One of the important strategies for improving recombinant protein expression in Pichia pastoris is optimizing control during the high density fermentation process. Some fermentation control has been used and got obviously higher expression level:optimization of basal salt medium composition; using two stage pH and two stage temperature control during cell growth and induction stage; appropriately increasing dissolved oxygen; selecting optimal cell density prior to induction (CWPTI) and specific growth rate prior to induction (?PTI),using limit amount of initial glycerolin the basic media and feeding it during the fermentation with exponential fed-batch Model,selecting appropriate methanol feeding strategy by methanol-limited feeding batch (MLFB),oxygen-limited feeding batch (OLFB),methanol-unlimited feeding batch (MNLFB) or temperature-limited feeding batch (TLFB). Some research suggested that the expression level of the recombinant protein increase 9 times after optimizing the fermentation control.

Objective To study corrective method for displacement in the procedure of electronic portal imaging device (EPID)-based intensity-modulated radiotherapy dose valuation by studying the relative mechanical displacement of different vendor EPID (aS1000,Varian; aS500,Varian; iViewGT,Elekta).Methods A 5 cm × 5 cm field was set up to acquire portal images for three kinds of EPID,then a in house software was used to analysis the portal images.The relative displacement was acquired via analyzing a series of comparation between center positions of gantry angle ranging from 0° to 360° and gantry angle at 0°.Results In the lateral direction,the maximum relative displacement of EPID with aS1000,S500 and iViewGT were (-0.23 ±0.17) mm,(2.94±0.17) mm and (0.35 ±0.09) mm,respectively.In the longitude direction,the displacements were (-4.16 ± 0.20) mm,(-4.15 ± 0.25) mm and (-1.66 ±0.11) mm,respectively.As to longitude direction,the displacements could be well fitted with the usage of quadruplicate empiric function.Conclusions There is a significant difference at the aspect of relative displacement between different vendors EPID at different gantry angles.And the displacement in the longitude direction is obviously larger than in the lateral direction.The relative displacement should be corrected when applying EPID to the intensity-modulated radiotherapy dose verification at different gantry angles.

Objective To optimize preparation process of Zushima Gel Cream. Methods The comprehensive evaluation set sensory evaluation, initial adhesive force, viscous force, and peeling strength score as indexes. The mixing time, refining temperature, mixing speed, and powder adding sequence were investigation factors. Orthogonal experiment was used to optimize forming process. Results Conditions of optimized preparation process were as following: add Zushima powder in Viscomate NP-700 and glycerol; mixing time was 5 min; refining temperature was 40 ℃; mixing speed was 100 r/min. Conclusion The preparation process is good and optimized Zushima Gel Cream has a good adhesive force, good glossiness and excipients. The preparation process is good.

We have developed a rapid and high throughput lipase-ANS (8-Anilino-l-naphthalenesulfonic acid) assay to evaluate the thermo-stability of lipases based on the ANS fluorescence signal's increasing and shifting when this small fluorescence probes binds to lipase. The testing lipase samples were incubated at a temperature range of 25 degrees C to 65 degrees C for 30 min before mixed with ANS solution (0.20 mg/mL lipase and 0.05 mmol/L ANS in the buffer of 20 mmol/L Tris-HCl, 100 mmol/L NaCl, pH 7.2) in a cuvette or microplate. Fluorescence signals of the samples were measured at EX 378 nm, EM 465 nm with a fluorescence photometer or a plate reader, and Tm was calculated with the software of GraphPad Prism5.0. The Tm values of several mutants of Penicillium expansum lipase (PEL) were measured with this ANS assay and conventional method simultaneously and the results show that Tm values are comparative and consistent between these methods, suggesting that the lipase-ANS assay is a reliable, rapid and high throughput method for lipase thermo-stability measurement.
Anilino Naphthalenesulfonates ; chemistry ; Enzyme Stability ; High-Throughput Screening Assays ; methods ; Hot Temperature ; Lipase ; metabolism ; Spectrometry, Fluorescence

The objectives of the study were to screen prostaglandin E2 receptor 4 antagonist from active compounds of Baeckea frutescens L. and to explore its anti-rheumatoid arthritis effect.The HEK293T-EP4 cell antagonist screening model was established in vitro. Homogeneous time-resolved fluorescence (HTRF) technique was used to screen the active compounds of Baeckea frutescens L..SPF grade ICR male mice were randomly divided into control group, model group, methotrexate group, and Baeckea frutescens L.compound BF-2 (100, 50 and 25 mg/kg) groups. The collagen-induced arthritis (CIA) mouse model was established in vivo. The swelling volume of the toes of mice was measured, and the pathological examination was analyzed by staining with hematoxylin and eosin.SPF grade ICR mice, male, were randomly divided into control group, BF-2 (100, 50 and 25 mg/kg) group, and aspirin group.The acetic acid-induced body writhing test was observed.The EP4 in vitro antagonist screening model was successfully established.The preliminary screening results found that BF-2, BF-20, BF-11 and BF-12 had strong EP4 antagonistic activity [(102.11 ± 3.45)%, (90.31 ± 3.59)%, (75.72 ± 1.79)% and (76.84 ± 1.64)%], and BF-2 had the strongest antagonistic activity (IC50 = 0.99 ± 0.08 μg/mL).BF-2 could significantly inhibit the toe swelling of CIA mice, and relieve the degradation of articular cartilage matrix and inflammatory cell infiltration.At the same time, compared with the control group, the writhing times of the mice in each dose of BF-2 were significantly reduced.In this study, BF-2 of Baeckea frutescens L.was selected as an EP4 antagonist, which has potential anti-rheumatoid arthritis activity.

