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β-glucosidase has important applications in food, pharmaceutics, biomass conversion and other fields, exploring β-glucosidase with strong adaptability and excellent properties thus has received extensive interest. In this study, a novel glucosidase from the GH1 family derived from Cuniculiplasma divulgatum was cloned, expressed, and characterized, aiming to find a better β-glucosidase. The amino acid sequences of GH1 family glucosidase derived from C. divulgatum were obtained from the NCBI database, and a recombinant plasmid pET-30a(+)-CdBglA was constructed. The recombinant protein was induced to express in Escherichia coli BL21(DE3). The enzymatic properties of the purified CdBglA were studied. The molecular weight of the recombinant CdBglA was 56.0 kDa. The optimum pH and temperature were 5.5 and 55 ℃, respectively. The enzyme showed good pH stability, 92.33% of the initial activity could be retained when treated under pH 5.5-11.0 for 1 h. When pNPG was used as a substrate, the kinetic parameters Km, Vmax and Kcat/Km were 0.81 mmol, 291.99 μmol/(mg·min), and 387.50 s-1 mmol-1, respectively. 90.33% of the initial enzyme activity could be retained when CdBglA was placed with various heavy metal ions at a final concentration of 5 mmol/L. The enzyme activity was increased by 28.67% under 15% ethanol solution, remained unchanged under 20% ethanol, and 43.68% of the enzyme activity could still be retained under 30% ethanol. The enzyme has an obvious activation effect at 0-1.5 mol/L NaCl and can tolerate 0.8 mol/L glucose. In conclusion, CdBglA is an acidic and mesophilic enzyme with broad pH stability and strong tolerance to most metal ions, organic solvents, NaCl and glucose. These characteristics may facilitate future theoretical research and industrial production.
Subject(s)
beta-Glucosidase , Sodium Chloride , Temperature , Glucose , Ethanol/chemistry , Ions , Hydrogen-Ion Concentration , Enzyme Stability , Substrate SpecificityABSTRACT
Tocilizumab has been reported to attenuate the "cytokine storm" in COVID-19 patients. We attempted to verify the effectiveness and safety of tocilizumab therapy in COVID-19 and identify patients most likely to benefit from this treatment. We conducted a randomized, controlled, open-label multicenter trial among COVID-19 patients. The patients were randomly assigned in a 1:1 ratio to receive either tocilizumab in addition to standard care or standard care alone. The cure rate, changes of oxygen saturation and interference, and inflammation biomarkers were observed. Thirty-three patients were randomized to the tocilizumab group, and 32 patients to the control group. The cure rate in the tocilizumab group was higher than that in the control group, but the difference was not statistically significant (94.12% vs. 87.10%, rate difference 95% CI-7.19%-21.23%, P = 0.4133). The improvement in hypoxia for the tocilizumab group was higher from day 4 onward and statistically significant from day 12 (P = 0.0359). In moderate disease patients with bilateral pulmonary lesions, the hypoxia ameliorated earlier after tocilizumab treatment, and less patients (1/12, 8.33%) needed an increase of inhaled oxygen concentration compared with the controls (4/6, 66.67%; rate difference 95% CI-99.17% to-17.50%, P = 0.0217). No severe adverse events occurred. More mild temporary adverse events were recorded in tocilizumab recipients (20/34, 58.82%) than the controls (4/31, 12.90%). Tocilizumab can improve hypoxia without unacceptable side effect profile and significant influences on the time virus load becomes negative. For patients with bilateral pulmonary lesions and elevated IL-6 levels, tocilizumab could be recommended to improve outcome.
