Objective To explore the efficacy of calmodulin antagonist berbamine(BBM) in down-regulating survivin mRNA. Methods Human breast cancer cell line MCF7 and its adriamycin-resistant counterpart MCF7/ADR were used in the study. The cells were cultured with different concentration of BBM for 72 hours. The mRNA expression level of survivin gene in both MCF and MCF7/ADR cells was detected by semi-quantitative RT-PCR. Results After treating MCF7 and MCF7/ADR cells by 20?mol/L BBM, the mRNA expression level of survivin gene decreased from 0.43?0.02 to 0.21?0.04 in MCF7 cells, and from 0.57?0.05 to 0.45?0.04 in MCF/ADR cells(P

Objective To investigate clinical effect of mild hypothermia therapy assisted intracranial hematoma evacuation in treatment of cerebral hemorrhage.Methods One hundred and ten patients with cerebral hemorrhage were selected in Affiliated Hospital of North China University of Science and Technology from December 2011 to December 2013,and were randomly divided into two groups.Fifty-five patients treated intracranial hematoma evacuation as control group.Another 55 patients treated mild hypothermia therapy assisted intracranial hematoma evacuation as observation group.Treatment effect was compared between two groups.Results Serum S100β,neuron specific enolization (NSE) enzyme,tumor necrosis factor α (TNF-α),creactive protein(CRP),cognitive function score,daily life ability score,neurological function defect score before and after treatment in control group were (0.82±0.12) μg/L and (0.53±0.09) μg/L,(19.42±2.30) μg/L and (10.36±1.07) μg/L,(3.62±0.57) mg/L and (1.54±0.30) mg/L,(29.43±4.36) g/L and (10.25± 1.07) g/L,(13.42± 1.58) points and (25.03± 1.19) points,(21.45± 3.27) points and (37.92 ± 5.83)points,(13.27± 1.35) points and (4.84 ± 1.08) points,the differences were significant (t =8.471,11.834,17.026,22.539,12.230,10.619,25.531,P ＜ 0.05).Serum S100β,NSE,TNF-α,CRP,cognitive function score,daily life ability score,neurological function defect score before and after treatment in observation group were (0.84±0.13)μg/L and (0.41±0.10) μg/L,(19.48±1.76) μg/L and (8.75±0.84) μg/L,(3.64± ±0.61) mg/Land (1.17±0.29) mg/L,(29.58±3.62) g/L and (6.02±1.18) g/L,(13.29±1.34) points and (27.58± 1.27) points,(21.68±4.02) points and (48.26±7.14) points,(13.46± 1.21) points and (3.57±0.85) points,the differences were significant(t=13.498,16.739,25.728,41.836,13.769,15.857,36.352,P＜0.05).Compared with serum S100β,NSE,TNF-α,CRP,cognitive function score,daily life ability score,neurological function defect score before treatment,there were no difference between two groups (P ＞0.05).Serum S100β,NSE,TNF-α,CRP,neurological function defect score after treatment in observation group were lower than control group(t =5.926,4.839,6.162,10.054,6.714,P＜0.05).Cognitive function score,daily life ability score after treatment in observation group were higher than control group (t =4.008,5.973,P ＜0.05).Postoperative Glasgow prognosis classification in observation group (14 cases of grade Ⅰ,27 cases of grade Ⅱ,11 cases of grade Ⅲ,2 cases of grade Ⅳ,1 case of grade Ⅴ) was better than control group(8 cases of grade Ⅰ,12 cases of grade Ⅱ,23 cases of grade Ⅲ,7 cases of grade Ⅳ,5 cases of grade Ⅴ),the differences were significant between the two groups (Z=17.085,P =0.002).Total effective rate in observation group 94.5％ (52/55) was higher than control group 78.2％ (43/55),the differences were significant between the two groups (Z =6.253,P=0.012).Conclusion Mild hypothermia therapy assisted intracranial hematoma evacuation in treatment of cerebral hemorrhage,can significantly reduce inflammatory factor and S100βlevel,improve neurological function,has significant effect and good prognosis.It is worthy of clinical use.

OBJECTIVE:To evaluate the utilization of traditional Chinese medicine(TCM) in our hospital and to provide reference for TCM pharmacy and rational use of TCM in the clinic.METHODS:1 052 outpatient prescriptions of TCM in our hospital from May to Oct.in 2009 were analyzed statistically by Microsoft Excel program.RESULTS:Prescriptions of respiratory system diseases in our hospital were mainly TCM prescription.TCM in middle-aged and aged group was a universal phenomenon.The ingredients in every prescription were excessive with high dosage.CONCLUSION:Clinicians and pharmacists should attach great importance on the application of TCM prescriptions to ensure safety and effectiveness of drug use.

