FK506 treatment inhibits caspase-3 activation and promotes oligodendroglial survival following traumatic spinal cord injury

The focus of this study is to examine the ability of FK506, an immunosuppressant that inhibits calcineurin activation, to limit caspase-3 activation in oligodendroglia following spinal cord injury (SCI). To better establish a role for calcineurin and caspase-3 activation in oligodendroglia following SCI, rats received a contusion injury to the spinal cord followed by treatment with FK506 or rapamycin (another immunosuppressant with no detectable inhibitory action on calcineurin activation). Animals were then sacrificed at 8 days postinjury and spinal cord tissue was processed using immunofluorescence histochemistry to examine cellular caspase-3 activation in ventral and dorsal white matter. In all treatment groups, numerous oligodendroglia were found to express the activated form of caspase-3 in regions proximal and distal to the injury epicenter. However, our findings suggest that treatment with FK506, but not rapamycin reduces the number of oligodendroglia expressing activated caspase-3 and increases the number of surviving oligodendroglia in dorsal white matter. These results provide initial evidence that agents that reduce the actions of calcineurin and subsequent caspase-3 activation may prove beneficial in the treatment of traumatic SCI.     

Olfactory ensheathing glia graft in combination with FK506 administration promote repair after spinal cord injury

The aim of this study was to determine whether a combination of olfactory ensheathing cell (OEC) graft with the administration of FK506, two experimental approaches that have been previously reported to exert protective/regenerative effects after spinal cord injury, promotes synergic restorative effects after complete or partial spinal cord injuries. In partial spinal cord injury, combination of an OEC graft and FK506 reduced functional deficits evaluated by the BBB score, motor-evoked potentials (MEPs) and H reflex tests, diminished cavitation, astrogliosis and increased sparing/regeneration of raphespinal fibers compared to untreated and single-treatment groups of rats. After complete spinal cord transection, the combined treatment significantly improved functional outcomes, promoted axonal regeneration caudal to the lesion, and diminished astrogliosis compared only to non-transplanted animals. Slightly, but non-significant, better functional and histological results were found in OEC-grafted animals treated with FK506 than in those given saline after spinal cord transection. Nevertheless, the combined treatment increased the percentage of rats that recovered MEPs and promoted a significant reduction in astrogliosis. In conclusion, this study demonstrates that OEC grafts combined with FK506 promote additive repair of spinal cord injuries to those exerted by single treatments, the effect being more remarkable when the spinal cord is partially lesioned.     

ApoE mimetic ameliorates motor deficit and tissue damage in rat spinal cord injury

Apolipoprotein E (apoE), a plasma protein responsible for transporting lipid and cholesterol, modulates responses of the central nervous system to injury. Small peptides derived from the receptor‐binding region of apoE can simulate some important bioactivities of apoE holoprotein and offer neuroprotection against brain injury. We tested whether COG1410, an apoE‐mimetic peptide, provides protection in a rat model of spinal cord injury (SCI). Traumatic injury was created at T8 by a cortical impact device. Injured rats were randomized to four treatment groups: vehicle, 0.15, 0.3, or 0.6 mg/kg COG1410; sham surgery rats received vehicle. Basso, Beattie, Bresnahan neurological score was evaluated prior to injury and at 1, 3, 7, and 14 days after injury. Histological changes were evaluated at 14 days. All injured rats lost body weight during the first week following injury. Body weight recovery was significantly improved in rats treated with COG1410. Mechanical impact resulted in severe motor deficit, and most animals had a BBB score of 0–1 at 24 hours postinjury. COG1410‐treated rats showed significantly improved functional recovery and ameliorated motor deficit at 14 days postinjury. Histological analysis showed that COG1410 groups had a significantly reduced lesion size at the site of injury, a larger preserved luxol fast blue‐stained area, and more visible neurons in the surrounding area of injury. Microglial activation was also significantly suppressed. These findings indicate that this apoE mimetic effectively improved neurological and histological outcome following SCI in rats, and the effect was associated with inhibition of microglial activation. © 2014 Wiley Periodicals, Inc.     

