The zinc chloride and cystine resistant strain of S.cerevisiae YZM-14(ZnCl2r,Cysr) was screened with the mutant processing of the protoplast of S.cerevisiae by combinative mutagens of ultraviolet and nitrite.The glutathione(GSH) production(84.72 mg/L),dry cell weight(7.63 g/L) and the intracellular GSH content(11.10 mg/g) of YZM-14 were 2.79,1.63 and 1.71 times compared with that of the initial strain.The biosynthetic process of GSH was divided into three phases according to the time course of the specific cell growth rate and GSH yielding coefficient.In the second phase,the metabolic flux of the pentose phosphate pathway and the GSH precursors biosynthetic pathway of the mutant strain increased by 8.1 mmol/(g?h),compared with that of the initial strain.Furthermore,the metabolic flux of the organic acids secretion of the mutant strain decreased.Through these mechanisms,the utilization efficiency of the carbon sources was enhanced and high production of GSH was obtained.

OBJECTIVETo assess the prevalence rate of Helicobacter pylori (H. pylori) in blood serum, its affecting factors and isoforms (CagA,VacA )infection in the elderly people in Beijing.

METHODS2006 residents were investigated through household questionnaire in different areas of Beijing (urban, suburban and mountainous district), who were older than 60 years old. Serum H. pylori CagA, VacA and Ureas antibody was detected by immunoblotting.

RESULTSThe total H. pylori infection rate was 83.4% and the infectious rate of I form pathogenic H. pylori was 56.0%. The incidence rate in urban or suburban district was higher than that of in mountainous district (P < 0.001). I form H. pylori infection rate in people with heavy labor or young elderly were higher than that of intelegent work or older elderly (P < 0.05 ). I form H. pylori infection rate in people of low diet was higher than that of high protein diet (P < 0.001).

CONCLUSIONThe rate of H. pylori infection in blood serum was high among the elderly people in Beijing with most of it belonged to type I . However, significant differences were noticed on the distribution of isoforms in different age groups, areas, professions and diet habit.

Age Distribution ; Aged ; China ; epidemiology ; Diet ; Female ; Helicobacter Infections ; blood ; epidemiology ; Helicobacter pylori ; classification ; physiology ; Humans ; Male ; Occupations ; Seroepidemiologic Studies

Adolescent ; Adult ; Athletes ; Boxing ; Electroencephalography ; Female ; Humans ; Physical Education and Training ; methods ; Young Adult

The high-copy-number plasmid pcDNA3.1+ is unstable within S almonella typhimurium. A novel plasmid pmcDNA3.1+ was constructed by removin g the promoter sequence of ampicillin resistance gene (bla gene) in plasmid pcDNA3.1+. In contrast to pcDNA3.1+, pmcDNA3.1+ was stable within Salmonel la typhimurium SL7207 in LB medium with or without ampicillin. Further experi ments showed the ?-lactamase activity of Salmonella typhimurium SL7207(pmc DNA3.1+) was apparently lowered than that of Salmonella typhimurium SL7207( pcDNA3.1+) and the high ampicillin concentration was maintained longer in LB me dium culturing Salmonella typhimurium SL7207(pmcDNA3.1+). When mice were a dministered with Salmonella typhimurium SL7207(pmcDNA3.1+) intraperitoneall y, more than 95% of Salmonella cells separated from the spleen still harbore d the plasmid pmcDNA3.1+ 7 days later; but 99% of Salmonella cells lost the plasmid pcDNA3.1+ at day 3 in mice innoculated with Salmonella typhimurium SL7207(pcDNA3.1+). By lowering the expression of bla gene, the rapid deco mposition of ampicillin in LB medium was avoided and the metabolic pressure was relieved for the host cells. This method offers a solution for the problem of t he instability of high-copy-number plasmid within Salmonella typhimurium.

