Following a small dose of nephrotoxic serum (NTS) WKY rats demonstrated crescentic glomerulonephritis, which was characterized by the early infiltration of CD8 positive cells in glomeruli. In vivo depletion of CD8 positive cells from WKY rats completely prevented proteinuria (4.6 +/- 4.8 mg/day vs. 105.3 +/- 11.6 mg/day on day 10; N = 19, P less than 0.001) and crescent formation (2.7 +/- 2.9% vs. 94.3 +/- 2.6%; P less than 0.001). Immunofluorescence revealed complete inhibition of the influx of CD8 positive cells and subsequent reduction of the infiltration of macrophages in the glomeruli. Glomerular binding of 125I-anti-rat glomerular basement membrane antibodies, host anti-rabbit IgG production and the C3 level in the circulation were the same as in the control. These data indicate that CD8 positive cells play a key role in glomerular injury and crescent formation. This model provides a useful system for studying the cellular mechanisms that lead to glomerular injury and subsequent crescent formation.

The effect of vasopressin on subcellular localization of AQP-CD and AQP3 water channels was examined in thirsted Brattleboro rats by immunohistochemistry and immunoelectron microscopy. AQP-CD was mainly present in the cytoplasm of the collecting duct cells in association with cytoplasmic vesicles but was sparse in the apical membrane in control vehicle-injected rats. In rats given vasopressin 15 min before death, the number of immunogold particles for AQP-CD in the apical membrane increased significantly (P < 0.002) from 1.8 +/- 0.2 to 10.0 +/- 0.4/microns with a significant decrease (P < 0.05) of cytoplasmic labeling from 32.6 +/- 6.4 to 24.6 +/- 5.6/microns 2, indicating that AQP-CD is the vasopressin-regulated water channel predicted by the "shuttle" hypothesis. In contrast, AQP3 was restricted to the basolateral membrane of the collecting duct cells, and the labeling density of AQP3 was unchanged by vasopressin treatment, indicating that AQP3 is constitutively expressed and may maintain high water permeability of the basolateral membrane.

A new water channel (aquaporin-8, gene symbol AQP8) was isolated from rat pancreas and liver by homology cloning. Ribonuclease protection assay showed intense expression of the gene in pancreas and liver, less intense in colon and salivary gland, and negligible in other organs. The full-length cDNA was obtained by ligation of ϳ1.4-kilobase (kb) cDNA isolated from the rat liver cDNA library to ϳ0.5 kb of the 5-end fragment obtained by the rapid amplification of cDNA ends method. A major transcript of ϳ1.45 kb was demonstrated in liver and colon by Northern blot analysis. Expression of the cRNA in Xenopus oocytes markedly enhanced osmotic water permeability in a mercury-sensitive manner, indicating a water channel function of this molecule. The open reading frame encoded a 263-amino acid protein with a predicted molecular size of 28 kDa. Hydropathy analysis represented six membranespanning domains and five connecting loops containing two sites of NPA motif as preserved in other aquaporins. Unlike other mammalian aquaporins, AQP8 has an unusual structure with a long N terminus and a short C terminus, which are found in plant aquaporin, ␥-tonoplast intrinsic protein. By in situ hybridization, AQP8 mRNA expression was assumed in hepatocytes, acinal cells of pancreas and salivary gland, and absorptive colonic epithelial cells. The physiological role(s) of AQP8 remain to be elucidated.The aquaporins are water-selective membrane channels found in many species of animals and plants as the family of major intrinsic protein (MIP) 1 (1-3). Aquaporin-1 (AQP1) is the first protein recognized as a channel-forming integral membrane protein of 28 kDa (CHIP-28) expressed in mammalian red blood cells (1, 4) and then as a water channel expressed in renal proximal tubules (5-7) and other water-permeable epithelia (8, 9). Thereafter, four other aquaporins have been cloned in mammals. AQP2 is the vasopressin-regulated water channel, exclusively present in apical membranes of principal cells of collecting ducts in the kidney (10 -13), whereas AQP3 is a water channel locating basolateral membranes of collecting duct cells and transporting water and small solutes such as glycerol and urea (14). AQP4 and AQP5 were cloned from brain and salivary gland, respectively, and were presumed to be implicated in the sensation of osmotic change in hypothalamus and secretion in exocrine glands (15,16). In plants, tonoplast intrinsic protein (TIP) is an integral membrane protein and belongs to the MIP family (17, 18). ␥-TIP was found in the vegetative organs of plant and not in seeds, although ␣-TIP and ␤-TIP were seed-specific (19).The previous studies showed the presence of unique aquaporins in various exocrine glands such as salivary gland and lacrimal gland (16). Since pancreas is a secretory organ, secreting various digestive enzymes such as lipase, amylase, and protease in a volume of about 2,000 ml/day in humans (20), it may be reasonable to speculate the presence of an aquaporin family in pancreas. In the present study, we attempted to isolate...

