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Tenascin supports lymphocyte rolling.

Clark RA, Erickson HP, Springer TA - J. Cell Biol. (1997)

Bottom Line: Tenascin has been reported to have both adhesive and anti-adhesive effects in static assays.When compared to rolling of the same cell type on E-selectin, rolling on tenascin was found to be smoother at all shear stresses tested, suggesting that cells formed a larger number of bonds on the tenascin substrate than on the E-selectin substrate.When protein plating densities were adjusted to give similar profiles of cell detachment under increasing shears, the density of tenascin was 8.5-fold greater than that of E-selectin.

View Article: PubMed Central - PubMed

Affiliation: The Center for Blood Research and Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA.

ABSTRACT
Tenascin is a large extracellular matrix molecule expressed at specific sites in the adult, including immune system tissues such as the bone marrow, thymus, spleen, and T cell areas of lymph nodes. Tenascin has been reported to have both adhesive and anti-adhesive effects in static assays. We report here that tenascin supports the tethering and rolling of lymphocytes and lymphoblastic cell lines under flow conditions. Binding was calcium dependent and was not inhibited by treatment of lymphocytes with O-glycoprotease or a panel of glycosidases including neuraminidase and heparitinase but was inhibited by treatment of cells with proteinase K. Binding was to the fibrinogen-like terminal domain of tenascin as determined by antibody blocking studies and binding to recombinant tenascin proteins. When compared to rolling of the same cell type on E-selectin, rolling on tenascin was found to be smoother at all shear stresses tested, suggesting that cells formed a larger number of bonds on the tenascin substrate than on the E-selectin substrate. When protein plating densities were adjusted to give similar profiles of cell detachment under increasing shears, the density of tenascin was 8.5-fold greater than that of E-selectin. Binding to tenascin was not dependent on any molecules previously identified as tenascin receptors and is likely to involve a novel tenascin receptor on lymphocytes. We postulate that the ability of tenascin to support lymphocyte rolling may reflect its ability to support cell migration and that this interaction may be used by lymphocytes migrating through secondary lymphoid organs.

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Binding of SKW3 cells to tenascin is not inhibited by  mAbs to known tenascin ligands or RGD peptide. SKW3 cells  were treated with the indicated mAbs or GRGDSP peptide before flow assays; M168 and GRGDSP were also added to the flow  chamber before the addition of cells. For GRGDSP peptide experiments, cells were maintained in the continued presence of the  peptide. Observed binding was compared to binding of cells  treated with control IgG CBRp150/2E1 (mAb experiments; control binding was 162 cells), normal rabbit serum (rabbit Ab; control binding was 171 cells), or untreated cells (GRGDSP peptide  experiments; control binding was 293 cells) to obtain percent control binding. The mean values of two experiments are shown;  ranges are indicated by bars.
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Figure 8: Binding of SKW3 cells to tenascin is not inhibited by mAbs to known tenascin ligands or RGD peptide. SKW3 cells were treated with the indicated mAbs or GRGDSP peptide before flow assays; M168 and GRGDSP were also added to the flow chamber before the addition of cells. For GRGDSP peptide experiments, cells were maintained in the continued presence of the peptide. Observed binding was compared to binding of cells treated with control IgG CBRp150/2E1 (mAb experiments; control binding was 162 cells), normal rabbit serum (rabbit Ab; control binding was 171 cells), or untreated cells (GRGDSP peptide experiments; control binding was 293 cells) to obtain percent control binding. The mean values of two experiments are shown; ranges are indicated by bars.

Mentions: More than a dozen molecules have been identified as tenascin receptors (Table I). Among them are a number of members of the integrin family. Most notably, binding via α2β1 may involve the fbg region of the tenascin molecule (35, 56). We therefore investigated whether treatment of SKW3 cells with blocking mAbs to these integrins would inhibit tethering and rolling on tenascin. Two different blocking mAbs to the β1 integrin had little or no effect on attachment of SKW3 cells to tenascin (Fig. 8), as did a third β1-blocking mAb, 4B4 (data not shown). Most significantly, the β1 mAb P4C10 obtained from two different sources did not inhibit binding of SKW3 cells in the shear flow assay despite reports that it blocks the binding of the α2β1 integrin to tenascin (56). A panel of blocking antibodies to the α1, α2, α3, α4, α5, α6, and αv integrin subunits also had no effect (Fig. 8); these included two different samples of P1E6, the α2 mAb reported to inhibit α2β1 interaction with tenascin (56). Furthermore, expression of the α2 integrin subunit did not correlate with the ability of cell lines to bind to tenascin. Hut 78 and Raji cell lines do not express the α2 integrin subunit (53) but bind efficiently to tenascin. The blocking αvβ3 mAb LM609 did not inhibit SKW3 cell binding, despite its ability to inhibit tenascin binding via αvβ3 (56), and a polyclonal antibody to the human αv subunit did not inhibit binding despite its ability to block αv-mediated binding of human cell lines to chicken tenascin (46). In addition, the RGD peptide GRGDSP, reported to block α2β1 and αvβ3 binding to tenascin (35, 56), also had no effect (Fig. 8). α2β1 is the only integrin thought to interact with the fbg region of tenascin; other integrins recognize tenascin's FN-III repeats (Table I). These integrins contain either αv or β1 subunits; however, multiple antibodies to these subunits failed to inhibit binding to tenascin in the flow assay.


Tenascin supports lymphocyte rolling.

