Objective To express the sucrose isomerase(SI) fused with the tetrameric coiled-coil domain of the cell surface protein tetrabrachion(TdoT),and study the enzymatic properties of the recombinant enzymes.Methods The gene of SI fused with TdoT at the N/C terminus was cloned into the expression vectors respectively to construct the recombinant expression vectors pET-24a-TdoT-SI and pET-24b-SI-TdoT,which were transformed into E.coli BL21(DE3) and induced to express recombinant enzymes.The enzymatic properties and product specificity of the purified recombinant enzymes were studied.Results TdoT-SI and SI-TdoT were expressed as inclusion bodies with catalytic activity,while SI inclusion bodies without TdoT showed no catalytic activity.The results of enzymatic property analysis showed that the optimum reaction temperature for TdoT-SI and SI-TdoT active inclusion bodies was 40 ℃,and the optimum reaction pH was 5.5 and 5.0,respectively.The K_m of TdoT-SI active inclusion bodies was(103.9±9.5) mmol/L and the k_(cat)/K_m was(0.06±0.002) L/(mmol·s),while the K_m of SI-TdoT active inclusion bodies was(54.4±6.6) mmol/L and the k_(cat)/K_m was(0.03±0.002) L/(mmol·s).The results of product specificity analysis exhibited that the proportion of isomaltulose in the product did not change significantly,while the proportion of trehalose decreased,and the proportion of monosaccharides increased with increasing reaction temperature.Conclusion The active inclusion bodies of SI fused with coiled-coil domain were successfully prepared by fusion expression technology.As a novel self-immobilized enzyme,it has the advantage of simultaneous expression and immobilization,which provides a new strategy for large-scale preparation and efficient utilization of recombinant SI.

Objective To explore the influence of integrated Chinese and western medicine therapy on anxiety,sugar metabolism and subjective well-being for type 2 diabetes Patients associated with anxiety.Methods Use glycated hemoglobin (HbAlc),trait anxiety inventory(TAI),and Memorial University of Newfoundland Scale of Happiness (MUNSH) to evaluate the condition of 60 patients with type 2 diabetes associated with anxiety.The patients participated in the study were randomly divided into Western medicine therapy group(control group)and integrated traditional Chinese and Western medicine therapy group (experimental group),with 30 cases in each group.The control group was treated with paroxetine while the experimental group was treated with paroxetine plus Jiawei-Xiaoyaowan.6 weeks after treatment,we tested the HbAlc level.TAI score and MUNSH score of those patients.Results After 6 weeks,the TAI score of patients in both experimental group and control group have obviously decreased (before treatment:54.07±6.41 and 56.33 ±6.01,respectively; after treatment:38.89±6.70 and 45.29±6.55,respectively) and the difference was statistically significant (P＜0.05).And the TAI score of the two groups after treatment was significantly different (P＜0.05).For both experimental and control groups,the HbA1e level were not obviously changed before and after the study with P＞0.05 (before treatment:8.14±2.69 and 8.07±2.11,respectively; after treatment:8.21±2.07 and 7.92± 1.90,respectively).Comparison between groups also showed no significant difference.MUNSH score have obviously increased in experimental group (before treatment:40.49±5.22; after treatment:44.53 ± 6.28,P＜0.05).The result of comparison between the two groups after treatment shows that MUNSH score of patients in the experimental group were obviously improved compared with patients in the control group (57.56 ±6.39),and the difference was statistically significant (P＜0.05).Conclusion The integrated traditional Chinese and western medicine therapy can obviously release anxiety and enhance subjective well-being in patients with cerebral infarction associated with anxiety.

Objective To study the blood supply of the flap accompanying vessels of the cutaneous nerves in the lower leg, and to design the reversed flap for clinical reference. Methods Anatomic observation was performed on 30 adults’ lower extremity specimens perfused via pressure with red latex through femoral arteries. Results Superficial peroneal nerves, sural nerves and saphenous nerves were all nourished by their accompanying arteries which, anastomosed with the cutaneous perforating branches of other arteries, also nourish the corresponding skin areas. Conclusion The blood supply of the reversed island flap accompanying vessels of the cutaneous nerves in the lower leg is reliable and so it is possible to design long flaps along the cutaneous nervous axis.

