Kinetic model analysis is a useful tool for understanding the regulation and control of cellular metabolism and thus offering a guideline for rational design of high efficiency cell factory. Based on previously published models and experimental measurement of enzyme kinetics data, we developed a kinetic model for the threonine biosynthesis pathway in Escherichia coli. This model integrates the central pathways that produce precursors, ATP and reducing power with the threonine biosynthesis pathway from aspartate. In contrast to the previous models, we considered the energy and reducing power balance rather than artificially set their concentrations. Metabolic control analysis of the model showed that enzymes PTS, G6PDH, HDH etc. have great flux control coefficients on the threonine biosynthesis flux. This indicates higher threonine synthesis flux could be achieved by overexpressing these enzymes.
Escherichia coli ; metabolism ; Industrial Microbiology ; Kinetics ; Metabolic Networks and Pathways ; Models, Biological ; Threonine ; biosynthesis

Objective:To study the inhibitory effect of salidroside on the proliferation of fibroblast-like synoviocytes with rheumatoid arthritis in human(HFLS-RA) induced by tomor necrossi factor-α(TNF-α),and to clarify the molecular mechanism of its control effect on rheumatoid arthritis(RA).Methods:The HFLS-RA were cultured in vitro,then treated with TNF-α and different concentrations of salidroside.The cells were divided into normal control group(0 μg·L-1TNF-α),model control group(10.0 μg·L-1TNF-α)and 12.5,25.0,50.0,and 100.0 μmol·L-1 salidroside groups(10.0 μg·L-1TNF-α+salidroside).The proliferation activity was detected by MTT mehthod;the expression levels of β-catenin,matrix metalloproteinase-7(MMP-7),and Cyclin-D1 in supernatant of the cells were detected by ELISA method;the expression level of β-catenin protein in cells was detected by Western blotting method.Results:Compared with normal control group,the proliferation activity of the HFLS-RA in model control group was significantly increased (P<0.05);compared with model control group,the proliferation activities of the HFLS-RA in 12.5 and 25.0 μmol·L-1 salidroside groups were decreased but there were no significant differences(P>0.05),and the proliferation activities of the HFLS-RA in 50.0 and 100.0 μmol·L-1 salidroside groups were significantly decreased (P<0.05).Compared with normal control group,the expression levels of β-catenin,MMP-7,and Cyclin-D1 in the supernatant of the cells in model control group were increased(P<0.05).Compared with model control group,the expression levels of β-catenin,MMP-7,and Cyclin-D1 in the supernatant of the cells in 12.5 and 25.0 μmol·L-1 salidroside groups were decreased,but there were no significant differences(P>0.05);the expression levels of β-catenin,MMP-7,and Cyclin-D1 in the supernatant of the cells in 50.0 and 100.0 μmol·L-1 salidroside groups were decreased(P<0.05).The Western blotting results showed that the expression level of β-catenin protein in the cell in model control group was higher than that in normal control group(P<0.01);the expression levels of β-catenin protein in the cells in 12.5 and 25.0 μmol·L-1 salidroside groups were lower than that in model control group,but there were no significant differences(P>0.05);the expression levels of β-catenin protein in the cells in 50.0 and 100.0 μmol·L-1 salidroside groups were lower than that in model control group(P<0.05).Conclusion:Salidroside could inhibit the proliferation of HFLS-RA,and its control effect might be related to the regulation of Wnt/β-catenin single pathway.

Objective To establish intellectual critique of nursing thesis course in postgraduate nurse,and to evaluate teaching effectiveness and the reasonableness of the course.Methods A total of 44 postgraduate nurses were involved in the study,which were carried out through action research of planning,acting and observing,and reflecting.After the course,researchers evaluated the reasonableness of the course and teaching effectiveness through the self-control.Results There was significant difference in the scientific research ability score of research object before and after the course:(60.09±16.26) scores vs.(71.27±13.22) scores,t=-3.264,P＜0.01.But there was no significant difference in the critical thinking ability scores of research object before and after the course:(295.55±29.71) scores vs.(292.71±26.67) scores,t=0.478,P＞0.05.Conclusions The course of intellectual critique of nursing thesis can improve the research ability of postgraduate nurse,but the role of the critical thinking ability remains to be discussed.And the course needs to be further improved.

