A steady-state labelling approach to the measurement of proteoglycan turnover in vivo and its application to glomerular proteoglycans. 1996

E L Akuffo, and J R Hunt, and J Moss, and D Woodrow, and M Davies, and R M Mason
Department of Biochemistry, Charing Cross and Westminster Medical School, London, UK.

Rats were implanted with mini osmotic pumps delivering sodium [35S]sulphate and their newly synthesized proteoglycans were labelled over a 146 h period (steady-state labelling). Proteoglycan turnover was measured in vivo using a chase protocol. Glomerular proteoglycans were recovered quantitatively and the perlecan present was isolated by immunoprecipitation. The procedure allows newly synthesized proteoglycans to be quantified in mass units (pmol of glycossminoglycan sulphate) after labelling and during the chase. Ultrastructural-immunogold experiments identified the location of perlecan as the glomerular basement membrane and mesangial matrix. Perlecan in the basement membrane was quantified using the ultrastructural-immunogold technique. Perlecan comprises about 10% of the total glomerular proteoglycans, which are otherwise associated with glomerular cells and the mesangium. Both the total glomerular heparan sulphate proteoglycans and perlecan turn over rapidly (t1/2 approximately 3-4 h and < 3 h respectively). In contrast, turnover of proteoglycans in other tissues was slow, except in the liver where the heparan sulphate and chondroitin sulphate t1/2 values were 16 h and 9 h respectively. Microalbuminuria was induced with a low-dose regimen of puromycin aminonucleoside. At the onset of microalbuminuria (5 days) there was no change in the level of newly synthesized perlecan, or in perlecan in the glomerular basement membrane detected by immunogold labelling. Newly synthesized perlecan had undergone a minimal change in turnover rate by day 5 in puromycin aminonucleoside-treated rats. In contrast, the total glomerular proteoglycan population showed a dramatic decrease in turnover by day 5. Since there was no evidence of accumulation of glomerular proteoglycans on either day 5 or day 6, it is likely that decreased turnover of cell-associated proteoglycans is accompanied by an equivalent decrease in their synthesis.

UI MeSH Term Description Entries
D007678 Kidney Glomerulus A cluster of convoluted capillaries beginning at each nephric tubule in the kidney and held together by connective tissue. Glomerulus, Kidney
D008854 Microscopy, Electron Microscopy using an electron beam, instead of light, to visualize the sample, thereby allowing much greater magnification. The interactions of ELECTRONS with specimens are used to provide information about the fine structure of that specimen. In TRANSMISSION ELECTRON MICROSCOPY the reactions of the electrons that are transmitted through the specimen are imaged. In SCANNING ELECTRON MICROSCOPY an electron beam falls at a non-normal angle on the specimen and the image is derived from the reactions occurring above the plane of the specimen. Electron Microscopy
D011509 Proteoglycans Glycoproteins which have a very high polysaccharide content. Proteoglycan,Proteoglycan Type H
D005260 Female Females
D000419 Albuminuria The presence of albumin in the urine, an indicator of KIDNEY DISEASES. Albuminurias
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D013462 Sulfur Radioisotopes Unstable isotopes of sulfur that decay or disintegrate spontaneously emitting radiation. S 29-31, 35, 37, and 38 are radioactive sulfur isotopes. Radioisotopes, Sulfur
D017208 Rats, Wistar A strain of albino rat developed at the Wistar Institute that has spread widely at other institutions. This has markedly diluted the original strain. Wistar Rat,Rat, Wistar,Wistar Rats
D051381 Rats The common name for the genus Rattus. Rattus,Rats, Laboratory,Rats, Norway,Rattus norvegicus,Laboratory Rat,Laboratory Rats,Norway Rat,Norway Rats,Rat,Rat, Laboratory,Rat, Norway,norvegicus, Rattus

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