810nm弱激光照射对大鼠急性脊髓损伤后巨噬细胞极化的作用研究

本研究构建急性大鼠脊髓夹伤模型,并将大鼠随机分为单纯脊髓损伤对照组及脊髓损伤联合弱激光照射组。照射组应用810 nm波长,150 m W照射功率,照射光斑0.3 cm^2的弱激光对脊髓损伤区进行经皮照射,连续照射3天,7天或14天。应用免疫荧光、免疫印迹实验方法,测定脊髓损伤区巨噬细胞及小胶质细胞的极化表达。应用酶联免疫吸附法测定脊髓损伤区白细胞介素4的表达情况。应用坚牢蓝髓鞘染色测定两组损伤脊髓中髓鞘保留的差异。采用BBB评分对两组大鼠后肢运动功能的恢复进行评估。结果表明,810 nm弱激光对脊髓损伤区连续照射3天,7天后,可显著减少M1型巨噬细胞及其标志物诱导型一氧化氮合酶的表达,在7天时间增加M2型巨噬细胞及其标志物精氨酸酶1的表达。弱激光照射组白细胞介素4的表达明显增加。损伤后14天,弱激光照射组脊髓损伤区髓鞘保留面积比值明显提高。损伤后7天及14天时,弱激光照射组大鼠的BBB评分明显升高。该实验结果表明,810 nm弱激光经皮照射,可增加大鼠急性脊髓损伤区M2型巨噬细胞及小胶质细胞的表达,并减少脊髓损伤后的髓鞘脱失,促进脊髓损伤大鼠运动功能的恢复。     

Involvement of acidic fibroblast growth factor in spinal cord injury repair processes revealed by a proteomics approach

Acidic fibroblast growth factor (aFGF; also known as FGF-1) is a potent neurotrophic factor that affects neuronal survival in the injured spinal cord. However, the pathological changes that occur with spinal cord injury (SCI) and the attribution to aFGF of a neuroprotective effect during SCI are still elusive. In this study, we demonstrated that rat SCI, when treated with aFGF, showed significant functional recovery as indicated by the Basso, Beattie, and Bresnahan locomotor rating scale and the combined behavior score (p < 0.01-0.001). Furthermore proteomics and bioinformatics approaches were adapted to investigate changes in the global protein profile of the damaged spinal cord tissue when experimental rats were treated either with or without aFGF at 24 h after injury. We found that 51 protein spots, resolvable by two-dimensional PAGE, had significant differential expression. Using hierarchical clustering analysis, these proteins were categorized into five major expression patterns. Noticeably proteins involved in the process of secondary injury, such as astrocyte activation (glial fibrillary acidic protein), inflammation (S100B), and scar formation (keratan sulfate proteoglycan lumican), which lead to the blocking of injured spinal cord regeneration, were down-regulated in the contusive spinal cord after treatment with aFGF. We propose that aFGF might initiate a series of biological processes to prevent or attenuate secondary injury and that this, in turn, leads to an improvement in functional recovery. Moreover the quantitative expression level of these proteins was verified by quantitative real time PCR. Furthermore we identified various potential neuroprotective protein factors that are induced by aFGF and may be involved in the spinal cord repair processes of SCI rats. Thus, our results could have a remarkable impact on clinical developments in the area of spinal cord injury therapy.     

Fas and FasL Expression in the Spinal Cord Following Cord Hemisection in the Monkey

The changes of endogenous Fas/FasL in injured spinal cord, mostly in primates, are not well known. In this study, we investigated the temporal changes in the expression of Fas and FasL and explored their possible roles in the ventral horn of the spinal cord and associated precentral gyrus following T(11) spinal cord hemisection in the adult rhesus monkey. A significant functional improvement was seen with the time going on in monkeys subjected to cord hemisection. Apoptotic cells were also seen in the ventral horn of injured spinal cord with TUNEL staining, and a marked increase presents at 7 days post operation (dpo). Simultaneously, the number of Fas and FasL immunoreactive neurons in the spinal cords caudal and rostral to injury site and their intracellular optical density (OD) in the ipsilateral side of injury site at 7 dpo increased significantly more than that of control group and contralateral sides. This was followed by a decrease and returned to normal level at 60 dpo. No positive neurons were observed in precentral gyrus. The present results may provide some insights to understand the role of Fas/FasL in the spinal cord but not motor cortex with neuronal apoptosis and neuroplasticity in monkeys subjected to hemisection spinal cord injury.     

