-We examined mechanisms of FITC-albumin uptake by alveolar type II epithelial cells using cultured RLE-6TN cells. Alkaline phosphatase activity and the expression of cytokeratin 19 mRNA, which are characteristic features of alveolar type II epithelial cells, were detected in RLE-6TN cells. The uptake of FITCalbumin by the cells was time and temperature dependent and showed the saturation kinetics of high-and low-affinity transport systems. FITC-albumin uptake was inhibited by native albumin, by chemically modified albumin, and by metabolic inhibitors and bafilomycin A 1, an inhibitor of vacuolar H ϩ -ATPase. Confocal laser scanning microscopic analysis after FITCalbumin uptake showed punctate localization of fluorescence in the cells, which was partly localized in lysosomes. FITC-albumin taken up by the cells gradually degraded over time, as shown by fluoroimage analyzer after SDS-PAGE. The uptake of FITC-albumin by RLE-6TN cells was not inhibited by nystatin, indomethacin, or methyl-␤-cyclodextrin (inhibitors of caveolae-mediated endocytosis) but was inhibited by phenylarsine oxide and chlorpromazine (inhibitors of clathrin-mediated endocytosis) in a concentration-dependent manner. Uptake was also inhibited by potassium depletion and hypertonicity, conditions known to inhibit clathrin-mediated endocytosis. These results indicate that the uptake of FITC-albumin in cultured alveolar type II epithelial cells, RLE-6TN, is mediated by clathrin-mediated but not by caveolae-mediated endocytosis, and intracellular FITC-albumin is gradually degraded in lysosomes. Possible receptors involved in this endocytic system are discussed. albumin clearance; caveolae-mediated endocytosis; endocytic receptor; modified albumin; protein transport THE DISTAL AIR SPACES OF THE lung (alveolar region) are lined with a continuous epithelium comprising two major types of epithelial cells, type I and type II. Type I epithelial cells have a squamous morphology and cover ϳ90 -95% of the alveolar surface area. Type II cells are cuboidal epithelial cells and cover ϳ5-10% of the surface area, although the number of type II cells in alveolar epithelia is similar to or more than that of type I cells. Type II cells also serve as progenitors of type I cells (13,40). Alveolar lining fluid contains various physiologically important proteins such as albumin, immunoglobulin G, and transferrin. The concentration of albumin in alveolar fluid is usually much lower than that in the blood, whereas the concentration would increase to 40 -65% of plasma level in hydrostatic pulmonary edema and to 75-95% in lung injury pulmonary edema. The clearance of serum proteins from the alveolar space is a critical process in recovery from pulmonary edema and in maintaining the normal alveolar milieu (17,22). Therefore, understanding the mechanisms of protein transport in alveolar epithelial cells is important for the development of better therapeutic approaches. In addition, such information could provide new insights into improved strategies for pulmonary delivery of ex...