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Skeletal muscle interleukin 15 promotes CD8(+) T-cell function and autoimmune myositis.

Huang PL, Hou MS, Wang SW, Chang CL, Liou YH, Liao NS - Skelet Muscle (2015)

Bottom Line: Consistently, deficiency of endogenous IL-15 affected neither skeletal muscle growth nor its responses to TNF-α and IFN-γ.On the other hand, the cytokine-stimulated skeletal muscle cells presented antigen and provided IL-15 to promote the effector function of memory-like CD8(+) T cells.These findings together indicate that skeletal muscle IL-15 directly regulates immune effector cells but not muscle cells and thus presents a potential therapeutic target for myositis.

View Article: PubMed Central - PubMed

Affiliation: Molecular Cell Biology, Taiwan International Graduate Program, Institute of Molecular Biology, Academia Sinica, and Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan ; Institute of Molecular Biology, Academia Sinica, Taipei, 11529 Taiwan.

ABSTRACT

Background: Interleukin 15 (IL-15) is thought to be abundant in the skeletal muscle under steady state conditions based on RNA expression; however, the IL-15 RNA level may not reflect the protein level due to post-transcriptional regulation. Although exogenous protein treatment and overexpression studies indicated IL-15 functions in the skeletal muscle, how the skeletal muscle cell uses IL-15 remains unclear. In myositis patients, IL-15 protein is up-regulated in the skeletal muscle. Given the supporting role of IL-15 in CD8(+) T-cell survival and activation and the pathogenic role of cytotoxic CD8(+) T cells in polymyositis and inclusion-body myositis, we hypothesize that IL-15 produced by the inflamed skeletal muscle promotes myositis via CD8(+) T cells.

Methods: Expression of IL-15 and IL-15 receptors at the protein level by skeletal muscle cells were examined under steady state and cytokine stimulation conditions. The functions of IL-15 in the skeletal muscle were investigated using Il15 knockout (Il15 (-/-) ) mice. The immune regulatory role of skeletal muscle IL-15 was determined by co-culturing cytokine-stimulated muscle cells and memory-like CD8(+) T cells in vitro and by inducing autoimmune myositis in skeletal-muscle-specific Il15 (-/-) mice.

Results: We found that the IL-15 protein was not expressed by skeletal muscle cells under steady state condition but induced by tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ) stimulation and expressed as IL-15/IL-15 receptor alpha (IL-15Rα) complex. Skeletal muscle cells expressed a scanty amount of IL-15 receptor beta (IL-15Rβ) under either conditions and only responded to a high concentration of IL-15 hyperagonist, but not IL-15. Consistently, deficiency of endogenous IL-15 affected neither skeletal muscle growth nor its responses to TNF-α and IFN-γ. On the other hand, the cytokine-stimulated skeletal muscle cells presented antigen and provided IL-15 to promote the effector function of memory-like CD8(+) T cells. Genetic ablation of Il15 in skeletal muscle cells greatly ameliorated autoimmune myositis in mice.

Conclusions: These findings together indicate that skeletal muscle IL-15 directly regulates immune effector cells but not muscle cells and thus presents a potential therapeutic target for myositis.

No MeSH data available.


