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Chronic thoracic spinal cord injury impairs CD8+ T-cell function by up-regulating programmed cell death-1 expression.

Zha J, Smith A, Andreansky S, Bracchi-Ricard V, Bethea JR - J Neuroinflammation (2014)

Bottom Line: Chronic SCI impaired both CD4+ and CD8+ T-cell cytokine production.The observed T-cell dysfunction correlated with increased expression of programmed cell death 1 (PD-1) exhaustion marker on these cells.Blocking PD-1 signaling in vitro restored the CD8+ T-cell functional defect.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Miami Project to Cure Paralysis, Department of Neurosurgery, Miller School of Medicine, University of Miami, Miami, FL 33136, USA. VBracchi@med.miami.edu.

ABSTRACT

Background: Chronic spinal cord injury (SCI) induces immune depression in patients, which contributes to their higher risk of developing infections. While defects in humoral immunity have been reported, complications in T-cell immunity during the chronic phase of SCI have not yet been explored.

Methods: To assess the impact of chronic SCI on peripheral T-cell number and function we used a mouse model of severe spinal cord contusion at thoracic level T9 and performed flow cytometry analysis on the spleen for T-cell markers along with intracellular cytokine staining. Furthermore we identified alterations in sympathetic activity in the spleen of chronic SCI mice by measuring splenic levels of tyrosine hydroxylase (TH) and norepinephrine (NE). To gain insight into the neurogenic mechanism leading to T-cell dysfunction we performed in vitro NE stimulation of T-cells followed by flow cytometry analysis for T-cell exhaustion marker.

Results: Chronic SCI impaired both CD4+ and CD8+ T-cell cytokine production. The observed T-cell dysfunction correlated with increased expression of programmed cell death 1 (PD-1) exhaustion marker on these cells. Blocking PD-1 signaling in vitro restored the CD8+ T-cell functional defect. In addition, we showed that chronic SCI mice had higher levels of splenic NE, which contributed to the T-cell exhaustion phenotype, as PD-1 expression on both CD4+ and CD8+ T-cells was up-regulated following sustained exposure to NE in vitro.

Conclusions: These studies indicate that alteration of sympathetic activity following chronic SCI induces CD8+ T-cell exhaustion, which in turn impairs T-cell function and contributes to immune depression. Inhibition of the exhaustion pathway should be considered as a new therapeutic strategy for chronic SCI-induced immune depression.

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Increased expression of exhaustion marker PD-1 on T-cells isolated from chronic spinal cord injury (SCI) mice. (A) Representative dot plots show the percentage of PD-1+ cells in gated CD4+ T-cells and CD8+ T-cells from uninjured (CT) and T9-SCI mice at chronic phase after injury (SCI). (B) Bar graphs show the mean ± SEM numbers of PD-1 expressing CD4+ T-cells and CD8+ T-cells. Twenty thousand events gated on live singlets were collected. n = 9 for CT mice, n = 11 for SCI mice. Data have been pooled across two independent experiments. *P < 0.05, one-tailed Student’s t-test.
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Figure 5: Increased expression of exhaustion marker PD-1 on T-cells isolated from chronic spinal cord injury (SCI) mice. (A) Representative dot plots show the percentage of PD-1+ cells in gated CD4+ T-cells and CD8+ T-cells from uninjured (CT) and T9-SCI mice at chronic phase after injury (SCI). (B) Bar graphs show the mean ± SEM numbers of PD-1 expressing CD4+ T-cells and CD8+ T-cells. Twenty thousand events gated on live singlets were collected. n = 9 for CT mice, n = 11 for SCI mice. Data have been pooled across two independent experiments. *P < 0.05, one-tailed Student’s t-test.

Mentions: T-cell exhaustion indicated by increased expression of exhaustion markers such as PD-1, cytotoxic T-lymphocyte antigen 4 (CTLA-4), T-cell immunoglobulin mucin-3 (TIM-3) and lymphocyte activation gene-3 (LAG-3) was shown to correlate with T-cell dysfunction in chronic viral infection models and aging animals [23,25,29,37-40]. To examine whether the T-cell impairment in cytokine production observed in chronic SCI mice was associated with T-cell exhaustion, we measured the expression of exhaustion markers on both CD4+ T-cells and CD8+ T-cells. The percentage of PD-1 expressing cells in both CD4+ T-cells and CD8+ T-cells was significantly higher in the spleen of chronic SCI mice compared with uninjured controls (CD4+PD1+ T-cells: uninjured: 12.9 ± 1.3%; chronic SCI: 18.0 ± 1.7%; P = 0.02. CD8+PD1+ T-cells: uninjured: 3.4 ± 0.2%; chronic SCI: 5.0 ± 0.4%; P = 0.003) (Figure 5A). The number of splenic CD8+PD-1+ cells was also significantly increased in the chronic SCI mice (uninjured: 0.43 ± 0.05 × 106; chronic SCI: 0.59 ± 0.04 × 106; P = 0.01) (Figure 5B). However, the number of splenic CD4+PD-1+ cells was not significantly changed by chronic SCI (uninjured: 1.9 ± 0.4 × 106; chronic SCI: 2.7 ± 0.3 × 106; P = 0.07) (Figure 5B). The expression of other exhaustion markers including CTLA-4, TIM-3 and LAG-3 on T-cells was not increased by chronic SCI (data not shown).


Chronic thoracic spinal cord injury impairs CD8+ T-cell function by up-regulating programmed cell death-1 expression.

