Photoinducible Azobenzene trimethylammonium bromide (AzoTAB)-mediated giant vesicle fusion compatible with synthetic protein translation reactions.

Lipid giant vesicles represent a versatile minimal model system to study the physicochemical basis of lipid membrane fusion. Membrane fusion processes are also of interest in synthetic cell research, where cell-mimicking behavior often requires dynamically interacting compartments. For these applications, triggered fusion compatible with transcription-translation systems is key in achieving complexity. Recently, a photosensitive surfactant, azobenzene trimethylammonium bromide (AzoTAB), has been reported to induce membrane fusion by a photoinduced conformational change. Using imaging flow cytometer (IFC) and confocal microscopy we quantitatively investigated photoinduced AzoTAB-mediated fusion of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine vesicles. The IFC analysis result showed that the fusion rate could reach about 40% following AzoTAB addition and UV irradiation in optimized conditions. We confirmed the compatibility between AzoTAB-induced vesicle fusion and a synthetic cell-free protein translation system using green fluorescent protein as reporter. With the techniques presented, cell-sized vesicle fusion can be quantitatively analyzed and optimized, paving the way to controllable synthetic cells with fundamental biological functions like the ability to express proteins from encapsulated plasmids.

Giant Vesicles Produced with Phosphatidylcholines (PCs) and Phosphatidylethanolamines (PEs) by Water-in-Oil Inverted Emulsions.

(1) Background: giant vesicles (GVs) are widely employed as models for studying physicochemical properties of bio-membranes and artificial cell construction due to their similarities to natural cell membranes. Considering the critical roles of GVs, various methods have been developed to prepare them. Notably, the water-in-oil (w/o) inverted emulsion-transfer method is reported to be the most promising, owning to the relatively higher productivity and better encapsulation efficiency of biomolecules. Previously, we successfully established an improved approach to acquire detailed information of 1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)-derived GVs with imaging flow cytometry (IFC); (2) Methods: we prepared GVs with different lipid compositions, including phosphatidylcholines (PCs), phosphatidylethanolamines (PEs), and PC/PE mixtures by w/o inverted emulsion methods. We comprehensively compared the yield, purity, size, and encapsulation efficiency of the resulting vesicles; (3) Results: the relatively higher productivities of GVs could be obtained from POPC, 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC), 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine (DLPE), DOPC: DLPE (7:3), and POPC: DLPE (6:4) pools. Furthermore, we also demonstrate that these GVs are stable during long term preservation in 4 °C. (4) Conclusions: our results will be useful for the analytical study of GVs and GV-based applications.

Observation of normal appearance and wall thickness of esophagus on CT images.

PURPOSE: This study sought to observe the appearance of normal esophagus, measure and record the thickness of esophageal wall in order to offer reference for estimating esophageal wall abnormalities and delineating gross tumor target of esophageal carcinomas on CT images. MATERIALS AND METHODS: From September 2006 to February 2007, 110 consecutive CT films from adult patients without esophageal diseases were collected and studied. On CT images the entire esophagus was divided into cervical, thoracic, retrocardiac and intraabdominal segments. The appearance of esophagus was described when the esophagus contracted or dilated. Thickness of esophageal wall and diameters of esophageal cavities were measured by hard-copy reading with a magnifying glass. Age, sex and the thickness of subcutaneous fat of each patient were recorded. RESULTS: It was observed that the esophagus presented both contracted and dilated status on CT images. In each segment there were certain portions of esophagus in complete contraction or dilatation. 47 images (42.7%) showed contracted esophagus in each segment available for measurement. The largest wall thickness when esophagus was in contraction and dilatation was 4.70 (95%CI: 4.44-4.95)mm and 2.11 (95%CI: 2.00-2.23)mm, respectively. When contracting, the intraabdominal esophagus was thicker than the cervical, thoracic and retrocardiac parts, and the average thickness was 5.68 (95%CI: 5.28-6.09)mm, 4.67 (95%CI: 4.36-4.86)mm, 4.56 (95%CI: 4.31-4.87)mm, and 4.05 (95%CI: 3.71-4.21)mm, respectively. When the esophagus was dilating, the average esophageal wall thickness was between 1.87 and 2.70 mm. The thickest part was cervical esophagus. Thickness of esophageal wall was larger in males than that of females (5.26 mm vs. 4.34 mm p<0.001). Age and the thickness of subcutaneous fat had no significant impact on the thickness of esophageal wall (p-value was 0.056 and 0.173, respectively). CONCLUSION: The Observation of normal appearance and wall thickness of esophagus helps us to identify thickened esophageal wall on CT images using new CT scan technologies. Thus it is probably helpful in judging esophageal diseases and delineating gross tumor target of esophageal carcinomas in modern radiotherapy.