Related in: MedlinePlus

Skeletal muscle IL-15 contributes to the progression of autoimmune myositis. a The expression of Il15 mRNA in various tissues of wt (n = 3), Il15f/f (n = 5), and ACTA-Il15−/− (n = 5) mice was detected by qPCR. WAT white adipose tissue. Data are mean ± SEM. ***p < 0.001. b The amount of IL-15/IL-15Rα complex protein in the serum of Il15f/f and ACTA-Il15−/− mice (n = 9–10) was measured by ELISA. Data are mean ± SEM. c Comparison of memory CD8+ T cell (mCD8) and NK cell (NK) level in the peripheral blood among wt, Il15−/−, Il15f/f, and ACTA-Il15−/− mice (n = 5 in each group). Fold change was calculated by normalizing the percentage of indicated cell type in mutant mice to that in wt mice. mCD8 were H57+CD19−CD8+CD44hiCD122hi, and NK were H57−CD19−NK1.1+. Data are mean ± SEM. ***p < 0.001. d Mononuclear cell infiltration in the quadriceps muscles of Syt7−/−Il15f/f and Syt7−/−ACTA-Il15−/− mice 14 days after C protein immunization (Left). Mononuclear cell infiltration was found in the endomysium (black square), perimysium (black arrowhead), and perivascular region (white arrowhead). Focal lymphatic invasion of myofibers were observed in C-protein-immunized Syt7−/−ACTA-Il15−/− mice as indicated by the white arrowhead (Middle). The histopathology scores were compiled in the right panel with each symbol representing one mouse (Right). Scale bar in left = 100 μm; middle = 25 μm. **p < 0.01. e Expression of mononuclear cell markers, Cd4, Cd8, and F4/80, mRNA in the quadriceps muscles of C-protein-immunized mice measured by qPCR. Each symbol representing one mouse. *p < 0.05, **p < 0.01. f Expression of immune-relevant genes mRNA in the quadriceps muscles of C-protein-immunized mice measured by qPCR. Each symbol representing one mouse. *p < 0.05, **p < 0.01
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Fig6: Skeletal muscle IL-15 contributes to the progression of autoimmune myositis. a The expression of Il15 mRNA in various tissues of wt (n = 3), Il15f/f (n = 5), and ACTA-Il15−/− (n = 5) mice was detected by qPCR. WAT white adipose tissue. Data are mean ± SEM. ***p < 0.001. b The amount of IL-15/IL-15Rα complex protein in the serum of Il15f/f and ACTA-Il15−/− mice (n = 9–10) was measured by ELISA. Data are mean ± SEM. c Comparison of memory CD8+ T cell (mCD8) and NK cell (NK) level in the peripheral blood among wt, Il15−/−, Il15f/f, and ACTA-Il15−/− mice (n = 5 in each group). Fold change was calculated by normalizing the percentage of indicated cell type in mutant mice to that in wt mice. mCD8 were H57+CD19−CD8+CD44hiCD122hi, and NK were H57−CD19−NK1.1+. Data are mean ± SEM. ***p < 0.001. d Mononuclear cell infiltration in the quadriceps muscles of Syt7−/−Il15f/f and Syt7−/−ACTA-Il15−/− mice 14 days after C protein immunization (Left). Mononuclear cell infiltration was found in the endomysium (black square), perimysium (black arrowhead), and perivascular region (white arrowhead). Focal lymphatic invasion of myofibers were observed in C-protein-immunized Syt7−/−ACTA-Il15−/− mice as indicated by the white arrowhead (Middle). The histopathology scores were compiled in the right panel with each symbol representing one mouse (Right). Scale bar in left = 100 μm; middle = 25 μm. **p < 0.01. e Expression of mononuclear cell markers, Cd4, Cd8, and F4/80, mRNA in the quadriceps muscles of C-protein-immunized mice measured by qPCR. Each symbol representing one mouse. *p < 0.05, **p < 0.01. f Expression of immune-relevant genes mRNA in the quadriceps muscles of C-protein-immunized mice measured by qPCR. Each symbol representing one mouse. *p < 0.05, **p < 0.01

Mentions: TNF-α and IFN-γ are commonly expressed in the skeletal muscle of patients suffering from inflammatory myopathies, in which CD8+ T cells infiltrate and play a critical role in disease progression [23, 25, 52]. The enhancement of memory-like CD8+ T-cell effector function by myoblast IL-15 in vitro prompted us to examine the role of skeletal muscle IL-15 in autoimmune myositis in vivo. We first generated skeletal-muscle-specific Il15−/− mice by crossing Il15f/f mice with ACTA-cre mice. The ACTA-Il15−/− mice showed an 80 % reduction of Il15 mRNA specifically in the skeletal muscle (Fig. 6a) with normal levels of IL-15/IL-15Rα complex and NK and memory CD8+ T cells in the peripheral blood (Fig. 6b, c).Fig. 6


Skeletal muscle interleukin 15 promotes CD8(+) T-cell function and autoimmune myositis.

