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Interleukin ‐ 23 Secreted by Activated Macrophages Drives γ δ T Cell Production of Interleukin ‐ 17 to Aggravate Secondary Injury After Intracerebral Hemorrhage

View Article: PubMed Central - PubMed

ABSTRACT

Background: Neuroinflammation plays a key role in intracerebral hemorrhage (ICH)–induced secondary brain injury, but the specific roles of peripheral inflammatory cells such as macrophages and lymphocytes remain unknown. The purpose of this study was to explore the roles of macrophages, T lymphocytes, and the cytokines they secrete as potential targets for treating secondary brain injury after ICH.

Methods and results: Our results showed that peripheral macrophages and T lymphocytes successively infiltrated the brain, with macrophage counts peaking 1 day after ICH and T‐lymphocyte counts peaking after 4 days. These peaks in cellular infiltration corresponded to increases in interleukin (IL)‐23 and IL‐17 expression, respectively. We found that hemoglobin from the hematoma activated IL‐23 secretion by infiltrating macrophages by inducing the formation of toll‐like receptor (TLR) 2/4 heterodimer. This increased IL‐23 expression stimulated γδT‐cell production of IL‐17, which increased brain edema and neurologic deficits in the model mice as a proinflammatory factor. Finally, we found that sparstolonin B (SsnB) could ameliorate brain edema and neurologic deficits in ICH model mice via inhibition of TLR2/TLR4 heterodimer formation, and notably, SsnB interacted with myeloid differentiation factor 88 Arg196.

Conclusions: Together, our results reveal the importance of the IL‐23/IL‐17 inflammatory axis in secondary brain injury after ICH and thus provide a new therapeutic target for ICH treatment.

No MeSH data available.


Related in: MedlinePlus

Macrophage and T‐lymphocyte infiltration into mouse brain after intracerebral hemorrhage (ICH). A through F, Temporal changes in absolute numbers of different inflammatory cell types in hemorrhagic hemispheres at 1, 4, and 7 days after ICH. Data were obtained for cells pooled from 6 mice, and the experiments were repeated 3 times. *P<0.05, **P<0.01 vs sham. G, Representative fluorescence microscopy images showing infiltrating F4/80+ cells in the perihematoma area at 1 day after ICH (blue=4′‐6‐diamidino‐2‐phenylindole [DAPI], red=F4/80, scale bars=100 μm). H, Representative fluorescence microscopy images showing CD3+ cell infiltration into the perihematoma area at 4 days after ICH (blue=DAPI, green=CD3, scale bars=100 μm).
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jah31823-fig-0001: Macrophage and T‐lymphocyte infiltration into mouse brain after intracerebral hemorrhage (ICH). A through F, Temporal changes in absolute numbers of different inflammatory cell types in hemorrhagic hemispheres at 1, 4, and 7 days after ICH. Data were obtained for cells pooled from 6 mice, and the experiments were repeated 3 times. *P<0.05, **P<0.01 vs sham. G, Representative fluorescence microscopy images showing infiltrating F4/80+ cells in the perihematoma area at 1 day after ICH (blue=4′‐6‐diamidino‐2‐phenylindole [DAPI], red=F4/80, scale bars=100 μm). H, Representative fluorescence microscopy images showing CD3+ cell infiltration into the perihematoma area at 4 days after ICH (blue=DAPI, green=CD3, scale bars=100 μm).

Mentions: After induction of ICH in WT C57BL/6 mice, the temporal changes in the numbers of different types of inflammatory cells in the hemorrhagic hemisphere were analyzed by FACS. Microglia and macrophages were gated according to the fluorescence intensity of CD45 and CD11b staining. The CD45high group was gated and further analyzed for the expression of CD3 and CD4. The CD45high CD3+ cells were gated and further analyzed for the expression of γδT (Figure S1). We observed the highest absolute numbers of CD45+ cells (Figure 1A), including T lymphocytes (CD3+ cells; Figure 1B), CD4+ T lymphocytes (Figure 1C), and γδT lymphocytes (Figure 1D), on day 4 after ICH, while the number of macrophages peaked at day 1 (Figure 1E). In contrast, no changes in the numbers of microglia were observed after ICH (Figure 1F). In confirmation of these FACS results, immunofluorescence staining showed that F4/80+ (macrophages/microglia; Figure 1G) and CD3+ cells (T lymphocyte; Figure 1H) were located along the perihematoma area after ICH.