Wound secretion from 20 patients with gaseous gangrene was collected for Gram staining,bacterial culture and drug sensitivity tests.The results indicated that gaseous gangrene was caused by the co-infection of both aerobic and anaerobic bacteria.Gram-negative bacilli were slishtly more common than other aerobic bacteria in gageous gangrene wound,which was different from the findings of ordinary gaseous gangrene.

Objective To explore the efficacy of calmodulin antagonist berbamine(BBM)on multidrug resistance(MDR)reversal and its mechanism. Methods Human breast cancer cell line MCF7 and its adriamycin-resistant counterpart MCF7/ADR were used in the study.The cells were cultured with ADR and different concentration of BBM. MTT assay was used to analyze the effect of BBM on cell growth inhibition.According to the MTT assay,the 50% inhibitory concentration(IC 50 ),the multiples of drug resistance and increased sensitivity of ADR were calculated.The concentration of intracellular ADR and expression level of P-glycoprotein(P-gp)were detected by flowcytometry(FCM).The mRNA expression level of mdr1 gene was detected by semi-quantitative reverse transcriptase polymerase chain reaction(RT-PCR)with ?-actin as internal reference. Results The IC 50 of ADR in MCF7 and MCF7/ADR cells were(0.98?0.06)?mol/L and(101.20?5.72)?mol/L,respectively.The resistant multiple of MCF/ADR cells to ADR was 103 folds higher than that of MCF7 cells.BBM increased the chemo-sensitivity of ADR in MCF7/ADR cells with dose-dependent relationship,i.e.when 5*!?mol/L ,10*!?mol/L and 20*!?mol/L BBM was added into the culture the chemo-sensitivity of ADR was increased to 2.76,5.88,and 28.26 folds(P

Objective To establish a method of competitive polymerase chain reaction combined with restrictive endonucleases to measure the methylation quantitatively in MDR1 promoter region.Methods One sense primer and two anti-sense primers were designed to amplify a fragment of MDR1 gene promoter region, which was located between -102 and +186 bp containing a methylation site. The internal reference DNA fragment, witch was less 30bp than target fragment was made by three times of polymerase chain reaction(PCR)using different anti-sense primer and then subcloned into the Pbluescriptsk+ plasmid. The genomic DNA digested by Hpa, a methylation-sensitive restrictive endonuclease and competitive internal reference DNA were competitively amplified for MDR1 promoter in the same tube by PCR. The PCR products were electrophoresed on agarose gel, stained by ethidium bromide and subjected to image analysis scanner. The amount of target fragment was calculated as following, the optical density ratio of target fragment to competitive internal reference fragment multiplied the amount of competitive internal reference DNA. The ratio of PCR products amplified from HpaⅡ digested DNA and undigested DNA was named the methylation rate.Results The genomic DNA serially diluted with optimal amount of competitive internal reference DNA were co-amplified for MDR1 promoter. The significant positive correlation between the ratio of two products and the amount of genomic DNA was demonstrated. The correlation coefficient was 0.992, P

Objective To express and purify recombinant human platelet glycoprotein Ⅵ extracellular domain and prepare the polyclonal antibody against it.Methods Human platelet glycoprotein Ⅵ extracellular domain fragment (123~268 residues) was amplified by polymerase chain reaction and cloned into the prokaryotic expression vector pGEX-3x.The recombinant plasmid was constructed and expressed in E.coli after induction with isopropyl-?-D-thiogalactopyranoside (IPTG).The fusion protein was identified by Western blot analysis after purification by affinity chromatography.Rabbits were immunized with the purified fusion protein, and the collected rabbit antiserum was evaluated by sandwich ELISA, Western blot and flow cytometry.Results The coding sequence of GPVI extracellular domain was successfully inserted into pGEX-3x.Sequencing result showed that the cloned gene was identical as reported.After induction, a Mr 42kd fusion protein was expressed and confirmed by Western blot, which was identical to that expected.The titers of the antisera were up to 1∶[KG-*2]128. Sandwich ELISA result demonstrated that the prepared antibody recognized GPVI in human platelet. Western blot and flow cytometry revealed that the prepared antibody reacted with GPVI of platelet lysate and the native GPVI on human platelet surface.Conclusions Using purified prokaryotic expressed the fusion protein as antigen, the specific antibody was elicited in the immunized animals. The prepared polyclonal antibodies react specially with GPVI on human platelet surface and can be used for further studies of GPVI.

