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Persistent inflammation and T cell exhaustion in severe sepsis in the elderly.

Inoue S, Suzuki K, Komori Y, Morishita Y, Suzuki-Utsunomiya K, Hozumi K, Inokuchi S, Sato T - Crit Care (2014)

Bottom Line: Fifty-five patients with severe sepsis and 30 healthy donors were prospectively enrolled, and 90-day survival was compared between elderly (≥ 65 years) and adult (18-64 years) septic patients with serial measurement of serum interleukin (IL)-6.Within 24 h after diagnosis of severe sepsis, peripheral blood mononuclear cells were stimulated ex vivo to measure expression of the activation maker CD25 in T cells, IL-2 levels in the supernatant, and proliferation.Expression of the negative co-stimulatory molecules, CD25, and IL-2 in CD4+ T cells was measured.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: Sepsis is known as a complex immunological response with hyperinflammation in the acute phase followed by immunosuppression. Although aging is crucial in sepsis, the impact of aging on inflammation and immunosuppression is still unclear. The purpose of this study was to investigate the relationship between inflammation and immunosuppression in aged patients and mice after sepsis.

Methods: Fifty-five patients with severe sepsis and 30 healthy donors were prospectively enrolled, and 90-day survival was compared between elderly (≥ 65 years) and adult (18-64 years) septic patients with serial measurement of serum interleukin (IL)-6. Within 24 h after diagnosis of severe sepsis, peripheral blood mononuclear cells were stimulated ex vivo to measure expression of the activation maker CD25 in T cells, IL-2 levels in the supernatant, and proliferation. In the mouse study, young (6-8 weeks) and aged (20-22 months) C57/B6 mice were subjected to cecal ligation and puncture (CLP), and survival was compared after 7 days with serial measurement of serum IL-6. Expression of the negative co-stimulatory molecules, CD25, and IL-2 in CD4+ T cells was measured.

Results: The survival rate in elderly sepsis patients and aged septic mice was significantly lower than that in adult patients and young septic mice (60% vs. 93% in septic patients, 0% vs. 63% in septic mice, P < 0.05). Serum IL-6 levels in elderly sepsis patients and aged septic mice were persistently higher than those in adult patients and young septic mice. Expression of negative co-stimulatory molecules in CD4+ T cells in the spleen, lymph nodes, and peripheral blood was significantly higher in aged mice than in young mice (P < 0.01). Ex vivo stimulation decreased CD25 expression, IL-2 production, and proliferation to a greater extent in CD4+ T cells from elderly patients and aged septic mice than in those from adult patients and young septic mice. Elderly patients demonstrated increased detection of gram-negative bacteria at days 14-16 and 28-32 after sepsis (P < 0.05).

Conclusions: Persistent inflammation and T cell exhaustion may be associated with decreased survival in elderly patients and mice after sepsis.

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Impaired ex vivo activation, IL-2 production, and proliferation of CD4 + T cells from elderly patients and aged mice with severe sepsis. (A) Dissociated peripheral blood mononuclear cells were stimulated ex vivo overnight with an anti-CD3/CD28 antibody, followed by flow cytometric analysis to quantify T cell activation. (B) IL-2 concentration in supernatants from peripheral blood mononuclear cells (PBMCs) with overnight stimulation by anti-CD3/CD28 antibody. (C) Profiles of carboxyfluorescein succinimidyl ester (CFSE) fluorescence intensity of activated PBMC cultures. Population doubling of CD4+ T cells can be clearly assessed by the decrease in CFSE fluorescence. Impaired proliferation was observed in CD4+ T cells from elderly healthy donors (HDs) and septic patients; n = 6 to 14 per group, *P <0.05, **P <0.01. (D) Dissociated splenocytes were stimulated overnight using the anti-CD3 antibody followed by staining for CD4+ T cells and quantification of the activation marker CD25 by flow cytometry ex vivo; n = 6 per group, *P <0.05, **P <0.01. (E) Percentage of IL-2-secreting CD4+ T cells on overnight incubation with an anti-CD3 antibody. (F) Population doubling of CD4+ T cells from young and aged splenocytes can be clearly assessed by the decrease in CFSE fluorescence. Impaired proliferation was observed in CD4+ T cells from aged sham and CLP mice; n = 6 to 8 per group, *P <0.05.
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Figure 6: Impaired ex vivo activation, IL-2 production, and proliferation of CD4 + T cells from elderly patients and aged mice with severe sepsis. (A) Dissociated peripheral blood mononuclear cells were stimulated ex vivo overnight with an anti-CD3/CD28 antibody, followed by flow cytometric analysis to quantify T cell activation. (B) IL-2 concentration in supernatants from peripheral blood mononuclear cells (PBMCs) with overnight stimulation by anti-CD3/CD28 antibody. (C) Profiles of carboxyfluorescein succinimidyl ester (CFSE) fluorescence intensity of activated PBMC cultures. Population doubling of CD4+ T cells can be clearly assessed by the decrease in CFSE fluorescence. Impaired proliferation was observed in CD4+ T cells from elderly healthy donors (HDs) and septic patients; n = 6 to 14 per group, *P <0.05, **P <0.01. (D) Dissociated splenocytes were stimulated overnight using the anti-CD3 antibody followed by staining for CD4+ T cells and quantification of the activation marker CD25 by flow cytometry ex vivo; n = 6 per group, *P <0.05, **P <0.01. (E) Percentage of IL-2-secreting CD4+ T cells on overnight incubation with an anti-CD3 antibody. (F) Population doubling of CD4+ T cells from young and aged splenocytes can be clearly assessed by the decrease in CFSE fluorescence. Impaired proliferation was observed in CD4+ T cells from aged sham and CLP mice; n = 6 to 8 per group, *P <0.05.