Objective To observe the changes of the cells of human hepatocellular carcinoma (HepG2)using RNA for silencing the expression of COMMD7 gene,and investigate related mechanism of COMMD7 gene promoting HepG2 proliferation.Methods COMMD7 gene shRNA was designed and constructed into COMMD7-shRNA plasmid.HepG2 cells were divided into the HepG2 group,control-shRNA group (empty vectors were infected) and COMMD7-shRNA group (positive vectors were infected).Cells shapes were observed by fluorescence microscope after infecting.The expression of COMMD7 and expression and phosphosylation of extracellular regulated protein kinase1/2 (ERK1/2) and MEK1/2 protein were measured by Western blot.The cell vitality was measured by cholecystokinin octapeptide (CCK-8),and the apoptosis of cell was detected by flow cytometry.The measurement data with normal distribution were presented as (x) ± s.The comparisons among groups were evaluated with the one-way ANOVA,and pairwise comparison was analyzed by the LSD-t test.Results The cells were oval or spindle shapes and displayed green fluorescent after infected successfully.The results of Western blot showed that the relative quantitative expression of COMMD7 protein in the HepG2 group,control-shRNA group and COMMD7-shRNA group were 0.90 ±0.18,1.03 ±0.05 and 0.23 ±0.03,respectively,with a significant difference among the 3 groups (F =152.08,P ＜ 0.05),and the expression of COMMD7 protein in the COMMD7-shRNA group was significantly lower than those in the other 2 groups (t =20.74,21.16,P ＜ 0.05).The results of CCK-8 showed that the scores of the HepG2 vitality in the HepG2 group,control-shRNA group and COMMD7-shRNA group were 1.193 ±0.024,1.225 ±0.034 and 1.147 ±0.021,respectively,with a significant difference among the 3 groups (F =6.90,P ＜ 0.05),and the HepG2 vitality in the COMMD7-shRNA group was significantly lower than those in the other 2 groups (t =3.53,3.69,P ＜ 0.05).The results of flow cytometry showed that the apoptosis rate of HepG2 in the HepG2 group,control-shRNA group and COMMD7-shRNA group were 6.1％ ± 0.3％,7.8％ ± 0.5％ and 20.9％ ± 1.4％,showing a significant difference among the 3 groups (F =270.80,P ＜0.05),and the apoptosis rate of HepG2 in the COMMD7-shRNA group was significant higher than those in the other 2 groups (t =21.77,19.36,P ＜0.05).The results of Western blot showed that the relative quantitative expression of phosphorylation (p)-ERK1/2 and p-MEK1/2 in the HepG2 group,control-shRNA group and COMMD7-shRNA group were 0.932 ±0.046,0.945 ±0.017,0.553 ±0.052 and 0.452 ±0.031,0.468±0.027,0.263 ± 0.022,respectively,showing significant differences among the 3 groups (F =93.61,49.16,P ＜ 0.05),and the relative quantitative expression of p-ERK1/2 and p-MEK1/2 in the COMMD7-shRNA group were significantly lower than those in the other 2 groups (t =11.94,12.17,9.33,8.65,P ＜ 0.05).Conclusions COMMD7 gene can promote HepG2 proliferation via activating ERK/mitogen-activated protein kinase (MAPK) signaling pathway,and its mechanism may be promoting the phosphorylation of expression of ERK1/2 and MEK1/2.