Objective To investigate the effects of rigid gas permeable contact lens (RGPCL) wearing 3 years on ocular surface in keratoconus.Design Retrospective case series.Participants 73 patients with keratoconus.Methods From July 2001 to July 2004,73 patients (142 eyes) wearing RGPCL for more than 3 years were collected in Peking University Optometry & Ophthalmology Center.Be- fore and at 1 year,2 years and 3 years of RGPCL wearing,density and morphologic changes of corneal endothelium were examined with non-contact specular microscope.Eye axial length,central and peripheral thickness of cornea were measured with A-scan pachymeter. All eyes were examined with slit-lamp microscope periodically.Main Outcome Measures Corneal endothelial density and morphology, corneal thickness,eye axial length,ocular surface changes.Results Before and at 1 year,2 years and 3 years of RGPCL wearing,aver- age corneal endothelial density was 2901.92?445.20,2862.78?497.13,2854.71?526.80,3015.61?421.22 (cells/mm~2)without significant difference statistically (F=1.571,P=0.20).Morphologic changes were not significant during 3 years.Eye axial length was 25.15?1.50, 24.93?1.36,24.78?1.25,25.39?1.31 (mm) without significant difference statistically (F=2.218,P=0.10).Corneal central thickness was 489.09?59.64,484.02?60.80,496.61?59.74,487.44?54.25(?m)without significant difference statistically (F=0.991,P=0.40).Peripheral thickness changes were also not significant statistically during 3 years.69 eyes with mild conjunctival congestion,12 eyes with corneal fluorescence stain and 6 eyes with corneal epithelial rough were found with slit-lamp microscope during follow-up.Conclusions For the patients with keratoconus,long term of RGPCL wear will not lead to significant ocular changes or severe ocular complications.

Background The surgery for femtosecond laser created laser in situ keratomileusis(LASIK)flaps has made great progression recent year,but the postoperative corneal wound healing and regeneration of nerve fibers after surgery are closely concerned.Objective This study was to compare and analyze the clinical outcomes between FEMTO LDV femtosecond laser flap and mechanical microkeratome Hansatome flap in LASIK.Methods A prospective case-controlled study was designed.The serial 38 myopic eyes of 38 patients were included from March through July,2010 in Henan Armed Police Force General Hospital.The patients were randomized into FEMTO LDV femtosecond laser assisted flap group(20 patients/20 eyes)and mechanical microkeratome Hansatome assisted flap group(18 patients/18 eyes)with the matched age,gender and refraction of spherical equivalent.HRT Ⅲ examinations were performed before surgery,1 week,1 month and 3 months after surgery to compare the morphological changes atthe center and margin of the flaps,and evaluate the similarities and differences of cellular morphology after surgery between the two approaches.Written informed consent was obtained from each patient prior to this medical trial.Results The best corrected visual acuity was ≥ 1.0 and the refract diopter was similar in both groups(+0.21 D±0.48 D and-0.04 D±0.54 D)1 month after LASIK.The corneal thickness was insignificant increased in the first week after LASIK,and the density of shallow stromal cells was decreased in 1 week,1 month and 3 months compared with pre-operation in the femtosecond laser assisted flap group(t =-27.99,-25.49,-28.87,P ＜ 0.01).In the Hansatome assisted flap group,significantly thickened corneal epithelium was seen in the first week after LASIK compared before LASIK(56.73 μm±2.47 μm versus 51.16 μm±1.11 μm)(t=9.29,P＜0.05),and the density of shallow stromal cells was decreased in 1 week,1 month and 3 months compared with pre-operation in the Hansatome assisted flap group(t =-17.57,-14.13,-19.63,P =0.00).The density of high reflective interface particles in cornea was lower in 1 week,1 month and 3 months after LASIK in the femtosecond laser assisted flap group than that in the Hansatome assisted flap group,showing significant differences between them(t =-13.505,-11.900,-14.084,P＜0.01).The active stromal cells were seen beneath the interface in both groups in the first week and gradually decreased after that time.Intact corneal nerve fibers were found in the femtosecond laser assisted flap group,but those in the Hansatome assisted flap group were shorter and smaller 3 months after LASIK.At 3 months after surgery,the flap margin showed stromal higher reflection and irregular secondary fibrosis in the femtosecond laser assisted flap group,and in contrast,the flap margin had the appearance of a unclearly identified fibrotic scar in the Hansatome assisted flap group.Conclusions Compared with the LASIK and Hansatome assisted flap,the LASIK with FEMTO LDV flap shows earlier nerve fiber regeneration and greater fibrotic scarring,which imply a good wound healing process in the LASIK with FEMTO LDV flap.

Objective:To compare the biomechanical effects between oblique Ban-pulling manipulation and lumbar erection-rotation manipulation in sitting position in treating lumbar intervertebral disc herniation (LIDH). Methods:A three-dimensional finite element model of L3-S1 was developed to carry out a comparative study between oblique Ban-pulling manipulation and lumbar erection and rotation manipulation in sitting position. The disc protrusion was assumed to be on the rear left of L4 disc, and the manipulations were performed on the right side. The loading process was simulated by two steps. In the first step, only the compression loading was imposed, and in the second step, both the compression loading and axial rotation moment were imposed. The displacement and stress distribution in L4 disc were investigated. Results:The values of stress and displacement in the second step were lower than those in the first step in each manipulation. The stress and displacement differences between the two steps were respectively 1.79 times and 3.03 times larger in oblique Ban-pulling manipulation than those in lumbar erection-rotation manipulation in sitting position. Conclusion: Oblique Ban-pulling manipulation may result in a better biomechanical effect than lumbar erection-rotation manipulation in sitting position for LIDH.

