Research ReportCortical expression of nuclear factor κB after human brain contusion
Introduction
Focal traumatic brain injury (TBI) induces primary neuronal degeneration in the site of injury and initiates a series of cellular and molecular events that lead to protracted secondary damage in the surrounding tissue and distally connected areas (Knoblach et al., 1999, Holmin et al., 1997, Holmin et al., 1998). In base to experimental studies performed in rodents, it is now known that secondary damage is exacerbated by the inflammatory response, commonly mediated and magnified by the upregulation or de novo expression of different genes (Yang et al., 1994, Yang et al., 1995, Hang et al., 2005, Arvin et al., 1995, Isaksson et al., 1997). Several studies have shown that the nuclear factor kappa B (NF-κB) family (cRel, RelA/p65, Rel B, p50 and p52) of transcription factor is one of the most important modulators of inflammatory gene expression in the nervous system as well as in non-neural tissues (Grilli and Memo, 1999, Baeuerle and Baltimore, 1996). NF-κB is one of the most important modulators of stress and inflammatory gene expression in the nervous system (CNS) and may be important determinants of cell death and disease of the CNS (Baeuerle and Baltimore, 1996, Salminen et al., 1995). NF-κB upregulation has been demonstrated in neurons and glial cells in response to experimental injury and neuropathological disorders, where it has been related to both neurodegenerative and neuroprotective activities (Yang et al., 1995, Hang et al., 2005, O'Neill and Kaltschmidt, 1997, Mattson et al., 2000, Kaltschmidt et al., 1994a, Bales et al., 1998, Nonaka et al., 1999). Activation of NF-κB in neurons has been generally involved in synaptic plasticity, neuronal function, development and survival (O'Neill and Kaltschmidt, 1997, Mattson et al., 2000), and in glial cells it has been mainly related to the inflammatory response (Kaltschmidt et al., 1994a, Bales et al., 1998) through induction of expression of proinflammatory genes such as TNF-α, IL-1β, IL-6, cyclooxygenase-2 (Cox-2), iNOS and adhesion molecules (VCAM-1 and ICAM-1) (Grilli and Memo, 1999).
Prototypical NF-κB heterodimers contain p50 and p65 subunits complexed with an IκB inhibitory protein to maintain an inactive cytosolic complex. Appropriate intracellular signals induce phosphorylation of IκB by an IκB kinase (IKK)-containing signalsome, dissociation of the inhibitory protein and its consequent degradation. The resulting nuclear migration of p65 and p50 modulates target gene expression by binding cognate GGGRNNYYCC DNA promoter sequences. Among the inducible transcription factors, members of the NF-κB family have been shown to play a role in the regulation of genes in the glial cells and neurons (Lipton, 1997). Some experiments have shown that p50 is involved in an adaptive response of neurons that survive injury (Meberg et al., 1996, Pennypacker et al., 2001). Selective activation of distinct subunits intrinsic to particular models of neurodegeneration may indeed promote cell death, with alternative dimer combinations conferring neuroprotection. However, till now it is unclear about the cellular localization of NF-κB subunits p65 and p50 in the different cell types and whether it changes among time in the human brain after TBI.
In this sense, in the last decade several studies have proposed a role of NF-κB activation in the degenerative and inflammatory response occurring after TBI in the rat brain (Yang et al., 1995, Nonaka et al., 1999, Nomoto et al., 2001). Till now, no study was found in the literature to investigate the NF-κB expression in the human brain after TBI. Therefore, little is known about the involvement of this transcription factor in the evolution of TBI-induced inflammatory response in the human brain contusion. Our previous studies have demonstrated that cortical brain damage is associated with increased activity of NF-κB and proinflammatory cytokine (Hang et al., 2004, Hang et al., 2005); therefore, the study of NF-κB along the evolution of TBI in the human brain may provide some insights into the importance of this signal transduction pathway in the degenerative/regenerative events that underlie poor brain lesion outcome. In this regard, the aim of the present study was to evaluate the binding activity and temporal and cellular expression pattern of NF-κB following a lesion biopsy in the human brain contusion.
Section snippets
EMSA autoradiography of NF-κB DNA binding activity
EMSA autoradiography of NF-κB DNA binding activity of 18 injured brain samples in 17 patients was shown in Fig. 1. Low NF-κB binding activity (weak EMSA autoradiography) was found in the patients who were early operated (≤ 6 h). NF-κB binding activity was progressively upregulated with the time from initial brain injury to operation. As compared with ≤ 6 h group, NF-κB binding activity measured by EMSA was significantly increased in the late operated patients (12–48 h, P < 0.05; ≥ 48 h, P < 0.01). In
Discussion
In this clinical study, we have first reported that upregulation of NF-κB occurred at human contused brain. The NF-κB binding activity was time dependent, showing progressively increased after brain injury. A weak expression of NF-κB was observed in the early stage of trauma, and the maximal expression occurred after 48 h postinjury. Immunohistochemical study showed that NF-κB subunits were localized in different cells. p65 was expressed at both neuroglial cells and vascular epithelial cells,
Patients
This study was approved by Najing University's Medical Institutional Review Board. Eighteen contused brain tissue biopsies were obtained from 17 consecutive patients undergoing surgery for brain contusions 5–80 h after trauma (Table 1). One patient underwent 2 operations at 12 h and 26 h postinjury, respectively (cases 10 and 14). No anti-inflammatory drugs, hypotension/hypovolemia or hypoxia either before hospital admission or during surgery was used in all patients. During surgery, following
Acknowledgments
We would like to thank Dr. Genbao Feng and Bo Wu for their technical assistance.
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