Objective To construct prokaryotic recombinant plasmids of transcriptional co-activator (TC) gene of Clonorchis sinensis, express and purify the recombinant protein and analyze its biological function. Methods A pair of primers was designed according to the known sequence of TC gene. The TC gene fragment was amplified by PCR. After purification and digestion with BamHⅠ and SalⅠ , the TC gene was connected to the prokaryotic expression vectors, pGEX-4T-1 and pET30a(+). By cloning target gene into these vectors, pGEX-4T-1 and pET30a(+), prokaryotic recombinant plasmids of TC gene were constructed and transferred into E.coli BL21. The positive expressed recombinants were detected by SDS-PAGE and Western blotting. Immobilized metal (Ni 2+ ) chelation affinity chromatography was used to purify His-TC produced by the expression of the recombinant protein pET30a(+)-TC. Results The recombinant plasmids, pGEX-4T-1-TC and pET30a(+)-TC, were constructed successfully. SDS-PAGE testified that the molecular weight of the recombinant protein was correct. Western blot analysis of GST-TC recombinant protein testified that the recombinant protein could be recognized by immunized rabbit serum, which means the protein is GST-immune active and the clone can express recombinant Clonorchis sinensis antigen. After affinity chromatography of the pET-TC protein, there was only one protein band with expected size on the SDS-PAGE gel. Conclusion The TC gene was screened from cDNA library of adult Clonorchis sinensis, cloned, expressed and purified. The purified protein of TC gene will be of importance for further research on the biological function of the gene.

Objective To observe the clinical effects of a new method:non-optic nerve transecring evisceration combined with first stage hydroxyapatite orbital implantation.Methods One hundred and twenty two eyes (122 cases) were randomly divided into group A and group B,evisceration was first undertaken,the scleral wall was superior-temporally to inferior-nasally dissected and double scleral petals were placed before the implanted hydroxyapatite artificial eyeball.The difference between group A and group B was:in group A,the optic nerve was transected behind the eyeball,but in group B,a 2 mm outside the optic nerve scleral circle cutting was taken.Hemostasis time before implantation,tensile force during conjunctival suture,pain during surgery,palpebral fissure height after eye speculum wearing,operation time consuming,pain after surgery,conjunctival edema and congestion,and ocular prosthesis wearing time were compared between the two groups.Results In groups A and B,the intra-operative hemostasis time were (23.46 ± 6.96)mins and (5.49 ± 1.72)mins,the tensile force score during conjunctiva suture were 3.39±0.74 and 0.45±0.59,the score of pain in the first day were 2.8 ±0.68 and 1.47 ± 0.67,the ocular prosthesis wearing time after surgery were (6.27±2.73) and (3.07 ± 2.11)weeks,respectively.The differences of all the parameters above were with statistical significance between groups A and B (P＜0.01).During 2 years follow-up,no complications such as orbital implant exposure or infection happened.Conclusions Compared to traditional methods,the non-optic nerve transecting evisceration method has merits of less impairment,short time-consuming,less pain,and quicker postoperative recovery.

Objective To observe the changes of intedeukin (IL)-17 in patients with maintenance hemodialysis (MHD) and its relationship with anemia. Methods Twenty sera from normal people(normal group) and sixty-eight sera from patients with MHD were sampled. Patients with MHD were divided into 4 groups by haemoglobin (Hb) levels: severity anemia group (12 cases), midrange anemia group(18 cases),light anemia group(18 cases) and non-anemia group(20 cases). IL-6 and IL-17 levels were measured in all the sera with ELISA. Meantime, C-reactive protein (CRP),Hb, ferritin and iron saturation ratio was observed. Results CRP,IL-17 and IL-6 levels in MHD patients were much higher than those in normal group. IL-17 levels in severity and midrange anemia group were higher than those in light anemia group,and IL-17 was positively correlated to Hb. Conclusions IL-17,IL-6 and CRP in patients is higher than that in normal people,and the microinflammation status exists in MHD. IL-17 may play a role in pathogenesis of microinflammation in MHD patients with anemia.