A microfluidic chip with micropillar arrays for three-dimensional (3D) cell culture was designed and validated.The chip consisted of a polydimethylsiloxane (PDMS) channel plate and a glass cover plate.One cell culture chamber composed of two rows of micropillar arrays and two lateral channels for transporting the culture medium were integrated on the PDMS channel plate.The spacing between micropillars directly affects the chip performance, which is critical for the design of the chip.In this work, the spacing between micropillars was optimized by numerical simulation and experimental validation.With the optimized microfluidic chip, the mixture of cells and extracellular matrix mimics could be steadily injected into the cell culture chamber, the nutrients in the culture medium from the lateral channels could quickly diffuse into the chamber, and the cell metabolites could also timely diffuse out of the chamber.To test the stability of the microenvironment in the microfluidic chip, neural stem cells were three-dimensionally cultured.

Studies showed that low fermentable oligosaccharides, disaccharides, monosaccharides and polyols (FODMAPs) diet can improve the symptoms of gastrointestinal dysfunction of inflammatory bowel disease (IBD) patients, the mechanism may be related to decrease of secretion of intestinal liquid and production of gas.Specific carbohydrate diet and paleolithic diet may be suitable for IBD patients.This article reviewed the advances in study on effects of different diets on gastrointestinal dysfunction in patients with IBD.

OBJECTIVE To evaluate the protective effect of Agaricus bisporus intracellular polysaccharides(IPS) and exopolysaccharides (EPS) on immunological liver injury induced by concanavalin A (Con A). METHODS Mice were pretreated with IPS and EPS (100, 200 and 400 mg kg- 1, ig) daily for 12 d. Immunological liver injury was induced by Con A 25 mg·kg-1 byinjection via the tail vein of mice.Eight hours after injection of Con A, the indexes of the liver, spleen and thymus, serum level of glutamicpyruvic transaminase (GPT), glutamic-oxalacetic transaminase (GOT), tumor necrosis factor-α (TNF-α) and interferon- γ (IFN- γ), splenic lymphocyte percentages of CD4 + and CD8 + , and liver homogenate content of superoxide dismutase (SOD) and malondialdehyde (MDA) were measured. Liver pathological changes were observed by HE staining. RESULTS Compared with normal group, the autoimmune liver injury in mice induced by Con A resulted in an increase in the liver index (P<0.01) , spleen index (P<0.01), the activity of GPT (P<0.01) and GOT (P<0.01), the content of TNF- α (P<0.01) and IFN- γ (P< 0.01), and the level of MDA (P<0.01), but a decrease in the thymus index (P<0.01), the percentage of CD4+ (P<0.01) , the ratio of CD4+/CD8+ (P<0.01), and SOD activity (P<0.01). Compared with model group, treatment with IPS (200 and 400 mg·kg-1) and EPS (200 and 400 mg·kg-1 ) respectively resulted in an increase in the thymus index (P<0.01) but in a decrease in the liver index and spleen index (P<0.01). Similarly, the activity of GOT and GPT was decreased obviously (P<0.01), and the content of TNF-α and IFN-γ in IPS and EPS 200 and 400 mg·kg-1 groups was decreased. Compared with model group, the activity of SOD in IPS and EPS (200 and 400 mg·kg- 1) group was increased (P<0.01) while MDA was decreased (P<0.01). Moreover, the percentage of CD4 + Iymphocytes decreased (P<0.01), whereas no significant difference was found in the ratio of CD4 +/CD8 + .Pathological changes of the liver were observed under a microscope. Pretreatment with IPS and EPS could effectively reduce the liver injury induced by Con A. CONCLUSION IPS and EPS have certain protective effect on immunological liver injury, which may be related to their ability to clean up free radicals, control lipid peroxidation and regulate the balance of the immune system.