Mentions: The patients’ blood samples were collected within 24 h after diagnosis of severe sepsis. The characteristics of the ex vivo stimulation assay described below are listed in Additional file 3. Upon activation using an anti-CD3/28 antibody, CD25 expression in CD4+ T cells was lower in adult and elderly patients than in HDs, which suggests that immunosuppression occurred even in the relatively early phase of sepsis. Importantly, the impact of aging on anti-CD3/28-mediated CD25 expression was evident in both HDs and in sepsis patients, which suggests that T-cell activation is impaired in aged individuals irrespective of infection status (Figure 6A). Similar to the results observed in human cells, both aging and sepsis induced a reduction of CD25 expression in mouse CD4+ T cells upon ex vivo stimulation by an anti-CD3 antibody (P <0.05; Figure 6D). Purified CD4+ T cells of aged mice also showed insufficient activation (Additional file 4).


Persistent inflammation and T cell exhaustion in severe sepsis in the elderly.

Inoue S, Suzuki K, Komori Y, Morishita Y, Suzuki-Utsunomiya K, Hozumi K, Inokuchi S, Sato T - Crit Care (2014)

Impaired ex vivo activation, IL-2 production, and proliferation of CD4 + T cells from elderly patients and aged mice with severe sepsis. (A) Dissociated peripheral blood mononuclear cells were stimulated ex vivo overnight with an anti-CD3/CD28 antibody, followed by flow cytometric analysis to quantify T cell activation. (B) IL-2 concentration in supernatants from peripheral blood mononuclear cells (PBMCs) with overnight stimulation by anti-CD3/CD28 antibody. (C) Profiles of carboxyfluorescein succinimidyl ester (CFSE) fluorescence intensity of activated PBMC cultures. Population doubling of CD4+ T cells can be clearly assessed by the decrease in CFSE fluorescence. Impaired proliferation was observed in CD4+ T cells from elderly healthy donors (HDs) and septic patients; n = 6 to 14 per group, *P <0.05, **P <0.01. (D) Dissociated splenocytes were stimulated overnight using the anti-CD3 antibody followed by staining for CD4+ T cells and quantification of the activation marker CD25 by flow cytometry ex vivo; n = 6 per group, *P <0.05, **P <0.01. (E) Percentage of IL-2-secreting CD4+ T cells on overnight incubation with an anti-CD3 antibody. (F) Population doubling of CD4+ T cells from young and aged splenocytes can be clearly assessed by the decrease in CFSE fluorescence. Impaired proliferation was observed in CD4+ T cells from aged sham and CLP mice; n = 6 to 8 per group, *P <0.05.
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Figure 6: Impaired ex vivo activation, IL-2 production, and proliferation of CD4 + T cells from elderly patients and aged mice with severe sepsis. (A) Dissociated peripheral blood mononuclear cells were stimulated ex vivo overnight with an anti-CD3/CD28 antibody, followed by flow cytometric analysis to quantify T cell activation. (B) IL-2 concentration in supernatants from peripheral blood mononuclear cells (PBMCs) with overnight stimulation by anti-CD3/CD28 antibody. (C) Profiles of carboxyfluorescein succinimidyl ester (CFSE) fluorescence intensity of activated PBMC cultures. Population doubling of CD4+ T cells can be clearly assessed by the decrease in CFSE fluorescence. Impaired proliferation was observed in CD4+ T cells from elderly healthy donors (HDs) and septic patients; n = 6 to 14 per group, *P <0.05, **P <0.01. (D) Dissociated splenocytes were stimulated overnight using the anti-CD3 antibody followed by staining for CD4+ T cells and quantification of the activation marker CD25 by flow cytometry ex vivo; n = 6 per group, *P <0.05, **P <0.01. (E) Percentage of IL-2-secreting CD4+ T cells on overnight incubation with an anti-CD3 antibody. (F) Population doubling of CD4+ T cells from young and aged splenocytes can be clearly assessed by the decrease in CFSE fluorescence. Impaired proliferation was observed in CD4+ T cells from aged sham and CLP mice; n = 6 to 8 per group, *P <0.05.
Mentions: The patients’ blood samples were collected within 24 h after diagnosis of severe sepsis. The characteristics of the ex vivo stimulation assay described below are listed in Additional file 3. Upon activation using an anti-CD3/28 antibody, CD25 expression in CD4+ T cells was lower in adult and elderly patients than in HDs, which suggests that immunosuppression occurred even in the relatively early phase of sepsis. Importantly, the impact of aging on anti-CD3/28-mediated CD25 expression was evident in both HDs and in sepsis patients, which suggests that T-cell activation is impaired in aged individuals irrespective of infection status (Figure 6A). Similar to the results observed in human cells, both aging and sepsis induced a reduction of CD25 expression in mouse CD4+ T cells upon ex vivo stimulation by an anti-CD3 antibody (P <0.05; Figure 6D). Purified CD4+ T cells of aged mice also showed insufficient activation (Additional file 4).

