| BackgroundSpinal cord injury (SCI), which refers to severe damage to the nerve system with potential consequences of permanent disability even to death, presents a clinical challenge to the current medical technology. Currently, an increasing number of people suffer diabetes in our country, Some studies show that diabetic patients have more trouble recovering from nerve damage as a result of SCI, but the molecule mechanism involved remains unknown. With regard to the nerve cell death in the injured segments of SCI, previous studies believe that it is a result of necrosis. Meanwhile, some recent studies show that it is caused by apoptosis of nerve cells. The endoplasmic reticulum (ER) is an organelle that ensures correct protein folding and assembly by expressing numerous molecular chaperones. The ER may be important for regulating intracellular apoptotic signals in neurons. Apoptosis also occurs upon endoplasmic reticulum (ER) stress. The research has proved that hyperglycemia may contribute to the apoptosis induction pathway associated with endoplasmic reticulum stress. The glucose-regulated protein (GRP78) is the chaperone of Caspase-12. The unfolded protein response can contribute to the salient expression of GPR78; integrate the misfolded and unfolded protein; restore the protein conformation and sustain inner environment. The signal transduction of GPR78is of great significance in protecting cells in ER stress. The precursor molecule of Caspase12, located on the surface of endoplasmic reticulum membrane, is activated by splitting certain locations in the process of endoplasmic reticulum stress, which triggers cell apoptosis. Therefore, Caspase-12and GRP78is a typical endoplasmic reticulum apoptin.We make a model of acute SCI of diabetic rats and non-diabetic rats to measure the expression changes of rake motor neuron Caspase-12and GRP78of T10gray substance spinal cord after injury24h and7d to explore the possible molecule mechanism of endoplasmic reticulum stress as diabetic rats suffer SCI, and the impact of hyperglycemia on rats’cell apoptosis after its spinal injury as well. We hope to provide animal experiment data to aid the treatment of diabetic patients with SCI.ObjectivesThe study aims to measure the expressions of Caspase-12and GRP78in rake motor neuron of injury T10gray substance spinal cord after diabetic rats sustain mechanical injury24h,7d to their spinal cords. In contrast to the rats with normal spinal injury, the study also intends to explore the function of endoplasmic reticulum stress in acute mechanical SCI to rats and the related impact from hyperglycemia.Methods60healthy adult SD rats are divided into3groups at random.(1) Sham operation control group,(2) SCI group,(3) Diabetic group. In sham operation control group, T10omni-posterior excision is performed without the spinal cord intact; in SCI group, the improved Allen method is employed to strike the T10segment of the spinal cord. In the group of diabetic rats, a diabetes model is developed by injecting STZ into the rats’ caudal veins and hyperglycaemia is maintained for6weeks before the improved Allen method is employed to strike the T10segment of the spinal cord. The BBB classification is used for grading, the morphology of gray substance spinal cord is employed for observation, and Immunohistochemistry are also involved in observing the changes in the motor function, pathomorphology of SCI and Caspase-12,GRP78 expressions after rats sustain mechanical injury24h and7d to their spinal cords.Results1. The control group of sham operation displays no motor dysfunction after the operation, scoring21points in terms of24hours post-operation; the group of diabetic rats with SCI and the group of non-diabetic rats with SCI display symptoms of paraplegia and severe motor dysfunction24hours after the operation, scoring from0to1in terms of BBB classification method. In terms of statistics, there’s no difference between the latter two groups (P=0.000, n=20) while both of them presents notable discrepancy with the sham group (P<0.01, n=20). After injury7d, the sham operation control group remains to score21the group of non-diabetic rats with SCI scores from6to8, accompanied by function recovery while the group of diabetic rats with SCI takes more time in function recovery, scoring from3to5in terms of BBB classification method.2. HE dyeing observation of spinal cord tissue slice shows that the control group of rats is close-knit in tissues, clear in distinction between the white matter and gray matter and intact in neuron pattern in terms of24h and7d post-operation. The group of non-diabetic rates with SCI is bleeding in rake gray matter, swelling in tissue and putresceing in neuron24h after operation. It develops a spindle necrotic area on the long axis of the spinal cord, cystic degeneration and spongiocyte hyperplasia7d post-operation. Patchy hemorrhage in rake gray matter, neuron apoptosis are seen in the group of diabetic rats24h after-operation, with less neuron compared with the non-diabetic rates. It develops immense vacuoles in anterior horn of spinal cord, accompanied with tissue looseness, neuron disappearance and macropages-infiltrated cavities7d after operation.3.Immunohistochemistry results show average numbers of Caspase-12positive cells in the control, non-diabetic and diabetic group are31.4±4.4/mm2,70.0±18.2/mm,90.0±5.8/mm respectively., which present a significant discrepancy in statistics (P<0.01, n=20)24hours after operation. Those figures become30.5±3.9/mm2,38.0±4.2/mm2,30.0±3.8/mm2respectively7d post-operation, showing no clear statistical differences. The average number of positive cells of GPR78in the three different groups are respectively10.1±2.8/mm2,35.0±13.2/mm2,12.0±3.3/mm224after SCI, which demonstrates an apparent discrepancy between non-diabetic and the other two (P<0.01,n=20). Those figures become12.1±3.4/mm2,14.2±3.2/mm2,13.0±3.8/mm2respectively7d post-operation, presenting no clear statistical differences(P>0.05,n=20).Conclusion1. Endoplasmic reticulum stress is triggered by SCI in view of the increasing expression of Caspase-12and GRP78in rats after the SCI. Meanwhile, the early ascent of GPR78is a self-protection mechanism initiated by cells and the increasing expression of Caspase-12promotes the spinal cell apoptosis.2. It is likely that the hyperglycemia of diabetic rats may associate with the descent of GPR78and ascent of Caspase-12considering those numbers in diabetic rats and non-diabetic rats respectively. |
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