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Interleukin 15 induces endothelial hyaluronan expression in vitro and promotes activated T cell extravasation through a CD44-dependent pathway in vivo.

Estess P, Nandi A, Mohamadzadeh M, Siegelman MH - J. Exp. Med. (1999)

Bottom Line: Here, we examine the effect of IL-15 on hyaluronan expression by endothelial cells, and investigate its role in vivo in promoting the extravasation of antigen-activated T cells through a CD44-dependent pathway.The results suggest that IL-15 can regulate endothelial cell function and thereby enables a CD44-initiated adhesion pathway that facilitates entry of activated T lymphocytes into inflammatory sites.They further demonstrate a novel role for IL-15 (distinct from any of IL-2) in regulating microvascular endothelial cell adhesive function help to understand the role of IL-15R expression on endothelium, and further support a central position for this cytokine in orchestrating multiple sequential aspects of T cell effector function and therefore chronic inflammatory processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, University of Texas, Southwestern Medical Center, Dallas 75235-9072, USA.

ABSTRACT
T cell recruitment to extralymphoid tissues is fundamental to the initiation and perpetuation of the inflammatory state during immune and autoimmune responses. Interleukin (IL)-15 is a proinflammatory cytokine whose described functions largely overlap with those of IL-2. The latter is attributable in large part to its binding of the heterotrimeric receptor that contains the beta and gamma chains of the IL-2R in combination with an unique IL-15R alpha chain. However, unlike IL-2, IL-15 and its receptor have a wide tissue and cell type distribution, including endothelial cells. Here, we examine the effect of IL-15 on hyaluronan expression by endothelial cells, and investigate its role in vivo in promoting the extravasation of antigen-activated T cells through a CD44-dependent pathway. The expression of hyaluronan on primary endothelial cells and microvascular endothelial cell lines is induced by IL-15, whereas IL-2 has no such activity. Moreover, intraperitoneal administration of IL-15 or TNF-alpha in the absence of other exogenous proinflammatory stimuli allows the extravasation of superantigen-stimulated T cells into this site in vivo in a CD44-dependent manner. T cell recruitment induced by IL-15 requires expression of an intact IL-2R beta chain, indicating that IL-15 operates in this context through the traditional IL-15R. The results suggest that IL-15 can regulate endothelial cell function and thereby enables a CD44-initiated adhesion pathway that facilitates entry of activated T lymphocytes into inflammatory sites. They further demonstrate a novel role for IL-15 (distinct from any of IL-2) in regulating microvascular endothelial cell adhesive function help to understand the role of IL-15R expression on endothelium, and further support a central position for this cytokine in orchestrating multiple sequential aspects of T cell effector function and therefore chronic inflammatory processes.

