Summary
We have employed the fluorescent dye nile red to distinguish between normal cells and cells containing lysosomal accumulations of phospholipids. When fibroblasts from an individual with a genetic deficiency in lysosomal sphingomyelinase activity (Niemann-Pick disease) were stained with nile red and visualized by fluorescence microscopy, orange-colored inclusions were observed throughout the cytoplasm. The orange fluorescent bodies could be distinguished from the neutral lipid droplets that fluoresce a brilliant yellow-gold in the presence of nile red. These inclusions were also observed in alveolar macrophages obtained from rats treated with amiodarone, an antiarrhythmic agent known to produce lysosomal phospholipidosis. Flow cytofluorometric analysis revealed that staining of these phospholipid-rich macrophages with nile red can distinguish them from control alveolar macrophages. These results demonstrate that nile red can be employed for the rapid staining of cellular phospholipid inclusions.
Similar content being viewed by others
References
Bligh EG, Dyer WJ (1959) A rapid method of total lipid extraction and purification. Can J Biochem Physiol 37:911–917
Brady RO (1983) Sphingomyelin lipidoses: Niemann-Pick disease. In: Stanbury JB, Wyngaarden JB, Frederickson DS, Goldstein JL, Brown MS (eds) The metabolic basis of inherited disease. McGraw-Hill, New York, pp 831–841
Brown WJ, Warfel J, Greenspan P (1988) Use of nile red in the detection of cholesteryl ester accumulation in acid lipase-deficient fibroblasts. Arch Pathol Lab Med 112:295–297
Greenspan P, Fowler SD (1985) Spectrofluorometric studies of the lipid probe, nile red. J Lipid Res 26:781–789
Greenspan P, Mayer EP, Fowler SD (1985) Nile red: a selective fluorescent stain for intracellular lipid droplets. J Cell Biol 100:965–973
Fowler SD, Mayer EP, Greenspan P (1985) Foam cells and atherogenesis. Ann NY Acad Sci 454:79–90
Fowler SD, Brown WJ, Warfel J, Greenspan P (1987) Use of nile red for the rapid in situ quantitation of lipids on thin-layer chromatograms. J Lipid Res 28:1225–1232
Heath MF, Costa-Jussa FR, Jacobs JM, Jacobson W (1985) The induction of pulmonary phospholipidosis and the inhibition of lysosomal phospholipases by amiodarone. Br J Exp Pathol 66:391–397
Hostetler KY, Reasor MJ, Walker ER, Yazaki PJ, Frazee BW (1986) Role of phospholipase A inhibition in amiodarone pulmonary toxicity in rats. Biochim Biophys Acta 875:400–405
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with Folin phenol reagent. J Biol Chem 193:265–275
Marchlinski FE, Gansler TS, Waxman HL, Josephson ME (1982) Amiodarone pulmonary toxicity. Ann Intern Med 97:839–845
Mason RJ, Williams MC, Greenleaf RD, Clements JA (1977) Isolation and properties of type II alveolar cells from rat lung. Am Rev Respir Dis 115:1015–1026
Mazière JC, Mazière C, Gardette J, Mora L, Polonovski J (1981) Changes in cholesterol metabolism in cultured fibroblasts from patients with Niemann-Pick disease. Biochem Biophys Res Commun 102:113–118
Miles PR, Ma JYC, Bowman L (1988) Degradation of pulmonary surfactant disaturated phosphatidylcholines by alveolar macrophages. J Appl Physiol 64:2474–2481
Nichols BA (1976) Normal rabbit alveolar macrophages. I. The phagocytosis of tubular myelin. J Exp Med 144:906–919
Reasor MJ, Ogle CL, Walker ER, Kacew S (1988) Amiodarone-induced phospholipidosis in rat alveolar macrophages. Am Rev Respir Dis 137:510–518
Takahashi K, Naito M (1985) Lipid storage disease: part II. Ultrastructural pathology of lipid storage cells in sphingolipidoses. Acta Pathol Jpn 35:385–408
Takahashi K, Naito M, Suziki Y (1987) Lipid storage disease: part III. Ultrastructural evaluation of cultured fibroblasts in sphingolipidoses. Acta Pathol Jpn 37:261–272
Weller PF, Ackerman SJ, Nicholson-Weller A, Dvorak AM (1989) Cytoplasmic lipid bodies of human neutrophilic leukocytes. Am J Pathol 135:947–959
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Brown, W.J., Sullivan, T.R. & Greenspan, P. Nile red staining of lysosomal phospholipid inclusions. Histochemistry 97, 349–354 (1992). https://doi.org/10.1007/BF00270037
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00270037