Objective To develop miniature pig model of coronary microembolization (CME) by easy and cost-effi-cient technique. Methods A total of 11 miniature pigs were divided into control group (n=5) and CME group (n=6). Femo-ral artery was punctured using 21 gauge needle that is normally used for transradial procedures. Microspheres were injected into the left anterior descending artery of the CME group by 5 F coronary radiography catheter and 1.8 F coronary micro-guide catheter. Serum concentrations of brain natriuretic peptide (BNP) and cardiac troponin I (cTnI) were evaluated just be-fore CME and 6 hours after CME. Apical myocardial pathological lesions were evaluated by optical microscope 6 hours after CME. Results All miniature pigs in control group survived, but one died in the CME group. 5 F coronary radiography cathe-ter and 1.8 F coronary micro-guide catheter reached designated location successfully. Before CME, serum BNP (ng/L:143.00 ± 13.51 vs 134.00 ± 15.57) and cTnI (μg/L:0.39 ± 0.09 vs 0.38 ± 0.10) showed no significant differences between these two groups (t values are 0.976 and 0.294 respectively,both P>0.05). By contrast, serum BNP (561.00 ± 80.65) and cTnI (2.75±0.58) were much higher in CME group than those (BNP 139.00±13.87;cTnI 0.54±0.14 ) in control group after CME (t values are 11.530 and 8.337 respectively,both P＜0.001). In CME group, microspheres, micro-infarction and inflammatory cell infiltration were seen under an optical microscope which are absent in control group. Conclusion Using new surgical consumables can successfully develop miniature pig model with CME. And the technique is simple, cost-efficient, practical so it is worth promoting.

A sensitive, rapid and accurate liquid chromatography-tandem mass spectrometric (LC-MS/MS) method with pre-column derivatization was developed for the simultaneous determination of erdosteine and its thiol-containing active metabolite in human plasma. Paracetamol and captopril were chosen as the internal standard of erdosteine and its active metabolite, respectively. Aliquots of 100 microL plasma sample were derivatized by 2-bromine-3'-methoxy acetophenone, then separated on an Agilent XDB-C18 (50 mm x 4.6 mm ID, 1.8 microm) column using 0.1% formic acid methanol--0.1% formic acid 5 mmol x L(-1) ammonium acetate as mobile phase, in a gradient mode. Detection of erdosteine and its active metabolite were achieved by ESI MS/MS in the positive ion mode. The linear calibration curves for erdosteine and its active metabolite were obtained in the concentration ranges of 5-3 000 ng x mL(-1) and 5-10 000 ng x mL(-1), respectively. The lower limit of quantification of erdosteine and its active metabolite were both 5.00 ng x mL(-1). The pharmacokinetic results of erdosteine and its thiol-containing active metabolite showed that the area under curve (AUC) of the thiol-containing active metabolite was 6.2 times of that of erdosteine after a single oral dose of 600 mg erdosteine tables in 32 healthy volunteers, The mean residence time (MRT) of the thiol-containing active metabolite was (7.51 +/- 0.788) h, which provided a pharmacokinetic basis for the rational dosage regimen.

ObjectiveTo investigate the feasibility of preventing tendon adhesion by using bFGF/ vitamin C composite biodegradable membrane.MethodsSixty Newzerland rabbits were divided into experimental and control groups randomly.After the animal model was established, the tendon autografts were encapsulated with the bFGF/vitamin C composite biodegradable membrane in experimental group, while no tendon autografts encapsulation in control.Three weeks after surgery, regular HE staining and AgNO3 staining were performed to observe the fibroblast nuclei and vitamin C.The quantity of collagen fibrils was measured by Luzex-F Image Analyzer.Eight weeks after surgery, the peritendinous adhesion and the maximum tensile load were analyzed.Results At 3 weeks after surgery, the numbers of vitamin C granules, fibroblast nuclei and collagen fibrils in the experimental group were significantly more than those in the control group(t = 11.78 ,P ＜0.001) .At 8 weeks after surgery, the peritendinous adhesion in the experimental group was significantly slighter than that in the control group(Z =3.922,P ＜0.005) ,and the maximum tensile load in the experimental group was significantly higher than that in the control group (t = 8.39, P ＜ 0.001) .ConclusionbFGF/vitamin C composite biodegradable membrane can stimulate the proliferation of fibroblasts and synthesis of collagen fibrils,improve the biomechanical property of tendon autografts and prevent the tendon adhesion.

