研究胜肽水凝膠在受損神經元細胞模型中的應用

Abstract

水凝膠可用於模擬細胞外基質（ECM）骨架結構，因此被廣泛應用於三維細胞培養。此研究中，我們從有自組裝特性的天然生物蛋白質挑選，並合成出同樣有自組裝特性的胜肽且應用在神經細胞培養的基材。在先前實驗室的研究中，我們發現胜肽（JTH202）能夠在生理條件下進行自組裝形成水凝膠。微觀下，水凝膠主要是由β-折疊組成的纖維狀結構。此外，我們使用流變儀探討了水凝膠的機械性質，並研究其自癒合及可注射性的能力。在確認水凝膠的性質後，我們在水凝膠上培養神經細胞（Neuron-2a cell），透過顯微鏡觀察，發現神經細胞可以在水凝膠上貼附以及生長。透過細胞存活率的測試，我們發現在水凝膠培養神經細胞具有高的存活率，說明水凝膠有良好的生物相容性。在三維細胞培養中，我們藉由高濃度谷氨酸誘導神經毒性，從而建立一個簡單的神經細胞受損模型。同時，我們將水凝膠當作藥物載體，用於包覆具有神經保護作用的小分子。實驗發現，包裹藥物的水凝膠可以有效的提升細胞存活率並降低谷氨酸誘導的細胞毒性。接著，我們透過清除自由基實驗（DPPH assay）測試發現，水凝膠具有高度抗氧化能力，因此在高濃度谷氨酸作用下，神經細胞在水凝膠上仍能有不錯的存活率。總括來說，JTH202胜肽水凝膠具有高度生物相容性，並可包覆小分子藥物，使水凝膠上貼附的細胞受益於其神經保護作用。這些結果表明，水凝膠在模擬細胞外基質（ECM）骨架結構方面具有潛在的應用價值，並在三維細胞培養中展現出良好的性能。Peptide hydrogels can be used to mimic the extracellular matrix (ECM), and they are widely applied in three-dimensional cell cultures nowadays. In this study, we synthesized a peptide derived from natural animal protein with self-assembly characteristics for application in neuronal cell culture. In our previous work, we found that the peptide (JTH202) can self-assemble to form a hydrogel under physiological conditions. Microscopically, the hydrogel consists mainly of β-sheet fibrous structures. Additionally, we investigated the mechanical properties of the hydrogel by rheometer and studied its self-healing and injectability abilities.After confirming the properties of the hydrogel, we cultured Neuron-2a cells on the hydrogel. It was observed that the neuronal cells could attach and grow on the hydrogel with very good cell viability, indicating the good biocompatibility of the hydrogel. In three-dimensional cell culture, we established a simple neuronal cell injury model by inducing neurotoxicity with a high concentration of glutamate. At the same time, we used hydrogel as a drug carrier to encapsulate small molecules with neuroprotective effects. The experiments showed that the drug-loaded hydrogel effectively enhanced cell viability and reduced glutamate-induced cytotoxicity. Furthermore, we discovered that the hydrogel exhibited high antioxidant capacity by DPPH assay. Therefore, even under the high concentrations of glutamate, the neuronal cells cultured on the hydrogel still exhibited good cell viability.In summary, the JTH202 peptide hydrogel demonstrated high biocompatibility and the ability to encapsulate small molecule drugs, thereby enabling the cells attached to the hydrogel to benefit from its neuroprotective effects. These results indicated the potential utility of hydrogels in mimicking the extracellular matrix (ECM) and their excellent performance in three-dimensional cell culture.