Objective: To explore the influence of different inner dressings in negative-pressure wound therapy (NPWT) on escharectomy wound of full-thickness burn rabbits. Methods: Eighteen Japanese white rabbits were inflicted with full-thickness burn on unilateral back. They were divided into polymer dressing group (PD), biological dressing group (BD), and silver biological dressing group (SBD), according to the random number table, with 6 rabbits in each group. On 3 days post burn, the wounds were performed with escharectomy, and then wounds of rabbits in group PD were covered with polyurethane foam. Wounds of rabbits in group BD were covered with porcine acellular dermal matrix (ADM) and wounds of rabbits in group SBD were covered with silver porcine ADM. Then continuous NPWT was performed on rabbits of the three groups for 7 days. Immediately after surgery and on post surgery day (PSD) 7, general observation of wound was conducted and tissue around the wound was harvested for determination of dry to wet weight ratio. The content of bacteria was counted and the content of tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β), and IL-6 in wound was determined by enzyme-linked immunosorbent assay. Fibroblasts in wound were counted after Masson staining and number of microvessels was counted after CD31 antibody immunohistochemical staining. Data were processed with analysis of variance for repeated measurement, LSD-t test, paired samples t test, and Bonferroni correction. Results: (1) Immediately after surgery, there was no granulation tissue in basal wound of rabbits in the three groups, with rich blood supply and obvious edema. On PSD 7, much granulation tissue was found in basal wound of rabbits in the three groups, with no or mild edema and no obvious redness and swelling in wound edge. (2) There were no significant differences in dry to wet weight ratios of tissue around the wound among and within the three groups immediately after surgery and on PSD 7 (with F values respectively 0.70 and 0.09, t values from 0.17 to 0.52, P values above 0.05). (3) Immediately after surgery, the content of bacteria in wounds of rabbits in groups PD, BD, and SBD was respectively (603.0±146.0) ×10^4, (573.0±63.0) ×10^4, and (590.0±100.0)×10⁴ colony-forming unit (CFU)/g, with no significant difference among them (F=0.13, P>0.05). On PSD 7, the content of bacteria in wounds of rabbits in groups PD, BD, and SBD were respectively (5.4±0.8) ×10^4, (4.6±0.9) ×10^4, and (3.5±0.9)×10⁴ CFU/g. Among them, the content of bacteria in wounds of rabbits in group SBD was lower than that in groups PD and BD, respectively (with t values respectively 3.78 and 2.29, P<0.05 or P<0.01). The content of bacteria in wounds of rabbits in the three groups on PSD 7 was decreased compared with that immediately after surgery (with t values from 10.05 to 21.81, P values below 0.01). (4) There was no significant difference in content of TNF-α, IL-1β, and IL-6 in wounds of rabbits in the three groups immediately after surgery and on PSD 7 (with F values from 0.10 to 1.89, P values above 0.05). The content of TNF-α in wounds of rabbits in the three groups on PSD 7 was significantly higher than that immediately after surgery (with t values from 2.93 to 5.01, P<0.05 or P<0.01). (5) There was no significant difference in amount of fibroblasts in wounds of rabbits in the three groups immediately after surgery and on PSD 7 (with F values respectively 0.01 and 0.81, P values above 0.05). The amount of fibroblasts in wounds of rabbits in the three groups on PSD 7 was larger than that immediately after surgery (with t values from 4.78 to 11.58, P values below 0.01). (6) There was no significant difference in number of microvessels in wounds of rabbits in the three groups immediately after surgery and on PSD 7 (with F values respectively 2.42 and 2.49, P values above 0.05). The number of microvessels in wounds of rabbits in the three groups on PSD 7 was larger than that immediately after surgery (with t values from 7.17 to 11.14, P values below 0.01). Conclusions SBD is better at inhibiting the growth of bacteria. PD, BD, and SBD have almost the same effects on reducing tissue edema and inflammatory reaction, and on promoting the accumulation of collagen fibers and tissue vascularization.

Objective:To explore the relationship between death indicators and unplanned return indicators on healthcare quality evaluation.Methods:A total of 836 976 medical record data were collected from 31 tertiary public general hospitals in a diagnosis-related groups(DRG) data platform in 2017. Multiple death indices(low and low-risk risk group mortality, high-risk group mortality, crude mortality, and risk adjusted mortality) and unplanned return indices(31-day unplanned readmission rate and 31-day unplanned return to surgery rate) were calculated. Pearson′s correlation coefficient was used to examine the relationships among those indices.Results:Death indicators were correlated with each other, but the unplanned readmission rate was not correlated with the unplanned reoperation rate( r=0.305). There was no correlation between unplanned re-entry rate and death rate. The correlation coefficients were as follows: unplanned readmission rate versus low and low-risk group mortality( r=-0.227), versus high-risk group mortality( r=-0.098), versus actual mortality( r=-0.130), versus risk adjusted mortality( r=0.010); unplanned reoperation rate versus low and low-risk group mortality( r=0.105), versus high-risk group mortality( r=0.030), versus actual mortality( r=-0.004), versus risk adjusted mortality( r=-0.141). Conclusions:The indicators of death and the indicators of unplanned return are not the same in terms of actual management technology and evaluation effect. They are complementary to each other and can form an ideal combination of quality evaluation indicators.