Insights into the catalytic and regulatory mechanisms of dihydroflavonol 4-reductase, a key enzyme of anthocyanin synthesis in Zanthoxylum bungeanum.

Accumulation of anthocyanins largely determines the fruit color, and dihydroflavonol 4-reductase (DFR) is a key enzyme involved in the formation of anthocyanins. However, the catalytic and regulatory mechanisms of DFR are unclear. In this study, the gene encoding DFR from Zanthoxylum bungeanum Maxim. was cloned and ZbDFR was analyzed in detail. The ZbDFR accepted dihydrokaempferol, dihydroquercetin and dihydromyricetin as substrates. Flavonols such as myricetin, quercetin and kaempferol significantly inhibited the activity of ZbDFR, while quercitrin and isoquercitrin slightly increased the activity. Quercetin was a competitive inhibitor at low concentrations, and it had a combined effect of competitive and noncompetitive inhibition at high concentrations, which was consistent with ZbDFR having two inhibitor binding sites. In addition, the content of different types of flavonoids in Z. bungeanum peel at green, semi-red and red stage was analyzed, and the in vivo results could be explained by the regulation of ZbDFR activity in vitro. Site-directed mutagenesis combined with enzyme activity experiments showed that Ser128, Tyr163, Phe164 and Lys167 are the key catalytic amino acid residues. The Ser128, Tyr163 and Lys167 were crucial for the hydrogen transfer reaction, and mutation of these amino acids resulted in the loss of all or most of the activity. Phe164 was found to be important for the regulation of ZbDFR by flavonols. Accordingly, ZbDFR is a node at which flavonoids regulate the synthesis of anthocyanins and proanthocyanins.

Multi-enzymatic recycling of ATP and NADPH for the synthesis of 5-aminolevulinic acid using a semipermeable reaction system.

5-Aminolevulinic acid (ALA) is an important cellular metabolic intermediate that has broad agricultural and medical applications. Previously, attempts have been made to synthesize ALA by multiple enzymes in cell free systems. Here we report the development of a semi-permeable system for ALA production using stable enzymes. Glucose, sodium polyphosphate, ATP, tRNA, glutamate and NADPH were used as substrates for ALA synthesis by a total of nine enzymes: adenylate kinase, polyphosphate kinase, glucose-6-phosphate dehydrogenase, phosphogluconolactonase, 6-phosphogluconate dehydrogenase, glutamyl-tRNA synthetase and glutamate-1-semialdehyde aminotransferase from E. coli, hexokinase from yeast, as well as glutamyl-tRNA reductase and its stimulator protein glutamyl-tRNA reductase binding protein (GBP) from Arabidopsis in a semi-permeable system. After reaction for 48 h, the glutamate conversion reached about 95%. This semi-permeable system facilitated the reuse of enzymes, and was helpful for the separation and purification of the product. The ALA production could be further improved by process optimization and enzyme engineering.Abbreviations: PPK: polyphosphate kinase; ADK: adenylate kinase; ALA: 5-Aminolevulinic acid; HK: hexokinase; ZWF: glucose-6-phosphatedehydrogenase; PGL: phosphogluconolactonase; GND: 6-phosphogluconate dehydrogenase; GTS: glutamyl-tRNA synthetase; GTR: glutamyl-tRNA reductase; GBP: GTR binding protein; GSAAT: glutamate-1-semialdehyde aminotransferase.

Comparison of Clarus video stylet,HPHJ-A video laryngoscope and Airtraq laryngoscope for guiding oral tracheal intubation in patients with snoring / 临床麻醉学杂志