Objective To observe changes of the pancreas after limb ischemia reperfusion, and the protective effect of taurine (Tau) postconditioning. Methods Twenty-four male SD rats were randomly divided into control group (Control), limb ischemia reperfusion (LIR) group, and taurine postconditioning (Tau) group (n=8 for each group). Plasma contents of xanthine oxidase (XOD), malondialdehyde (MDA), reactive oxygen species (ROS), superoxide dismutase (SOD), pancreatic amylase (AMS), pancreatic lipase (LPS) and trypsin were detected in three groups. Morphological changes of the pancreas were observed using optical microscopy. The expressions of anti-C/enhancer-binding protein homologous protein (CHOP) in pancreas were analyzed by immunohistochemistry assay. TUNEL staining was performed to estimate the apoptosis of pancre-atic cells. Results Compared with the control group and Tau group, plasma contents of MDA, XOD, ROS, AMS, LPS, and trypsin were significantly increased in LIR group, but the level of SOD was significantly lower in LIR group ( P<0.05). HE staining showed that acinar structure was disrupted , pancreatic lobule gap widened, stromal vascular dilatation and conges-tion were observed in LIR group. The perivascular infiltration of inflammatory cells appeared, islet cells were damaged with irregular islet. The immunohistochemical analysis showed the increased expression of CHOP in pancreas in LIR group than that of Tau group. And the pancreatic apoptosis was enhanced in LIR group detected by TUNEL staining. Conclusion Tau-rine postconditioning can ameliorate pancreatic injury following limb ischemia reperfusion, which may be related to the inhi-bition of oxidative stress, down-regulation of CHOP expression, thereby reducing endoplasmic reticulum stress-induced apoptosis.

ObjectiveTo analyze the literature characteristics on essential hypertension treated by external therapy of traditional Chinese medicine, summarize the current research situation and trend in the field, and to provide a reference for relative researches.MethodsThe papers relevant to treatingessential hypertension with external therapy of TCM included inSinoMedwere statistically analyzed from the aspects of publishing year, journals distribution,author’s unit and districtdistribution,research funds and literature content with bibliometrics method.ResultsThe total number of the literature for analyzing was 226, the number of papers increased gradually.Authors of the papers were mainly from TCM universities or colleges.Papersdistributedmainly in the more economically developed regions,which issued the largest amountinGuangdongprovince.Papers supported by research funds accounted for 16.81% in all the literature. Most literaturewasclinical research and the most commonly used for external therapywasacupuncture.ConclusionThe research and clinical work of essential hypertension treatment with external therapy of traditional Chinese medicinewerepaid more close attention in recent years, but therewerestill some problems that need to be solvedto form a viable, effective treatment system.

Objective To study the effects of different types of exercise training on learning and memory, as well as on the expression of synaptophysin (SYP) and on postsynaptic density protein 95 (PSD-95) in rats in which a model of vascular dementia had been created.Methods Forty male Wistar rats were divided randomly into a voluntary exercise group (V-EX) , a forced exercise group (F-EX) , an involuntary exercise group (I-EX) , a vascular dementia group (VD) and a sham-operation group (Sham) , with 8 rats in each group.Two-vessel occlusion (2-VO) of the arteria carotis communis was used to create a model of vascular dementia in all of the rats except those in the sham-operation group.Beginning one week after the surgery, the V-Ex rats were free to run in a running wheel.The F-EX rats were forced to run 270 m a day in an electric wheel.The I-EX rats were stimulated to imitate the gait pattern of their forelimbs running at 9 m/min three times a day for l0 minutes each time.No special training was given to the rats in the other 2 groups.Three weeks after the surgery, their learning and memory were tested using a novel object recognition test.Immediately after the test, their prefrontal cortex was sampled and the expression of SYP and PSD-95 was detected using western blotting.Results The average novel object recognition indices of the rats in the V-EX, F-EX and I-EX groups were all significantly higher than that of the VD group.Average PSD-95 expression was also significandy higher than in the VD group.Conclusion Exercise, whether voluntary, forced or induced by functional electrical stimulation can improve learning and memory in vascular dementia, at least in rats.The mechanism is possibly that the training can increase the expression of PSD-95 in the prefrontal cortex, though not SYP.