Expression of Glial Fibrillary Acidic Protein and Neurofilament mRNA in Gliosis Induced by Experimental Autoimmune Encephalomyelitis

We have previously shown that the content of glial fibrillary acidic protein (GFAP) gradually increases in the spinal cord of Lewis rats with acute experimental autoimmune encephalomyelitis (EAE), reaching a level 1.5-2 times greater than that in controls by 35 days postimmunization (dpi). We report here that the increase in GFAP mRNA level followed a completely different time course and reached higher levels relative to controls than did that of the protein. Total RNA was isolated using a modified version of current methods using phenol/chloroform extractions to ensure optimal recovery from spinal cord. Control animals yielded 323 +/- 35 micrograms (mean +/- SD; n = 34) of total RNA/spinal cord throughout the experimental period. EAE animals contained up to three times as much total RNA during 11-14 dpi, a finding largely reflecting the infiltration of inflammatory cells. By 65 dpi, total RNA levels closely approached control values. As early as 10 dpi, increased amounts of GFAP mRNA were detected in EAE animals relative to controls. During 11-14 dpi, GFAP mRNA levels reached six- to eightfold greater than values in controls and then slowly declined throughout the remainder of the time course, with a fourfold increase still detected at 65 dpi. However, coinciding with the height of inflammation and clinical signs at 12 dpi, the GFAP mRNA content dropped to approximately 50% of the level at 11 dpi but rose again at 13 dpi. This dip was mirrored by a similar decrease in neurofilament mRNA content, but otherwise the level of this message remained relatively constant and equal to that in controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

Alterations in the expression of the apurinic/apyrimidinic endonuclease-1/redox factor-1 (APE/ref-1) and DNA damage in the caudal region of acute and chronic spinal cord injured rats treated by embryonic neural stem cells

The oxidative mechanisms of injury-induced damage of neurons within the spinal cord are not very well understood. We used a model of T8-T9 spinal cord injury (SCI) in the rat to induce neuronal degeneration. In this spinal cord injury model, unilateral avulsion of the spinal cord causes oxidative stress of neurons. We tested the hypothesis that apurinic/apyrimidinic endonuclease (or redox effector factor-1, APE/Ref-1) regulates this neuronal oxidation mechanism in the spinal cord region caudal to the lesion, and that DNA damage is an early upstream signal. The embryonic neural stem cell therapy significantly decreased DNA-damage levels in both study groups - acutely (followed up to 7 days after SCI), and chronically (followed up to 28 days after SCI) injured animals. Meanwhile, mRNA levels of APE/Ref-1 significantly increased after embryonic neural stem cell therapy in acutely and chronically injured animals when compared to acute and chronic sham groups. Our data has demonstrated that an increase of APE/Ref-1 mRNA levels in the caudal region of spinal cord strongly correlated with DNA damage after traumatic spinal cord injury. We suggest that DNA damage can be observed both in lesional and caudal regions of the acutely and chronically injured groups, but DNA damage is reduced with embryonic neural stem cell therapy.     

