Sorting in early endosomes reveals connections to docking- and fusion-associated factors

The early endosomes constitute a major sorting platform in eukaryotic cells. They receive material through fusion with endocytotic vesicles or with trafficking vesicles from the Golgi complex and later sort it into budding vesicles. While endosomal fusion is well understood, sorting is less characterized; the 2 processes are generally thought to be effected by different, unrelated machineries. We developed here a cell-free assay for sorting/budding from early endosomes, by taking advantage of their ability to segregate different cargoes (such as transferrin, cholera toxin subunit B, and low-density lipoprotein, LDL) into different carrier vesicles. Cargo separation required both carrier vesicle formation and active maturation of the endosomes. Sorting and budding were insensitive to reagents perturbing clathrin coats, coatomer protein complex-I (COPI) coats, dynamin, and actin, but were inhibited by anti-retromer subunit antibodies. In addition, the process required Rab-GTPases, phosphatidylinositol-3-phosphate, and, surprisingly, the docking factor early endosomal autoantigen 1 (EEA1). Sorting also required the function of the N-ethylmaleimide-sensitive factor (NSF), a well-known fusion cofactor, while it did not depend on preceding fusion of endosomes. We conclude that fusion, docking, and sorting/budding are interconnected at the molecular level.     

LigaSure在阴式子宫切除术中的应用

目的探讨LigaSure在阴式子宫切除术中的临床应用效果。方法2007年1月～2008年1月,应用LigaSure行阴式子宫全切术200例。子宫肌瘤86例,子宫增大如孕10～14周;子宫腺肌病73例,子宫增大如孕8～11周;宫颈上皮内瘤变Ⅲ级10例;子宫脱垂Ⅱ～Ⅲ度15例;子宫内膜不典型增生6例;功能失调性子宫出血10例。用LigaSure钳夹、电凝、切断骶韧带、主韧带、子宫血管、圆韧带、输卵管及卵巢固有韧带。结果200例手术全部顺利完成。手术时间20～50min,术中出血量15～150ml,术后住院时间3～5d,无严重并发症发生。200例均术后3个月复诊,检查阴道残端无线头及肉芽组织生成,愈合良好。结论应用LigaSure行阴式子宫切除术,手术操作步骤简化,手术难度系数降低,手术时间短,术中出血少。     

Degradation of cytoplasmic substrates by FtsH,a membrane-anchored protease with many talents

Control of cellular processes by regulated proteolysis is conserved among all organisms. FtsH, the only membrane-anchored AAA protease in bacteria, fulfills a variety of regulatory functions. This review focuses on soluble FtsH substrates in Escherichia coli and in other bacteria and outlines emerging substrate recognition principles.     

SecA2-dependent secretion of autolytic enzymes promotes Listeria monocytogenes pathogenesis

Pathogenic bacteria secrete proteins that promote invasion of host tissues and resistance to immune responses. However, secretion mechanisms that contribute to the enormous morbidity and mortality of Gram-positive bacteria are largely undefined. An auxiliary protein secretion system (SecA2) has recently emerged in Listeria monocytogenes and eight other Gram-positive pathogens. Here, a proteomics approach identified seventeen SecA2-dependent secreted and surface proteins of L. monocytogenes, the two most abundant of which [the p60 and N-acetylmuramidase (NamA) autolysins] hydrolyze bacterial peptidoglycan (PGN) and contribute to host colonization. SecA2-deficient (DeltaSecA2) bacteria were rapidly cleared after systemic infection of murine hosts, and in cultured cells showed reduced cell-cell spread. p60 or NamA deficiencies (Deltap60 and DeltaNamA) caused intermediate reductions in bacterial virulence in vivo, yet showed no defect for infection of cultured cells. Restoration of virulence in Deltap60 bacteria required full-length p60 with an intact catalytic domain, suggesting that PGN hydrolysis by p60 is crucial for L. monocytogenes virulence. Coordinated PGN hydrolysis by p60 and NamA activities is predicted to generate a muramyl glycopeptide, glucosaminylmuramyl dipeptide (GMDP), which is known to modify host inflammatory responses. Thus, SecA2-dependent secretion may promote release of muramyl peptides that subvert host pattern recognition.     

Coronary hybrid revascularization from January 1997 to January 2001: a clinical follow-up

Background. Hybrid revascularization (HyR), combining minimally invasive left internal mammary artery (LIMA) bypass grafting to the left anterior descending coronary artery (LAD) and catheter interventional treatment of the remaining coronary lesions, avoids the disadvantages associated with cardiopulmonary bypass (CPB). We investigated the clinical follow-up of 57 patients with multivessel disease undergoing this procedure in the last 4 years.

Methods. Between January 1997 and January 2001, 57 consecutive patients (41 men and 16 women, aged 65.7 ± 7.9 years) with coronary artery disease (two-vessel, n = 34; three-vessel, n = 23) were treated with off-pump LIMA-to-LAD bypass combined with balloon angioplasty and stenting of the remaining significantly obstructed (> 50%) coronary vessels. Clinical follow-up data included a early postoperative and a 6-month control angiography and a patient interview in January 2001.

Results. All patients underwent LIMA-to-LAD bypass-grafting and balloon angioplasty in 72 coronary lesions without procedural-related complications. However, one early LIMA bypass occlusion was documented during coronary angiography. Postoperatively no deterioration of preexistent organ dysfunction was observed in any patient. The mean follow-up was 100.7 ± 37.9 weeks in 55 of 57 patients (97%). Control angiography 6 months after HyR (n = 34) revealed a patent LIMA bypass in 33 patients and 8 in-stent restenoses (> 50%) in the coronary arteries that were treated interventionally by re-PTCA (n = 6) or by conventional CABG (n = 1). In 1 patient medical treatment resulted in significant reduction of angina so no further intervention was considered necessary. After HyR 1 patient died 18 months later of an intracerebral hemorrhage. All other patients are alive and doing well.

Conclusions. Our results indicate that in selected patients with multivessel disease including left main stem stenosis HyR is an effective and secure procedure with excellent early and good midterm results. Especially elderly patients with severe concomitant diseases appear to benefit from this approach by avoiding CPB.     

Correction of traumatic tricuspid regurgitation using the double orifice technique

We present a case of acute traumatic tricuspid regurgitation in a 39-year-old man who was involved in a motor vehicle accident. A large ecchymotic region over the anterior chest wall prompted evaluation by both transthoracic and transesophageal echocardiography which confirmed the valvular injury. At surgery, valvular incompetence was found to be the result of a flail anterior leaflet due to papillary muscle rupture. The valve was successfully repaired using a single stitch double orifice technique in combination with a ring annuloplasty. The valve remains competent 18 months after surgery.     

Longitudinal spinal cord sections as substratum for anti-neurofilament antibody detection

A rapid and technically simple method for demonstrating anti-neurofilament antibodies using longitudinal sections of rat spinal cord as substratum and indirect immunofluorescent technique is reported. The results compare well with those obtained by the technically more difficult and time-consuming methods using as substratum central neurons cultivated in vitro. A total of 195 serum specimens from different neurological disorders and healthy subjects were studied. Immunofluorescent autoantibodies to neurofilaments were found in specimens of serum from patients with Creutzfeldt-Jakob disease (CJD), kuru, amyotrophic lateral sclerosis, parkinsonism dementia (Guam), Alzheimer's disease, and multiple sclerosis but in higher frequency in CJD and kuru than in the other disease or in healthy control subjects.