Organization of Hyaluronan and Versican in the Extracellular Matrix of Human Fibroblasts Treated With the Viral Mimetic, poly I:C
Stephen P. Evanko 1, Susan Potter-Perigo 1, Pamela Y. Johnson 1 and Thomas N. Wight 1*
1 Hope Heart Program, Benaroya Research Institute at Virginia Mason, Seattle, Washington
* To whom correspondence should be addressed. E-mail: twight{at}benaroyaresearch.org.
Submitted on March 6, 2009
Accepted on 6 June 2009
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Abstract |
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We have examined structural details of hyaluronan- and versican-rich pericellular matrices in human lung fibroblasts, as well as fixation effects after treatment with the viral mimetic, poly I:C. Lateral aggregation of hyaluronan chains was promoted by acid-ethanol-formalin fixation compared to a network appearance with formalin alone. However, hyaluronidase-sensitive cable structures were seen in live cells, suggesting they are not a fixation artifact. With all fixatives, versican and hyaluronan probes bound alternately along strands extending from the plasma membrane. However, a yellow co-localization signal required aggregation/overlap of several hyaluronan/versican strands and was more pronounced after acid-ethanol-formalin fixation. In addition to the main cell surface, hyaluronan and versican were also associated with fine actin-positive membrane protrusions, retraction fibers, and surface blebs. After wounding plus treatment with poly I:C, cells displayed larger hyaluronan coats and cable-like structures, as well as more membrane protrusions. However, treated cells did not migrate and had increased stress fibers compared to control wounded cells. Deposition of hyaluronan into cable-like structures in response to poly I:C was diminished but still apparent following actin filament disruption with cytochalasin D, suggesting the protrusions only partially facilitate cable formation. As seen by scanning electron microscopy, the membrane protrusions may participate in poly I:C-induced binding of monocytes to hyaluronan- and versican-rich matrices. These results suggest that poly I:C-induced hyaluronan- and versican-rich cable structures are not deposited during migration, and cellular protrusions partially contribute to hyaluronan cable formation. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.
Key Words:
hyaluronan, versican, extracellular matrix, pericellular space, monocytes, migration, poly I:C
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