Chun-Hua Hang

Inhibitory Effect on Cerebral Inflammatory Response following Traumatic Brain Injury in Rats: A Potential Neuroprotective Mechanism of N-Acetylcysteine

Although N-acetylcysteine (NAC) has been shown to be neuroprotective for traumatic brain injury (TBI), the mechanisms for this beneficial effect are still poorly understood. Cerebral inflammation plays an important role in the pathogenesis of secondary brain injury after TBI. However, it has not been investigated whether NAC modulates TBI-induced cerebral inflammatory response. In this work, we investigated the effect of NAC administration on cortical expressions of nuclear factor kappa B (NF-κB) and inflammatory proteins such as interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and intercellular adhesion molecule-1 (ICAM-1) after TBI. As a result, we found that NF-κB, proinflammatory cytokines, and ICAM-1 were increased in all injured animals. In animals given NAC post-TBI, NF-κB, IL-1β, TNF-α, and ICAM-1 were decreased in comparison to vehicle-treated animals. Measures of IL-6 showed no change after NAC treatment. NAC administration reduced brain edema, BBB permeability, and apoptotic index in the injured brain. The results suggest that post-TBI NAC administration may attenuate inflammatory response in the injured rat brain, and this may be one mechanism by which NAC ameliorates secondary brain damage following TBI.

Simvastatin reduces secondary brain injury caused by cortical contusion in rats: Possible involvement of TLR4/NF-κB pathway

Simvastatin, a cholesterol-lowering agent, has demonstrated neuroprotective effects against brain injury, but the underlying mechanisms remain unclear. This study was undertaken to evaluate the effect of simvastatin on the Toll-like receptor 4 (TLR4) and nuclear factor-kappa B (NF-kappaB) related signaling pathway and secondary brain injury in rats after traumatic brain injury (TBI). Adult male Wistar rats were divided into four groups: (1) Sham group (n=25); (2) Sham+vehicle group (n=25); (3) TBI+vehicle group (n=30); and (4) TBI+simvastatin group (n=30). Right parietal cortical contusion was made by using a weight-dropping method. In TBI+simvastatin group, simvastatin was administered orally at a dose of 37.5 mg/kg at 1 and 6 h after TBI. Brain samples were extracted at 24 h after trauma. As a result, we found that treatment with simvastatin markedly inhibited the mRNA and protein expressions of TLR4, NF-kappaB and the downstream inflammatory agents, such as interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), and intercellular adhesion molecule-1 (ICAM-1). Administration of simvastatin following TBI significantly ameliorated the secondary brain damage, such as cortical apoptosis, brain edema, blood-brain barrier (BBB) impairment, and motor deficits. In conclusion, post-TBI simvastatin administration may attenuate TLR4/NF-kappaB-mediated inflammatory response in the injured rat brain, and this may be one mechanism by which simvastatin improves outcome following TBI.

Effect of systemic LPS injection on cortical NF-κB activity and inflammatory response following traumatic brain injury in rats

The aim of current study is to investigate the effect of systemic administration of lipopolysaccharide (LPS) on the temporal pattern of cortical nuclear factor kappa B (NF-kappaB) binding activity, inflammatory response and secondary damage in the injured brain following traumatic brain injury (TBI). Right parietal cortical contusion in rats was made by using weight-dropping method. The rats were randomly divided into sham, LPS, TBI and TBI-LPS groups, with LPS injected intraperitoneally. NF-kappaB binding activity, cytokines, intercellular adhesion molecule-1 (ICAM-1) and brain damage were detected by electrophoretic mobility shift assay (EMSA), enzyme-linked immunosorbent assay (ELISA), immunohistochemistry and terminal deoxynucleotidyl-transferase-mediated biotin-dUTP nick end labeling (TUNEL) apoptosis, respectively. The results showed that systemic administration of LPS following TBI could induce an immediate, strong and persistent upregulation of NF-kappaB, tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and ICAM-1 in the area surrounding the injured brain. As compared with rats of sham, LPS and TBI groups, NF-kappaB binding activity, TNF-alpha and IL-6 were significantly upregulated in the surrounding cortex of injured site as early as 3 h postinjury when challenged with LPS, kept at high level up to 7-days postinjury. ICAM-1-positive vessels and apoptotic TUNEL-positive cells in the injured brain were also significantly increased in TBI-LPS rats. It was concluded that inflammatory response and secondary brain damage occurred in the injured brain could be highly exacerbated by endotoxemia.