Subject(s)
Humans , Antibodies, Monoclonal, Humanized , COVID-19/drug therapy , SARS-CoV-2 , Treatment OutcomeABSTRACT
Objective? To?estimate?the?incidence?and?risk?factors?for?deep?venous?thrombosis?(DVT)?in?patients?with?severe?traumatic?brain?injury?(TBI)?treated?in?the?intensive?care?unit?(ICU).? Methods? 105?patients?with?TBI?admitted?to?the?First?Affiliated?Hospital?of?University?of?Science?and?Technology?of?China?from?January?2016?to??June?2018?were?enrolled?[Glasgow?coma?scale?(GCS)?3-8;?concise?injury?score?for?other?parts≤3].?All?patients?did?not?receive?any?medication?or?physical?measures?to?prevent?DVT?during?hospitalization.?Bilateral?compression?Doppler?ultrasounds?of?the?double?lower?limbs?and?upper?limbs?were?performed?to?clarify?the?occurrence?of?DVT?on?the?first?day?of?admission?and?twice?a?week?until?ICU?discharge?or?the?death?of?patient.?The?examination?was?performed?by?a?senior?ultrasound?doctor.?It?was?defined?as?DVT?as?long?as?any?deep?vein?had?thrombosis.?Patients?were?divided?into?two?groups?according?to?whether?DVT?occurred?or?not?during?hospitalization.?Clinical?data?such?as?body?mass?index?(BMI),?coagulation?index,?platelet?count?(PLT)?and?deep?venous?catheterization?were?obtained?from?the?clinical?chemistry?laboratory?database?and?patient?files.?Logistic?regression?was?used?to?analyze?the?risk?factors?of?DVT.?Binary?Logistic?regression?was?used?to?calculate?the?predictive?probability?of?risk?factors.?The?predictive?value?of?risk?factors?and?predictive?probability?to?the?occurrence?of?DVT?was?analyzed?by?receiver?operating?characteristic?(ROC)?curve.? Results? In?105?patients?with?simple?TBI,?42?patients?developed?DVT?during?hospitalization,?and? the?incidence?of?DVT?was?40%.?Univariate?Logistic?regression?showed?that?high?BMI?[odds?ratio?(OR)?=?1.490,?95%?confidence?interval?(95%CI)?=?1.174-1.891,?P?=?0.001],?high?PLT?(OR?=?1.023,?95%CI =?1.006-1.040,?P =?0.007),?shorten?activated?partial?thromboplastin?time?(APTT;?OR?=?0.913,?95%CI?=?0.853-0.978,?P?=?0.010)?and?catheterization?in?deep?vein?(OR?=?0.114,?95%CI =?0.026-0.493,?P?=?0.004)?were?risk?factors?for?DVT.?It?was?shown?by?multivariate?regression?analysis?that?high?BMI?(OR?=?1.378,?95%CI =?1.411-1.665,?P?=?0.001),?high?PLT?(OR?=?1.017,?95%CI =?1.003-1.032,?P?=?0.020),?low?APTT?(OR =?0.920,?95%CI =?0.860-0.982,?P?=?0.012)?and?catheterization?in?deep?vein??(OR?=?0.113,?95%CI =?0.029-0.443,?P?=?0.002)?were?independent?risk?factors?for?DVT.?The?predictive?probability?formula:?Logit?P?=?-4.673+0.321×BMI-0.083×APTT+0.017×PLT-2.181×catheterization?in?deep?vein.?It?was?shown?by?ROC?curve?analysis?that?high?BMI,?high?PLT,?low?APTT?and?catheterization?in?deep?vein?could?predict?the?occurrence?of?DVT?in?severe?TBI?patients,?and?the?area?under?ROC?curve?(AUC)?was?0.775,?0.709,?0.709?and?0.680,?respectively.?The?AUC?of?prediction?probability?was?0.890,?and?its?sensitivity?and?specificity?were?respectively?increased?to?88.10%?and?85.71%.? Conclusions? The?incidence?of?DVT?was?higher?in?patients?with?simple?severe?TBI.?High?coagulation,?high?BMI,?high?PLT?and?catheterization?in?deep?vein?are?the?independent?risk?factors?for?DVT.