Elamipretide reduces pyroptosis and improves functional recovery after spinal cord injury

Aims

Elamipretide (EPT), a novel mitochondria-targeted peptide, has been shown to be protective in a range of diseases. However, the effect of EPT in spinal cord injury (SCI) has yet to be elucidated. We aimed to investigate whether EPT would inhibit pyroptosis and protect against SCI.

Methods

After establishing the SCI model, we determined the biochemical and morphological changes associated with pyroptosis, including neuronal cell death, proinflammatory cytokine expression, and signal pathway levels. Furthermore, mitochondrial function was assessed with flow cytometry, quantitative real-time polymerase chain reaction, and western blot.

Results

Here, we demonstrate that EPT improved locomotor functional recovery following SCI as well as reduced neuronal loss. Moreover, EPT inhibited nucleotide-binding oligomerization domain-like receptor 3 (NLRP3) inflammasome activation and pyroptosis occurrence and decreased pro-inflammatory cytokines levels following SCI. Furthermore, EPT alleviated mitochondrial dysfunction and reduced mitochondrial reactive oxygen species level.

Conclusion

EPT treatment may protect against SCI via inhibition of pyroptosis.     

Valproic acid reduces autophagy and promotes functional recovery after spinal cord injury in rats

Secondary damage is a critical determinant of the functional outcome in patients with spinal cord injury (SCI), and involves multiple mechanisms of which the most important is the loss of nerve cells mediated by multiple factors. Autophagy can result in cell death, and plays a key role in the development of SCI. It has been recognized that valproic acid (VPA) is neuroprotective in certain experimental animal models, however, the levels of autophagic changes in the process of neuroprotection by VPA treatment following SCI are still unknown. In the present study, we determined the extent of autophagy after VPA treatment in a rat model of SCI. We found that both the mRNA and protein levels of Beclin-1 and LC3 were significantly increased at 1, 2, and 6 h after SCI and peaked at 2 h; however, Western blot showed that autophagy was markedly decreased by VPA treatment at 2 h post-injury. Besides, post-SCI treatment with VPA improved the Basso-Beattie-Bresnahan scale, increased the number of ventral horn motoneurons, and reduced myelin sheath damage compared with vehicle-treated animals at 42 days after SCI. Together, our results demonstrated the characteristics of autophagy expression following SCI, and found that VPA reduced autophagy and enhanced motor function.     

他克莫司对骨髓间充质干细胞移植治疗大鼠脊髓损伤的影响

目的骨髓间充质干细胞(BMSC)移植同时应用他克莫司(FK506),观察两者对脊髓损伤后大鼠恢复的影响。方法用钳夹法制作大鼠急性脊髓损伤动物模型30只,随机分成3组。A组为单纯损伤组,B组为单纯骨髓干细胞移植组,C组为骨髓干细胞联合FK506组。于伤后1w、2w、4w、6w、8w采用BBB评分。8周后取材,行免疫组化及病理学检查。结果B、C两组大鼠后肢运动功能均有较明显恢复,C组较B组为快,两组统计学上有差异(P<0.05)。A组亦有所恢复,但程度较轻。A组切片,未见神经轴索通过,B、C组可见少量神经轴索样结构。免疫组化显示:A组NF200阳性细胞,GFAP阳性细胞均较B组C组少(P<0.05)。C组Nogo阳性神经元较A、B两组明显减少(P<0.05)。B组NF200阳性细胞,GFAP阳性细胞均较C组少(P<0.05)。结论骨髓间质干细胞移植对于后肢功能的恢复有促进作用,联合应用FK506有协同效果。     

Systemic hypothermia improves histological and functional outcome after cervical spinal cord contusion in rats