AIM: To study the action of Yuyang Powder(Herba Swertiae, Corium Elephantis, Rhizoma Bletillae, etc.) on the experimental gastric ulcer. METHODS: The rat gastric ulcer models were induced by burned acetic acid, irritabillity stimulus, ethanol injure gastric mucosa and pyloric ligation, respectively. The ulcer was obsered and the content of gastric juice, gastric acidity and pepsin were measured. RESULTS: Yuyang Powder could significantly reduce the areas of ulcers, restrain gastric ulcer, promote gastric ulcer heal, reduce gastric juice volume, restrain gastric secretion (in pyloric ligation). CONCLUSION: Yuyang Powder has the preventive and therapeutic in gastric ulcer.

Objective To study the clinical evaluation of the effect of topical application of Periocline ointment in the adjuvant treatment of periodontitis.Methods100 patients with periodontitis as the research object in this study in Xi'an Jiaotong university school hospital from April 2015 to August 2016 were randomly divided into the control group and the experimental group, 50 cases in each group.The control group were treated with the conventional treatment, at this basis, the experimental group were given Periocline ointment topical application as the auxiliary treatment.Eight weeks after treatment, the clinical treatment and related indicators were compared in the two groups.ResultsThe bleeding index, probing depth and attachment level in the experimental group were significantly better than those in the control group (P<0.05).In the experimental group of 50 patients, the total effective number of cases in the treatment of 48 cases, The total effective rate in the experimental group was 94.0%, which was significantly higher than 88.0% in the control group (P<0.05).ConclusionIt can improve the effective rate that Periocline ointment was local applicated in the adjuvant treatment of periodontitis;it is worthy of clinical application.

AIM:To observe the effects of cholecystokinin octapeptide(CCK-8) on the expression of proinflammatory cytokines IL-1?,IL-6 and anti-inflammatory cytokines IL-10,IL-4 in LPS-attacked mice.METHODS: Kunming mice were randomly assigned and injected intraperitoneally with LPS alone or/and CCK-8 at different time points.The expression of IL-1?,IL-6,IL-10 and IL-4 in the serum and lung tissues were assayed by ELISA and RT-PCR.RESULTS: The expression of IL-1?,IL-6,IL-10 and IL-4 were upregulated in LPS-attacked mice.Pre-treatment of CCK-8 decreased both IL-1? and IL-6 expression and augmented IL-10 and IL-4 expression in LPS-attacked mice.CONCLUSIONS: CCK-8 exerts an anti-inflammatory effect by inhibiting the expression of IL-1?,IL-6 and increasing the expression of IL-10,IL-4 in LPS-attacked mice,which could alleviate the inflammatory response in lung tissue.

Aim To investigate the effects and mechanisms of CCK-8 on IL-1? induced proliferation of RSC-364, a rat fibroblast-like synovial cell line. Methods MTT colorimetric assay and Western blot were used to measure cell proliferation and p38MAPK phosphorylation level to elucidate the mechanism of CCK-8 in IL-1? induced RSC-364 proliferation. Results CCK-8 significantly inhibited IL-1?-induced RSC-364 proliferation at 10 -12 , 10 -10 , 10 -8 , 10 -6 mol ? L -1 , and IL-1?-activated p38MAPK activity at 10 -10 , 10 -8 , 10 -6 mol?L -1 in a dose-dependent manner. The effect of CCK-8 was blocked by CR1409 (a CCKA-receptor antagonist) and CR2945 (a CCKB-receptor antagonist). Conclusion CCK-8 inhibits IL-1?-induced RSC-364 proliferation, probably by reducing p38MAPK activity through CCKA and CCKB receptors.