Potential contribution of nuclear factor-κB to cerebral vasospasm after experimental subarachnoid hemorrhage in rabbits

Nuclear factor-κB (NF-κB) plays a key role in inflammation, which is involved in the development of cerebral vasospasm after subarachnoid hemorrhage (SAH). In the present study, we assessed the potential role of NF-κB in regulation of cerebral vasospasm. Nuclear factor-κB DNA-binding activity was measured in cultured vascular smooth muscle cells (VSMCs) treated with hemolysate and pyrrolidine dithiocarbamate (PDTC, 80 μmol/L), an inhibitor of NF-κB. Forty-two rabbits were divided into three groups: control, SAH, and PDTC groups (n = 14 for each group). The caliber of the basilar artery was evaluated. Nuclear factor-κB DNA-binding activity and the gene expression levels of cytokines and adhesion molecules in the basilar artery were measured. Immunohistochemical study was performed to assess the expression and localization of tumor necrosis factor (TNF)-α, intercellular adhesion molecule (ICAM)-1, and myeloperoxidase (MPO). It was observed that NF-κB DNA-binding activity was significantly increased by treatment with hemolysate in cultured VSCMs, but this increase was suppressed by pretreatment with PDTC. Severe vasospasm was observed in the SAH group, which was attenuated in the PDTC group. Subarachnoid hemorrhage could induce increases of NF-κB DNA-binding activity and the gene expression levels of TNF-α, interleukin (IL)-1β, ICAM-1, and vascular cell adhesion molecule (VCAM)-1, which were reduced in the PDTC group. Immunohistochemical study demonstrated that the expression levels of TNF-α, ICAM-1, and MPO were all increased in the SAH group, but these increases were attenuated in the PDTC group. Our results suggest that NF-κB is activated in the arterial wall after SAH, which potentially leads to vasospasm development through induction of inflammatory response.

Comparison between one- and two-hemorrhage models of cerebral vasospasm in rabbits.

Injection of blood into the cisterna magna is one of the most frequently used methods to produce subarachnoid hemorrhage (SAH) models in animals. Although the two-hemorrhage model of vasospasm is frequently used in canine and rat models, most studies with rabbits only use the one-hemorrhage model. In the present study, we accomplished a side-by-side comparison between one- and two-hemorrhage models in rabbits. A total of 38 rabbits were randomly divided into three groups, i.e. control group (n = 5), one (n = 15)- and two (n = 18)-hemorrhage model groups. The degree of cerebral vasospasm, the time course of cerebral vasospasm, the clinical behavior, and the residual amount of subarachnoid blood clots were measured on days 3, 5 and 7 after the establishment of the models. Compared with one-hemorrhage model, the time course of vasospasm in the two-hemorrhage model was more coincident with that observed in humans, produced more severe vasospasm after SAH, and had an acceptable low mortality. In conclusion, the two-hemorrhage model in rabbits is more appropriate than the one-hemorrhage model for the research on SAH or cerebral vasospasm, and thus can be used for the investigation of the mechanisms of and therapeutic approaches for cerebral vasospasm.

Effects of recombinant human erythropoietin (rhEPO) on JAK2/STAT3 pathway and endothelial apoptosis in the rabbit basilar artery after subarachnoid hemorrhage

Previous studies have shown that recombinant human erythropoietin (rhEPO) can attenuate the degree of cerebral vasospasm following experimental subarachnoid hemorrhage (SAH). However, the mechanisms for this beneficial effect are still poorly understood. SAH-induced endothelial apoptosis may trigger, aggravate, and maintain cerebral vasospasm. We, therefore, tried to analyze whether rhEPO administration influenced the endothelial cell apoptosis in the basilar artery after SAH. Another aim of the current study was to investigate the modulation of rhEPO on the activity of Janus kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3), which played an important role in the signaling of apoptosis. A total of 48 rabbits were randomly divided into four groups; control group, SAH group, SAH+vehicle group, and SAH+rhEPO group. All SAH animals were subjected to injection of autologous blood into cisterna magna twice on day 0 and day 2. The rhEPO was administered i.p. starting 5 min after the induction of SAH on day 0 and repeated every 8 h for 120 h. The basilar arteries were extracted on day 5 after SAH. As a result, we found that administration of rhEPO could activate JAK2 and STAT3 in the basilar artery and decrease the apoptosis index of endothelial cells following SAH. Moreover, the anti-apoptotic genes such as bcl-2 and bcl-xL were up-regulated after the injections of rhEPO. In conclusion, the therapeutic effect of rhEPO on the subsequent vasospasm after SAH may relate to its inhibition on the endothelial apoptosis in the cerebral arteries, which may be mediated in part by JAK2/STAT3 signaling pathway.