Huang PL, Hou MS, Wang SW, Chang CL, Liou YH, Liao NS - Skelet Muscle (2015)

Skeletal muscle IL-15 contributes to the progression of autoimmune myositis. a The expression of Il15 mRNA in various tissues of wt (n = 3), Il15f/f (n = 5), and ACTA-Il15−/− (n = 5) mice was detected by qPCR. WAT white adipose tissue. Data are mean ± SEM. ***p < 0.001. b The amount of IL-15/IL-15Rα complex protein in the serum of Il15f/f and ACTA-Il15−/− mice (n = 9–10) was measured by ELISA. Data are mean ± SEM. c Comparison of memory CD8+ T cell (mCD8) and NK cell (NK) level in the peripheral blood among wt, Il15−/−, Il15f/f, and ACTA-Il15−/− mice (n = 5 in each group). Fold change was calculated by normalizing the percentage of indicated cell type in mutant mice to that in wt mice. mCD8 were H57+CD19−CD8+CD44hiCD122hi, and NK were H57−CD19−NK1.1+. Data are mean ± SEM. ***p < 0.001. d Mononuclear cell infiltration in the quadriceps muscles of Syt7−/−Il15f/f and Syt7−/−ACTA-Il15−/− mice 14 days after C protein immunization (Left). Mononuclear cell infiltration was found in the endomysium (black square), perimysium (black arrowhead), and perivascular region (white arrowhead). Focal lymphatic invasion of myofibers were observed in C-protein-immunized Syt7−/−ACTA-Il15−/− mice as indicated by the white arrowhead (Middle). The histopathology scores were compiled in the right panel with each symbol representing one mouse (Right). Scale bar in left = 100 μm; middle = 25 μm. **p < 0.01. e Expression of mononuclear cell markers, Cd4, Cd8, and F4/80, mRNA in the quadriceps muscles of C-protein-immunized mice measured by qPCR. Each symbol representing one mouse. *p < 0.05, **p < 0.01. f Expression of immune-relevant genes mRNA in the quadriceps muscles of C-protein-immunized mice measured by qPCR. Each symbol representing one mouse. *p < 0.05, **p < 0.01
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Fig6: Skeletal muscle IL-15 contributes to the progression of autoimmune myositis. a The expression of Il15 mRNA in various tissues of wt (n = 3), Il15f/f (n = 5), and ACTA-Il15−/− (n = 5) mice was detected by qPCR. WAT white adipose tissue. Data are mean ± SEM. ***p < 0.001. b The amount of IL-15/IL-15Rα complex protein in the serum of Il15f/f and ACTA-Il15−/− mice (n = 9–10) was measured by ELISA. Data are mean ± SEM. c Comparison of memory CD8+ T cell (mCD8) and NK cell (NK) level in the peripheral blood among wt, Il15−/−, Il15f/f, and ACTA-Il15−/− mice (n = 5 in each group). Fold change was calculated by normalizing the percentage of indicated cell type in mutant mice to that in wt mice. mCD8 were H57+CD19−CD8+CD44hiCD122hi, and NK were H57−CD19−NK1.1+. Data are mean ± SEM. ***p < 0.001. d Mononuclear cell infiltration in the quadriceps muscles of Syt7−/−Il15f/f and Syt7−/−ACTA-Il15−/− mice 14 days after C protein immunization (Left). Mononuclear cell infiltration was found in the endomysium (black square), perimysium (black arrowhead), and perivascular region (white arrowhead). Focal lymphatic invasion of myofibers were observed in C-protein-immunized Syt7−/−ACTA-Il15−/− mice as indicated by the white arrowhead (Middle). The histopathology scores were compiled in the right panel with each symbol representing one mouse (Right). Scale bar in left = 100 μm; middle = 25 μm. **p < 0.01. e Expression of mononuclear cell markers, Cd4, Cd8, and F4/80, mRNA in the quadriceps muscles of C-protein-immunized mice measured by qPCR. Each symbol representing one mouse. *p < 0.05, **p < 0.01. f Expression of immune-relevant genes mRNA in the quadriceps muscles of C-protein-immunized mice measured by qPCR. Each symbol representing one mouse. *p < 0.05, **p < 0.01
Mentions: TNF-α and IFN-γ are commonly expressed in the skeletal muscle of patients suffering from inflammatory myopathies, in which CD8+ T cells infiltrate and play a critical role in disease progression [23, 25, 52]. The enhancement of memory-like CD8+ T-cell effector function by myoblast IL-15 in vitro prompted us to examine the role of skeletal muscle IL-15 in autoimmune myositis in vivo. We first generated skeletal-muscle-specific Il15−/− mice by crossing Il15f/f mice with ACTA-cre mice. The ACTA-Il15−/− mice showed an 80 % reduction of Il15 mRNA specifically in the skeletal muscle (Fig. 6a) with normal levels of IL-15/IL-15Rα complex and NK and memory CD8+ T cells in the peripheral blood (Fig. 6b, c).Fig. 6