Hypothermia has been employed during the past 30 years as a therapeutic modality for spinal cord injury (SCI) in animal models and in humans. With our newly developed rat cervical model of contusive SCI, we investigated the therapeutic efficacy of transient systemic hypothermia (beginning 5 minutes post‐injury for 4 hours, 33°C) with gradual rewarming (1°C per hour) for the preservation of tissue and the prevention of injury‐induced functional loss. A moderate cervical displacement SCI was performed in female Fischer rats, and behavior was assessed for 8 weeks. Histologically, the application of hypothermia after SCI resulted in significant increases in normal‐appearing white matter (31% increase) and gray matter (38% increase) volumes, greater preservation (four‐fold) of neurons immediately rostral and caudal to the injury epicenter, and enhanced sparing of axonal connections from retrogradely traced reticulospinal neurons (127% increase) compared with normothermic controls. Functionally, a faster rate of recovery in open field locomotor ability (BBB score, weeks 1–3) and improved forelimb strength, as measured by both weight‐supported hanging (43% increase) and grip strength (25% increase), were obtained after hypothermia. The current study demonstrates that mild systemic hypothermia is effective for retarding tissue damage and reducing neurological deficits following a clinically relevant contusive cervical SCI. J. Comp. Neurol. 514:433–448, 2009. © 2009 Wiley‐Liss, Inc.     

Effect of methylprednisolone on motor function and spinal cord blood flow after spinal cord compression in rats

The effect of methylprednisolone (MP) on neurologic recovery and spinal cord blood flow (SCBF) was investigated up to 4 days after a spinal cord compression injury in rats. The injury was produced at midthoracic level by applying a load of 35 g on a 2.2 x 5.0 mm compression plate for 5 min, which resulted in transient paraparesis. MP was given as a bolus dose of 30 mg/kg i.v. 60 min after injury (n = 20) and controls were given saline (n = 10). The motor performance was assessed daily as the capacity angle on the inclined plane and SCBF was measured by 14C-iodoantipyrine autoradiography on Days 1 or 4. On Day 1 the capacity angle was reduced from about 63 degrees preoperatively to 33 +/- 2 degrees (mean +/- SEM) in the control group and to 50 +/- 1 degrees in the group treated with MP (p less than 0.05). Thereafter there was a slight improvement in both groups, but the difference persisted throughout the observation period. On Day 4 both gray and white matter SCBF was better preserved in MP-treated animals than in the control group (59 +/- 4 versus 49 +/- 3 ml/min/100 g tissue for gray matter and 13.6 +/- 0.6 versus 10.7 +/- 0.8 ml/min/100 g tissue for white matter). Posttraumatic treatment with MP, thus, improved both the neurologic recovery during the first 4 days and SCBF as measured on Day 4. It is speculated that the effect of MP is at least partly exerted on the vascular bed.     

Chemokine antagonist infusion promotes axonal sparing after spinal cord contusion injury in rat

