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The Protective Effects and Its Mechanisms of Extract of Ginkgo Biloba on Blood-spinal Cord Barrier After Acute Spinal Cord Injury in Rats

Author: ShaoWei
Tutor: LuoZhuoJing
School: Fourth Military Medical University
Course: Surgery
Keywords: Spinal cord injury EGb761 Blood - spinal cord barrier Interleukin -1β Intercellular adhesion molecule-1 Apoptosis TUNEL Caspase-3
CLC: R285.5
Type: Master's thesis
Year: 2011
Downloads: 24
Quote: 0
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Abstract


High incidence of spinal cord injury (Spinal cord injury, SCI), the prognosis is poor, not only caused huge patient's physical and psychological trauma, but also brought a heavy burden to families and society, we need urgently to make unremitting efforts to find the right treatment strategies to improve the quality of life of these patients, and to reduce the burden on the society and the family. SCI's primary damage is irreversible; combined with secondary damage caused by spinal cord tissue, expanding the scope of the original trauma aggravating the nerve cell loss thereby affecting an important prognostic factor. How to suppress or even reverse the secondary injury is the of SCI treatment research priorities and hot. The study showed that the blood - spinal cord barrier (Blood-spinal cord barrier, BSCB) damage and subsequent inflammatory response, Spreading and apoptosis is an important part of the secondary injury mechanisms [1]. EGb761 (Ginkgo biliba extract) is the international standard of Ginkgo biloba extract, the main ingredient terpene lactones (6%), and ginkgo flavonoid glycosides (24%); study confirmed that it has strong antioxidant and neuroprotective role and can promote nerve regeneration, ischemic brain damage and central nervous system (Central nervous system, CNS) has been applied to the clinical treatment of degenerative diseases [2, 3]. However EGb761 similar role in the SCI has little or nothing reported. This study be adopted of EGb761 on acute SCI model applied intervention aimed at the observed protective effect the EGb761 of nerve tissue in rats with acute SCI BSCB permeability changes from the early stage, the expression of inflammatory cytokines and nerve cell apoptosis , chronic spinal cord tissue cavity formation to explore the possible mechanism of action, in order to provide experimental evidence for EGb761 used in the treatment of SCI. Purposes: (1) observation of the protective effect of Ginkgo biloba extract (EGb761) on the blood-spinal cord barrier after acute spinal cord injury in rats, and to explore the possible protective mechanism. (2) To observe the characteristics of the different stages of spinal cord tissue pathological changes and neuronal apoptosis in rats with acute spinal cord injury, and observe EGb761 early neuronal apoptosis after SCI, the late void formation and recovery of motor function, explore the blood - spinal cord barrier and nerve tissue protective effect of possible mechanisms. Methods: This study consists of two parts experiments. (1) experiment Ⅰ: 120 female SD rats were randomly divided into the simple laminectomy group (A), the damage to the control group (B), the effect of EGb761 treatment group (C) n = 40. A group only T9 laminectomy, not injured spinal cord. B, C group modified Allen's production T9 spinal cord injury model in rats given intraperitoneally daily by the C group was to put to death before that EGb761 with 100mg/kg, A, B group were given normal saline at the same time. After 3h, 6h, 24h, 72h sacrificed animals, take T9 spinal cord by fluorescence microscopy microscopy and dimethyl formamide (DMF) extraction was observed after SCI BSCB Evans blue protein complex ( EBA) changes in the permeability of the content of interleukin-1β (Interleukin-1β, IL-1β) were measured by ELISA, immunohistochemistry detected intercellular adhesion molecule -1 (Intercellular adhesion molecular, ICAM-1) in the spinal cord expression changes in the organization. (2) Experiment II: 60 female SD rats were randomly divided into the simple laminectomy group (A), the damage to the control group (B), the effect of EGb761 treatment group (C) n = 20. Postoperative 1d, 3d, 7d, 14d animals in each group the BBB behavioral score and footprint experimental batches were killed, and the T9 segmental spinal cord HE staining for histological observation, the injured area and its adjacent area neuron count and measure the 14d group empty area of ??the damage zone; TUNEL and Caspase-3 immunohistochemical staining nerve cell apoptosis. Results: (1) A group each time point the spinal cord without apparent leakage of Evans blue content of IL-1β and ICAM-1 expression is maintained at a basic level; leakage of Evans blue content of, IL-1β, and ICAM-B group at each time point 1 expression were significantly higher than in group A; 6h, 24h, 72h Evans blue through the BSCB leakage into the spinal cord tissue was significantly lower than that in group B, C group EGb761 intervention at all time points of IL-1β, ICAM-1 expression than those in group B was significantly reduced. (2) A group each time point BBB score and hind legs compared to walking the footprint with normal rats were no obvious abnormalities; B, C group behavior Evaluation of no significant differences in postoperative 1d, 3d and 7d, but in the 14d , BBB score was significantly higher in group C (15.40 ± 0.89 vs 13.60 ± 0.55, P lt; 0.05). 14d after footprint analysis experiment: step C group was significantly higher than that in the control group (10.44 ± 0.26 vs 9.81 ± 0.33cm, P lt; 0.05), step width, the effect of EGb761 treatment group was significantly less than the control group (4.52 ± 0.14 vs 5.58 ± 0.11cm, P lt; 0.05); histological observation of the formation of voids in the SCI 14d C group after spinal cord tissue degeneration and necrosis area was significantly smaller than in group B (2.92 ± 0.18 vs 3.63 ± 0.27mm2, P lt; 0.05). (3) HE staining neuron cell counts and TUNEL staining of apoptotic cells count confirmed the existence of the phenomenon of widespread neuronal loss and apoptosis of spinal cord tissue after SCI. Apoptosis of nerve cells in the gray matter in the SCI 1d peaked apoptosis in white matter peaked after 7d, and glial cells; and B was significantly higher than in the SCI 7d C neurons count group (14.04 ± 1.80 vs 7.44 ± 1.15, P lt; 0.05) and apoptotic cell count was significantly lower than that in group B (58.12 ± 2.96 vs 69.52 ± 4.74, P lt; 0.05), and Caspase-3 positive cells distribution number of trends coincide. Conclusions: (1) Allen's rat model of acute spinal cord injury, EGb761 can reduce the content of IL-1β in the local spinal cord, reduced ICAM-1 expression alleviate BSCB destruction. (2) SCI pathological changes of spinal cord tissue and nerve cell apoptosis has a specific time and space distribution characteristics; early after injury (3) the use of a significant neuroprotective effect EGb761 early pathological changes of the spinal cord tissue can reduce SCI inhibit the injured area surrounding neuronal apoptosis, reduce injury Ministry spinal cord necrosis area, and the forward eventually improve hindlimb motor function.

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