A neutrophil elastase inhibitor (ONO-5046) reduces neurologic damage after spinal cord injury in rats

In view of a cytoprotective effect of elastase inhibitor on chemokine-mediated tissue injury, we examined the neuroprotective effect of ONO-5046, a specific inhibitor of neutrophil elastase, in rats with spinal cord injury. Standardized spinal cord compression markedly increased cytokine-induced neutrophil chemo-attractant (CINC)-1 mRNA and protein. Their increases correlated with neurologic severity of injured rats. Immunohistochemically, CINC-1 protein was detected sequentially in vascular endothelial cells at 4 h, in perivascular neutrophils at 8 h, and in neutrophils infiltrating into cord substance at 12 h. Pretreatment with ONO-5046 (50 mg/kg) markedly ameliorated motor disturbance in injured rats, and reduced CINC-1 protein and mRNA expression. ONO-5046 also significantly reduced the increase of neutrophil accumulation or infiltration estimated by myeloperoxidase activity, and the extent of vascular permeability by Evans blue extravasation in the injured cord segment in comparison to control animals receiving vehicle. These results suggest that CINC-1 contributed to inflammation in rat spinal cord injury and ONO-5046 attenuated neurologic damage partly by blocking CINC-1 production of the chemoattractant, preventing neutrophil activation and vascular endothelial cell injury.     

Spatial and temporal relationship between monocyte chemoattractant protein-1 expression and spinal glial activation following peripheral nerve injury

Peripheral nerve injury can induce spinal microglial/astrocyte activation. Substances released by activated glial cells excite spinal nociceptive neurons. Pharmacological disruption of glial activation or antagonism of substances released by activated glia prevent or reverse pain hypersensitivity. It is not known, however, what causes spinal cord glia to shift from a resting to an activated state. In an attempt to understand the potential role of monocyte chemoattractant protein-1 (MCP-1) in triggering spinal glial activation and its contribution to the development of neuropathic pain, we investigated the effect of peripheral nerve injury on MCP-1 expression in dorsal root ganglia (DRG) and the spinal cord, and established its temporal relationship with activation of spinal microglia and astrocytes. We observed that MCP-1 was induced by chronic constriction of the sciatic nerve in DRG sensory neurons, spinal cord motor neurons and in the superficial dorsal horn, ipsilateral to the injury. Neuronal MCP-1 induction was followed by surrounding microglial activation. After peaking at day 7 after injury, MCP-1 levels began to decline rapidly and had returned to baseline by day 150. In contrast, microglial activation peaked by day 14 and declined afterwards to reach a lower, yet significantly raised level beyond day 22 and remained increased until the end of the test period. Astrocyte activation became detectable later, progressed more slowly and also remained increased until the end of the test period, in parallel with a decreased nociceptive threshold. Our results suggest that neuronal MCP-1 may serve as a trigger for spinal microglial activation, which participates in the initiation of neuropathic pain. Delayed, sustained astrocyte activation may participate with microglia in the persistent phase of pain hypersensitivity.     

Differential activation of innate immune responses by adenovirus and adeno-associated virus vectors

Adenovirus vectors induce acute inflammation of infected tissues due to activation of the innate immune system and expression of numerous chemokines and cytokines in transduced target cells. In contrast, adeno-associated virus (AAV) vectors are not associated with significant inflammation experimentally or clinically. We tested the ability of AAV vectors to induce the expression of chemokines in vitro and to activate the innate immune system in vivo. In human HeLa cells and murine renal epithelium-derived cells (REC cells) the adenovirus vector AdlacZ induced the expression of multiple inflammatory chemokines including RANTES, interferon-inducible protein 10 (IP-10), interleukin-8 (IL-8), MIP-1beta, and MIP-2 in a dose-dependent manner. The use of AAVlacZ did not induce the expression of these chemokines above baseline levels despite 40-fold-greater titers than AdlacZ and greater amounts of intracellular AAVlacZ genomes according to Southern and slot blot analysis. This finding confirmed that the lack of AAVlacZ induction of chemokine was not due to reduced transduction. In DBA/2 mice, the intravenous administration of 2.5 x 10(11) particles of AAVlacZ resulted in the rapid induction of liver tumor necrosis factor alpha (TNF-alpha), RANTES, IP-10, MIP-1beta, MCP-1, and MIP-2 mRNAs. However, 6 h following injection, chemokine mRNA levels returned to baseline. As expected, administration of 10-fold less AdlacZ caused an induction of liver TNF-alpha and chemokine mRNAs that persisted for more than 24 h posttransduction. Whereas intravenous administration of 2.5 x 10(11) particles of AAVlacZ triggered a transient infiltration of neutrophils and CD11b(+) cells into liver, this response stood in contrast to widespread inflammation and toxicity induced by AdlacZ. Kupffer cell depletion abolished AAVlacZ but not AdlacZ-induced chemokine expression and neutrophil infiltration. In summary, these results show that AAV vectors activate the innate immune system to a lesser extent than do adenovirus vectors and offer a possible explanation for the reduced inflammatory properties of AAV compared to adenovirus vectors.     

