Abstract
Bone resorbing osteoclasts require tight attachment of their plasma membrane to the bone
surface in order to retain the specific microenvironment and thus to be able to dissolve the
bone matrix underneath. Cadherins are transmembrane glycoproteins usually mediating
homophilic calcium-dependent cell-cell adhesion. In the present work we have studied the
effects of the cadherin CAR sequence HAV-containing hexapeptide AHAVSE on osteoclasts. The
primary attachment of osteoclasts to bone surface is not affected by the peptide, suggesting
that it is not mediated by cadherins. Treatment of osteoclast cultures with AHAVSE decreased
the number of resorption pits and the total resorbed area. Furthermore, we show rapid
inactivation of osteoclasts with AHAVSE, which is seen as a decrease in the percentage of
osteoclasts with actin rings. Pan-cadherin antibodies localized cadherin-like molecule in
the sealing zone area of osteoclasts. These results suggest that cadherin-like molecules may
mediate the tight attachment of osteoclasts in the sealing zone area and that the decrease
of resorption in AHAVSE-treated osteoclast cultures is due to prevention of sealing zone
formation.
We studied the polarity of mesenchymal osteoblasts using osteosarcoma cell line
UMR-108 and endosteal osteoblasts in situ in bone tissue cultures.
Immunofluorescence confocal microscopy revealed that the vesicular stomatitis virus
glycoprotein (VSV G) was targeted to the culture medium-facing surface. In endosteal
osteoblasts, VSV G protein was found in the surface facing the bone marrow and circulation.
On the contrary, Influenza virus hemagglutinin (HA) was localized to the bone
substrate-facing surface of the UMR-108 cells. Electron microscopy showed that VSV particles
were budding from the culture medium-facing surface, whereas Influenza viruses budded from
the bone substrate-facing plasma membrane. These findings suggest the bone attaching plasma
membrane of osteoblasts is apical, and the circulation or bone marrow facing plasma membrane
is basolateral in nature.
Gap junctions often mediate communication between different cells and cell types. In the
present work, we demonstrate that rat osteoclasts show connexin-43 staining localizing in
the plasma membrane of the cells in cell-cell contacts and over the basolateral membrane of
osteoclasts. The effects of heptanol and Gap 27, known gap- junctional inhibitors, were
studied using the well-characterized pit formation assay. The inhibitors decreased the
number and activity of osteoclasts, suggesting a defect in the fusion of mononuclear
osteoclast precursors to multinucleated mature osteoclasts. Furthermore, the total resorbed
area and the number of resorption pits also decreased in the cultures. These results suggest
that gap-junctional connexin-43 plays a functional role in osteoclasts, and that the
blocking of gap junctions decreases both the number and the activity of osteoclasts.
| Identifer | oai:union.ndltd.org:oulo.fi/oai:oulu.fi:isbn951-42-5935-1 |
| Date | 26 March 2001 |
| Creators | Ilvesaro, J. (Joanna) |
| Publisher | University of Oulu |
| Source Sets | University of Oulu |
| Language | English |
| Detected Language | English |
| Type | info:eu-repo/semantics/doctoralThesis, info:eu-repo/semantics/publishedVersion |
| Format | application/pdf |
| Rights | info:eu-repo/semantics/openAccess, © University of Oulu, 2001 |
| Relation | info:eu-repo/semantics/altIdentifier/pissn/0355-3221, info:eu-repo/semantics/altIdentifier/eissn/1796-2234 |
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