Objective To compare the effectiveness and feasibility of oral tracheal intubation with Clarus video stylet,HPHJ-A video laryngoscope and Airtraq laryngoscope and in snoring pa-tients.Methods Ninety patients undergoing elective snoring surgery with general anesthesia,3 9 males and 51 females,aged 22-55 years,BMI 25-29 kg/m2,ASA physical status Ⅰ-Ⅲ,were ran-domly divided into three equal-sized groups (n=30):Clarus Video Stylet group (group C),HPHJ-A video laryngoscope group (group H)and Airtraq laryngoscope group (group A).The time of success-ful endotracheal intubation and the success rate of initial intubation of all groups were observed.The mean arterial pressure (MAP),heart rate(HR)and Rate-pressure product(RPP)were also monitored before induction of anesthesia(T0),before tracheal intubation(T1),at 0 min(T2),1 min (T3),and 3 min (T4)after intubation,as well as the throat injury and hemorrhage were noted. Results Patients in group C were successful intubated.Two patients in group H and one patient in group A with failed intubation were successfully intubated by using Clarus video stylet.The time re-quired for successful intubation in group C was longer than groups H and A (P<0.05).Compared with T0,the increasing of hemodynamic parameters (MAP,HR,RPP)after induction of all groups were significant(P<0.05).Compared with T1,the HR,MAP and RPP were increased significantly at T2in all groups (P<0.05).Compared with T1,the MAP and RPP were increased significantly at T3in all groups,and the HR were increased significantly at T3in groups H and A (P<0.05).There was no significant difference in the HR between T1and T3in group C.The increase in the HR and RPP at T2 was lower in group C than that in both the groups H and A (P <0.05).The changes of hemodynamic parameters were no significant among the three groups at other time.The differences of sore throat score and hemorrhage were no significant.Conclusion Compared with HPHJ-A video laryngoscope and Airtraq laryngoscope,Clarus video stylet for guiding oral tracheal intubation in snoring patients have less influence on hemodynamic parameters,and have no limited mouth opening. But Clarus Video Stylet spends longer intubating times,and has no obvious advantages on preventing throat injury.

Comparison of intubation of Shikani optical stylet and Clarus Video Stylet in patients with cervical spine immobilization / 实用医学杂志

Objective To compare the application of Shikani optical stylet(SOS)and Clarus Video Sty-let(Tracway)in patient with cervical spine immobilization in tracheal intubation. Method Sixty patients,ASAⅠ~Ⅱ,undergoing cervical internal fixation operation,were randomly divided into Shikani optical stylet group (Group S,n = 30)and Clarus Video Stylet group(Group T,n = 30). MAP,HR and RPP(The rate-pressure product)were recorded at the point before induction of anesthesia(T1),before intubation(T2),at the immediate time of intubation(T3)and 1 min(T4),3 mins(T5),and 5 mins after intubation(T6).The intubation time,one-time success rate of intubation,the number of intubation times and the incidence of sore throat and other complica-tions were observed. Results The one-time intubation time in group S was obviously shorter than that in group T (P<0.05).The incidence of mild sore throat and intubation throat injury rate were lower in group S than those in group T(P<0.05).Compared with those at T1,MAP and RPP decreased significantly at point of T2~T6in both of two groups(P<0.05).There was no significantly difference in MAP,HR and RPP at any points of time between the two groups. The one-time success rate of intubation,the number of intubation times and the incidence of air-way complications in two groups(P>0.05)were no significantly different. Conclusions Compared with Clarus Video Stylet(Tracway),Shikani optical stylet can shorten the intubation time in patients with cervical spine immo-bilization,but no difference was found in regard to the hemodynamic influence on intubation,success rate of intu-bation,the intubation times,the sore throat and the other related complications.

Effects of different depths of anesthesia on postoperativecognitive dysfunction:a meta-analysis / 实用医学杂志

Objective To investigate the effects of different depths of anesthesia on incidence of postopera-tive cognitive dysfunction (POCD). Methods We systematically retrievedPubmed,OVID,CNKI,CBM and Wanfang database and VIP database for randomized controlled trials(RCTs)from inceptionto December 312016, comparing different depths of anesthesia for their impacts on incidence of early POCD. After data extraction and quality evaluation,Revman 5.3 software was used for statistical data analysis. Results A total of 714 patients in 8 eligible RCTs were identified. Results of meta-analysis were as follows.(1)Incidence of POCD of depth anesthesia (NTS=E0-E1)was lower than general anesthesia(NTS=D0-D1)1 d after surgery(OR=0.21,95%CI 0.13~0.35,P < 0.00001).(2)Incidence of POCD of depth anesthesia(NTS = E1)was lower than general anesthesia (NTS=D0)7 d after surgery(OR=0.45,95%CI 0.23~0.91,P=0.03).(3)Incidence of POCD of NTS=E1 was lower than NTS=D07d after surgery(OR=0.42,95%CI 0.24~0.71,P=0.001). Conclusion Comparedwith general anesthesia,depth anesthesia is associated with a lower incidence of early POCD.