Chemokines in proliferative diabetic retinopathy and proliferative vitreoretinopathy

PURPOSE: To determine levels of the chemokines CCL1/I-309, CCL2/MCP-1, CCL3/MIP-1alpha, CCL4/MIP-1beta, CCL7/MCP-3, CCL8/MCP-2, CXCL5/ENA-78, CXCL6/GCP-2, CXCL10/IP-10, and CXCL11/I-TAC in the vitreous humor and serum, from patients with proliferative diabetic retinopathy (PDR), proliferative vitreoretinopathy (PVR), and rhegmatogenous retinal detachment with no PVR (RD), and to investigate the expression of MCP-1, CXCL12/SDF-1, and the chemokine receptor CXCR3 in epiretinal membranes. METHODS: Paired vitreous humor and serum samples were obtained from patients undergoing vitrectomy for the treatment of RD (57 specimens), PVR (32 specimens), and PDR (88 specimens). The levels of chemokines were measured by enzyme-linked immunosorbent assays. Eighteen PDR and 5 PVR membranes were studied by immunohistochemical techniques. RESULTS: Of all the chemokines studied, only MCP-1 and IP-10 were detected in vitreous humor samples. MCP-1 levels in vitreous humor samples were significantly higher than in serum samples (p < 0.001). MCP-1 levels were significantly higher in vitreous humor samples from patients with PVR and PDR compared with RD (p = 0.0002). MCP-1 levels in vitreous humor samples from patients with active PDR were significantly higher than in inactive PDR cases (p = 0.0224). IP-10 levels in vitreous humor samples were significantly higher than in serum samples (p = 0.0035). IP-10 levels were significantly higher in vitreous humor samples from patients with PVR and PDR compared with RD (p = 0.0083). The incidence of IP-10 detection in vitreous humor samples was significantly higher in active PDR cases compared with inactive cases (p = 0.0214). There was a significant association between the incidence of IP-10 detection and increased levels of MCP-1 in vitreous humor samples from all patients, and patients with RD and PDR (p < 0.001 for all comparisons). MCP-1, and SDF-1 were localized in myofibroblasts in PVR and PDR membranes and in vascular endothelial cells in PDR membranes. CXCR3 was expressed by vascular endothelial cells in PDR membranes. CONCLUSION: MCP-1, IP-10 and SDF-1 may participate in pathogenesis of PVR and PDR. Myofibroblasts and vascular endothelial cells are the major cell types expressing MCP-1, SDF-1, and CXCR3 in epiretinal membranes.     

Induction of multiple chemokine and colony-stimulating factor genes in experimental Burkholderia pseudomallei infection