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Objective To investigate the effect of early rehabilitation physiotherapy on muscle quality and function in critical patients. Methods A prospective randomized controlled study was performed. Adult critically ill patients admitted to intensive care unit (ICU) of Anhui Provincial Hospital from October 1st, 2016 to August 31st, 2017 who had been hospitalized for more than 7 days and had acute physiology and chronic health evaluation Ⅱ(APACHE Ⅱ ) > 8 were enrolled, and they were divided into treatment group and control group according to random number table method. All patients were given routine treatment, and on this basis, the treatment group was given rehabilitation therapy within 24 hours after admission, including limb active / passive activities, respiratory muscle function training and transcutaneous electrical nerve stimulation, etc. Bedside ultrasound was used to measure the area and cross sectional thickness of left rectus femoris muscle and the cross sectional thickness of middle thigh muscle of patients at 1, 4 and 7 days after treatment; at the same time, the muscle strength of sober patients was evaluated by medical research council (MRC) muscle strength evaluation method, and the mechanical ventilation time, ICU hospitalization time and ICU expenses were recorded. Results Forty patients were enrolled in this study, with 20 in each group. Compared with the control group, the difference of left rectus femoris muscle area between 1 day and 4 days, 4 days and 7 days, 1 day and 7 days (cm2: 0.19±0.02 vs. 0.31±0.19, 0.02±0.01 vs. 0.08±0.05, 0.04±0.02 vs. 0.38±0.23), and the difference in left rectus femoris thickness (cm: 0.01±0.01 vs. 0.14±0.13, 0.03±0.03 vs. 0.16±0.14) and the difference in middle thigh muscle thickness (cm: 0.02±0.02 vs. 0.11±0.09, 0.03±0.02 vs. 0.16±0.12) between 1 day and 4 days, 1 day and 7 days in the treatment group were significantly reduced (all P <0.01). The MRC strength score in the treatment group was significantly higher than that of the control group at 7 days (52.06±3.52 vs. 47.94±3.96, P < 0.05). The mechanical ventilation time in the treatment group (n = 15) and the control group (n = 13) were (138.5±34.5) hours and (185.0±40.9) hours, respectively, and the difference between two groups were statistical significance (P < 0.05). Compared with the control group, the incidence rate of ICU acquired muscle weakness (ICUAW) in the treatment group was significantly decreased [5.0% (1/20) vs. 40.0% (8/20), P < 0.05], the length of ICU stay was significantly shortened (days: 17.67±4.91 vs. 22.06±5.94, P < 0.05), and the ICU expenses were significantly reduced (ten thousand yuan: 7.53±2.09 vs. 9.55±1.73, P < 0.05). Conclusion Early rehabilitation physiotherapy can improve the muscle quality and function in critical patients, and decrease the length of ICU stay.
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Objective To observe the variation tendency of laboratory indexes after delivery in pregnant women with acute fatty liver of pregnancy (AFLP) and the effects of plasma exchange (PE) on the patients prognoses and those parameters. Methods The data of 37 patients with AFLP admitted to the Department of Intensive Care Unit (ICU) of Anhui Provincial Hospital from January 2009 to June 2017 were retrospectively analyzed and the authors tried to clarify the natural course of AFLP; all the selected patients met the Swansea criteria of the AFLP diagnosis. The patients were divided into a PE group (9 cases) and a non-PE group (28 cases) according to whether they were treated with PE or not. At the same time, the effects of PE on liver function and coagulation parameters in the patients were analyzed. Results The most common clinical symptoms of 37 AFLP patients were as follows: nausea and vomiting (21 cases, incidence 56.8%), jaundice (21 cases, 56.8%), upper abdominal pain (14 cases, 37.8%), and more than 90% of patients had more than one symptom. All 37 patients with AFLP had abnormal liver function, their differences in the severity degrees were great, and the level of aspartate aminotransferase (AST) was (271.65±265.90) U/L;the levels of creatinine (SCr) of 72.9% patients (27 cases) were more than 140 mmol/L; the prothrombin times (PT) of 72.97% patients (27 cases) were more than 15 seconds; all patients had received blood product transfusion. Liver function and blood coagulation parameters mostly returned to normal within 1 week after delivery: AST rapidly decreased on the 1 - 2 days after delivery and then continued to decrease; all the cholesterol levels in the two groups decreased to the lowest levels within 3 - 4 days after delivery and then began to rise; the total bilirubin (TBil) levels kept on basically unchanging after delivery; in the patients with levels of platelet counts (PLT) on admission ≥ 150×109/L, on the second day after delivery, their counts returned to approximately 100×109/L, while in patients with PLT < 150×109/L on admission, on the sixth day after delivery, the counts began to slowly increase to about 100×109/L; the patients with fibrinogen (Fib) levels > 1.5 g/L on admission, the levels slightly decreased 2 days before delivery, then gradually recovered and stabilized after delivery, while the patients with Fib < 1.5 g/L, the levels began to rise slowly on the second day after delivery; prothrombin time (PT) and activated partial thromboplastin time (APTT) were basically return to the normal reference ranges on the fifth day after delivery. The PE therapy could significantly improve the patients' laboratory indexes: the levels of AST, TBil, PLT, PT were significant lower immediately and on the second day after PE compared with those levels before the PE [AST (U/L): 197 (114, 383), 116 (65, 311) vs. 239 (125, 430), TBil (μmol/L):109 (67, 126), 125 (103, 178) vs. 164 (99, 198), PLT (×109/L): 96.44±33.10, 89.89±33.03 vs. 126.22±40.83, PT (s): 17.82±5.93, 18.36±3.19 vs. 22.67±8.44, all P < 0.05]. Conclusion After terminating the delivery in time for about one week, the AFLP patients' various indexes may basically return to normal; PE therapy can improve the patients' liver function and coagulation indexes.
