CD8 T cell-derived perforin aggravates secondary spinal cord injury through destroying the blood-spinal cord barrier

https://doi.org/10.1016/j.bbrc.2019.03.002Get rights and content

Highlights

  • CD8 T cell-derived perforin is detrimental to trauma spinal cord injury recovery.

  • Perforin disrupts tight junction proteins in the blood-spinal cord barrier.

  • Increased permeability of the barrier allows infiltration of inflammatory cytokines.

  • Secondary inflammation is detrimental to trauma spinal cord injury recovery.

  • Evidences from both an animal model and humans lead to same conclusion.

Abstract

Perforin plays an important role in autoimmune and infectious diseases, but its function in immune inflammatory responses after spinal cord injury (SCI) has received insufficient attention. The goal of this study is to determine the influence of perforin after spinal cord injury (SCI) on secondary inflammation. Compared recovery from SCI in perforin knockout (Prf1−/−) and wild-type(WT)mice, WT mice had significantly lower the Basso mouse score (BMS), CatWalk XT, and motor-evoked potentials (MEPs) than Prf1−/− mice. Spinal cord lesions were also more obvious through glial fibrillary acidic protein (GFAP), Nissl, and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining. Furthermore, the blood-spinal cord barrier (BSCB) disruption was more severe and inflammatory cytokine levels were higher. Flow cytometry indicated that perforin mainly originated from CD8 T cells. With flow cytometry and enzyme-linked immunosorbent assay (ELISA), human cerebrospinal fluid (CSF) yielded similar results. Together, this study firstly demonstrated that CD8 T cell-derived perforin is detrimental to SCI recovery in the mouse model. Mechanistically, this effect occurs because perforin increases BSCB permeability, causing inflammatory cells and related cytokines to infiltrate and disrupt the nervous system.

Introduction

Spinal cord injury (SCI) causes primary vascular damage and initiates a cascade of events that alters blood-spinal cord barrier (BSCB) permeability [1]. However, little attention has been directed at how BSCB integrity affects post-SCI regeneration processes. Interestingly, after disrupting tight junction (TJ) proteins, perforin can increase BSCB permeability [2], promote inflammatory cytokine entry from peripheral blood to the injury site, as well as aggravate secondary inflammation at and around the injury site in viral infections and autoimmune diseases [3,4]. As previously reported, perforin is primarily secreted by CD8 cytotoxic T cell [5]. In peripheral blood and the central nervous system (CNS), CD8 T cell-derived perforin participates in a series of inflammatory and immune responses [6]. Overly strong action from CD8 T cells can cause lethal inflammation, and removing them specifically reduces the chances of mice dying from an immune response [7,8]. Nevertheless, it is unknown whether perforin functions similarly during acute trauma to the spinal cord.

This study firstly reveals that CD8 T cell-derived perforin destroys BSCB post-SCI, thus recruiting inflammatory cytokines to the injury site and aggravating secondary SCI. The data presented here expand on the well known functions of CD8T cells and the current realization that these lymphocytes contribute to injuries thought to only trigger an innate immune response.

Section snippets

Animals

C57BL/6J, CD8 T knockout(CD8T−/−), and Prf−/− mice were purchased from Jackson Laboratory and propagated by the experimental animal center of Jinan University Medical College. All mice aged 7–8 weeks weighed 17–22 g during surgery. The center was not pathogen-controlled. All procedures were approved and implemented by the Commission for Animal Protection and Use of Jinan University.

Contusive SCI model

An experimental model of acute SCI was established using the New York University spinal cord hitter [9]. Mice were

CD8-derived perforin harms recovery of spinal cord function

More CD8 T cells and perforin appeared in the injured sites after SCI than in the sham operation group (Fig. S1). At 3 d post-SCI, BMS continued to rise before peaking in Week 6 post-SCI (Fig. 1 A), while we did not find differences in BMS of WT and Prf1−/− mice without SCI (Fig. S2). Furthermore, compared with WT mice, regularity index, stride length, and cadence in Prf1−/− mice increased significantly in CatWalk-assisted gait analysis at 6 wk post-SCI (Fig. 1B–D). Meanwhile, Electromyography

Discussion

Inflammatory disruptions are more pronounced in CNS than other tissues, because neurons and gliocytes have limited regenerative capacity, leading to irreversible dysfunction after injury [29]. Therefore, controlling targeted inflammation is a valuable method to promote post-SCI functional recovery [30].

Perforin is a good candidate for targeted therapy because the protein is involved in numerous inflammatory responses [31,32], including the CD8 T cell apoptosis pathway [33]. Indeed, here we

Acknowledgements

This work was supported by the Natural Science Foundation of Guangdong Province (2018A030313576), and the Natural Science Foundation of Guangzhou city (201803010001), China. Key Program of Traditional Chinese Medicine of Guangdong Province (20173018), China.

References (34)

  • Y. Yao et al.

    Antigen-specific CD8(+) T cell feedback activates NLRP3 inflammasome in antigen-presenting cells through perforin

    Nat. Commun.

    (2017)
  • A.J. Johnson et al.

    Antigen-specific CD8+ T cells mediate a peptide-induced fatal syndrome

    J. Immunol.

    (2005)
  • M.A. Huggins et al.

    Perforin expression by CD8 T cells is sufficient to cause fatal brain edema during experimental cerebral malaria

    Infect. Immun.

    (2017)
  • D.M. Basso et al.

    Basso Mouse Scale for locomotion detects differences in recovery after spinal cord injury in five common mouse strains

    J. Neurotrauma

    (2006)
  • G.C. Koopmans et al.

    The assessment of locomotor function in spinal cord injured rats: the importance of objective analysis of coordination

    J. Neurotrauma

    (2005)
  • G. Sun et al.

    γδ T cells provide the early source of IFN-γ to aggravate lesions in spinal cord injury

    J. Exp. Med.

    (2018)
  • J. Lee et al.

    Fluoxetine inhibits matrix metalloprotease activation and prevents disruption of blood-spinal cord barrier after spinal cord injury

    Brain

    (2012)
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