Clark RA, Erickson HP, Springer TA - J. Cell Biol. (1997)

Binding of SKW3 cells to tenascin is not inhibited by  mAbs to known tenascin ligands or RGD peptide. SKW3 cells  were treated with the indicated mAbs or GRGDSP peptide before flow assays; M168 and GRGDSP were also added to the flow  chamber before the addition of cells. For GRGDSP peptide experiments, cells were maintained in the continued presence of the  peptide. Observed binding was compared to binding of cells  treated with control IgG CBRp150/2E1 (mAb experiments; control binding was 162 cells), normal rabbit serum (rabbit Ab; control binding was 171 cells), or untreated cells (GRGDSP peptide  experiments; control binding was 293 cells) to obtain percent control binding. The mean values of two experiments are shown;  ranges are indicated by bars.
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Related In: Results  -  Collection

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Figure 8: Binding of SKW3 cells to tenascin is not inhibited by mAbs to known tenascin ligands or RGD peptide. SKW3 cells were treated with the indicated mAbs or GRGDSP peptide before flow assays; M168 and GRGDSP were also added to the flow chamber before the addition of cells. For GRGDSP peptide experiments, cells were maintained in the continued presence of the peptide. Observed binding was compared to binding of cells treated with control IgG CBRp150/2E1 (mAb experiments; control binding was 162 cells), normal rabbit serum (rabbit Ab; control binding was 171 cells), or untreated cells (GRGDSP peptide experiments; control binding was 293 cells) to obtain percent control binding. The mean values of two experiments are shown; ranges are indicated by bars.
Mentions: More than a dozen molecules have been identified as tenascin receptors (Table I). Among them are a number of members of the integrin family. Most notably, binding via α2β1 may involve the fbg region of the tenascin molecule (35, 56). We therefore investigated whether treatment of SKW3 cells with blocking mAbs to these integrins would inhibit tethering and rolling on tenascin. Two different blocking mAbs to the β1 integrin had little or no effect on attachment of SKW3 cells to tenascin (Fig. 8), as did a third β1-blocking mAb, 4B4 (data not shown). Most significantly, the β1 mAb P4C10 obtained from two different sources did not inhibit binding of SKW3 cells in the shear flow assay despite reports that it blocks the binding of the α2β1 integrin to tenascin (56). A panel of blocking antibodies to the α1, α2, α3, α4, α5, α6, and αv integrin subunits also had no effect (Fig. 8); these included two different samples of P1E6, the α2 mAb reported to inhibit α2β1 interaction with tenascin (56). Furthermore, expression of the α2 integrin subunit did not correlate with the ability of cell lines to bind to tenascin. Hut 78 and Raji cell lines do not express the α2 integrin subunit (53) but bind efficiently to tenascin. The blocking αvβ3 mAb LM609 did not inhibit SKW3 cell binding, despite its ability to inhibit tenascin binding via αvβ3 (56), and a polyclonal antibody to the human αv subunit did not inhibit binding despite its ability to block αv-mediated binding of human cell lines to chicken tenascin (46). In addition, the RGD peptide GRGDSP, reported to block α2β1 and αvβ3 binding to tenascin (35, 56), also had no effect (Fig. 8). α2β1 is the only integrin thought to interact with the fbg region of tenascin; other integrins recognize tenascin's FN-III repeats (Table I). These integrins contain either αv or β1 subunits; however, multiple antibodies to these subunits failed to inhibit binding to tenascin in the flow assay.

Bottom Line: Tenascin has been reported to have both adhesive and anti-adhesive effects in static assays.When compared to rolling of the same cell type on E-selectin, rolling on tenascin was found to be smoother at all shear stresses tested, suggesting that cells formed a larger number of bonds on the tenascin substrate than on the E-selectin substrate.When protein plating densities were adjusted to give similar profiles of cell detachment under increasing shears, the density of tenascin was 8.5-fold greater than that of E-selectin.

View Article: PubMed Central - PubMed

Affiliation: The Center for Blood Research and Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA.

ABSTRACT
Tenascin is a large extracellular matrix molecule expressed at specific sites in the adult, including immune system tissues such as the bone marrow, thymus, spleen, and T cell areas of lymph nodes. Tenascin has been reported to have both adhesive and anti-adhesive effects in static assays. We report here that tenascin supports the tethering and rolling of lymphocytes and lymphoblastic cell lines under flow conditions. Binding was calcium dependent and was not inhibited by treatment of lymphocytes with O-glycoprotease or a panel of glycosidases including neuraminidase and heparitinase but was inhibited by treatment of cells with proteinase K. Binding was to the fibrinogen-like terminal domain of tenascin as determined by antibody blocking studies and binding to recombinant tenascin proteins. When compared to rolling of the same cell type on E-selectin, rolling on tenascin was found to be smoother at all shear stresses tested, suggesting that cells formed a larger number of bonds on the tenascin substrate than on the E-selectin substrate. When protein plating densities were adjusted to give similar profiles of cell detachment under increasing shears, the density of tenascin was 8.5-fold greater than that of E-selectin. Binding to tenascin was not dependent on any molecules previously identified as tenascin receptors and is likely to involve a novel tenascin receptor on lymphocytes. We postulate that the ability of tenascin to support lymphocyte rolling may reflect its ability to support cell migration and that this interaction may be used by lymphocytes migrating through secondary lymphoid organs.

Show MeSH
Related in: MedlinePlus