No abstract

Here we report a new isolation method for mouse glomeruli. The method is fast and simple and allows for the isolation of virtually all glomeruli present in the adult mouse kidney with minimal contamination of nonglomerular cells. Mice were perfused through the heart with magnetic 4.5-m diameter Dynabeads. Kidneys were minced into small pieces, digested by collagenase, filtered, and collected using a magnet. The number of glomeruli retrieved from one adult mouse was 20,131 ؎ 4699 (mean ؎ SD, n ‫؍‬ 14) with a purity of 97.5 ؎ 1.7%. The isolated glomeruli retained intact morphology, as confirmed by light and electron microscopy, as well as intact mRNA integrity, as confirmed by Northern blot analysis. The method was applicable also to newborn mice, which allows for the isolation of immature developmental stage glomeruli. This method makes feasible transcript profiling and proteomic analysis of the developing, healthy and diseased mouse glomerulus. As the genome projects are near completion, 1,2 an important step in the functional analysis of genome data are the determination of transcriptomes corresponding to specific cellular functions and states of differentiation. Such analyses require methods allowing for the isolation of highly homogenous population of cells and/or microorgans from in vivo situations. One such microorgan is the kidney glomerulus. Glomeruli constitute ϳ10% of whole kidney tissues and are unique structures of microvasculature mainly made up of three highly specialized cell types; fenestrated endothelial cells, mesangial cells, and podocytes. These cell types together with the glomerular basement membrane form the permeable barrier across which blood is filtered to produce primary urine. During the past decade several gene products have been shown to play essential roles in glomerulus development, 3-5 function, and pathology. 6 However, our knowledge of the molecular mechanisms governing glomerulus morphogenesis and development of the specialized features of its individual cells is still very limited. An obvious difficulty in addressing these issues stems from the low abundance of the glomerulus cells and the inability of the glomerulus cell types to retain their differentiated features in cell culture. Podocytes, for example, make up less than 2% of kidney tissues. Although endothelial cells and pericytes exist outside the glomerulus, their phenotype within the glomerulus is quite distinct from related cells elsewhere. 7 We describe a new protocol for the isolation of glomeruli from mice. The protocol is fast and allows for the isolation of virtually all glomeruli present in a mouse kidney at 97% purity. The method thus allows for transcript profiling and proteomic analysis of the glomerulus using standard procedures.

No abstract

IL-23 and Th17 cells not only induce Th17-cell-mediated neutrophilic airway inflammation but also up-regulate Th2-cell-mediated eosinophilic airway inflammation.

Data replication is a key technology in distributed data sharing systems, enabling higher availability and performance. This paper surveys optimistic replication algorithms that allow replica contents to diverge in the short term, in order to support concurrent work practices and to tolerate failures in low-quality communication links. The importance of such techniques is increasing as collaboration through wide-area and mobile networks becomes popular.Optimistic replication techniques are different from traditional "pessimistic" ones. Instead of synchronous replica coordination, an optimistic algorithm propagates changes in the background, discovers conflicts after they happen and reaches agreement on the final contents incrementally.We explore the solution space for optimistic replication algorithms. This paper identifies key challenges facing optimistic replication systems -ordering operations, detecting and resolving conflicts, propagating changes efficiently, and bounding replica divergence -and provides a comprehensive survey of techniques developed for addressing these challenges.