A family of water-selective channels, aquaporins (AQP), has been demonstrated in various organs and tissues. However, the localization and expression of the AQP family members in the gastrointestinal tract have not been entirely elucidated. This study aimed to demonstrate the expression and distribution of several types of the AQP family and to speculate on their role in water transport in the rat gastrointestinal tract. By RNase protection assay, expression of AQP1–5 and AQP8 was examined in various portions through the gastrointestinal tract. AQP1 and AQP3 mRNAs were diffusely expressed from esophagus to colon, and their expression was relatively intense in the small intestine and colon. In contrast, AQP4 mRNA was selectively expressed in the stomach and small intestine and AQP8 mRNA in the jejunum and colon. Immunohistochemistry and in situ hybridization demonstrated cellular localization of these AQP in these portions. AQP1 was localized on endothelial cells of lymphatic vessels in the submucosa and lamina propria throughout the gastrointestinal tract. AQP3 was detected on the circumferential plasma membranes of stratified squamous epithelial cells in the esophagus and basolateral membranes of cardiac gland epithelia in the lower stomach and of surface columnar epithelia in the colon. However, AQP3 was not apparently detected in the small intestine. AQP4 was present on the basolateral membrane of the parietal cells in the lower stomach and selectively in the basolateral membranes of deep intestinal gland cells in the small intestine. AQP8 mRNA expression was demonstrated in the absorptive columnar epithelial cells of the jejunum and colon by in situ hybridization. These findings may indicate that water crosses the epithelial layer through these water channels, suggesting a possible role of the transcellular route for water intake or outlet in the gastrointestinal tract.

T he podocyte is a highly differentiated cell that has characteristic interdigitating foot processes that cover the outer surface of the glomerular basement membrane (GBM) in the kidney (1). The turnover rate of podocytes is very low under normal and various pathologic conditions compared with that of other glomerular cells (2,3). Meanwhile, podocytes contribute to the hydraulic permeability of the glomerulus and play a crucial role as a filter for macromolecules (1). Because of these biologic and morphologic characteristics of podocytes, injuries to podocytes are accompanied by characteristic changes in morphology, as observed by electron microscopy (EM), including effacement of foot processes, microvillous transformation, and occasional detachment from the GBM (4 -7). In several immunologic and nonimmunologic forms of glomerulonephritis, the podocyte is the primary target of injury (8,9). Podocyte injury is also a key event leading to glomerular sclerosis. Recent studies have revealed that the denuded GBM left behind after a podocyte becomes detached and subsequently adheres to parietal epithelial cells, resulting in the formation of a synechia of the glomerular tuft to Bowman's capsule, which represents the earliest stage of segmental sclerosis (10,11).We recently demonstrated the presence of podocytes and their cell fragments in the urinary sediment of patients with glomerular diseases, in an immunofluorescence (IF) study using a specific monoclonal antibody against podocalyxin (PCX), a glycoprotein that is prominently expressed on podocytes (12). Quantification of urinary podocytes has clinical significance in its ability to predict acute glomerular lesions (13). In addition to urinary podocytes, urine sediments from nephritic patients contain PCX-positive granular structures (PPGS) in or around the urine casts. We hypothesized that these structures represent urinary podocytes and their cell debris. We subsequently found that PPGS are excreted in the urine in far greater numbers compared with urinary podocytes. However, because we also found PPGS in the urine of patients without any urinary podocytes, we questioned whether these structures truly represent cell debris from detached podocytes. Thus, the purpose of the present study was to trace PPGS to their origin immunohistochemically. Materials and Methods Patients, Urine Samples, and Kidney SpecimensUrine samples voided in the morning were obtained from 50 healthy children and adolescents (25 male and 25 female; mean age, 12.3 yr; range, 3 to 20 yr) and 53 patients with active glomerulonephritis or nephrotic syndrome (29 male and 24 female; mean age, 11.3 yr; range,