Acute Disease ; Case-Control Studies ; Child ; Child, Preschool ; Female ; GATA3 Transcription Factor ; genetics ; Humans ; Infant ; Interferon-gamma ; genetics ; Interleukin-4 ; genetics ; Male ; Mucocutaneous Lymph Node Syndrome ; blood ; genetics ; immunology ; RNA, Messenger ; Reverse Transcriptase Polymerase Chain Reaction ; T-Box Domain Proteins ; genetics ; Th1 Cells ; immunology ; Th2 Cells ; immunology

Objective: To study the influence of 5′-untranslated region modification of pPIC9 on expression of LL-37 in Pichia pastoris. Methods: The sequence GGATCCAA was deleted from 5′-UTR of pPIC9 and the modified product was transformed into E. coli DH5α to construct a modified eukaryotic vector pPIC9-EDIT. After PCR and sequencing, pPIC9-EDIT was ligated with LL-37 sequence coded by the biased codon of yeast, the product was then transformed into E. coli DH5α to construct the recombinant expression vector pPIC9-EDIT-LL-37, the latter was transformed into P. pastoris GS115 by spheroplasting and the insert was confirmed by PCR. The bacteriolytic activity to E. coli. DH5α was analyzed to screen the highest expressing strain and to determine the best inducing time and concentration of methanol. The fermentation product was analyzed by Tricine-SDS-PAGE and Western blotting. The antibacterial activities of expression products of pPIC9-LL-37 and pPIC9-EDIT-LL-37 were compared, and the changes of LL-37 protein expression were determined before and after modification. Results: pPIC9-EDIT and pPIC9-EDIT-LL-37 were successfully constructed. Expression of LL-37 gene was confirmed by PCR in P. pastoris after pPIC9-EDIT-LL-37 transformation. The highest expressing strain was identified; the best inducing time was 72 h and the best concentration of methanol was 0.5%. Tricine-SDS-PAGE and Western blotting analysis showed that the expression product was LL-37. The expression level of LL-37 protein increased by 35 times after modification. Conclusion: Modification of pPIC9 5′-UTR can obviously improve expression of LL-37 protein in P. pastoris; it is worth to be used in the research of other heterogenous protein.

Objective: To construct a eukaryotic expression vector of human antimicrobial peptide LL-37 (pPIC9-LL-37) and to express it in P. pastoris. Methods: The full-length gene encoding antimicrobial peptide LL-37 was synthesized by overlap extension-PCR method using the sequence of LL-37 and P. pastoris biased codon. The full-length gene was cloned into pPIC9 vector and the product was transformed into E. coli DH5α to construct expression vector pPIC9-LL-37. After identification by PCR and sequencing, pPIC9-LL-37 was used to transfect P. pastoris. The expression of LL-37 was induced by methanol and the highest expressing strain was screened. The concentrated fermentation product was analyzed by Tricine-SDS-PAGE and Western-blot, and the antibacterial activity of the expression product on E. coli. DH5α was tested. Results: The eukaryotic expression vector pPIC9-LL-37 was successfully constructed. The fusion of LL-37 gene into P. Pastoris was confirmed by PCR. High expression of LL-37 was expressed by 0.5% methanol and the highest expressing strain was screened out. The fermentation supernatant contained 0.5 μg/ml LL-37 and had strong antibacterial activity against E. coli. Tricine-SDS-PAGE and Western-blot analysis confirmed that the product was LL-37. Conclusion: We have successfully constructed pPIC9-LL-37 for transfecting P. pastoris. Methanol can induce the high expression of LL-37 and the expressed LL-37 has strong antimicrobial activity.