Bottom Line: Consistently, deficiency of endogenous IL-15 affected neither skeletal muscle growth nor its responses to TNF-α and IFN-γ.On the other hand, the cytokine-stimulated skeletal muscle cells presented antigen and provided IL-15 to promote the effector function of memory-like CD8(+) T cells.These findings together indicate that skeletal muscle IL-15 directly regulates immune effector cells but not muscle cells and thus presents a potential therapeutic target for myositis.

View Article: PubMed Central - PubMed

Affiliation: Molecular Cell Biology, Taiwan International Graduate Program, Institute of Molecular Biology, Academia Sinica, and Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan ; Institute of Molecular Biology, Academia Sinica, Taipei, 11529 Taiwan.

ABSTRACT

Background: Interleukin 15 (IL-15) is thought to be abundant in the skeletal muscle under steady state conditions based on RNA expression; however, the IL-15 RNA level may not reflect the protein level due to post-transcriptional regulation. Although exogenous protein treatment and overexpression studies indicated IL-15 functions in the skeletal muscle, how the skeletal muscle cell uses IL-15 remains unclear. In myositis patients, IL-15 protein is up-regulated in the skeletal muscle. Given the supporting role of IL-15 in CD8(+) T-cell survival and activation and the pathogenic role of cytotoxic CD8(+) T cells in polymyositis and inclusion-body myositis, we hypothesize that IL-15 produced by the inflamed skeletal muscle promotes myositis via CD8(+) T cells.

Methods: Expression of IL-15 and IL-15 receptors at the protein level by skeletal muscle cells were examined under steady state and cytokine stimulation conditions. The functions of IL-15 in the skeletal muscle were investigated using Il15 knockout (Il15 (-/-) ) mice. The immune regulatory role of skeletal muscle IL-15 was determined by co-culturing cytokine-stimulated muscle cells and memory-like CD8(+) T cells in vitro and by inducing autoimmune myositis in skeletal-muscle-specific Il15 (-/-) mice.

Results: We found that the IL-15 protein was not expressed by skeletal muscle cells under steady state condition but induced by tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ) stimulation and expressed as IL-15/IL-15 receptor alpha (IL-15Rα) complex. Skeletal muscle cells expressed a scanty amount of IL-15 receptor beta (IL-15Rβ) under either conditions and only responded to a high concentration of IL-15 hyperagonist, but not IL-15. Consistently, deficiency of endogenous IL-15 affected neither skeletal muscle growth nor its responses to TNF-α and IFN-γ. On the other hand, the cytokine-stimulated skeletal muscle cells presented antigen and provided IL-15 to promote the effector function of memory-like CD8(+) T cells. Genetic ablation of Il15 in skeletal muscle cells greatly ameliorated autoimmune myositis in mice.

Conclusions: These findings together indicate that skeletal muscle IL-15 directly regulates immune effector cells but not muscle cells and thus presents a potential therapeutic target for myositis.

No MeSH data available.


Related in: MedlinePlus