Melioidosis is a disease of the tropics caused by the facultative intracellular bacterium Burkholderia pseudomallei. In human infection, increased levels of IFN-gamma in addition to the chemokines interferon-gamma-inducible protein 10 (IP-10) and monocyte interferon-gamma-inducible protein (Mig) have been demonstrated. However, the role of these and other chemokines in the pathogenesis of melioidosis remains unknown. Using BALB/c and C57BL/6 mice as models of the acute and chronic forms of human melioidosis, the induction of mRNA was assessed for various chemokines and CSF (G-CSF, M-CSF, GM-CSF, IP-10, Mig, RANTES, MCP-1, KC and MIP-2) in spleen and liver following B. pseudomallei infection. Patterns of chemokine and CSF induction were similar in liver and spleen; however, responses were typically greater in spleen, which reflected higher tissue bacterial loads. In BALB/c mice, high-level expression of mRNA for all chemokines and CSF investigated was demonstrated at day 3 postinfection, correlating with peak bacterial load and extensive infiltration of leucocytes. In contrast, increased mRNA expression and bacterial numbers in C57BL/6 mice were greatest between 4 and 14 days following infection. This paralleled increases in the size and number of abscesses in liver and spleen of C57BL/6 mice at days 3 and 14 postinfection. Earlier induction of cytokine-induced neutrophil chemoattractant (KC), macrophage inflammatory protein-2 (MIP-2), monocyte chemoattractant protein-1 (MCP-1), granulocyte-macrophage CSF (GM-CSF) and macrophage CSF (M-CSF) mRNA was demonstrated in spleen, while MIP-2, MCP-1, IP-10 and Mig were demonstrated in liver of BALB/c mice when compared to spleen and liver of C57BL/6. The magnitude of cellular responses observed in the tissue correlated with increased levels of the chemokines and CSF investigated, as well as bacterial load. Compared with C57BL/6 mice, greater infiltration of neutrophils was observed in liver and spleen of BALB/c mice at day 3. In contrast, early lesions in C57BL/6 mice predominantly comprised macrophages. These results suggest that the inability of BALB/c mice to contain the infection at sites of inflammation may underlie the susceptible phenotype of this mouse strain towards B. pseudomallei infection.     

Physiological and immunological changes in the brain associated with lethal eastern equine encephalitis virus in macaques

Aerosol exposure to eastern equine encephalitis virus (EEEV) can trigger a lethal viral encephalitis in cynomolgus macaques which resembles severe human disease. Biomarkers indicative of central nervous system (CNS) infection by the virus and lethal outcome of disease would be useful in evaluating potential medical countermeasures, especially for therapeutic compounds. To meet requirements of the Animal Rule, a better understanding of the pathophysiology of EEEV-mediated disease in cynomolgus macaques is needed. In this study, macaques given a lethal dose of clone-derived EEEV strain V105 developed a fever between 2–3 days post infection (dpi) and succumbed to the disease by 6 dpi. At the peak of the febrile phase, there was a significant increase in the delta electroencephalography (EEG) power band associated with deep sleep as well as a sharp rise in intracranial pressure (ICP). Viremia peaked early after infection and was largely absent by the onset of fever. Granulocytosis and elevated plasma levels of IP-10 were found early after infection. At necropsy, there was a one hundred- to one thousand-fold increase in expression of traumatic brain injury genes (LIF, MMP-9) as well as inflammatory cytokines and chemokines (IFN-γ, IP-10, MCP-1, IL-8, IL-6) in the brain tissues. Phenotypic analysis of leukocytes entering the brain identified cells as primarily lymphoid (T, B, NK cells) with lower levels of infiltrating macrophages and activated microglia. Massive amounts of infectious virus were found in the brains of lethally-infected macaques. While no infectious virus was found in surviving macaques, quantitative PCR did find evidence of viral genomes in the brains of several survivors. These data are consistent with an overwhelming viral infection in the CNS coupled with a tremendous inflammatory response to the infection that may contribute to the disease outcome. Physiological monitoring of EEG and ICP represent novel methods for assessing efficacy of vaccines or therapeutics in the cynomolgus macaque model of EEEV encephalitis.     

Alterations in tissue Mg++, Na+ and spinal cord edema following impact trauma in rats

Alterations in water content and total tissue Na+ and Mg++ of rat spinal cord tissue were followed over time after a 100 g-cm impact injury to the T-9 spinal cord segment. Rats subjected to laminectomy but not trauma served as controls. In the injured segment there was a progressive increase in water content with increased Na+ and decreased Mg++ at 1 hour and 24 hours after trauma. At seven days, water and Na+ content remained elevated, whereas Mg++ levels had returned to preinjury baseline values. Because of its important role in many metabolic and physiological regulatory processes the early decline in Mg++ concentration after trauma may contribute to the development of secondary tissue damage after spinal cord injury.     