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Induction of bPG staining on ECs in response to cytokine treatment. (A) Levels of cell surface HA increase following treatment of SVEC4-10 cells with TNF-α or IL-15. Cells were incubated for 4 h with TNF-α (10 ng/ml), IL-15 (50 ng/ml), IL-2 (50 ng/ml), or IFN-γ (10 ng/ml), harvested in EDTA, stained with bPG-biotin plus SA-PE, and analyzed by FACS® for bPG binding. Untreated cells, which constitutively express some HA, bind background levels of bPG (solid line). After treatment with TNF-α or IL-15, cells show even greater levels of staining (stippled line), whereas IL-2 has no effect. As previously shown, binding to bPG was not increased by incubation with IFN-γ 4. Baseline staining of SVEC4-10 cells with SA-PE alone is indicated by the dashed line in the TNF-α panel. Preincubating TNF-α–treated cells with soluble HA reduced bPG staining to near background levels (heavy line). (B) Expression of ICAM-1, VCAM, P-selectin, and E-selectin by SVEC4-10 cells before (stippled line) and after (solid line) IL-15 treatment. (C) TME-3H3, 1°LNEC, LEII, and HUVEC cells were treated with human IL-15 (top) or IL-2 (bottom), as in A. TME-3H3 and 1°LNEC respond to IL-15 treatment by expressing increased levels of HA, whereas LEII and HUVEC cells are not affected. In contrast to IL-15, IL-2 had no effect on any of the EC lines or primary ECs. Solid lines, untreated cells; stippled lines, treated cells. (D) Dose dependence of increased HA levels in response to IL-15 treatment. SVEC4-10 cells were incubated for 4 h with IL-15 or IL-2 at varying concentrations, as shown. Results are reported as the mean fluorescence intensity (MFI) of cells after staining with bPG-biotin plus SA-PE, as determined by FACS® analysis. IL-2 had no effect on HA expression by SVEC4-10 cells, even at 200 ng/ml, whereas IL-15 had a dose-dependent effect on levels of surface HA, with changes seen at 10 ng/ml. Data shown are the mean ± SD from three separate experiments.
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Figure 1: Induction of bPG staining on ECs in response to cytokine treatment. (A) Levels of cell surface HA increase following treatment of SVEC4-10 cells with TNF-α or IL-15. Cells were incubated for 4 h with TNF-α (10 ng/ml), IL-15 (50 ng/ml), IL-2 (50 ng/ml), or IFN-γ (10 ng/ml), harvested in EDTA, stained with bPG-biotin plus SA-PE, and analyzed by FACS® for bPG binding. Untreated cells, which constitutively express some HA, bind background levels of bPG (solid line). After treatment with TNF-α or IL-15, cells show even greater levels of staining (stippled line), whereas IL-2 has no effect. As previously shown, binding to bPG was not increased by incubation with IFN-γ 4. Baseline staining of SVEC4-10 cells with SA-PE alone is indicated by the dashed line in the TNF-α panel. Preincubating TNF-α–treated cells with soluble HA reduced bPG staining to near background levels (heavy line). (B) Expression of ICAM-1, VCAM, P-selectin, and E-selectin by SVEC4-10 cells before (stippled line) and after (solid line) IL-15 treatment. (C) TME-3H3, 1°LNEC, LEII, and HUVEC cells were treated with human IL-15 (top) or IL-2 (bottom), as in A. TME-3H3 and 1°LNEC respond to IL-15 treatment by expressing increased levels of HA, whereas LEII and HUVEC cells are not affected. In contrast to IL-15, IL-2 had no effect on any of the EC lines or primary ECs. Solid lines, untreated cells; stippled lines, treated cells. (D) Dose dependence of increased HA levels in response to IL-15 treatment. SVEC4-10 cells were incubated for 4 h with IL-15 or IL-2 at varying concentrations, as shown. Results are reported as the mean fluorescence intensity (MFI) of cells after staining with bPG-biotin plus SA-PE, as determined by FACS® analysis. IL-2 had no effect on HA expression by SVEC4-10 cells, even at 200 ng/ml, whereas IL-15 had a dose-dependent effect on levels of surface HA, with changes seen at 10 ng/ml. Data shown are the mean ± SD from three separate experiments.

Mentions: IL-15 has not been reported previously to induce increased levels of adhesion molecule expression. To examine the effect of IL-15 on the level of surface HA expression, the LN-derived EC line SVEC4-10 was incubated for 4 h in the presence of IL-15, TNF-α, IL-2, or IFN-γ. Using a biotinylated form of the HA-binding proteoglycan bPG to detect cell surface HA, we found that IL-15 treatment resulted in a marked increase in the amount of surface HA expression, which was comparable to levels seen with TNF-α treatment (reference 4 and Fig. 1 A). Moreover, HA surface expression appeared selectively induced, as other endothelial adhesion molecules showed no substantial alterations in levels of expression after IL-15 treatment (Fig. 1 B), although VCAM-1 and P-selectin were both increased on this cell line after LPS treatment (data not shown). Kinetic studies further demonstrated that surface HA expression was maximal at 4 h, consistent with the time course previously demonstrated with other proinflammatory cytokines 4. In contrast to IL-15, IL-2 had no effect on the level of bPG binding on these ECs, although these two cytokines have overlapping functions in many biological systems 15. IFN-γ (Fig. 1 A) and IL-12 4 had no effect on HA expression, as previously reported. Also, bPG staining of TNF-α–treated SVEC4-10 cells was blocked by preincubating cells with soluble HA before staining with bPG 1, indicating specificity of this reagent 17. Thus, the proinflammatory cytokine IL-15 acts as a potent and selective regulator of surface HA expression.


