O-GlcNAc糖基转移酶在原核和真核细胞中的表达和提纯

目的 :本文为进一步研究蛋白 O- Glc NAc糖基化修饰提供重要资源。方法 :为了探索制备纯化具有生物学活性的重组 OGT的有效方法 ,本文用大肠杆菌和 Hi5昆虫细胞表达并纯化具有活性的重组 OGT。携带人 OGTc DNA的 PET32 a质粒在大肠杆菌中表达出带有 1个 S- Tag的重组人 OGT融合蛋白 ,然后用与 S-蛋白质偶联的琼脂糖凝胶颗粒从细菌裂解液中亲和纯化 OGT融合蛋白。结果 :其纯化的 OGT在电泳中显现单一条带并具有生物活性 ,但其活性在洗脱后丧失。在 Hi5昆虫细胞中表达的鼠肝 OGT带有一 His6 tag,经镍离子螯合柱纯化后 ,其比活性达到 0 .88nmol· min- 1 · mg- 1 OGT。结论 :在这两个表达系统中制备重组 OGT各有利弊。     

Feasibility of Surgical Treatment for Laryngomalacia Using the Spontaneous Respiration Technique

Objectives.In this study, we review our institutional experience with pediatric laryngomalacia (LM) and report our experiences of patients undergoing supraglottoplasty using the spontaneous respiration using intravenous anesthesia and high-flow nasal oxygen (STRIVE Hi) technique.Methods.The medical records of 29 children with LM who visited Seoul National University Hospital between January 2017 and March 2019 were retrospectively reviewed. Surgical management was performed using the STRIVE Hi technique. Intraoperative findings and postoperative surgical outcomes, including complications and changes in symptoms and weight, were analyzed.Results.Of the total study population of 29 subjects, 20 (68.9%) were female. The patients were divided according to the Onley classification as follows: type I (n=13, 44.8%), II (n=10, 34.5%), and III (n=6, 20.7%). Twenty-five patients (86.2%) had comorbidities. Seventeen patients (58.6%) underwent microlaryngobronchoscopy under STRIVE Hi anesthesia. Four patients with several desaturation events required rescue oxygenation by intermittent intubation and mask bagging during the STRIVE Hi technique. However, the procedure was completed in all patients without any severe adverse effects. Overall, 15 children (51.7%) underwent supraglottoplasty, of whom 14 (93.3%) showed symptom improvement, and their postoperative weight percentile significantly increased (P=0.026). One patient required tracheostomy immediately after supraglottoplasty due to associated neurological disease.Conclusion.The STRIVE Hi technique is feasible for supraglottoplasty in LM patients, while type III LM patients with micrognathia or glossoptosis may have a higher risk of requiring rescue oxygenation during the STRIVE Hi technique.     

全髋股骨头假体无骨水泥翻修术的中期疗效

目的 报道采用长柄全干式或半干式多孔表涂股骨头假体加自体植骨行无骨水泥翻修术(RTHA)20例20髋的中期疗效。方法 均为首次RTHA的20例中诊断股骨头假体骨水泥固定后松动11髋，无骨水泥固定后松动9髋，其中3例伴假体干端骨折。骨缺损(Mulliken 分类)：Ⅱ期8例，Ⅲ期12例。手术采用国产仿AML长柄半干式表涂和进口长柄全干式表涂股骨头假体各10例，后者包括Bi-Metric(Biomet．uSA)3例、AML(Depuy，uSA)4例，REF(Lima，Italy)3例，并取自体髂骨剪成火柴梗状植入假体周围间隙。结果 平均随访7年，功能优13例(65％)，良6例(30％)，尚可l例(5％)。X线表现：所有病例股骨近段骨缺损植骨区修复良好，表现为骨皮质增厚，致密。假体呈骨性固定16例，纤维性稳定4例，无1例松动，至今无1例再翻修。术后第1年内，骨性固定组假体下沉均＜3mm，纤维性稳定组假体下沉均＞3mm，下沉最多达6mm．以后未继续下沉。远段假体髓内充填率在骨性固定组均＞90％，而纤维性稳定组均＜80％。结论 采用植骨修复结构性骨缺损，并选用长柄多孔表涂全髋股骨头假体施行翻修术，是获得显著疗效的关键。     