Interleukin-1 receptor antagonist (IL-1ra), a member of IL-1 family, is a naturally occurring IL-1 inhibitor as "receptor antagonist", which blocks biological responses mediated by IL-1. Recombinant human IL-1ra (rhIL-1ra, Kineret) was introduced in clinical trials involving patients with RA. Between 2001 to approximately 2002, rhIL-1 ra was approved by the US Food and Drug Administration and the European Agency for the Evaluation of Medicine Procedure. Unfortunately, 10,000 to 100,000-fold excess amounts of IL-1ra are needed to relieve disease because minimal IL-1 can induce complete biological responses, and the dosage of 100 to approximately 150mg/day in a RA patient is so big that it greatly influence patients' physical, psychological and economical situation. In this study, IL-1ra mutants were established by site-specific mutagenesis to improve its stability. The sites of mutagenesis included R6 K7-AA,R93 K94-AA and K97 R98-AA. IL-1ra and its mutants were expressed in E. coli BL21 (DE3) using pTIG-Trx expressing system with the induction of IPTG. The recombinant proteins were purified by Ni2+ chelate chromatography and Sephadex G75 gel filtration chromatography. The activity of mutants is as high as IL-1ra. We characterized the pharmacokinetic profile of IL-1ra and its mutants. The third mutant's half life is 2.26 times than wt IL-1ra. The study has provided some approaches and experience for further research to improve the metabolism stability of IL-1ra.
Animals ; Escherichia coli ; genetics ; metabolism ; Female ; Humans ; Interleukin 1 Receptor Antagonist Protein ; biosynthesis ; genetics ; pharmacokinetics ; Mutagenesis, Site-Directed ; methods ; Mutant Proteins ; biosynthesis ; pharmacokinetics ; Recombinant Proteins ; biosynthesis ; genetics ; pharmacokinetics

Objective To develop a clinically useful assay for detecting the mutations of HBV pre-C/BCP based on the pyrosequencing and accuracy, reproducibility and reliability of this assay was evaluated. Methods The pyrosequencing primers for HBV pre-C/BCP mutation were designed through the cluster analysis among one hundred HBV gene sequences. After the amplification of the fragment of pre-C/BCP with the template of pre-C/BCP mutation plasmids, the pyrosequencing method for pre-C/BCP detection was initially set up with this standard sample. The accuracy, reliability and reproducibility of the pre-C/BCP pyrosequencing were confirmed through the pre-C/BCP plasmids as a standard sample when compared with Sanger/genechip sequencing method pre-C/BCP pyrosequencing assay was applied for detecting pre-C/BCP mutation types of 60 chnical serum samples in HBV patients. Results The pre-C/BCP mutation detection assay based on pyrosequencing has been established in our study. The coincidence rate between pyrosequencing and Sanger squencing was 100%. The coincidence rate between the result of pyrosequencing and of genechip method was 91.7%. The reproducibility of this assay was 97. 8%. It indicates the pre-C/BCP pyrosequencing is a high-accurate method with, good-reproducibility and high-reliability. And multi-site detection can be achieved by pyrosequencing one time. A rare mutation T1758C was also detected. Conclusion Pyrosequencing for pre-C/BCP mutations assay is high-throughout method for simultaneous detection of multi-site mutation.

Objective To investigate the clinical significance of external obturation with cotton ball in treatment of digestive tract lip shape fistula. Methods A prospevtive randomixed controllde study was conducted in 30 patients with digestive tract lip shape fistula admitted from January 2001 to November 2007,which were divided into experimental group ( 15 patienets) and control griup ( 15 patjients). The control group received traditional fistula mouth continuous drainage,with fastomg or part enteral nutrition and parenteral nutrition. The experimernal group received external obturatiion with cotton ball,with controling spills of oineteatinal fluid and resuming normal eating and activetes, The patients general information, fistuala mouth sixe, volume, weight, albumin, determinstic operation time, postoperative complications, hospital, signficantly reduced fistula, flow, postoperative compared between the two groups. Results Compared with the control group , significantly redced fistula, flow, postoperative complivations and cost, increased wdighe and albumin,as well as shortened operation time and hospital stay were found oin the experimetal grou. Conclusin External obturation with cotton ball is safe effective methde in treatment of digestive tract lip shape fistula, obviusly shortening the course of disease, redcing patients'cost and pain, and rapidly improving patients nutritional status.