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. 2019 Oct;18(5):e12980.
doi: 10.1111/acel.12980. Epub 2019 Jun 14.

Advanced age promotes colonic dysfunction and gut-derived lung infection after stroke

Affiliations

Advanced age promotes colonic dysfunction and gut-derived lung infection after stroke

Shu Wen Wen et al. Aging Cell. 2019 Oct.

Abstract

Bacterial infection a leading cause of death among patients with stroke, with elderly patients often presenting with more debilitating outcomes. The findings from our retrospective study, supported by previous clinical reports, showed that increasing age is an early predictor for developing fatal infectious complications after stroke. However, exactly how and why older individuals are more susceptible to infection after stroke remains unclear. Using a mouse model of transient ischaemic stroke, we demonstrate that older mice (>12 months) present with greater spontaneous bacterial lung infections compared to their younger counterparts (7-10 weeks) after stroke. Importantly, we provide evidence that older poststroke mice exhibited elevated intestinal inflammation and disruption in gut barriers critical in maintaining colonic integrity following stroke, including reduced expression of mucin and tight junction proteins. In addition, our data support the notion that the localized pro-inflammatory microenvironment driven by increased tumour necrosis factor-α production in the colon of older mice facilitates the translocation and dissemination of orally inoculated bacteria to the lung following stroke onset. Therefore, findings of this study demonstrate that exacerbated dysfunction of the intestinal barrier in advanced age promotes translocation of gut-derived bacteria and contributes to the increased risk to poststroke bacterial infection.

Keywords: aging; bacteria; colon; infection; stroke.

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Conflict of interest statement

None declared.

Figures

Figure 1
Figure 1
Advanced age exacerbates neurological impairment and lung infection poststroke. The following assessments were performed 24 hr after mid‐cerebral artery occlusion induction on young (7–10 weeks) and older (12–15 months) mice: (a) preclinical magnetic resonance imaging (MRI) to assess volume of brain oedema (n = 6/group). Oedema region indicated within yellow outline; (b) neurological assessment (n = 14–16/group); (c) bacteriological analysis of lung homogenates to assess poststroke lung infection (n = 6–8/group). Data represent the mean ± SEM. Significance was determined by Mann–Whitney U test, and a p‐value ≤0.05 was considered statistically significant: *p ≤ 0.05, **p ≤ 0.01
Figure 2
Figure 2
Colonic permeability increases with age after stroke. Serum levels of orally gavaged fluorescein‐isothiocyanate‐labelled (FITC)‐dextran were quantified from mid‐cerebral artery occlusion (MCAO) or sham mice to examine permeability of the (a) small intestine and (b) colon. (c) Extravasation of Evans blue dye from the colon was quantified 24 hr after MCAO or sham surgery as an indicator of vascular permeability. (d) Representative H&E images of colonic crypts at 100× magnification (scale bar = 2 mm). Histological scores for various parameters were totalled to indicate the degree of colonic pathology. A higher score indicates more visible clinical signs of colonic damage (n = 4–8/group). Black arrows denote examples of crypt damage and loss of architecture. Data represent the mean ± SEM. Significance determined by Mann–Whitney U test. A p‐value ≤0.05 was considered statistically significant: *p ≤ 0.05
Figure 3
Figure 3
Stroke induces robust mucosal changes in older animals. (a) Representative images of colonic goblet cells at 100× magnification (scale bar = 2 mm; black arrow), stained using PAS‐Alcian blue at 24 hr postsurgery (n = 4–5/group). (b) Gene expression of mucin 2 (Muc2), Muc4 and Muc13 from the colon of young and older mice was analysed by qPCR 24 hr after mid‐cerebral artery occlusion (MCAO) and expressed as a fold change relative to that of young sham‐operated controls (n = 7/group). (c) Protein levels of secretory IgA in the serum and (d) colon 24 hr after MCAO or sham surgery (n = 7–8/group). Data represent the mean ± SEM. Significance was determined by one‐way ANOVA with post hoc comparison and Holm–Sidak multiple testing correction. A p‐value ≤0.05 was considered statistically significant: *p ≤ 0.05, **p ≤ 0.01
Figure 4
Figure 4
Age‐dependent disruption of colonic tight junctions after stroke. (a) Representative immunofluorescence images of colonic cross sections at magnification of 400× (scale bar = 50 µm), stained for nuclei (DAPI; blue), EpCAM (Alexa Fluor 488; green) and ZO‐1 (Alexa Fluor 568; red). The area of ZO‐1 staining relative to DAPI staining was quantified 24 hr after mid‐cerebral artery occlusion (MCAO) or sham surgery (n = 5/group). The gene expression of (b) occludin (Ocldn), (c) junctional adhesion molecule‐A (JAM‐A), (d) claudin 3 (Cldn3) and (e) claudin 5 (Cldn5) from the colon of young and older mice was analysed by qPCR 24 hr after MCAO and expressed as a fold change relative to that of young sham‐operated controls (n = 7/group). Data represent the mean ± SEM. Significance was determined by unpaired t test. A p‐value ≤0.05 was considered statistically significant: *p ≤ 0.05
Figure 5
Figure 5
Tumour necrosis factor (TNF)‐α may facilitate the breakdown of colonic barriers poststroke. Protein expression of TNF‐α and IL‐10 in the (a–b) colon and (c–d) serum from young and older mice was quantified 5 hr after mid‐cerebral artery occlusion (MCAO) or sham surgery (n = 5–8/group). (e) Colon cross sections from naïve older mice were treated with recombinant TNF‐α for 1 hr in an ex‐vivo setting, while untreated tissues acted as controls. Gene expression of claudin 3 (Cldn3), claudin 5 (Cldn5), junctional adhesion molecule‐A (JAM‐A) and occludin (Ocldn) was assessed by qPCR and expressed as a fold change relative to untreated older colons (n = 5/group). (f) To examine whether TNF‐α can alter intestinal permeability in vivo, recombinant TNF‐α (20 µg/kg) or saline as control was administered (i.p.) immediately following blood reperfusion to young MCAO and sham‐operated mice. Serum levels of orally gavaged FITC‐dextran were quantified to examine colon permeability. Data represent the mean ± SEM. Significance was determined by one‐way ANOVA with post hoc comparison and Holm–Sidak multiple testing correction. A p‐value ≤ 0.05 was considered statistically significant: *p ≤ 0.05, ***p ≤ 0.001 and § p ≤ 0.1 considered a significant trend
Figure 6
Figure 6
Stroke results in the translocation of intestinal bacteria for systemic dissemination in older animals. (a) Three hours after mid‐cerebral artery occlusion (MCAO) or sham surgery, young and older mice were orally inoculated with streptomycin‐resistant derivative of the Escherichia coli strain DLL206, at a time point when gut permeability was evident. The (b) caecum, (c) faeces, (d) colon and (e) lung were assessed 24 hr later for the presence and load of streptomycin‐resistant E. coli. n = 5–6/group. Data represent the mean ± SEM (log‐scale). Significance was determined by one‐way ANOVA with post hoc comparison and Holm–Sidak multiple testing correction. A p‐value ≤0.05 was considered statistically significant: *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001 and § p ≤ 0.1 considered a significant trend

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