A reliable method for simultaneous determinition of eleven representative components (neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, isochlorogenic acid B, isochlorogenic acid A, isochlorogenic acid C, shanzhiside, geniposidic acid, genipin-1-β-D-gentiobioside, geniposide and secoxyloganin) in combination of chromatographic fingerpint analysis for Reduning injection was developed by ultra high-performance liquid chromatography (UPLC). The method was performed on an Agilent ZORBAX SB-C18 anlytical column (3. 0 mm x 100 mm, 1. 8 µm) with a guard column of Agilent UPLC Guard ZORBAX SB-C18 (3.0 mm x 5 mm) at the column temperature of 30 °C. The gradient mobile phase consisted of acetonitrile (A)-0. 1% phosphoric acid (B) with a flow rate of 0. 4 mL . min-1. The injection volumn was 2 µL. The detection wavelengths were set at 324 nm and 238 nm for quantit tive analysis and 225 nm for fingerpint chromatography. Neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, isochlorogenic acid B, isochlorogenic acid A, isochlorogenic acid C, shanzhiside, geniposidic acid, genipin-1-β-D-gentiobioside, geniposide and secoxyloganin were baseline seperated with good linearity relationships (r >0. 999) between concentration and peak areas over the linear ranges. The average recoverys of the investigated compounds were 103.5%, 100. 2%, 103. 3%, 102. 8%, 101. 3%, 102. 8%, 97. 36%, 99. 62%, 98. 16%, 102. 8%, 99. 27%, respectively. Reduning injection of forty-five batches was analyzed by UPLC finge print chromatography. Thirty batches were selected to generate the reference fringerprint chromatography with fourteen common peaks. The similarity values between the reference fringerprint chromatography and the remaining fifteen batches were higher than 0. 99. The developed method was fast, accurate and sensitive. It could be used as a reference for the quality control of multiple components determination and fingerprint chromatography for Reduning injection in future.
Chlorogenic Acid ; chemistry ; isolation & purification ; standards ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; standards ; Glucosides ; chemistry ; isolation & purification ; standards ; Iridoids ; chemistry ; isolation & purification ; standards ; Quality Control ; Reference Standards ; Reproducibility of Results ; Sensitivity and Specificity ; Time Factors

Objective:To study effects of neuroligin (NLG) on the proliferation and apoptosis in human neuroblastoma SH-SY5Y cells.Methods:The SH-SY5Y cells were cultured in vitro for 24 hours,and then transfected with NLG siRNA at dose of 50,100,200 μmol/L,respectively.MTT procedure was used to detect the cell proliferation,and expression levels of apoptosis gene including Bax or Bcl-2 and Bcl-xl were measured by RT-PCR.Results:Compared to control groups proliferation of SH-SY5Y cells were distinctly inhibited after NLG siRNA transfection accompany with a dose-dependent,which was caused by activation of apoptosis.Conclusions:NLG protect neuron by inhibiting apoptosis.

Objective To construct the recombinant plasmid pYES6-S and express and purify spike protein of severe acute respiratory syndrome(SANS)coronavirus in Saccharomyces cerevisiae. Methods DNA fragments of SANS coronavirus were obtained by reverse transeription.Four over- lapped fragments of spike protein genes were amplified by polymerase chain reaction(PCR)and ligated into an integral spike protein gene by restriction enzyme digestion.The spike protein gene recombined with pYES6 and cloned into E.coll.The recombinant plasmid pYES6-S was induced and expressed in Saccharomyces cerevisiae(INVScl)by galactose.Results The recombinant plasmid pYES6-S was confirmed that inserted fragment was right in length,direction and base matching by restriction enzyme digestion and sequencing.The purified protein encoded by the whole spike protein gene was about Mr 110?10~3 identified by electrophoresis.Conclusion The whole spike protein gene of SARS coronavirus is cloned into E.coli and the protein is expressed in Saccharomyces cerevisiae successful ly.which can be helpful in SARS vaccine research.