Spinal cord injury produced by mechanical contusion causes the onset of acute and chronic degradative events. These include blood brain barrier disruption, edema, demyelination, axonal damage and neuronal cell death. Posttraumatic inflammation after spinal cord injury has been implicated in the secondary injury that ultimately leads to neurologic dysfunction. Studies after spinal cord contusion have shown expression of several chemokines early after injury and suggested a role for them in the ordered recruitment of inflammatory cells at the lesion site (McTigue et al. [1998] J. Neurosci. Res. 53:368-376; Lee et al., [2000] Neurochem Int). We have demonstrated previously that infusion of the broad-spectrum chemokine receptor antagonist (vMIPII) in the contused spinal cord initially attenuates leukocyte infiltration, suppresses' gliotic reaction and reduces neuronal damage after injury. These changes are accompanied by increased expression of bcl-2, the endogenous apoptosis inhibitor, and reduced neuronal apoptosis (Ghirnikar et al. [2000] J. Neurosci. Res. 59:63-73). We demonstrate that 2 and 4 weeks of vMIPII infusion in the contusion-injured spinal cord also results in decreased hematogenous infiltration and is accompanied by reduced axonal degeneration in the gray matter. Luxol fast blue and MBP immunoreactivity indicated reduced myelin breakdown in the dorsal and ventral funiculi. Increased neuronal survival in the ventral horns of vMIPII infused cords was seen along with increased bcl-2 staining in them. Immunohistochemical identification of fiber phenotypes showed increased presence of calcitonin gene related peptide, choline acetyl transferase and tyrosine hydroxylase positive fibers as well as increased GAP43 staining in treated cords. These results suggest that sustained reduction in posttraumatic cellular infiltration is beneficial for tissue survival. A preliminary report of this study has been published (Eng et al. [2000] J. Neurochem. 74(Suppl):S67B). In contrast to vMIPII, infusion of MCP-1 (9-76), a N-terminal analog of the MCP-1 chemokine showed only a modest reduction in cellular infiltration at 14 and 21 dpi without significant tissue survival after spinal cord contusion injury. Comparing data on tissue survival obtained with vMIPII and MCP-1 (9-76) further validate the importance of the use of broad-spectrum antagonists in the treatment of spinal cord injury. Controlling the inflammatory reaction and providing a growth permissive environment would enhance regeneration and ultimately lead to neurological recovery after spinal cord injury. J. Neurosci. Res. 64:582-589, 2001. Published 2001 Wiley-Liss, Inc.     

The Effects of FK506 on Dorsal Column Axons Following Spinal Cord Injury in Adult Rats: Neuroprotection and Local Regeneration

There is considerable evidence that immunophilin ligands can promote the regeneration of axons in peripheral nerves and act as neuroprotective agents in the CNS. We have examined the effects of FK506 and GPI 1046 on the responses to partial transection of ascending spinal dorsal column axons at T9, in some cases combined with crush of one sciatic nerve. FK506 (0.5 or 2.0 mg/kg) and GPI 1046 (10 or 40 mg/kg) was administered subcutaneously immediately after surgery and five times a week thereafter. Some animals received methylprednisolone (MP) (two subcutaneous doses of 30 mg/kg) in addition to, or instead of, FK506. After survival times of 1-12 weeks, dorsal column axons were labeled transganglionically with cholera toxin B-HRP. There was massive axonal sprouting at the lesion sites in animals with sciatic nerve injury and immunophilin ligand treatment. In FK506-treated animals a few severed sensory axons regenerated for up to 10 mm rostral to the lesion. Of greater significance, 30% of 71 FK506-treated animals had spared axons in the dorsal column, extending to the nucleus gracilis, versus 8% of 50 control animals (P < 0.05), showing that FK506 reduces the likelihood of axonal destruction due to secondary injury. A combination of FK506 and MP afforded greater protection than MP alone (P < 0.05), but axonal survival was not affected by sciatic nerve crush, dose of FK506, or survival time after injury. GPI 1046 (n = 11) did not promote axonal survival. Thus FK506 protects axons from secondary injury following spinal cord trauma, and in this experimental model, its neuroprotective effect is greater than that of MP.     

Efficacy of methylprednisolone therapy for the injured rat spinal cord

Currently the synthetic glucocorticosteroid methylprednisolone sodium succinate (MPSS) is the standard therapy after acute spinal cord injury (SCI) in humans based on reported neurological improvements. The mechanisms for its beneficial actions are not entirely clear, but experimental evidence suggests MPSS affords some degree of neuroprotection. As many studies with rat models of SCI have been unable to demonstrate improved behavioral outcome or tissue sparing after MPSS treatment, we chose to stereologically assess whether it alters lesion volume and tissue sparing over time, as well as long-term behavioral recovery. Adult rats subjected to contusion SCI with the NYU impactor were administered either MPSS or saline for 24 hr beginning 5 min post injury. Over time the lesion dimensions were extremely dynamic, such that by 6 weeks post injury the volumes were reduced to a third of those seen after the first week. MPSS marginally reduced lesion volumes across time vs. controls, but the amount of spared gray and white matter remained unaltered between the two groups. Behavioral results further showed that MPSS failed to improve recovery of hind-limb function. These findings add to the emerging scrutiny of MPSS as the standard therapy for acute SCI, as well as indicate the existence of a therapeutic window for tissue sparing restricted to the first several days after this type of SCI in rats. Equally important, our results caution the use of lesion volume dimensions or percent tissue sparing at the epicenter as indicators of therapeutic efficacy because neither reflects the actual amount of tissue sparing.     