Progesterone neuroprotection in spinal cord trauma involves up-regulation of brain-derived neurotrophic factor in motoneurons

Progesterone (PROG) provides neuroprotection to the injured central and peripheral nervous system. These effects may be due to regulation of myelin synthesis in glial cells and also to direct actions on neuronal function. Both types of cells express classical intracellular PROG receptors (PR), while neurons additionally express the PROG membrane-binding site called 25-Dx. In motoneurons from rats with spinal cord injury (SCI), PROG restores to normal the deficient levels of choline acetyl-transferase and of alpha3 subunit Na,K-ATPase mRNA, while levels of the growth associated protein GAP-43 mRNA are further stimulated. Recent studies suggest that neurotrophins are possible mediators of hormone action, and in agreement with this assumption, PROG treatment of rats with SCI increases the expression of brain-derived neurotrophic factor (BDNF) at both the mRNA and protein levels in ventral horn motoneurons. In situ hybridization (ISH) has shown that SCI reduces BDNF mRNA levels by 50% in spinal motoneurons, while PROG administration to injured rats (4mg/kg/day during 3 days, s.c.) elicits a three-fold increase in grain density. In addition to enhancement of mRNA levels, PROG increases BDNF immunoreactivity in perikaryon and cell processes of motoneurons of the lesioned spinal cord, and also prevents the lesion-induced chromatolytic degeneration of spinal cord motoneurons as determined by Nissl staining. Our findings strongly indicate that motoneurons of the spinal cord are targets of PROG, as confirmed by the expression of PR and the regulation of molecular parameters. PROG enhancement of endogenous neuronal BDNF could provide a trophic environment within the lesioned spinal cord and might be part of the PROG activated-pathways to provide neuroprotection. Thus, PROG treatment constitutes a new approach to sustain neuronal function after injury.     

Rapid Calpain I Activation and Cytoskeletal Protein Degradation Following Traumatic Spinal Cord Injury: Attenuation with Riluzole Pretreatment

Abstract: Immunocytochemical and immunoblotting techniques were used to investigate calpain I activation and the stability of the calpain-sensitive cytoskeletal proteins microtubule-associated protein 2 (MAP2) and spectrin at 1, 4, and 24 h after contusion injury to the spinal cord. Spinal cord injury resulted in the activation of calpain I at all time points examined, with the highest level of activation occurring at 1 h. At the same early time point, there was a loss of dendritic MAP2 staining in spinal cord sections, accompanied by pronounced perikaryal accumulation. The loss in MAP2 staining in the injured spinal cord progressed over the 24-h survival period to affect regions 3 mm distant to the site of injury. The presence of calpain I-specific spectrin degradation was apparent in neuronal cell bodies and fibers as early as 1 h after injury, with the most intense staining occurring within and juxtaposed to the injury site. Spectrin breakdown products in neuronal cell bodies declined rapidly at 4 h and were nearly undetectable at 24 h after injury. Immunoblot studies confirmed the immunocytochemical results by demonstrating a significant increase in calpain I activation, a significant decrease in MAP2 levels, and a significant increase in spectrin breakdown. Finally, treatment of animals with riluzole, an inhibitor of glutamate release, before surgery reduced significantly the loss of MAP2 levels observed at 24 h after injury. These results demonstrate that Ca²⁺-dependent protease activation and degradation of critical cytoskeletal proteins are early events after spinal cord injury and that treatments that minimize the actions of glutamate may limit their breakdown.     

