A kind of pharmaceutical composition for treating spinal cord injury and repairing and application thereof

A technology for spinal cord injury and therapeutic effect, applied in the field of pharmaceutical compositions for treating spinal cord injury and repairing

Active Publication Date: 2022-07-22
JINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to solve the technical problems existing in the treatment process of spinal cord injury repair in the prior art, thereby providing a new treatment target for spinal cord injury repair, and investigating the function and mechanism of Rab3A in the process of spinal cord injury repair In-depth research has revealed the key role of Rab3A in the repair and treatment of spinal cord injury, providing a practical theoretical basis and direction for the clinical treatment of spinal cord injury

Method used

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  • A kind of pharmaceutical composition for treating spinal cord injury and repairing and application thereof
  • A kind of pharmaceutical composition for treating spinal cord injury and repairing and application thereof
  • A kind of pharmaceutical composition for treating spinal cord injury and repairing and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Example 1 Preparation of plasmids and constructs

[0071] (1) Rab3A and spastin sequences were obtained for preparation of cDNA, which were then cloned into pEGFP-C1 (Clontech, CA, USA), pGEX-5x-3 (Amer-sham Pharmacia Biotech, NJ, USA) and pCMV-Tag2 (Stratagene, CA, USA) vector, and the constructed construct was confirmed by DNA sequencing method;

[0072] (2) Pull-down analysis using glutathione S-transferase (GST): Spinal cord tissue sections were ground and lysed, then GST-agarose beads (Invitrogen, CA, USA) were rinsed and mixed with lysis buffer at 4 Incubate for 1 h at °C, then centrifuge at 1000 g for 10 min. 4°C. The supernatant was then collected and these steps were repeated one more time. Appropriate bead fusion proteins were then added to the spinal cord tissue, and the samples were incubated overnight at 4°C. After spinning at 1000 g for 5 min at 4°C, unbound protein was washed with 1 mL of wash buffer, and then the samples were spun at 1000 g for 1 min...

Embodiment 2

[0076] Example 2 Construction of rat SCI model

[0077] The Sprague-Dawley (SD) rats used in the experiment were purchased from the Laboratory Animal Center of Sun Yat-Sen University. The rats were housed individually in a facility at 25 ± 3 °C and provided with regular food and water. Specifically, the following steps were included:

[0078] (1) SD rats were anesthetized by intraperitoneal injection of 10% chloral hydrate (0.35 mL / 100 g body weight), after which a 2.5 cm longitudinal dorsal incision was made to expose the T9-11 spinous process and lamina;

[0079] (2) The entire T10 lamina is removed, and the exposed area of ​​the spine is about 2.5mm × 3mm;

[0080] (3) Use a stabilizer to fix the T10 section bilaterally, and set the nitrogen tank to control the impact head to 18psi or 124kPa. Load a U-shaped stabilizer with a rat onto the platform of the Louisville Injury System Equipment (LISA) and adjust the dura / spinal height directly below the impactor while monitoring...

Embodiment 3

[0085] Example 3 Rab3A regulates neuron growth and development

[0086] Firstly, the research on the growth and development of neurons by Rab3A overexpression includes the following steps:

[0087] (1) GFP-Rab3A and GFP (blank control group) were transfected into primary cultured hippocampal neurons for 72 hours, respectively;

[0088] (2) After 48 hours, the hippocampal neurons were fixed and photographed, and the scale was 100 μm.

[0089] The result is as Figure 1-3 shown, where figure 1 Schematic diagram of the immunofluorescence staining results of hippocampal neurons, figure 2 Schematic diagram of the quantitative statistics of the length of axons and neurites in hippocampal neurons, image 3 Schematic diagram of the quantitative statistical results of the number of axons and processes in hippocampal neurons, n=20 / group, the results are represented by Mean±SD, and * represents p <0.05. The scale bar is 100 μm. According to the results, overexpression of Rab3A in...

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Abstract

The present invention relates to a pharmaceutical composition for treating and repairing spinal cord injury and its application. The present invention finds through research that Rab3A plays a key role in SCI, and interacts with Spastin to regulate neurite outgrowth. By identifying proteins that were differentially expressed in SCI, it was demonstrated that Rab3A expression levels were downregulated during SCI. In addition, it was also found that Rab3A can physically interact with Spastin and regulate Spastin degradation through the lysosomal pathway, thereby affecting Spastin function. Collectively, these findings highlight a signal transduction pathway in which Rab3A-mediated spastin degradation regulates neurite branch formation and growth. Since SCI is a major cause of disability, repairing the structural defects of the spinal cord caused by injury or degeneration is crucial in the modern field of regenerative medicine. A variety of pharmacological interventions can be designed to treat SCI and assist with associated tissue repair, and can be combined with other cellular interventions.

Description

technical field [0001] The present invention relates to the field of biomedicine, in particular, the present invention relates to a pharmaceutical composition for treating and repairing spinal cord injury and its application. Background technique [0002] Spinal cord injury (SCI) can lead to permanent spinal cord dysfunction, seriously affecting the quality of life of patients. The main cause of dysfunction after SCI is the direct mechanical damage leading to neuronal damage, and subsequent secondary damage, including inflammatory response, oxidative stress response and excitatory damage, etc., resulting in neuronal axonal damage and glial damage. Plasma cells proliferate, eventually leading to neuronal cell death. Neurological dysfunction caused by SCI is permanent because after spinal cord tissue damage, the ability of axons to regenerate is inhibited and it is difficult to pass through the damaged area, thereby irreversibly impairing the transmission of motor and sensory...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K45/00A61K31/713A61K38/17A61K49/00A61P25/00G01N33/68G01N33/53
CPCA61K45/00A61K31/713A61K38/1709A61K49/0008A61P25/00G01N33/6893G01N33/53G01N2800/28
Inventor 张国威陈云杨裕豪林宏生
Owner JINAN UNIVERSITY
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