Zha J, Smith A, Andreansky S, Bracchi-Ricard V, Bethea JR - J Neuroinflammation (2014)

Increased expression of exhaustion marker PD-1 on T-cells isolated from chronic spinal cord injury (SCI) mice. (A) Representative dot plots show the percentage of PD-1+ cells in gated CD4+ T-cells and CD8+ T-cells from uninjured (CT) and T9-SCI mice at chronic phase after injury (SCI). (B) Bar graphs show the mean ± SEM numbers of PD-1 expressing CD4+ T-cells and CD8+ T-cells. Twenty thousand events gated on live singlets were collected. n = 9 for CT mice, n = 11 for SCI mice. Data have been pooled across two independent experiments. *P < 0.05, one-tailed Student’s t-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4230802&req=5

Figure 5: Increased expression of exhaustion marker PD-1 on T-cells isolated from chronic spinal cord injury (SCI) mice. (A) Representative dot plots show the percentage of PD-1+ cells in gated CD4+ T-cells and CD8+ T-cells from uninjured (CT) and T9-SCI mice at chronic phase after injury (SCI). (B) Bar graphs show the mean ± SEM numbers of PD-1 expressing CD4+ T-cells and CD8+ T-cells. Twenty thousand events gated on live singlets were collected. n = 9 for CT mice, n = 11 for SCI mice. Data have been pooled across two independent experiments. *P < 0.05, one-tailed Student’s t-test.
Mentions: T-cell exhaustion indicated by increased expression of exhaustion markers such as PD-1, cytotoxic T-lymphocyte antigen 4 (CTLA-4), T-cell immunoglobulin mucin-3 (TIM-3) and lymphocyte activation gene-3 (LAG-3) was shown to correlate with T-cell dysfunction in chronic viral infection models and aging animals [23,25,29,37-40]. To examine whether the T-cell impairment in cytokine production observed in chronic SCI mice was associated with T-cell exhaustion, we measured the expression of exhaustion markers on both CD4+ T-cells and CD8+ T-cells. The percentage of PD-1 expressing cells in both CD4+ T-cells and CD8+ T-cells was significantly higher in the spleen of chronic SCI mice compared with uninjured controls (CD4+PD1+ T-cells: uninjured: 12.9 ± 1.3%; chronic SCI: 18.0 ± 1.7%; P = 0.02. CD8+PD1+ T-cells: uninjured: 3.4 ± 0.2%; chronic SCI: 5.0 ± 0.4%; P = 0.003) (Figure 5A). The number of splenic CD8+PD-1+ cells was also significantly increased in the chronic SCI mice (uninjured: 0.43 ± 0.05 × 106; chronic SCI: 0.59 ± 0.04 × 106; P = 0.01) (Figure 5B). However, the number of splenic CD4+PD-1+ cells was not significantly changed by chronic SCI (uninjured: 1.9 ± 0.4 × 106; chronic SCI: 2.7 ± 0.3 × 106; P = 0.07) (Figure 5B). The expression of other exhaustion markers including CTLA-4, TIM-3 and LAG-3 on T-cells was not increased by chronic SCI (data not shown).

Bottom Line: Chronic SCI impaired both CD4+ and CD8+ T-cell cytokine production.The observed T-cell dysfunction correlated with increased expression of programmed cell death 1 (PD-1) exhaustion marker on these cells.Blocking PD-1 signaling in vitro restored the CD8+ T-cell functional defect.

View Article: PubMed Central - HTML - PubMed

Affiliation: The Miami Project to Cure Paralysis, Department of Neurosurgery, Miller School of Medicine, University of Miami, Miami, FL 33136, USA. VBracchi@med.miami.edu.

ABSTRACT

Background: Chronic spinal cord injury (SCI) induces immune depression in patients, which contributes to their higher risk of developing infections. While defects in humoral immunity have been reported, complications in T-cell immunity during the chronic phase of SCI have not yet been explored.

Methods: To assess the impact of chronic SCI on peripheral T-cell number and function we used a mouse model of severe spinal cord contusion at thoracic level T9 and performed flow cytometry analysis on the spleen for T-cell markers along with intracellular cytokine staining. Furthermore we identified alterations in sympathetic activity in the spleen of chronic SCI mice by measuring splenic levels of tyrosine hydroxylase (TH) and norepinephrine (NE). To gain insight into the neurogenic mechanism leading to T-cell dysfunction we performed in vitro NE stimulation of T-cells followed by flow cytometry analysis for T-cell exhaustion marker.

Results: Chronic SCI impaired both CD4+ and CD8+ T-cell cytokine production. The observed T-cell dysfunction correlated with increased expression of programmed cell death 1 (PD-1) exhaustion marker on these cells. Blocking PD-1 signaling in vitro restored the CD8+ T-cell functional defect. In addition, we showed that chronic SCI mice had higher levels of splenic NE, which contributed to the T-cell exhaustion phenotype, as PD-1 expression on both CD4+ and CD8+ T-cells was up-regulated following sustained exposure to NE in vitro.

Conclusions: These studies indicate that alteration of sympathetic activity following chronic SCI induces CD8+ T-cell exhaustion, which in turn impairs T-cell function and contributes to immune depression. Inhibition of the exhaustion pathway should be considered as a new therapeutic strategy for chronic SCI-induced immune depression.

Show MeSH
Related in: MedlinePlus