Methylprednisolone and tetrandrine in combination with direct current electrical field for treating acute spinal cord injury

BACKGROUND: The direct current electrical field can effectively promote the regeneration of the spinal cord; moreover, methylprednisolone (MP) can relieve secondary edema after spinal cord injury. Tetrandrine (Tet) is an effective component of hanfangji and can protect the effect of spinal cord and axis-cylinder. Whether direct current electrical field combining with MP or Tet has synergic or strengthening effect on treating complete spinal cord injury or not should be studied further. OBJECTIVE:To study the effect of direct current electrical field assisted by MP and Tet on treating spinal cord injury. DESIGN: Randomized controlled animal study. SETTING: People's Hospital of Hainan Province. MATERIALS: A total of 45 healthy hybrid dogs, of both genders, weighing 10–12 kg, aged 1.5–2 years, were provided by Animal Center of Hainan Province. Somatosensory evoked potential meter (DANTEC Company), IBAS-2.0 imaging analysis meter (Germany), and self-made electronic stimulator. METHODS: The experiment was carried out in Hainan People's Hospital from May 2001 to June 2004. All experimental dogs were randomly divided into 4 groups: control group (n =9), electrostimulating group (n =12), MP + electrostimulating group (n =12) and Tet + electrostimulating group (n =12). ① After anesthesia, Allen WD method was used to induce complete spinal cord injury. The metal bar, which was 10 cm in height fell freely and vertically hit the spinal cord to provide a complete spinal cord injury. Dogs in control group and electrostimulating group were implanted electrical stimulators 6 hours after spinal cord injury (no electricity in control group); dogs in MP + electrostimulating group were injected 30 mg/kg MP for 15 minutes at 2 hours after spinal cord injury and electrical stimulators implanted at 6 hours after injury; dogs in Tet + electrostimulating group were intravenously injected with 7.5 mg/kg Tet at 2 hours after spinal cord injury and electrical stimulators implanted at 6 hours after injury; and then, 7.5 mg/kg Tet injected at days 2 and 3 after injury. ② Specimens were taken from control group from three dogs of every month; from the injured segments of spinal cords at 1 month, 2 months and 3 months; and from electrostimulating group, MP + electrostimulating group and Tet + electrostimulating group of 4 dogs for histological examinations. ③ Detection of neurological function: Neurological function was evaluated with the functional 10 grading system. The scores ranged from 0 to 10 (0: complete paraplegia; 10: normality). ④ Detection of cortical somatosensory evoked potential (CSEP): According to the scheme formulated by the International Electroencephalographical Association, the patterns of the fundamental waves were P1–N1–P1 waves. The latency of the P1 wave and the amplitude of P1–N1 waves were mainly observed individually at 1, 2 and 3 months after the injury. ⑤ Histological detection: All spinal cord specimens of the injuried segment were harvested at 1, 2 and 3 months after injury. They were stained with hematoxylin and Nissl staining methods, and then were observed under an optical microscope, and the neurons were counted. The sectional areas of the neurons and the density of the Nissl bodies were measured by a system image pattern analysis (IBAS-2.0, Germany). MAIN OUTCOME MEASURES: The neurological function, cortical somatosensory evoked potential, neuronal amount, sectional area of neurons and Nissl body density at 1 to 3 months after injury. RESULTS: All 45 experimental dogs were involved in the final analysis. ① Detection of neurological function: One month later, the dogs in MP + electrostimulating group could walk, but the dogs in electrostimulating group and Tet + electrostimulating group could stand. Two months after injury, the dogs in MP + electrostimulating group almost recovered to normal, but the dogs in electrostimulating group could walk and those in Tet + electrostimulating group could run. Those in control group had no parent recovery. ② Detection of P1 latency and P1–N1 amplitude: Changes of P1 latency in control group were long and P1–N1 amplitude was very low at 1 month later. Compared to electrostimulating group, MP + electrostimulating group and Tet + electrostimulating group, there were significant differences (P < 0.05). P1 latency was manifestly shortened and amplitude were raised in electrostimulating group, MP + electrostimulating group and Tet + electrostimulating group. Those in MP + electrostimulating group and Tet + electrostimulating group were superior to those in electrostimulating group and there were significant differences (P < 0.05). ③ Sectional areas of neurons and Nissl body density: At 1–3 months after injury, sectional areas of neurons were larger in electrostimulating group [(170.14±7.45), (209.60±14.80), (312.47±12.63) μm2], MP + electrostimulating group [(282.18±15.25), (418.18±16.27), (515.25±15.10) μm2] and Tet + electrostimulating group [(231.81±7.38), (322.67±8.45), (386.82±10.42) μm2] than control group[(98.12±4.93), (113.50±6.74), (122.59±8.03) μm2, P < 0.05]; especially, sectional area was the largest in MP + electrostimulating group. At 1–3 months after injury, Nissl body density was more in electrostimulating group (170.14±7.45, 209.60±14.80, 312.47±12.63), MP + electrostimulating group (282.18±15.25, 418.18±16.27, 515.25±15.10) and Tet + electrostimulating group (231.81±7.38, 322.67±8.45, 386.82±10.42) than control group (98.12±4.93, 113.50±6.74, 122.59±8.03, P < 0.05); especially, Nissl body density was the most in MP + electrostimulating group. CONCLUSION: The direct current electrical field can effectively promote spinal cord regeneration. The combination of direct current electrical field with large dose MP or Tet has synergistic effects for treating spinal cord injury. The curative effects of direct current electrical field with large dose MP are much better than those with Tet.     

