2024年6月7日
In this study, nerve-mimetic composite hydrogels were developed and characterized that can be utilized to investigate and capitalize on the pro-regenerative behavior of adipose-derived stem cells for spinal cord injury repair.
In this study, a combinatorial cell delivery platform is developed for spinal cord injury using decellularized tissue-derived hydrogels. The efficacy of decellularization and cell viability in the composite hydrogels is analyzed to determine their potential as a 3D culture platform. The results will allow further investigation of therapeutic potentials for spinal cord injury.
Recently, in the field of spinal cord injury repair, decellularization has been extensively utilized in the context of hydrogels. This process removes all the cellular and nuclear debris from tissues or organs to prevent a negative immune response, while preserving the extracellular matrix components. Now that the composite nerve hydrogels are optimized, characterization of the platform will be continued to maximize the neuro-regenerative behavior of ASCs for spinal cord repair.
This work will contribute to combinatorial therapeutic development for spinal cord injury repair, especially enhancing stem cell therapy for potential clinical translation. To begin, thaw the frozen spinal cord at four degrees Celsius in the refrigerator for 18 to 24 hours before decellularization. Using sterile scissors, carefully remove the dura mater.
Cut the spinal cord into small pieces, about one centimeter in length. Place three pieces into a 50-milliliter tube. Rinse the spinal cord with deionized water at four degrees Celsius for 18 to 24 hours at 60 RPM.
Next, rinse the spinal cord with 0.025%trypsin EDTA, followed by PBS for 15 minutes twice. Then, rinse the spinal cord with the following solutions for decellularization. Finally, rinse the spinal cord with deionized water for one hour twice, followed by PBS for one hour.
Then, lyophilize the spinal cord at 0.01 millibar and 56 degrees Celsius for three days before storing it until use. Thaw the previously frozen porcine sciatic nerve at four degrees Celsius in the refrigerator for 18 to 24 hours before decellularization. Now, cut the sciatic nerve into small pieces, approximately one centimeter in length.
Place three pieces into a 50-milliliter tube. Rinse the sciatic nerve with deionized water for seven hours. Then, rinse the sciatic nerve with the following solutions for decellularization.
Then, lyophilize it at 0.01 millibar and 56 degrees Celsius for three days before storing it until use. Begin by using autoclave scissors to chop the previously decellularized porcine spinal cord and sciatic nerve into powder. Digest the tissues separately in 0.01 normal hydrochloric acid solution containing one milligram per milliliter of pepsin at a concentration of 15 milligrams per milliliter.
Place a magnetic bar in the solution and stir at 500 RPM for at least four days at four degrees Celsius to generate pregel solutions. Mix sciatic nerve and spinal cord pregel. Then, dilute the hydrogel to the desired concentration using M199 media and PBS.
Use one normal sodium hydroxide and hydrochloric acid to adjust the pH to 7.4. Once the pH is adjusted, resuspend human adipose-derived stem cells in the pregel at a density of 1 million cells per milliliter. Dilute the pregel to 12 milligrams per milliliter using PBS.
Place it into the wells, and then place the PDMS lid onto the pregel. Incubate the plate for 30 minutes at 37 degrees Celsius. Then, add PBS to the wells.
Remove the PDMS lid and the solution. Add ASC growth media to ASC-laden hydrogels and incubate at 37 degrees Celsius for culturing.
本研究开发了一种利用脱细胞组织来源水凝胶的组合式细胞递送平台,用于脊髓损伤修复。对这些水凝胶进行了表征,以评估其作为脂肪来源干细胞三维培养平台的功效和潜力。
整合脱细胞神经基质与干细胞递送的组合型水凝胶平台,可应对脊髓损伤修复中的多种复杂挑战。该方法能够构建与疾病相关的三维系统,用于评估神经再生策略,从而提高早期治疗开发的可预测性。该平台的模块化特性使其适用于再生医学研发领域的广泛产品组合。
这种基于水凝胶的平台可融入从发现到临床前的研究连续体,支持对神经再生疗法进行迭代假设验证和机制性风险降低。