低能量光照對骨髓間葉幹細胞抗衰老之影響探討

The Effect of Low Level Light Irradiation on Senescence Status of Bone Marrow Derived Mesenchymal Stem Cells

劉家亨(Jia-Heng Liu)

指導教授 ： 李文婷

中原大學/工學院/生物醫學工程研究所/碩士(2017年)

https://doi.org/10.6840/cycu201700846

摘要

本研究以D-半乳糖 (D-galactose, D-gal) 誘導骨髓間葉幹細胞 (Mesenchymal stem cells, MSCs) 老化為模型，探討低能量光照 (Low level light irradiation, LLLI) 對幹細胞抗老化之影響。首先檢測不同濃度D-gal 處理 MSCs 後的細胞存活率、老化相關β-半乳糖苷酶 (Senescence-associated-β-galactosidase, SA-β-gal) 活性、活性氧物種 (Reactive oxygen species, ROS) 與丙二醛 (Malondialdehyde, MDA) 含量，結果發現 SA-β-gal 活性、ROS 與MDA皆隨著 D-gal 的濃度提高而增加，細胞存活率則隨之下降。接著，以紅光 (630 nm) 與近紅外光 (850 nm) 照射，發現與控制組相比，SA-β-gal 活性與 MDA 含量下降，細胞存活率則有些微提升。顯示 D-gal 可提高細胞內氧化壓力，誘導幹細胞趨向老化；而光照治療可降低氧化壓力，延緩細胞老化發生。另外也探討 LLLI 對 MSCs 老化相關基因 p53、p21、p16 表達的影響以及硬骨分化能力的檢測。結果發現，p53、p21、p16 老化相關基因的表達會隨著 D-gal 濃度提高而增加，並且經由 LLLI 後，其老化相關基因的表達則會下降。而在硬骨分化方面，隨著 D-gal 濃度的增加，鹼性磷酸酶 (Alkaline phosphatase, ALP) 的活性及鈣沉積量皆會受到抑制；經由 LLLI 後，早期硬骨分化指標 ALP 的活性雖然低於控制組，但晚期分化指標鈣沉積量高於控制組。綜合以上結果，D-gal 可以誘導 MSCs 趨向老化，進而降低其生長及分化能力，而經由 LLLI 光照治療後，可有效延緩細胞老化發生，並且促進 MSCs 的增生及硬骨分化能力。

關鍵字

低能量光照 ； 間葉幹細胞 ； D-半乳糖 ； 抗老化 ； 硬骨分化

並列摘要

The study used D-galactose (D-gal) treated bone marrow mesenchymal stem cells (MSCs) as an aging model, and aimed to explore the influence of low level light irradiation (LLLI) on anti-senescence of stem cells. We first determined cell viability, activity of senescence-associated- β-galactosidase (SA-β-gal), reactive oxygen species (ROS) and malondialdehyde (MDA) level on D-gal treated MSCs. SA-β-gal activity, ROS and MDA level were found to increase with increasing concentration of D-gal, whereas cell viability decreased. After red (630 nm) and near-infrared (850 nm) light irradiation, MDA concentration and SA-β-gal activity declined, whereas cell viability elevated as compared to the control group. The results suggest that D-gal treatment could induce senescence of MSCs via oxidative stress. LLLI could delay MSCs senescence by reducing oxidative stress. In addition, the effect of LLLI to MSCs senescence gene, p53, p21, p16 expression and osteogenic marker expression was tested. The results showed p53, p21, and p16 senescence gene expression increased when increasing D-gal concentration while it decreased after LLLI. In osteogenic marker expression, increasing D-gal concentration inhibited alkaline phosphatase (ALP) and calcium deposition expression. Although ALP expression was lower than control group, calcium deposition expression was higher than control group after LLLI. Overall, D-gal is able to induce MSCs senescence and reduces MSCs growing and osteogenic differentiation ability. LLLI treatment can effectively delay MSCs senescence and induced MSCs growing and osteogenic differentiation.

並列關鍵字

low level light irradiation ； mesenchymal stem cell ； D-galactose ； anti-senescence ； osteogenesis differentiation

參考文獻

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