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Objective To evaluate the efficacy and safety of colistimethate sodium (CMS) for the treatment of critical patients infected by pan-drug resistantAcinetobacter baumannii (PDR-AB) or pan-drug resistant Pseudomonas aeruginosa (PDR-PA).Methods 321 isolates of PDR-AB and 204 isolates of PDR-PA from critical patients admitted to 35 intensive care units (ICUs) of grade two or above were collected from the Anhui Antimicrobial Resistance Investigation Net (AHARIN) program from September 2012 to September 2015, while the minimal inhibitory concentrations (MIC) of colistin were determined by the E-test. A series of Monte Carlo simulations was performed for CMS regimens (1 MU q8h, 2 MU q8h, and 3 MU q8h, and MU meant a million of unit), and the probability of achieving a 24-hour area under the drug concentration time curve (AUC24)/MIC ratio > 60 and risk of nephrotoxicity for each dosing regimen was calculated. Each simulation was run over three CLCr ranges: 32.51%). Moreover, low value of PTA ( 89.24%) even in patients with CLCr ≥ 90-120 mL/min, and PTA was 33.68%).Conclusion Measurement of MIC, individualized CMS therapy and therapeutic drug-level monitoring should be considered to achieve the optimal drug exposure and ensure the safety of CMS.
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Objective To investigate the possible mechanism of natural killer cells(NK cells)in immune dysfunction in sepsis by monitoring the phenotype and function of periphery NK cells in patients with sepsis. Methods A retrospective study was conducted. The patients with systemic inflammatory response syndrome(SIRS,n=59)or sepsis(n=65)admitted to Department of Critical Care Medicine of Anhui Provincial Hospital from August 2011 to August 2013 were enrolled. Blood samples were collected within 48 hours after intensive care unit(ICU)admission,the phenotype and function of periphery NK cells were determined by flow cytometry. Twenty-eight healthy people served as controls. Results The proportion and number of peripheral blood CD3-CD56+NK cells in SIRS and sepsis groups were normal,and no statistical difference was found when compared with those of the healthy control group〔cell proportion:0.102±0.019,0.102±0.108 vs. 0.106±0.018,F=0.018,P=0.982;cell number(×106/L):182.46±65.98, 172.97±63.51 vs. 179.25±60.44,F=0.349,P=0.706〕. It was shown by NK cell degranulation detection that there was no significant difference in the expression of CD107 and interferon-γ(IFN-γ)secretion〔CD107:0.135±0.050,0.140±0.058,0.128±0.070,F=0.583,P=0.560;IFN-γ(kU/L):14.36±4.74,12.49±4.21, 13.45±5.04,F=1.616,P=0.202〕among healthy control group,SIRS group,and sepsis group. It was shown by antibody dependent cytotoxic effect(ADCC)test that there was no difference in the expression of CD107 among healthy control group,SIRS group,and sepsis group(0.574±0.166,0.643±0.165,0.581±0.157,F=0.808,P=0.448). When compared with healthy controls,the secretion of IFN-γwas increased in SIRS patients(kU/L:40.5±13.2 vs. 28.4±9.6,P=0.001),while reduced in sepsis patients(kU/L:19.8±6.7 vs. 28.4±9.6,P<0.01). Compared with SIRS group,only NK cell surface inhibitory receptors CD158e(KIR 3DL1)expression in sepsis group was significantly increased(0.203±0.057 vs. 0.079±0.021,t=15.762,P<0.001),and there were no significant differences in the other phenotype between the two groups. Compared with SIRS group,the IFN-γproduction of the sepsis group was significantly lowered(kU/L:0.280±0.040 vs. 0.310±0.038,t=3.390,P=0.009),and the level of IL-12 was also significantly decreased(ng/L:0.15±0.03 vs. 0.30±0.08,t=32.832,P<0.001). Conclusion It was showed by NK cell phenotype and function assay that the function of NK cells in patients with sepsis was impaired and led to a poor production of IFN-γ. The IFN-γmediated immune dysfunction may be a main reason for the disorder of NK cell function,which laid the foundation of the clinical immune intervention practice to improve to NK cell function.
