Role of proteoglycans and cytoskeleton in the effects of TGF-beta 1 on renal proximal tubule cells

Transforming growth factor-beta (TGF-beta) is a critical cell regulatory protein which influences cell growth, cell differentiation and cell chemotaxis. TGF-beta 1 has been previously shown to promote a migratory and adherent transformation of monolayers of renal proximal tubule cells in primary cul...

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Bibliographic Details
Published in:Kidney international 1993-03, Vol.43 (3), p.575-584
Main Authors: Humes, H D, Nakamura, T, Cieslinski, D A, Miller, D, Emmons, R V, Border, W A
Format: Article
Language:English
Subjects:
Actins - metabolism
Animals
Cells, Cultured
Cytochalasin B - pharmacology
Cytoskeleton - metabolism
DNA - biosynthesis
Fibronectins - biosynthesis
Glycosides - pharmacology
Kidney Tubules, Proximal - drug effects
Kidney Tubules, Proximal - metabolism
Kidney Tubules, Proximal - ultrastructure
Phenotype
Proteoglycans - biosynthesis
Rabbits
Transforming Growth Factor beta - pharmacology
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Summary:Transforming growth factor-beta (TGF-beta) is a critical cell regulatory protein which influences cell growth, cell differentiation and cell chemotaxis. TGF-beta 1 has been previously shown to promote a migratory and adherent transformation of monolayers of renal proximal tubule cells in primary culture to form solid clusters of cells. To better understand the cellular basis of this TGF-beta 1 effect, these studies evaluated the influence of TGF-beta 1 on the synthesis of proteoglycans and on cytoskeleton rearrangement in rabbit renal proximal tubule cells in primary culture, and their role in this transformation effect of TGF-beta 1. Biosynthetic labeling of proteoglycans with 35S sulfate and enzyme digestion studies demonstrated that TGF-beta 1 promoted the synthesis of heparan sulfate proteoglycans in these cells. The importance of proteoglycan synthesis induced by TGF-beta 1 in this migration and aggregation process was demonstrated with the use of two chemically-dissimilar proteoglycan synthesis inhibitors: xyloside and galactosamine. Both compounds inhibited TGF-beta 1 stimulation of proteoglycan synthesis and diminished TGF-beta 1 promoted transformation of proximal tubule cells as assessed by quantitative morphometry. Further experiments evaluated the influence of TGF-beta 1 on actin microfilaments with the use of rhodamine conjugated phalloidin staining and immunofluorescent microscopy, and demonstrated that TGF-beta 1 provoked a dramatic rearrangement of actin microfilaments into stress fibers. The use of actin microfilament disrupting agents, cytochalasin B and D, attenuated the stress fiber formation promoted by TGF-beta 1 and inhibited the TGF-beta 1-induced morphologic transformation of these cells. Further studies evaluated these effects on the rate of DNA synthesis in these cells, as assessed with 3H-thymidine incorporation. Proteoglycan synthesis inhibitors significantly diminished the maximal proliferative response of these epithelial cells to epidermal growth factor (EGF). In contrast, actin microfilament disaggregation with cytochalasin B or D did not change the rate of DNA synthesis in response to EGF but did attenuate the antiproliferative effect of TGF-beta 1 on EGF-induced DNA synthesis cells. These studies demonstrate that the TGF-beta 1 promoted synthesis cells. These studies demonstrate that the TGF-beta 1 promoted an increase in the production of proteoglycans and a higher ordered structure of the cytoskeleton. Both effects were
ISSN:0085-2538
1523-1755
DOI:10.1038/ki.1993.85