Comparison of the Manual Assessment of Respiratory Motion (MARM) and the Hi Lo Breathing Assessment in determining a simulated breathing pattern

AimThe aim of this study was to assess and compare the validity and potential utility of two manual breathing assessment procedures: the Manual Assessment of Respiratory Motion (MARM) and the Hi Lo Breathing Assessment. A secondary aim was to gauge the relationships between experience and the ability to perform these assessment techniques, by comparing the performance of students with practitioners.Method56 osteopaths and osteopathic students were taught the MARM and the Hi Lo Breathing Assessment and trained to simulate breathing patterns. The participants, acting alternatively as breathers and examiners, then attempted to accurately determine whether the breathing patterns simulated by their partner were predominately abdominal, thoracic or, in the case of the Hi Lo, paradoxical. Participants were surveyed on their confidence in the use of each technique, their perceived ease in using each technique, and their intended future use of the techniques. Student and practitioner abilities to detect simulated breathing patterns were compared for the MARM and Hi Lo.ResultsOverall scores for correctly determining breathing patterns were not significantly different for the MARM or the Hi Lo, and there was no notable moderation of this effect according to experience, with both practitioners and students demonstrating a high level of performance on both techniques. There were some differences in accuracy of performance across different breathing styles, with Hi Lo assessment of paradoxical breathing being more difficult to identify correctly. Ease of learning was similar for MARM and Hi Lo but confidence in using the techniques, and intended future use was higher for the MARM. There were some significant relationships between these utility measures and performance, particularly on the MARM.ConclusionsThis study builds on our previous study to strengthen the evidence for the validity of the MARM and also supports the validity of the Hi Lo. Responses to the survey indicate that, overall, participants preferred the MARM to the Hi Lo. This study is a preliminary investigation of these techniques. Future studies to test the validity of these techniques should be performed in a clinical setting on individuals with actual rather than simulated breathing pattern disturbances.     

Persistent gastric intramucosal ischemia in patients with sepsis following resuscitation from shock.

STUDY OBJECTIVES: (1) To determine the effects of resuscitation of patients with severe sepsis to conventional hemodynamic end points and normal blood lactate levels on postresuscitation sequential assessments of gastric intramucosal pH (pHi). (2) To determine whether trends in pHi are reflected in trends in systemic hemodynamic, oxygen utilization, and acid-base assessments. DESIGN: Prospective cohort study. SETTING: Medical ICU in an inner-city, university-based medical center. PATIENTS: Twelve recently admitted patients with severe sepsis and signs of circulatory shock who were successfully resuscitated to normal hemodynamic end points and lactate levels and who were also monitored with pulmonary artery catheters and gastric tonometers. INTERVENTIONS: Because of the observational nature of this study, no specific interventions were employed. The physician staff administered i.v. fluids and pharmacologic agents, during and after the resuscitative period, to treat infection and to achieve and maintain hemodynamic stability. Mechanical ventilation and supplemental oxygen were provided as needed. The hemodynamic and physiologic monitoring employed was determined by the managing physicians and established medical ICU routines. MEASUREMENTS AND RESULTS: A total of 12 patients were studied. Systemic hemodynamic, oxygen utilization, and acid-base assessments and pHi were recorded following resuscitation, and every 12 h thereafter. pHi decreased from 7.33 +/- 0.08 (mean +/- SD) following resuscitation to 7.26 +/- 0.04 at 24 h, 7.20 +/- 0.07 at 36 h (p < 0.05), and 7.24 +/- 0.08 at 48 h. Corresponding statistically significant and clinically relevant changes in systemic hemodynamic, oxygen utilization, and acid-base variables were not observed. The hospital mortality of this patient group was high (10 of 12; 83%). CONCLUSIONS: Gastric intramucosal acidosis develops and persists for at least 48 h in patients resuscitated from septic shock to conventional resuscitative end points, including the normalization of lactate levels. These regional changes were not reflected in corresponding changes in systemic acid-base and oxygen utilization variables. Direct determinations of pHi and therapy directed toward the resolution of splanchnic ischemia may be required to improve the outcome in these patients.     

