Organization of hyaluronan and versican in the extracellular matrix of human fibroblasts treated with the viral mimetic poly I:C.
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 with a network appearance with formalin alone. However, hyaluronidase-sensitive cable structures were seen in live cells, suggesting that 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 colocalization 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 with 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 that 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 that 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.
Evanko
Stephen P
SP
Hope Heart Program, Benaroya Research Institute at Virginia Mason, Seattle, WA 98101, USA.
Potter-Perigo
Susan
S
Johnson
Pamela Y
PY
Wight
Thomas N
TN
D000199:Actins / Q000378:metabolism; D001692:Biological Transport / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D022081:Cell Surface Extensions / Q000187:drug effects / Q000378:metabolism; D005109:Extracellular Matrix / Q000187:drug effects* / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D006820:Hyaluronic Acid / Q000378:metabolism*; D008855:Microscopy, Electron, Scanning; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism; D011070:Poly I-C / Q000494:pharmacology*; D022502:Stress Fibers / Q000187:drug effects / Q000378:metabolism; D053675:Versicans / Q000378:metabolism*; D014780:Viruses
D000199:Actins; D053675:Versicans; D006820:Hyaluronic Acid; D011070:Poly I-C