Interleukin 15 induces endothelial hyaluronan expression in vitro and promotes activated T cell extravasation through a CD44-dependent pathway in vivo.

Estess P, Nandi A, Mohamadzadeh M, Siegelman MH - J. Exp. Med. (1999)

Induction of bPG staining on ECs in response to cytokine treatment. (A) Levels of cell surface HA increase following treatment of SVEC4-10 cells with TNF-α or IL-15. Cells were incubated for 4 h with TNF-α (10 ng/ml), IL-15 (50 ng/ml), IL-2 (50 ng/ml), or IFN-γ (10 ng/ml), harvested in EDTA, stained with bPG-biotin plus SA-PE, and analyzed by FACS® for bPG binding. Untreated cells, which constitutively express some HA, bind background levels of bPG (solid line). After treatment with TNF-α or IL-15, cells show even greater levels of staining (stippled line), whereas IL-2 has no effect. As previously shown, binding to bPG was not increased by incubation with IFN-γ 4. Baseline staining of SVEC4-10 cells with SA-PE alone is indicated by the dashed line in the TNF-α panel. Preincubating TNF-α–treated cells with soluble HA reduced bPG staining to near background levels (heavy line). (B) Expression of ICAM-1, VCAM, P-selectin, and E-selectin by SVEC4-10 cells before (stippled line) and after (solid line) IL-15 treatment. (C) TME-3H3, 1°LNEC, LEII, and HUVEC cells were treated with human IL-15 (top) or IL-2 (bottom), as in A. TME-3H3 and 1°LNEC respond to IL-15 treatment by expressing increased levels of HA, whereas LEII and HUVEC cells are not affected. In contrast to IL-15, IL-2 had no effect on any of the EC lines or primary ECs. Solid lines, untreated cells; stippled lines, treated cells. (D) Dose dependence of increased HA levels in response to IL-15 treatment. SVEC4-10 cells were incubated for 4 h with IL-15 or IL-2 at varying concentrations, as shown. Results are reported as the mean fluorescence intensity (MFI) of cells after staining with bPG-biotin plus SA-PE, as determined by FACS® analysis. IL-2 had no effect on HA expression by SVEC4-10 cells, even at 200 ng/ml, whereas IL-15 had a dose-dependent effect on levels of surface HA, with changes seen at 10 ng/ml. Data shown are the mean ± SD from three separate experiments.
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Related In: Results  -  Collection

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Figure 1: Induction of bPG staining on ECs in response to cytokine treatment. (A) Levels of cell surface HA increase following treatment of SVEC4-10 cells with TNF-α or IL-15. Cells were incubated for 4 h with TNF-α (10 ng/ml), IL-15 (50 ng/ml), IL-2 (50 ng/ml), or IFN-γ (10 ng/ml), harvested in EDTA, stained with bPG-biotin plus SA-PE, and analyzed by FACS® for bPG binding. Untreated cells, which constitutively express some HA, bind background levels of bPG (solid line). After treatment with TNF-α or IL-15, cells show even greater levels of staining (stippled line), whereas IL-2 has no effect. As previously shown, binding to bPG was not increased by incubation with IFN-γ 4. Baseline staining of SVEC4-10 cells with SA-PE alone is indicated by the dashed line in the TNF-α panel. Preincubating TNF-α–treated cells with soluble HA reduced bPG staining to near background levels (heavy line). (B) Expression of ICAM-1, VCAM, P-selectin, and E-selectin by SVEC4-10 cells before (stippled line) and after (solid line) IL-15 treatment. (C) TME-3H3, 1°LNEC, LEII, and HUVEC cells were treated with human IL-15 (top) or IL-2 (bottom), as in A. TME-3H3 and 1°LNEC respond to IL-15 treatment by expressing increased levels of HA, whereas LEII and HUVEC cells are not affected. In contrast to IL-15, IL-2 had no effect on any of the EC lines or primary ECs. Solid lines, untreated cells; stippled lines, treated cells. (D) Dose dependence of increased HA levels in response to IL-15 treatment. SVEC4-10 cells were incubated for 4 h with IL-15 or IL-2 at varying concentrations, as shown. Results are reported as the mean fluorescence intensity (MFI) of cells after staining with bPG-biotin plus SA-PE, as determined by FACS® analysis. IL-2 had no effect on HA expression by SVEC4-10 cells, even at 200 ng/ml, whereas IL-15 had a dose-dependent effect on levels of surface HA, with changes seen at 10 ng/ml. Data shown are the mean ± SD from three separate experiments.
Mentions: IL-15 has not been reported previously to induce increased levels of adhesion molecule expression. To examine the effect of IL-15 on the level of surface HA expression, the LN-derived EC line SVEC4-10 was incubated for 4 h in the presence of IL-15, TNF-α, IL-2, or IFN-γ. Using a biotinylated form of the HA-binding proteoglycan bPG to detect cell surface HA, we found that IL-15 treatment resulted in a marked increase in the amount of surface HA expression, which was comparable to levels seen with TNF-α treatment (reference 4 and Fig. 1 A). Moreover, HA surface expression appeared selectively induced, as other endothelial adhesion molecules showed no substantial alterations in levels of expression after IL-15 treatment (Fig. 1 B), although VCAM-1 and P-selectin were both increased on this cell line after LPS treatment (data not shown). Kinetic studies further demonstrated that surface HA expression was maximal at 4 h, consistent with the time course previously demonstrated with other proinflammatory cytokines 4. In contrast to IL-15, IL-2 had no effect on the level of bPG binding on these ECs, although these two cytokines have overlapping functions in many biological systems 15. IFN-γ (Fig. 1 A) and IL-12 4 had no effect on HA expression, as previously reported. Also, bPG staining of TNF-α–treated SVEC4-10 cells was blocked by preincubating cells with soluble HA before staining with bPG 1, indicating specificity of this reagent 17. Thus, the proinflammatory cytokine IL-15 acts as a potent and selective regulator of surface HA expression.