组织工程支架材料在脊髓损伤修复中的应用*

背景：脊髓损伤最初往往会导致细胞和组织的不断丢失,组织工程支架可以模拟细胞外基质的生理状态,从而有利于细胞的黏附、迁移、扩增和分化。 目的：总结近年来组织工程支架材料联合细胞和/或细胞因子修复脊髓损伤的新进展。 方法：应用计算机检索PubMed、Ovid Medline及CBM数据库中2000-10/2010-10 与组织工程支架材料修复脊髓损伤相关的文章。 结果与结论：组织工程材料治疗脊髓损伤需要3 因素：种子细胞、组织工程支架、细胞因子。组织工程支架对于损伤脊髓断端起到桥接作用,而种植于材料的种子细胞和/或细胞因子可以促进神经轴突的生长和迁移。可用于组织工程支架的材料可分为天然材料和人工合成材料,包括胶原、壳聚糖、琼脂糖/藻酸盐、聚乳酸、纤连蛋白、聚羟基乙酸/聚乳酸、聚β羟丁酸等,动物实验已经取得一些成果,显示组织工程支架材料联合细胞移植修复效果更好,但临床上目前尚无开展组织工程支架材料修复脊髓损伤的研究。     

甲基强的松龙对大鼠急性脊髓损伤截瘫后肠道细菌移位的抑制作用

背景：我们以往的研究显示急性脊髓损伤截瘫后,自主神经紊乱导致的肠道动力障碍促使细菌移位的发生。目前认为大剂量甲基强的松龙可促进脊髓损伤患者神经功能的恢复。但不清楚甲基强的松龙能否通过促进神经功能的恢复,从而抑制急性脊髓损伤后肠道细菌的移位？本实验的目的为明确上述问题。方法：建立大鼠脊髓损伤截瘫模型。实验组大鼠脊髓损伤截瘫后立即给予大剂量甲基强的松龙,对照组大鼠脊髓损伤截瘫后立即给予生理盐水作为对照,在急性脊髓损伤后24、72小时及1周评估BBB运动功能得分,采血行细菌培养和内毒素检测。同时采集肠系膜淋巴结、脾脏、肝脏标本行细菌培养。脊髓损伤后1周行肠系膜淋巴结、脾脏、肝脏、空肠和回肠组织学观察。结果：急性脊髓损伤后1周,实验组大鼠BBB运动得分显著高于对照组(10.36±0.86 vs 6.32±1.02, P <0.05)。两组动物在脊髓损伤后24小时均发现内毒素血症及细菌生长。然而,脊髓损伤后72小时及1周,实验组大鼠血浆内毒素水平显著低于对照组(损伤后72小时：216.15±12.90 vs 435.54±10.76 , P <0.05,损伤后1周：106.58±18.56 vs 368.85±17.35, P <0.05)。脊髓损伤截瘫后移位细菌主要为大肠杆菌、阴沟杆菌、大肠埃希氏菌、普通变形杆菌、肠粪球菌等。脊髓损伤后1周实验组大鼠肠系膜淋巴结、脾脏、肝脏、空肠和回肠组织学变化程度较对照组轻。结论：大剂量甲基强的松龙冲击疗法可抑制急性脊髓损伤后肠道细菌的移位。急性脊髓损伤患者立即给予甲基强的松龙和抗生素可能有助于抑制潜在的细菌移位。     

Effects of tacrolimus （FK506） on sciatic nerve regeneration in rats

BACKGROUND： Tacrolimus （FK506） protects peripheral nerves located in damaged regions by inhibiting T lymphocyte proliferation and activation. OBJECTIVE： To evaluate the effect of FK506 on promoting regeneration of rat sciatic nerve. DESIGN, TIME AND SETTING： A randomized, controlled, animal study was performed at the Laboratory of the Department of Orthopedic Surgery, Dalian Medical University, China, from September 2007 to September 2008. MATERIALS： A total of 60 adult, male, Sprague-Dawley rats were equally and randomly divided into model, local administration and systemic administration groups. All rats received a neurotomy of bilateral sciatic nerves to establish models of nerve regeneration