Interleukin ‐ 23 Secreted by Activated Macrophages Drives γ δ T Cell Production of Interleukin ‐ 17 to Aggravate Secondary Injury After Intracerebral Hemorrhage
Macrophage and T‐lymphocyte infiltration into mouse brain after intracerebral hemorrhage (ICH). A through F, Temporal changes in absolute numbers of different inflammatory cell types in hemorrhagic hemispheres at 1, 4, and 7 days after ICH. Data were obtained for cells pooled from 6 mice, and the experiments were repeated 3 times. *P<0.05, **P<0.01 vs sham. G, Representative fluorescence microscopy images showing infiltrating F4/80+ cells in the perihematoma area at 1 day after ICH (blue=4′‐6‐diamidino‐2‐phenylindole [DAPI], red=F4/80, scale bars=100 μm). H, Representative fluorescence microscopy images showing CD3+ cell infiltration into the perihematoma area at 4 days after ICH (blue=DAPI, green=CD3, scale bars=100 μm).
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jah31823-fig-0001: Macrophage and T‐lymphocyte infiltration into mouse brain after intracerebral hemorrhage (ICH). A through F, Temporal changes in absolute numbers of different inflammatory cell types in hemorrhagic hemispheres at 1, 4, and 7 days after ICH. Data were obtained for cells pooled from 6 mice, and the experiments were repeated 3 times. *P<0.05, **P<0.01 vs sham. G, Representative fluorescence microscopy images showing infiltrating F4/80+ cells in the perihematoma area at 1 day after ICH (blue=4′‐6‐diamidino‐2‐phenylindole [DAPI], red=F4/80, scale bars=100 μm). H, Representative fluorescence microscopy images showing CD3+ cell infiltration into the perihematoma area at 4 days after ICH (blue=DAPI, green=CD3, scale bars=100 μm).
Mentions: After induction of ICH in WT C57BL/6 mice, the temporal changes in the numbers of different types of inflammatory cells in the hemorrhagic hemisphere were analyzed by FACS. Microglia and macrophages were gated according to the fluorescence intensity of CD45 and CD11b staining. The CD45high group was gated and further analyzed for the expression of CD3 and CD4. The CD45high CD3+ cells were gated and further analyzed for the expression of γδT (Figure S1). We observed the highest absolute numbers of CD45+ cells (Figure 1A), including T lymphocytes (CD3+ cells; Figure 1B), CD4+ T lymphocytes (Figure 1C), and γδT lymphocytes (Figure 1D), on day 4 after ICH, while the number of macrophages peaked at day 1 (Figure 1E). In contrast, no changes in the numbers of microglia were observed after ICH (Figure 1F). In confirmation of these FACS results, immunofluorescence staining showed that F4/80+ (macrophages/microglia; Figure 1G) and CD3+ cells (T lymphocyte; Figure 1H) were located along the perihematoma area after ICH.

View Article: PubMed Central - PubMed

ABSTRACT

Background: Neuroinflammation plays a key role in intracerebral hemorrhage (ICH)&ndash;induced secondary brain injury, but the specific roles of peripheral inflammatory cells such as macrophages and lymphocytes remain unknown. The purpose of this study was to explore the roles of macrophages, T lymphocytes, and the cytokines they secrete as potential targets for treating secondary brain injury after ICH.

Methods and results: Our results showed that peripheral macrophages and T lymphocytes successively infiltrated the brain, with macrophage counts peaking 1&nbsp;day after ICH and T&#8208;lymphocyte counts peaking after 4&nbsp;days. These peaks in cellular infiltration corresponded to increases in interleukin (IL)&#8208;23 and IL&#8208;17 expression, respectively. We found that hemoglobin from the hematoma activated IL&#8208;23 secretion by infiltrating macrophages by inducing the formation of toll&#8208;like receptor (TLR) 2/4 heterodimer. This increased IL&#8208;23 expression stimulated &gamma;&delta;T&#8208;cell production of IL&#8208;17, which increased brain edema and neurologic deficits in the model mice as a proinflammatory factor. Finally, we found that sparstolonin B (SsnB) could ameliorate brain edema and neurologic deficits in ICH model mice via inhibition of TLR2/TLR4 heterodimer formation, and notably, SsnB interacted with myeloid differentiation factor 88 Arg196.

Conclusions: Together, our results reveal the importance of the IL&#8208;23/IL&#8208;17 inflammatory axis in secondary brain injury after ICH and thus provide a new therapeutic target for ICH treatment.

No MeSH data available.


Related in: MedlinePlus