Changes in expression of ciliary neurotrophic factor (CNTF) and CNTF-receptor α after spinal cord injury

To determine if ciliary neurotrophic factor (CNTF) is involved in the response to spinal cord injury, we studied changes in the expression of CNTF and that of its receptor, CNTF-receptor α (CNTFRα), in the rat spinal cord after a unilateral spinal cord hemisection. Using in situ hybridization, we found that CNTFRα mRNA levels in spinal cord motoneurons increased dramatically by 1 day after hemisecting the spinal cord at T2. This increase in expression was present only in motoneurons caudal, but not rostral, to the lesion. In addition, we detected increased levels of CNTF mRNA in the spinal cord white matter, also by 1 day following injury. Unlike CNTFRα, however, the increase in CNTF mRNA was evident both rostral and caudal to the lesion. Levels of both CNTF and CNTFRα mRNA declined between 1 and 5 days, and by 10 days they were not significantly different from normal animals. These findings suggest that CNTF may play a local role in the response to spinal cord injury. © 1997 John Wiley & Sons, Inc. J Neurobiol 32: 251–261, 1997.     

CpG oligodeoxynucleotides induce murine macrophages to up-regulate chemokine mRNA expression

Intramuscular injection of synthetic oligodeoxynucleotides (ODN) expressing unmethylated CpG motifs trigger the rapid development of a local inflammatory response. In vitro studies demonstrate that macrophages exposed to CpG ODN up-regulate expression of mRNA encoding the chemokines MIP-1alpha, MIP-1beta, MIP-2, RANTES, JE/MCP-1, and IP-10. Within 6 h of in vivo administration, CpG ODN induce a significant increase in chemokine mRNA levels at the site of injection and draining lymph nodes. These chemokines may contribute to the migration and stimulation of inflammatory cells that contribute to the development of CpG ODN-induced immune responses.     

自组装短肽在脊髓急性创伤早期减少细胞凋亡的研究

目的 为研究在脊髓急性损伤中,自组装短肽溶液对创伤后神经组织的保护作用及创伤后早期神经组织细胞凋亡的时间分布规律.方法 实验使用了手术切断大鼠脊髓胸段T8至T10作为脊髓损伤动物模型,使用末端脱氧核苷酸转移酶介导生物素标记(TUNEL)技术,检测实验后不同时间点的损伤脊髓组织细胞凋亡情况.结果 伤后2 h,损伤区及邻近段开始出现末端标记阳性细胞;伤后8 h,凋亡阳性细胞数达高峰;同时,自组装短肽实验组细胞凋亡较空白对照组少.结论 自组装短肽溶液对急性脊髓创伤组织具有保护作用.     

Induction of the chemokines interleukin-8 and IP-10 by human immunodeficiency virus type 1 tat in astrocytes

A finding commonly observed in human immunodeficiency virus type 1 (HIV-1)-infected patients is invasion of the brain by activated T cells and infected macrophages, eventually leading to the development of neurological disorders and HIV-1-associated dementia. The recruitment of T cells and macrophages into the brain is likely the result of chemokine expression. Indeed, earlier studies revealed that levels of different chemokines were increased in the cerebrospinal fluid of HIV-1-infected patients whereas possible triggers and cellular sources for chemokine expression in the brain remain widely undefined. As previous studies indicated that HIV-1 Tat, the retroviral transactivator, is capable of inducing a variety of cellular genes, we investigated its capacity to induce production of chemokines in astrocytes. Herein, we demonstrate that HIV-1 Tat(72aa) is a potent inducer of MCP-1, interleukin-8 (IL-8), and IP-10 expression in astrocytes. Levels of induced IP-10 protein were sufficiently high to induce chemotaxis of peripheral blood lymphocytes. In addition, Tat(72aa) induced IL-8 expression in astrocytes. IL-8 mRNA induction was seen less then 1 h after Tat(72aa) stimulation, and levels remained elevated for up to 24 h, leading to IL-8 protein production. Tat(72aa)-mediated MCP-1 and IL-8 mRNA induction was susceptible to inhibition by the MEK1/2 inhibitor UO126 but was only modestly decreased by the inclusion of the p38 mitogen-activated protein kinase (MAPK) inhibitor SB202190. In contrast, Tat-mediated IP-10 mRNA induction was suppressed by SB202190 but not by the MEK1/2 inhibitor UO126. These findings indicate that MAPKs play a major role in Tat(72aa)-mediated chemokine induction in astrocytes.