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Objective To investigate the effects of positive end-expiratory pressure (PEEP) on hemodynamics and intra-abdominal pressure in patients with acute respiratory distress syndrome (ARDS) with normal intra-abdominal pressure (IAP) and intra-abdominal hypertension (IAH) during mechanical ventilation under a incremental PEEP.Methods Eighteen ARDS patients with normal IAP or IAH treated in intensive care unit of Anhui Provincial Hospital were enrolled for study.At different PEEP levels (5,10,15,20 cmH2O),hemodynamic parameters including extravascular lung water index (EVLWI),intrathoracic blood volume index (ITBVI),stroke volume variation (SVV) and IAP were measured with PICCO technology.Results Compared with ARDS patients with normal IAP,the ARDS patients with IAH were characterized by:(1) EVLWI was decreasing under the increment of PEEP,but there was no significant difference between 15 cmH2O and 20 cmH2O (t =0.593,P =0.572).As PEEP was gradually escalated,EWLVI had no significant changes in patients with normal IAP.(2) SVV increased as PEEP was being titrated to higher level in both groups but in normal IAP group such effect was observed till PEEP above 10cmH2O (PEEP5 vs.PEEP10,t=0.326,P=0.752; PEEP,10 vs.PEEP 15,t=-20.032,P< 0.01).(3) There was no statistical difference in ITBVI between two groups at varied levels of PEEP.(4) PEEP could increase IAP levels and had much more profound effects on patients with IAH.Conclusions PEEP is a contributing factor changing IAP.It is necessary to take the level of PEEP into account as IAP variation is interpreted in patients under mechanical ventilation.Different PEEP levels can significantly affect the EVLWI in patients with IAH but not do not in ones with nomal IAP.
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Traditional Chinese medicine placebo has been put more and more attention to. However, there is no ac-cepted quality evaluation method for TCM placebo. How to evaluate TCM placebo objectively and quantitatively is a common problem in the industry of Chinese medicine. New technologies such as the Intelligent Sensory Technique have been used to establish the placebo evaluation methods which are suitable for TCM characteristics. This article provided the basis for establishing scientific, rational and objective evaluation guiding principles for TCM.
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Objective To analyze the clinical features and the effect of therapy on cutaneous emphysema of chest wall and/or pneumomediastinum complicated in severe acute triphosgene poisoning patients.Methods Among 81 triphosgene poisoning patients,5 complicated with cutaneousemphysema of chest wall and/or pneumomediastinum were analyzed in respect of the clinical data including age,gender,arterial blood gas (ABG),modes of mechanical ventilation support and so on.Results Five patients consisting of 3 males and 2 females,aged (23.20 ± 5.17) years,were complicated with cutaneous emphysema of chest wall and/or pneumomediastium with a prevalence rate of 0.06%.Of them,4 were alleviated completely and 1 died of acute respiratory distress syndrome (ARDS).There was no significant difference in arterial blood gas analysis (ABG) between patients with cutaneousemphysema and/or pneumomediastinum and patients without ( P > 0.05 ).Conclusions Triphosgene-induced acute lung injury treated with mechanical ventilation support with high PEEP is highly suggested as high risk factor for the formation of cutaneous emphysema of chest wall and/or pneumomediastinum in severe acute Triphosgene poisoning patients.It is very important to set the PEEP level of mechanical ventilation support as low as possible for avoidance of alveolar rupture.