Expression of myeloid differentiation primary response protein 88 (Myd88) in the cerebral cortex after experimental traumatic brain injury in rats

A growing body of evidence indicates that Toll-like receptors (TLRs) and Interleukin-1 (IL-1) family have been shown to be involved in the damaging inflammatory processes associated with stroke, infection, neoplasia, and other diseases in the central nervous system. Myeloid differentiation primary response protein 88 (Myd88) is a critical adaptor protein that transmits signals for TLRs and IL-1 family. Therefore, this study aimed to detect the expression of Myd88 protein and mRNA in a rat weight-dropping trauma model and to clarify the role of Myd88 after traumatic brain injury (TBI). A total of fifty-four Sprague Dawley (SD) rats were randomly divided into control group and TBI groups at hours 6, 12 and on day 1, day 2, day 3, and day 7. The TBI groups suffered experimental TBI by improved Feeney model. Myd88 expression is measured by Reverse Transcription PCR (RT-PCR), Western blot analysis and immunohistochemistry; and nuclear factor-kappaB (NF-κB) binding activity by electrophoretic mobility shift assay (EMSA); The levels of tumor necrosis factor-α (TNF-α) and Interleukin 1β (IL-1β) were measured by enzyme linked immunosorbent assay (ELISA) and the intercellular adhesion molecule-1 (ICAM-1) expression by immunohistochemistry. The expression of Myd88 in the injured brain was dramatically increased through 6 h and 7 days postinjury, and peaked on 3days. NF-κB, TNF-α, IL-1β and ICAM-1 also ascended significantly after TBI. Our data demonstrated that Myd88 was increasingly expressed in a parallel time course to the up-regulation of NF-κB, proinflammatory cytokines and ICAM-1 and there was a highly positive relationship among them. These findings might have important implications during the administration of specific Myd88 antagonists in order to prevent or reduce inflammatory response after TBI.

Progesterone prevents traumatic brain injury-induced intestinal nuclear factor kappa B activation and proinflammatory cytokines expression in male rats.

We have previously shown that traumatic brain injury (TBI) can induce an upregulation of nuclear factor kappa B (NF-κB) and proinflammatory cytokines in the gut, which play an important role in the pathogenesis of acute gut mucosal injury mediated by inflammation. In this work, we investigated whether progesterone administration modulated intestinal NF-κB activity and proinflammatory cytokines expression after TBI in male rats. As a result, we found that administration of progesterone following TBI could decrease NF-κB binding activity, NF-κB p65 protein expression, and concentrations of interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α) in the gut. TBI-induced damages of gut structure were ameliorated after progesterone injections. The results of the present study suggest that the therapeutic benefit of post-TBI progesterone injections might be due to its inhibitory effects on intestinal NF-κB activation and proinflammatory cytokines expression.

Intraoperative indocyanine green angiography in intracranial aneurysm surgery: Microsurgical clipping and revascularization.

BACKGROUND The goal of this report is to illustrate the use of intraoperative indocyanine green (ICG) angiography in the surgical management of intracranial aneurysms, including microsurgical clipping and revascularization. METHODS This study included a series of 45 patients who were surgically treated between June 2007 and May 2008 for intracranial aneurysms. Fourty-three of the patients had anterior circulation aneurysms, and 2 had posterior circulation aneurysms. Forty-one patients were treated with microsurgical clipping. Four patients underwent revascularization combined with aneurysm dissection or trapping. Intraoperative ICG angiography was used to visualize the aneurysm clipping, patency of parent artery or graft. The ICG angiography technique is described, with particular reference to evaluation of the aneurysm clipping and revascularization. RESULTS Eighty-nine ICG angiography procedures were performed in 45 patients with intracranial aneurysms. The aneurysms were completely obliterated for all patients, and the grafts were patented for all except 1 patient. Pre-clipping ICG angiography showed the relationship of aneurysm and its parent artery clearly. After aneurysms being clipped, intraoperative ICG angiography found remnant of aneurysms, stenosis or occlusion of parent arteries and grafts in 8 cases, which were revised in the same surgical procedure. The results of ICG angiography correlated well with postoperative DSA in 97% patients. CONCLUSION ICG angiography can provide real-time information and guide revision in the same surgical procedure for the management of intracranial aneurysms.

Expression of Toll-like receptor 4 in the brain in a rabbit experimental subarachnoid haemorrhage model.

Abstract.Objective:To investigate the expression of the Toll-like receptor (TLR) 4 in the brain after experimental subarachnoid haemorrhage (SAH) in rabbits.Methods:A total of 52 rabbits were randomly divided into four groups: control group; day 3, day 5, and day 7 groups. Day 3, day 5, and day 7 groups were all SAH groups in which the rabbits were killed on day 3, 5, and 7, respectively. In SAH groups, autologous arterial blood was injected into cisterna magna twice on day 0 and day 2. Immunostaining and immunoblotting experiments were performed to detect the expression of TLR4 protein. Reverse-transcriptase polymerase chain reaction was used to analyze the presence and quantity of TLR4 mRNA.Results:The expressions of TLR4 protein and mRNA were increased remarkably in SAH groups compared with the control group. The immunohistochemical staining demonstrated high level expression of TLR4 was present mainly in the endothelial cells of capillaries in the brain.Conclusion:Our results indicate that TLR4 expression is upregulated in the brain after experimental SAH.