chambers. The powder and injection of FK506 were supplied by Fujisawa Pharmaceutical, Japan. METHODS： The regeneration chambers of the model group were infused with 0.2 mL saline. The systemic group were injected with 0.2 mL saline, followed by daily subcutaneous injections of FK506 （1 mg/kg）, for 14 days. The local administration group was infused with 0.2 mL FK506 （1 μg/mL）. MAIN OUTCOME MEASURES： Local immune response was observed using hematoxylin-eosin staining. Myelinated nerve fiber number, myelin sheath and nerve fiber thickness were observed using toluidine blue staining. Wet weight of gastrocnemius was evaluated. Compound muscle action potential amplitude, latency, and conduction time were recorded, and motor nerve conduction velocity was calculated using electrophysiology. RESULTS： The total number of myelinated nerve fibers in the local and systemic administration groups was significantly higher than in the model group. The density of myelinated nerve fibers, myelin sheath thickness and mean axon diameter were significantly increased in the systemic administration group compared with the model group （P 〈 0.05）. Lymphocyte infiltration was decreased in the local and systemic administration groups compared with the model group. The wet weight of rat gastrocnemius in the local and systemic administration groups were significantly greater compared with the model group （P 〈 0.05）. Motor nerve conduction velocity was the fastest in the systemic administration group, and the slowest in the model group. Compound muscle action potential amplitude was larger in the systemic administration group compared with the local administration and model groups （P 〈 0.05）. CONCLUSION： Systemic administration of FK506 can promote regeneration of rat sciatic nerve and recovery of neural function. Systemic administration produced better regeneration and recovery of function than local administration of FK506.     