Objective To deepen the understanding of chronic eosinophilic leukemia (CEL).Methods The course of diagnosis and treatment in a case of FIP1L1/PDGFRα fusion gene negative CEL was reported. Flow cytometry was used to analyze the immunophenotype of the cells in peripheral blood and pleural fluid. Karyotype was analyzed with G-banding. The expression of FIP1L1/PDGFRα fusion gene was detected by RT-PCR technique. Routine pathological examination of the tissues from bone marrow, lung and spleen were performed. Result A sixteen-year-old girl had severe anemia, fever, splenomegaly,thrombocytopenia and dominant hypereosinophilia lasting for 22 months. Trephine biopsy showed a hypercellular marrow with eosinophilic proliferation and moderate reticular fibrosis. Eosinophilic infiltration was found in lung and spleen and embolism was also found in spleen. She had a clonal chromosomal abnormality t(5;12)(q31;p13). The expression of FIP1L1/PDGFRα was negative. An abnormal clone of T cells expressing CD3-,CD4-,CD8- was found in peripheral blood and pleural fluid, in which the cional T cell accounted for 5.43% and 1.66% of the total lymphocytes respectively. The patient was refractory to treatment with hydroxyurea, prednisone and interferon alpha. She had poor response to a combination of therapy with low dose cytosine arabinoside, mitoxantrone, vincristine, cyclophosphamide, methotrexate and prednisone. She did not respond to imatinib and died of multiple organ failure. Conclusion The present case fulfilled the WHO diagnostic criteria of FIP1L1/PDGFRα(-) CEL which did not respond to routine treatment and imatinib. Allogenic stem cell transplantation should be considered as early as possible in this case. It is noteworthy that clonal CD3-,CD4-,CD8- T-cell abnormality is related to the pathogenesis of CEL.

To explore the correlation between nucleated red blood cell (NRBC) count and perinatal asphyxia in neonates.Full-term newborns born from May 2011 to November 2012 were recruited and divided into perinatal asphyxia (n =40) and normal (n =30) groups.Apgar score was recorded immediately at delivery.The umbilical arterial blood was also collected into anticoagulant-treated tube and NRBC was counted by Japan OlympusCX41 biological microscope.NRBC count for perinatal asphyxia group [(10.70 ± 2.61)/100 WBC] was significant higher than that for normal group [(2.67 ± 0.35)/100 WBC].A statistically significant negative correlation existed between NRBC and umbilical arterial blood pH,Apgar score at 1 min,BE value (r =-0.802,P ＜ 0.05 ; r =-0.639,P ＜ 0.05 ; r =-0.566,P ＜ 0.05).Associated with perinatal asphyxia in neonates,NRBC may be used as a simple index for assessing the severity of neonatal perinatal asphyxia.

Objective To clone and express the human asialoglycoprotein receptor(ASGPR) H1 subunit, purify and identify the immunoreactivity of the recombinant protein, and establish the enzyme linked immunosorbent assay (ELISA) to detect anti-ASGPR antibodies in diagnosis of autoimmune hepatitis. Methods The CRDHI cDNA (435 bp) was subcloned into eukaryotic vector PEGH, and the recombinant protein expression was induced by D (+)-Galactose. The recombinant CRDH1 was purified with Glutathione Sepharose 4B, and its immunoreactivity was identified by SDS-PAGE and western blot as well as MALDI-TOF. ELISA was established to detect the anti-ASGPR antibodies in serum samples of 45 patients with AIH, 30 patients with SLE, 30 patients with RA, 10 patients with SS and 30 normal controls. Results The sequencing of recombinant plasmid showed the CRDH1 gene was successfully inserted to the eukaryotic expression vector with correct sequence and open reading frame. The fusion protein showed a molecular weight of 42 500 Da on SDS-PAGE gel and confirmed to be the human ASGPR by MALDI-MS through peptide mass fingerprint analysis with Mascot in human protein database. It shared 98. 34% homology with ASGPR H1 subunit. Western blot analysis showed that the fusion protein had the same immunoreactivity as human ASGPR. The results of ELISA indicated that the positive rate of anti-ASGPR was 35.6% ( 16/45 ), but the ELISA was negative in other control. There was significant difference of positivity of the autoantibodies between AIH and non-AIH controls (χ2 = 31.85,P < 0. 01 ). Conclusions The human plasmid containing ASGPR is successfully clone into Saccharomyces cerevisiae Y258. The recombinant autoantigen owns good antigenicity and specificity. ELISA established with the purified protein shows good specificity for diagnosis of AIH.