Bottom Line: Here, we examine the effect of IL-15 on hyaluronan expression by endothelial cells, and investigate its role in vivo in promoting the extravasation of antigen-activated T cells through a CD44-dependent pathway.The results suggest that IL-15 can regulate endothelial cell function and thereby enables a CD44-initiated adhesion pathway that facilitates entry of activated T lymphocytes into inflammatory sites.They further demonstrate a novel role for IL-15 (distinct from any of IL-2) in regulating microvascular endothelial cell adhesive function help to understand the role of IL-15R expression on endothelium, and further support a central position for this cytokine in orchestrating multiple sequential aspects of T cell effector function and therefore chronic inflammatory processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, University of Texas, Southwestern Medical Center, Dallas 75235-9072, USA.

ABSTRACT
T cell recruitment to extralymphoid tissues is fundamental to the initiation and perpetuation of the inflammatory state during immune and autoimmune responses. Interleukin (IL)-15 is a proinflammatory cytokine whose described functions largely overlap with those of IL-2. The latter is attributable in large part to its binding of the heterotrimeric receptor that contains the beta and gamma chains of the IL-2R in combination with an unique IL-15R alpha chain. However, unlike IL-2, IL-15 and its receptor have a wide tissue and cell type distribution, including endothelial cells. Here, we examine the effect of IL-15 on hyaluronan expression by endothelial cells, and investigate its role in vivo in promoting the extravasation of antigen-activated T cells through a CD44-dependent pathway. The expression of hyaluronan on primary endothelial cells and microvascular endothelial cell lines is induced by IL-15, whereas IL-2 has no such activity. Moreover, intraperitoneal administration of IL-15 or TNF-alpha in the absence of other exogenous proinflammatory stimuli allows the extravasation of superantigen-stimulated T cells into this site in vivo in a CD44-dependent manner. T cell recruitment induced by IL-15 requires expression of an intact IL-2R beta chain, indicating that IL-15 operates in this context through the traditional IL-15R. The results suggest that IL-15 can regulate endothelial cell function and thereby enables a CD44-initiated adhesion pathway that facilitates entry of activated T lymphocytes into inflammatory sites. They further demonstrate a novel role for IL-15 (distinct from any of IL-2) in regulating microvascular endothelial cell adhesive function help to understand the role of IL-15R expression on endothelium, and further support a central position for this cytokine in orchestrating multiple sequential aspects of T cell effector function and therefore chronic inflammatory processes.

Show MeSH
Related in: MedlinePlus