Pathogenesis of delayed radiation injury in the rat spinal cord after X-ray irradiation

The pathogenesis of delayed radiation injury was examined experimentally. Exposure fields in the thoracic and lumbar spinal cords of 54 adult female specific pathogen-free Wistar rats were X-ray irradiated. The rats were observed clinically and pathologically for up to 6 months after irradiation and the relationships between the chronological progress of paralysis and pathological findings and between vascular and parenchymal changes were analyzed by reconstructing serial spinal cord sections.     

Expression and detrimental role of hematopoietic prostaglandin D synthase in spinal cord contusion injury

Prostaglandin D(2) (PGD(2) ) is a potent inflammatory mediator, which is implicated in both the initiation and resolution of inflammation in peripheral non-neural tissues. Its role in the central nervous system has not been fully elucidated. Spinal cord injury (SCI) is associated with an acute inflammatory response, which contributes to secondary tissue damage that worsens functional loss. We show here, with the use of hematopoietic prostaglandin D synthase (HPGDS) deficient mice and a HPGDS selective inhibitor (HQL-79), that PGD(2) plays a detrimental role after SCI. We also show that HPGDS is expressed in macrophages in the injured mouse spinal cord and contributes to the increase in PGD(2) in the contused spinal cord. HPGDS(-/-) mice also show reduced secondary tissue damage and reduced expression of the proinflammatory chemokine CXCL10 as well as an increase in IL-6 and TGFβ-1 expression in the injured spinal cord. This was accompanied by a reduction in the expression of the microglia/macrophage activation marker Mac-2 and an increase in the antioxidant metallothionein III. Importantly, HPGDS deficient mice exhibit significantly better locomotor recovery after spinal cord contusion injury than wild-type (Wt) mice. In addition, systemically administered HPGDS inhibitor (HQL-79) also enhanced locomotor recovery after SCI in Wt mice. These data suggest that PGD(2) generated via HPGDS has detrimental effects after SCI and that blocking the activity of this enzyme can be beneficial.     

Delayed administration of dapsone protects from tissue damage and improves recovery after spinal cord injury

After spinal cord injury (SCI), a complex cascade of pathophysiological processes increases the primary damage. The inflammatory response plays a key role in this pathology. Recent evidence suggests that myeloperoxidase (MPO), an enzyme produced and released by neutrophils, is of special importance in spreading tissue damage. Dapsone (4,4'-diaminodiphenylsulfone) is an irreversible inhibitor of MPO. Recently, we demonstrated, in a model of brain ischemia/reperfusion, that dapsone has antioxidant, antiinflammatory, and antiapoptotic effects. The effects of dapsone on MPO activity, lipid peroxidation (LP) processes, motor function recovery, and the amount of spared tissue were evaluated in a rat model of SCI. MPO activity had increased 24.5-fold 24 hr after SCI vs. the sham group, and it had diminished by 38% and 19% in the groups treated with dapsone at 3 and 5 hr after SCI, respectively. SCI increased LP by 45%, and this increase was blocked by dapsone. In rats treated with dapsone, a significant motor function recovery (Basso-Beattie-Bresnahan score, BBB) was observed beginning during the first week of evaluation and continuing until the end of the study. Spontaneous recovery 8 weeks after SCI was 9.2 ± 1.12, whereas, in the dapsone-treated groups, it reached 13.6 ± 1.04 and 12.9 ± 1.17. Spared tissue increased by 42% and 33% in the dapsone-treated groups (3 and 5 hr after SCI, respectively) vs. SCI without treatment. Dapsone significantly prevented mortality. The results show that inhibition of MPO by dapsone significantly protected the spinal cord from tissue damage and enhanced motor recovery after SCI.