紅景天與紅景天苷對於NMDA所引起視網膜傷害之預防效果

Protective effects of Rhodiola rosea and Salidroside against NMDA-induced retinal injuries

林宜蓁(Yi-Jen Lin)

指導教授 ： 張菡馨

中山醫學大學/健康管理學院/營養學系碩士班/碩士(2017年)

摘要

大花紅景天(Rhodiola crenulata)為常食用的中藥材，其中富含紅景天苷（Salidroside）、苷元酪醇（Tryosol）、没食子酸(Gallic acid)及其他酚類化合物，這些多酚類物質具有極佳抗氧化效果。近幾年青光眼的治療方向不再只是降低眼壓，轉而朝向減少神經細胞損傷成為主要的治療目標，而造成神經細胞損傷並且逐漸死亡的其中原因是興奮性毒性(Excitoxicity)。興奮性毒性之作用乃是透過麩氨酸(Glutamate)活化細胞上的麩氨酸受器(如NMDA)，造成過多的鈣離子流入細胞，使細胞出現粒線體的功能障礙(Mitochondrial Dysfunction)，進而產生強烈的氧化壓力(Oxdiative stress)而促使細胞走上凋亡路徑。
根據各項文獻研究結果，本研究推測大花紅景天中所含之各種功效成分，因其強抗氧化能力，能幫助清除組織細胞中過多的自由基而減少氧化傷害，可能幫助減緩青光眼的病程進展。本研究以小鼠預先餵食紅景天萃取物及其主要功效成份Salidroside，再以玻璃體注射40 mM NMDA誘導老鼠視網膜產生傷害，探討紅景天與Salidroside是否對NMDA所誘發的視網膜損傷具有保護功效。首先觀察NMDA傷害24小時、7天後視網膜電位圖(Electroretinogram,ERG)、視網膜病理、及蛋白質表現量，發現NMDA傷害24小時後視網膜視神經節細胞(RGC)數量明顯減少，以及內叢狀層(IPL)的厚度明顯變厚，另外於視神經節細胞層(GCL)與內核層(INL)皆可觀察到氧化指標8-OHdG與MDA的訊號表現，且視網膜細胞凋亡相關因子Caspase-8、Capase-3被大量活化、發炎因子Cox-2表現量也明顯增加；受傷後7天視網膜的內叢狀層(IPL)與內核層的厚度顯著變薄，在視網膜電位圖測試中，傷害組的a、b波的波幅均有下降之現象。
接著測試紅景天及其主要功效成分Salidroside，是否可預防因NMDA引起的視網膜損傷，預先給予紅景天(100mg/kg)與Salidroside(50mg/kg)的傷害組，可減少視網膜損傷後24小時細胞凋亡數量，以及避免內叢狀層的厚度變薄，並且改善視網膜氧化壓力情形、降低8-OHdG與MDA的訊號表現，此外對於細胞凋亡路徑與發炎反應的變化均有減緩之趨勢。另一方面預先食用紅景天或Salidroside的對照組，對視網膜的各項檢測並未造成任何影響。上述結果可推論紅景天與Salidroside的補充對視網膜因興奮性神經毒性而引起之損傷具保護作用。

關鍵字

青光眼 ； NMDA ； 神經興奮性毒性 ； 視網膜 ； 紅景天 ； 紅景天苷

並列摘要

Rhodiola crenulata is widely used in traditional Chinese medicine. It is rich in salidroside, tryosol, gallic acid, and other polyphenols that are considered to have high antioxidant ability. The main goal of glaucoma treatment has been to reduce the intraocular pressure. Recently, however, prevention against neuronal cell damage becomes a major therapeutic target.
The mechanism of neuronal cell death in glaucoma is excitotoxicity triggered by N-methyl-D-aspartate receptor (NMDA receptor) overexpression, followed byexcessive Ca2+ influx inside cells. The accumulation of Ca2+ causes mitochondrial dysfunction, increase of oxidative stress, and finally leads to neuronal apoptosis. This study aims to investigate whether oral administration of Rhodiola crenulata and its effective ingredient salidroside may help to reduce glaucomatous damages caused by NMDA receptor overexpression in a mouse model.

Firstly, the visual function as represented by electroretinogram (ERG), retinal pathology, and protein expression were assessed after intravitreal injection of 40 mM NMDA after 24 hours and 7 days. At 24 hours after injection, ganglion cell layer (GCL) cell number was significantly decreased and the thickness of inner plexiform layer (IPL) was increased. The numbers of 8-OHdG and MDA positive cells were increased in GCL and inner nuclear layer (INL). With the injection, elevated expression of Caspase-8, Caspase-3 and Cox-2 was also observed as detected by Western blot analysis. By 7 days after the injection, the thickness of IPL and INL was reduced significantly. In the aspect of visual function, NMDA injury caused lower a-wave and b-wave amplitude in ERG performance.

Next, the protective effect of Rhodiola and salidroside against NMDA-induced retinal damages and their overdose effects were tested. Treatment with Rhodiola (100 mg/kg) and salidroside (50 mg/kg) significantly decreased cell death in the retina, restored the thickness of IPL, and improved the status of high retinal oxidative pressure, as shown by decreased MDA and 8-OHdG positive cells. In addition, Rhodiola and salidroside treatment reduced both apoptosis related protein expression and inflammatory reaction. On the other hand, Rhodiola and Salidroside did not exert any negative impacts on the retina. These results indicate that Rhodiola and Salidroside protect retinal neurons against excitotoxic effects induced by NMDA intravitreal injection in mice.

並列關鍵字

Glaucoma ； NMDA ； Excitotoxicity ； Retina ； Rhodiola ； Salidroside

參考文獻

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