Effects of different depths of anesthesia on postoperativecognitive dysfunction:a meta-analysis / 实用医学杂志

Objective To investigate the effects of different depths of anesthesia on incidence of postopera-tive cognitive dysfunction (POCD). Methods We systematically retrievedPubmed,OVID,CNKI,CBM and Wanfang database and VIP database for randomized controlled trials(RCTs)from inceptionto December 312016, comparing different depths of anesthesia for their impacts on incidence of early POCD. After data extraction and quality evaluation,Revman 5.3 software was used for statistical data analysis. Results A total of 714 patients in 8 eligible RCTs were identified. Results of meta-analysis were as follows.(1)Incidence of POCD of depth anesthesia (NTS=E0-E1)was lower than general anesthesia(NTS=D0-D1)1 d after surgery(OR=0.21,95%CI 0.13~0.35,P < 0.00001).(2)Incidence of POCD of depth anesthesia(NTS = E1)was lower than general anesthesia (NTS=D0)7 d after surgery(OR=0.45,95%CI 0.23~0.91,P=0.03).(3)Incidence of POCD of NTS=E1 was lower than NTS=D07d after surgery(OR=0.42,95%CI 0.24~0.71,P=0.001). Conclusion Comparedwith general anesthesia,depth anesthesia is associated with a lower incidence of early POCD.

Effect of dexmedetomidine on gastric mucosal injury induced by intestinal ischemia-reperfusion in rats / 中华麻醉学杂志

Objective To evaluate the effect of dexmedetomidine on gastric mucosal injury induced by intestinal ischemia-reperfusion (I/R) in rats.Methods Thirty-six healthy male Sprague-Dawley rats,aged 4-5 months,weighing 200-250 g,were randomly divided into 4 groups (n =9 each) using a random number table:sham operation group (group S),intestinal I/R group (group I/R),low-dose dexmedetomidine group (group LD),and high-dose dexmedetomidine group (group HD).Intestinal I/R was produced by occlusion of the superior mesenteric artery for 1 h followed by 2 h reperfusion in anesthetized rats.Dexmedetomidine 2.5 and 5.0 μg · kg-1 · h-1 were infused via the caudal vein for 1 h starting from 1 h before ischemia in LD and HD groups,respectively.The rats were sacrificed at 2 h of reperfusion,and the gastric mucosa was obtained for examination of the pathological changes (with light microscope) and for determination of the expression of serine/threonine kinase (Akt),phosphorylated Akt (p-Akt),activated caspase-3 and caspase-3 (by Western blot).The ratio of p-Akt to Akt (p-Akt/Akt)was calculated to reflect the phosphorylation of Akt.The ratio of activated caspase-3 to easpase-3 (activated caspase-3/caspase-3) was calculated to reflect the activation of caspase-3.Results Compared with group S,the expression of Akt was significantly up-regulated,the expression of p-Akt was significantly downregulated,the phosphorylation of Akt was significantly decreased,and the activation of caspase-3 was significantly increased in group I/R,and the expression of p-Akt was significantly up-regulated,and the phosphorylation of Akt and activation of caspase-3 were significantly increased in LD and HD groups (P＜0.05).Compared with group I/R,the expression of Akt was significantly down-regulated,the expression of p-Akt was significantly up-regulated,the phosphorylation of Akt was significantly increased,and the activation of caspase-3 was significantly decreased in LD and HD groups (P＜0.05).The degree of gastric mucosal injury was significantly lower in LD and HD groups than in group I/R,and there was no significant difference in the degree of gastric mucosal injury between group LD and group HD.Conclusion Dexmedetomidine can attenuate gastric mucosal injury induced by intestinal I/R,and the mechanism may be related to activation of PI3K/Akt signaling pathways and inhibition of cell apoptosis in rats.