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Anti-Inflammatory Mechanism of Neural Stem Cell Transplantation in Spinal Cord Injury

View Article: PubMed Central - PubMed

ABSTRACT

Neural stem cell (NSC) transplantation has been proposed to promote functional recovery after spinal cord injury. However, a detailed understanding of the mechanisms of how NSCs exert their therapeutic plasticity is lacking. We transplanted mouse NSCs into the injured spinal cord seven days after SCI, and the Basso Mouse Scale (BMS) score was performed to assess locomotor function. The anti-inflammatory effects of NSC transplantation was analyzed by immunofluorescence staining of neutrophil and macrophages and the detection of mRNA levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6) and interleukin-12 (IL-12). Furthermore, bone marrow-derived macrophages (BMDMs) were co-cultured with NSCs and followed by analyzing the mRNA levels of inducible nitric oxide synthase (iNOS), TNF-α, IL-1β, IL-6 and IL-10 with quantitative real-time PCR. The production of TNF-α and IL-1β by BMDMs was examined using the enzyme-linked immunosorbent assay (ELISA). Transplanted NSCs had significantly increased BMS scores (p < 0.05). Histological results showed that the grafted NSCs migrated from the injection site toward the injured area. NSCs transplantation significantly reduced the number of neutrophils and iNOS+/Mac-2+ cells at the epicenter of the injured area (p < 0.05). Meanwhile, mRNA levels of TNF-α, IL-1β, IL-6 and IL-12 in the NSCs transplantation group were significantly decreased compared to the control group. Furthermore, NSCs inhibited the iNOS expression of BMDMs and the release of inflammatory factors by macrophages in vitro (p < 0.05). These results suggest that NSC transplantation could modulate SCI-induced inflammatory responses and enhance neurological function after SCI via reducing M1 macrophage activation and infiltrating neutrophils. Thus, this study provides a new insight into the mechanisms responsible for the anti-inflammatory effect of NSC transplantation after SCI.

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Related in: MedlinePlus

NSC transplantation inhibited macrophages’ activation after spinal cord injury. One week post-transplantation, the injured site was double-stained for iNOS and Mac-2 via immunofluorescent staining. (A) Representative images of spinal cord sections immunostained for iNOS (red) and Mac-2 (green) at the injury epicenter; (B) quantitative analysis of iNOS and Mac-2 double-positive cells by ImageJ (n = 5). Results are displayed as the mean ± standard error. * p < 0.05. Scale bar: 20 μm.
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ijms-17-01380-f004: NSC transplantation inhibited macrophages’ activation after spinal cord injury. One week post-transplantation, the injured site was double-stained for iNOS and Mac-2 via immunofluorescent staining. (A) Representative images of spinal cord sections immunostained for iNOS (red) and Mac-2 (green) at the injury epicenter; (B) quantitative analysis of iNOS and Mac-2 double-positive cells by ImageJ (n = 5). Results are displayed as the mean ± standard error. * p < 0.05. Scale bar: 20 μm.

Mentions: To evaluate the effects of NSC treatment on macrophages and neutrophils after SCI, we identified Mac-2-positive microglia/macrophages and anti-neutrophil-positive neutrophils by immunofluorescence staining at seven days after transplantation. As reported, M1 (iNOS-positive) macrophages, pro-inflammatory macrophages, could lead to further damage following primary mechanical injury by secreting inflammatory cytokines, such as TNF-α and chemokines [22]. To determine whether NSC transplantation could promote functional recovery following SCI via regulating the activation of macrophages, we detected the expression of macrophages’ activation marker iNOS. The activated M1 macrophages were identified by double Mac-2 and iNOS staining, and the mean density of the INOS+/Mac-2+ signal was significantly reduced in the NSC group (0.10 ± 0.020) compared to the control group (0.06 ± 0.015) (p < 0.05; Figure 4). It suggested that NSC transplantation was able to inhibit the activation of the M1 macrophage, which has been reported to be capable of inhibiting axon extension and to be neurotoxic.