Objective To synthesize glycylproline p-nitroanilide tosylate as a substrate for glycylprolyl dipeptidyl-aminopeptidase(GPDA),and to study its clinical application.Methods The title compound was prepared by DCC-HOBt synthesis.GPDA in human serum was detected by continuous monitoring method using substrate. Results The substrate with purity of 98.5% was obtained.The linearity of the method was up to 358.1 U/L.Intraassay CV and interassay CV of GPDA in diverse serum samples were 3.01% and 5.04%,respectively.It was shown that GPDA level in patients with hepatic cancer was significantly increased,while those in patients with gastric carcinoma and chronic gastritis were significantly decreased compared with that in the control group in clinical detecting(P

Animals ; Drugs, Chinese Herbal ; pharmacology ; Female ; Heart Failure ; drug therapy ; physiopathology ; Hemodynamics ; Male ; Myocardial Ischemia ; drug therapy ; physiopathology ; Phytotherapy ; Rats ; Rats, Wistar ; Rhodiola

Adult ; Aged ; Cohort Studies ; Dust ; analysis ; Female ; Humans ; Male ; Manganese ; analysis ; Manganese Poisoning ; diagnosis ; Middle Aged ; Occupational Exposure ; analysis ; Retrospective Studies ; Young Adult

ObjectiveTo explore the influence of electrical stimulation on nervous cells.MethodsOn 5th day the brain primitive nervous cells of rat brain, in vitro culturing, were divided into 4 groups, in order of control group, electrical pole control group, low stimulation group, and high stimulation group. The cells were stimulated with electro-acupuncture apparatus for 7 days, 30 minutes once, twice a day. On 10th day, the cells were observed microscopically for 3 days and taken photos.ResultsElectrical stimulation is helpful to raise the survival rate of cell.Conclusions Electrical stimulation influences morphological change of nervous cells during culturing.

ObjectiveTo study the effects of aqueous extract of Trigonella foenum-graecum(AET) on the blood glucose in normal and alloxan(ALX)-diabetic mice.MethodsFasting blood glucose and glucose tolerance in normal and ALX-diabetic mice were measured respectively 7 days after AET had been given.ResultsAET had not significantly effected the fasting blood glucose of normal mice, but improved their glucose toleranc. Otherwise, AET reduced fasting blood glouse of diabetic mice induced by ALX significantly.ConclusionAET can be used on treatment of diabetes mellitus.

OBJECTIVETo explore the diagnostic value of the radiologic characteristics of osteoporotic Kummell's disease.

METHODSTotal 16 patients with pathologically confirmed osteoporotic Kummell's diseases were reviewed from May 2010 to May 2012, including 4 males and 12 females with the mean age of 73.4 years (ranged, 67 to 83 years old). Radiologic imagings of all patients, including X-ray, CT and MRI, were analyzed retrospectively.

RESULTSIntravertebral linear clefts could be seen on the AP and lateral X-ray films of vertebrae. Sagittal and axial CT scans demonstrated the vacuum cleft phenomenon with liquid and air was identified within the vertebral body. Sagittal MRI showed the callapsed vertebral segment and the area of fluid signal with clear and intact border within the vertebral body. The fluid signal was low on T1-weighted images and high on T2-weighted images and stir images, which was corresponding to an intravertebral vacuum cleft.

CONCLUSIONThe radiologic characteristics of Kurmmell's diseases can provide valuable evidences for the early diagnosis.

Aged ; Aged, 80 and over ; Female ; Humans ; Magnetic Resonance Imaging ; Male ; Osteonecrosis ; diagnosis ; diagnostic imaging ; pathology ; Retrospective Studies ; Spinal Fractures ; diagnosis ; diagnostic imaging ; pathology ; Tomography, X-Ray Computed

OBJECTIVE:To observe the changes of insoluble particles in traditional Chinese drugs injections(TCDI)mixed with infusion fluid and to study the way to solve METHODS:61 kinds of TCDI in therapeutic dosages were mixed in 0 9% sodium chloride solution,then the insoluble particles formed with diameters of 2 5?5 0?10 0 and 25 0?m were counted with Coulter counter,and determined with physical method and microscope The precise filter for liquid medicine were investigated in respect to its flow rate,quantity of flow,adsorbability and scavenging action RESULTS:(1)The number of insoluble particles in 26 kinds of TCDI exceeded the standard in ChP accounting for 42 6% of total samples observed (2)The insoluble particles included glass fragments,active carbon,rubber particles,soft flocks and residue of drugs (3)The flow rate and quantity of flow met the clinical requirement with a scavenging rate of 88 5%,and no adsorbability was found CONCLUSION:Precise infilter for liquid medicine can scavenge the particles in TCDI so as to ensure the safe use of drugs for patients