoking index,pathological type,clinical stage,lymph node metastasis and with chronic diseases between case group and control group(P<0.05),while no difference in pleural effusion,comprehensive treatment etc(P>0.05).There were differences in the base level of Cyfra21-1,SCCA,TK1;and there were differences in Cyfra21-1,SCCA,TK1 distribution between case group and control group(P<0.05).There was difference in the base level of Cyfra21-1(P<0.05),while no differences in SCCA,TK1(P>0.05) in patients with different stages of lung cancer.There were differences in the base levels of Cyfra21-1 and SCCA(P<0.05),while no differences in TK1(P>0.05) in patients with different pathological types of lung cancer.Logistic regression analysis results showed that the OR value of SCCA,TK1 with lung cancer prognosis were respective 7.235(1.674-14.613),5.009(0.973-10.778),5.816(0.879-16.235).Conclusion: The baseline level of Cyfra21-1 can reflect the prognosis of lung cancer patients,while SCCA,TK1 not.

0.05).CONCLUSION:Gatifloxacin exhibited no effect on the pharmacokinetics of doxofylline in healthy human body when the two drugs used concomitantly.