Experimental Study On Spinal Cord Ischemic Injury

inflammatory cascade. Studies in a number of inflammatory states demonstrated that pentoxifylline markedly reduced accumulation of TNF-a mRNA in monocytes and T cells and so the production.This study is aim to investigate the effect of necrosis and apoptosis of neuron, the possible role of TNF-a in spinal cord ischemic injury, and to evaluate the functional protection of PTX against spinal cord ischemic injury in rabbits.This paper include two part. Part I is conserning double role of tumor necrosis factor-a(TNF-a) of inducing neuron necrosis and apoptosis in the spinal cord ischemic injury. Part II is about functional protection by pentoxifylline against spinal cord ischemic injury.MethodsPart I The rabbits sustained spinal cord ischemia with 45 minutes cross-clamping of the infrarenal aorta. Groups were as follows: sham operation(n=18); ischemia(n=30),receiving only vehicle; anti TNF-a mAb(n=30),receiving anti TNF-a mAb before clamping. At different time points after injury, serum was assayed with ELISA for TNF-a and spinal cords were harvest for MPO activity, histopathologic analysis, TUNEL staining, immunohistochemistry for PEC AM-1 and Caspase-3. The number of necrotic and apoptosic neuron were counted and then to be analysed. Morphological change of neuron was observed with transmission electron microscope. DNA was extracted and observed by gel electrophoresis.Part II The rabbits also sustained spinal cord ischemia with 45 minutes cross-clamping of the infrarenal aorta. Groups were as follows: sham operational 2); ischemic control(n=20), receiving normal salin before clamping; PTX I (n=20), receiving PTX before clamping and PTX II (n=20), receiving PTX at the onset of reperfusion. Rabbits were evaluated for hind-limb motor function with the modified Tarlov scoring system at 48h. The active area was measured by TTC staining. Serum was assayed with ELISA for TNF-a and spinal cords were harvest for MPO activity,histopathologic analysis, TUNEL staining, immunohistochemistry for PEC AM-1 and Caspase-3, and the number of necrotic and apoptosic neuron were counted and then the results were analysed at 12h, 24h, 48h and 72h after injury. Blood gas and glucose also measured at preischemia, intraischemia and postischemia time.ResultsPart I In ischemia group, TNF-a in serum increased at 1 h and reached the peak at 4 h after injury. The active of MPO in tissue increased from 1 h to 72 h after injury. In immunohistochemistry observation, PECAM-1 was found in the endothelial cells mainly in the medial membrance and the junction of cells; Caspase-3 expressed in the neuron cytoplasm only at 12 h and reduced markedly at 72 h after injury. Necrotic neurons were founded at 1 h but apoptosic neurons only were founded at 24 h and reduced markedly at 72 h. The number of necrotic neuron was higher than the apoptosic neuron (P<0.05 ). In the anti TNF-a mAb group, the previous detection was not serious (P<0.05) . Morphological change of neuron necrosis and apoptosis was observed with transmission electron microscope. DNA Ladder was not found by gel electrophoresis.Part II Improved Tarlov scores were seen in PTX-treated rabbits as compared with ischemic control rabbits at 48h after injury. A significant reduction of TNF-a was found in serum, active of MPO and immunoreactive of the PECAM-1 and Caspase-3 in PTX-treated rabbits. The number of necrotic and apoptosic neuron was higher in PTX-treated rabbits than in the ischemic control rabbits(P<0.05). No necrotic and apoptosic neuron were founded in the sham operation group. The active area in TTC staining slice is higher in PTX-treated rabbits than in the ischemic control rabbits (P<0.05). Blood gas and glucose changed markedly at postischemia time.Conclusions 1. A major mode of neuron death in spinal cord ischemic injury was necrosis, butdenying the role of apoptosis completely was also false.2. TNF-a can induce both neuron necrosis and apoptosis in spinal cord ischemic injury.3. PTX can induce protection against ischemic injury in the spinal cord by preventing both necrosis and apoptos...