Anti-Inflammatory Mechanism of Neural Stem Cell Transplantation in Spinal Cord Injury
NSC transplantation inhibited macrophages’ activation after spinal cord injury. One week post-transplantation, the injured site was double-stained for iNOS and Mac-2 via immunofluorescent staining. (A) Representative images of spinal cord sections immunostained for iNOS (red) and Mac-2 (green) at the injury epicenter; (B) quantitative analysis of iNOS and Mac-2 double-positive cells by ImageJ (n = 5). Results are displayed as the mean ± standard error. * p < 0.05. Scale bar: 20 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5037660&req=5

ijms-17-01380-f004: NSC transplantation inhibited macrophages’ activation after spinal cord injury. One week post-transplantation, the injured site was double-stained for iNOS and Mac-2 via immunofluorescent staining. (A) Representative images of spinal cord sections immunostained for iNOS (red) and Mac-2 (green) at the injury epicenter; (B) quantitative analysis of iNOS and Mac-2 double-positive cells by ImageJ (n = 5). Results are displayed as the mean ± standard error. * p < 0.05. Scale bar: 20 μm.
Mentions: To evaluate the effects of NSC treatment on macrophages and neutrophils after SCI, we identified Mac-2-positive microglia/macrophages and anti-neutrophil-positive neutrophils by immunofluorescence staining at seven days after transplantation. As reported, M1 (iNOS-positive) macrophages, pro-inflammatory macrophages, could lead to further damage following primary mechanical injury by secreting inflammatory cytokines, such as TNF-α and chemokines [22]. To determine whether NSC transplantation could promote functional recovery following SCI via regulating the activation of macrophages, we detected the expression of macrophages’ activation marker iNOS. The activated M1 macrophages were identified by double Mac-2 and iNOS staining, and the mean density of the INOS+/Mac-2+ signal was significantly reduced in the NSC group (0.10 ± 0.020) compared to the control group (0.06 ± 0.015) (p < 0.05; Figure 4). It suggested that NSC transplantation was able to inhibit the activation of the M1 macrophage, which has been reported to be capable of inhibiting axon extension and to be neurotoxic.

View Article: PubMed Central - PubMed

ABSTRACT

Neural stem cell (NSC) transplantation has been proposed to promote functional recovery after spinal cord injury. However, a detailed understanding of the mechanisms of how NSCs exert their therapeutic plasticity is lacking. We transplanted mouse NSCs into the injured spinal cord seven days after SCI, and the Basso Mouse Scale (BMS) score was performed to assess locomotor function. The anti-inflammatory effects of NSC transplantation was analyzed by immunofluorescence staining of neutrophil and macrophages and the detection of mRNA levels of tumor necrosis factor-&alpha; (TNF-&alpha;), interleukin-1&beta; (IL-1&beta;), interleukin-6 (IL-6) and interleukin-12 (IL-12). Furthermore, bone marrow-derived macrophages (BMDMs) were co-cultured with NSCs and followed by analyzing the mRNA levels of inducible nitric oxide synthase (iNOS), TNF-&alpha;, IL-1&beta;, IL-6 and IL-10 with quantitative real-time PCR. The production of TNF-&alpha; and IL-1&beta; by BMDMs was examined using the enzyme-linked immunosorbent assay (ELISA). Transplanted NSCs had significantly increased BMS scores (p &lt; 0.05). Histological results showed that the grafted NSCs migrated from the injection site toward the injured area. NSCs transplantation significantly reduced the number of neutrophils and iNOS+/Mac-2+ cells at the epicenter of the injured area (p &lt; 0.05). Meanwhile, mRNA levels of TNF-&alpha;, IL-1&beta;, IL-6 and IL-12 in the NSCs transplantation group were significantly decreased compared to the control group. Furthermore, NSCs inhibited the iNOS expression of BMDMs and the release of inflammatory factors by macrophages in vitro (p &lt; 0.05). These results suggest that NSC transplantation could modulate SCI-induced inflammatory responses and enhance neurological function after SCI via reducing M1 macrophage activation and infiltrating neutrophils. Thus, this study provides a new insight into the mechanisms responsible for the anti-inflammatory effect of NSC transplantation after SCI.

No MeSH data available.


Related in: MedlinePlus