人造血管内瘘进行高通量血液透析的临床应用

目的 报道采用人造血管移植动—静脉内瘘术进行高通量血液透析的临床应用效果。方法 尿毒症患24例，行聚四氟乙烯(FrrFE)人造血管移植动—静脉内瘘术，进行高通量血液透析(HFD)，透析器采用Fresenius F60。观察血流量和并发症发生率。结果人造血管通畅率100％，HFD时血流量均可达到300～450ml／min。术后感染发生率4．17％，血栓形成发生率8．33％，假性动脉瘤发生率4．17％；HFD低血压发生率37．5％，反超滤发生率12．5％。结论 人造血管具有生物相容性好、通畅率高、使用时间长和易穿刺等优点，是血液透析患移植内瘘的理想选择；人造血管动—静脉内瘘能为HFD提供足够的血流。     

Reevaluation of the efficacy and safety of the neutrophil elastase inhibitor, Sivelestat, for the treatment of acute lung injury associated with systemic inflammatory response syndrome; a phase IV study

Introduction

Sivelestat, a neutrophil elastase inhibitor, has been approved in Japan for the treatment of patients with acute lung injury (ALI) associated with systemic inflammatory response syndrome (SIRS). The Pharmaceuticals and Medical Devices Agency (PMDA) has ordered to conduct a postmarket clinical study in order to reevaluate the efficacy and safety of Sivelestat in actual clinical settings in Japan.

Methods

According to the PMDA’s order, we evaluated the efficacy and safety of Sivelestat in Japanese patients with ALI associated with SIRS using ventilator-free days (VFD) as the primary endpoint. The surrogate endpoints are ventilator-weaning rate, ICU discharge rate, and 180-day survival rate. Study design was an open-label, non-randomized, multi-center clinical trial. Sivelestat was intravenously administered at 0.2 mg/kg/h continuously for a maximum of 14 days. Sivelestat group and control group were compared by adjusting the outcome values using an inverse probability of treatment weighted method based on the propensity scores.

Results

Four hundred and four Sivelestat group patients and 177 control group patients were enrolled. The adjusted mean number of VFD was 15.7 and 12.1 in the Sivelestat group and control group, respectively (P = 0.0022). Both the adjusted ventilator-weaning rate and ICU discharge rate were significantly higher in the Sivelestat group than in the control group (P = 0.0028 and P = 0.019, respectively). The adjusted 180-day survival rate was significantly higher in the Sivelestat group than in the control group (71.8 percent vs. 56.3 percent).

Conclusions

Sivelestat contributed to early weaning from the mechanical ventilation, while showing no negative effect on the long-term outcomes of ALI associated with SIRS. The results of this study suggest the clinical usefulness of Sivelestat in this patient population.     

Short-term and low-dose IL-2 therapy increases the reduced Treg cells in patients with microscopic polyangiitis

ObjectiveThe breakdown of immune tolerance mediated by the reduced regulatory T (Treg) cell contributes to autoimmune diseases, which can be recovered by the short-term and low-dose interleukin 2 (IL-2). However, the role of Treg cells in microscopic polyangiitis (MPA) and the efficacy of short-term and low-dose IL-2 for MPA remain unclear. Therefore, we performed a retrospective study to explore the role of Treg cells and evaluate the efficacy of short-term and low-dose IL-2 therapy in MPA.Methods52 MPA were collected as research objects, and 15 of them voluntarily received short-term and low-dose IL-2 subcutaneous injection combined with conventional therapy. 60 volunteers were recruited as health controls (HC) according to the inclusion and exclusion criteria. The number of circulating CD4 + T cell subsets was detected by flow cytometry.ResultsPatients with MPA had reduced circulating Treg cells than HCs (P < 0.001), and the level of Treg cells were reduced in MPA-activity and ANCA-positive group (P = 0.018 and P = 0.008 respectively). The patients with lower Treg cells had the higher incidence of the organ involvement (P = 0.006). The level of Treg cells in MPA was doubled after the short-term and low-dose IL-2 combined with conventional therapy (P = 0.001), and the disease activity indicators such as ESR and CRP were improved (P < 0.05) with no apparent side effects.ConclusionPatients with MPA had reduced circulating Treg cells, especially the MPA-activity and ANCA-positive patients. And the patients with lower Treg cells were more likely to exhibit the organ involvement. Short-term and low-dose IL-2 therapy increased the reduced Treg cells and promoted the remission of the disease at a certain extent with well tolerance.