Objective To analyse the spectral patterns of complementarity determining region 3 (CDR3) length distribution of T lymphocyte receptor beta chain variable (TRBV) gene families in infiltrating T cells of the liver tissues and the peripheral blood samples of patients with chronic hepatitis B (CHB) in order to evaluate the characteristics of T cell clonal expansion. Methods The spectral patterns drift of TRBV gene families (the monoclonal/oligoclonal TCR β T cells) in the peripheral blood and hepatic tissues from 11 cases of CHB patients were analyzed by the real-time fluorescence quantitative reverse transcription polymerase chain reaction (FQ-PCR) with DNA melting curve analysis, and abnormal rates of TRBV gene families were compared between CHB patients and healthy control. The comparison of rates was done by chi square test. Results The gene melting spectral pattern of 26 TRBV families of the 11 CHB patients, no matter in the peripheral blood or hepatic tissue, showed either a single peak or prominent melting peaks, even disappeared for certain TRBV families. The abnormal rate of TRBV gene families in the hepatic tissues was significantly higher than that in the peripheral blood samples (x2 = 23. 246, P＜0. 01). What is more interesting was that some parts of TRBV families were identical in both the peripheral blood and the hepatic tissue in certain patients. TCR BV13.1, TCR BV17 and TCR BV22 fragments were found to be restricted used in both the peripheral blood and hepatic tissue by some CHB patients. Conclusions T cells in the peripheral blood and the hepatic tissues of CHB patients can develop clonal expansion to some extent.Parts of TRBV families are restricted used in the peripheral blood and hepatic tissue in some CHB patients, which offers a foundation for further studying the common specific spectral drift patterns of TRBV CDR3 gene in CHB patients.

Objective To investigate the risk factors, clinical and angiographic features of women aged 50 or less with coronary artery disease(CAD). Methods One hundred and seventy-three female CAD patients comifrmed by coronary angiographic aged 50 or less were classiifed as group A, while another 494 non-CAD women aged 50 or less as group B. The differences in CAD risk factors, clinical and angiographic features between the 2 groups were analyzed. Results There were more women with diabetes, positive CAD family history, dyslipidemia or hypertension (especially diastolic hypertension) in group A than in group B. Patient in group A had higher diastolic pressures and serum glucose level than those in group B but both groups had similar body weights, systolic pressures and menopause ages. The serum total cholesterol and triglyceride levels were higher in patients in group A than those in group B while high-density-lipoprotein (HDL) cholesterol and apolipoprotein A levels were lower in group A. The low-density-lipoprotein (LDL) and apolipoprotein B were higher in group A than in group B but without signiifcance. There were more women with positive urine protein in group A than in group B. In group A, more than 50%of patients were with single diseased artery while another 15%with slight coronary artery atherosclerosis or even normal arteries. Most of the lesions were found in left anterior descending artery (LAD) and its branches. Conclusions Risk factors of CAD included diabetes, positive CAD family history, dyslipidemia, hypertension(especially diastolic hypertension) and positive urine protein in women aged 50 or less Menopause alone, without other CAD risk factors, would not lead to CAD. Single vessel disease was more commonly found in this group of patients.

Objective To evaluate the CT and MRI characteristics of Primary Central Nervous System Lymphoma(PCNSL)in immunocompetent patients,and enhance its diagnosis level.Methods CT and MRI data of 20 patients with PCNSL confirmed by histo-pathology were analyzed retrospectively.MRI scans were performed with and without Gadolinium contrast.Two of them had contrast-enhanced CT scan;six had CT scan without contrast administration;1 had CT scan with both non-contrast and contrast enhancement.Re- suits Totally,38 lesions were found in all patients:14 lesions of them were single and 24 lesions were found in 6 patients.Generally,the lesions were located in the surface and/or midline of the brain.The signal features and density were similar to meningioma,and strongly enhancing after contrast administration.Thirty-six of the 38 lesions had spicular sign peripheral to the lesion.Conclusion Although the manifestations of the PCNSL are variety,there are still many characteristics in the medical imaging,especially in the locations,the signal features,and spicular sign in the edge of the lesions after contrast material injection.

0.05)and the TNM staging (P=0.55).A mild elevated compared other pathological classification was found in small cell lung cancer (0.191?0.275).Conclusions The results showed that RFQ-PCR was suitable for measurement of the mRNA level of PLKI in bronchoscopic bioptic specimens.This study suggest elevated expression of PLK1 might play a important role in development of lung cancer,so that PLK1 might be a potential tumor marker for Lung cancers.Advanced studies will be needed to clarify that PLKI mRNA level do not relate to TNM staging and pathological classification.