Bottom Line: Fifty-five patients with severe sepsis and 30 healthy donors were prospectively enrolled, and 90-day survival was compared between elderly (≥ 65 years) and adult (18-64 years) septic patients with serial measurement of serum interleukin (IL)-6.Within 24 h after diagnosis of severe sepsis, peripheral blood mononuclear cells were stimulated ex vivo to measure expression of the activation maker CD25 in T cells, IL-2 levels in the supernatant, and proliferation.Expression of the negative co-stimulatory molecules, CD25, and IL-2 in CD4+ T cells was measured.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Introduction: Sepsis is known as a complex immunological response with hyperinflammation in the acute phase followed by immunosuppression. Although aging is crucial in sepsis, the impact of aging on inflammation and immunosuppression is still unclear. The purpose of this study was to investigate the relationship between inflammation and immunosuppression in aged patients and mice after sepsis.

Methods: Fifty-five patients with severe sepsis and 30 healthy donors were prospectively enrolled, and 90-day survival was compared between elderly (≥ 65 years) and adult (18-64 years) septic patients with serial measurement of serum interleukin (IL)-6. Within 24 h after diagnosis of severe sepsis, peripheral blood mononuclear cells were stimulated ex vivo to measure expression of the activation maker CD25 in T cells, IL-2 levels in the supernatant, and proliferation. In the mouse study, young (6-8 weeks) and aged (20-22 months) C57/B6 mice were subjected to cecal ligation and puncture (CLP), and survival was compared after 7 days with serial measurement of serum IL-6. Expression of the negative co-stimulatory molecules, CD25, and IL-2 in CD4+ T cells was measured.

Results: The survival rate in elderly sepsis patients and aged septic mice was significantly lower than that in adult patients and young septic mice (60% vs. 93% in septic patients, 0% vs. 63% in septic mice, P < 0.05). Serum IL-6 levels in elderly sepsis patients and aged septic mice were persistently higher than those in adult patients and young septic mice. Expression of negative co-stimulatory molecules in CD4+ T cells in the spleen, lymph nodes, and peripheral blood was significantly higher in aged mice than in young mice (P < 0.01). Ex vivo stimulation decreased CD25 expression, IL-2 production, and proliferation to a greater extent in CD4+ T cells from elderly patients and aged septic mice than in those from adult patients and young septic mice. Elderly patients demonstrated increased detection of gram-negative bacteria at days 14-16 and 28-32 after sepsis (P < 0.05).

Conclusions: Persistent inflammation and T cell exhaustion may be associated with decreased survival in elderly patients and mice after sepsis.

Show MeSH
Related in: MedlinePlus