褐藻醣膠改善餵食低蛋白飲食老年小鼠肌肉萎縮之影響

肌少症(Sarcopenia)已被認為是一種新的老年性疾病，其特徵在於骨骼肌質量和功能的漸行性和普遍性喪失，65歲以上國人罹患肌少症比例為7-10%，80歲以上男性罹患率更高達近30%。國民營養調查發現，衰弱老人六大類食物中，奶類及蔬果類為攝取不足盛行率最高的前2名，與肌少症發生息息相關。研究發現，肌肉流失主要原因是膳食蛋白質缺乏與發炎，攝取較高蛋白質的老人，肌肉質量降低速度較慢，衰弱風險及症候都較少。骨骼肌肉流失與泛素-蛋白酶體途徑(Ubiquitin-proteasome pathway, UPP)與自噬作用-溶酶體途徑(Autophagy-lysosome pathway, ALP)增加蛋白質的異化有密切關係。小分子褐藻醣膠(Fucoidan)是一種天然硫酸化多醣，存在於不同種類的褐藻，具有多種生物活性。由於動物實驗已證實，小分子褐藻醣膠可顯著減緩癌症合併化學治療所產生的肌肉耗損，故本實驗目的為評估褐藻醣膠對於改善餵食低蛋白飲食老年小鼠肌肉萎縮之功效。92週大C57BL/6J小鼠(相當於人類的64歲)，分成給予一般飼料控制組、低蛋白飲食組及低蛋白飲食合併管灌餵食褐藻醣膠(300 mg/kg)，12週後犧牲取腓腸肌、股四頭肌、脛前肌、伸趾長肌和比目魚肌。結果顯示，相較控制組，低蛋白飲食的小鼠體重和肌肉重量顯著降低，且後腿骨骼肌肉束橫切面積顯著減少，而褐藻醣膠的補充顯著改善低蛋白飲食小鼠造成的體重減輕與肌肉流失 (p<0.05)。此外，相較控制組，低蛋白飲食的小鼠腓腸肌Myosin heavy chian (MyHC)蛋白質表現降低，UPP相關因子MuRF1和ALP相關因子LC3Ⅱ、Beclin1和Caspase-3的表現、磷酸化p38及轉錄因子Nuclear factor kappa B (NF-кB)、signal transducer and activator of transcription 3 (STAT3)及CCAAT/enhancer binding protein-δ (C/EBP-δ)核蛋白質表現增加，而補充褐藻醣膠可顯著改善上述分子表現。與低蛋白飲食小鼠相比，補充褐藻醣膠可提高小鼠肌肉內抗氧化酵素glutathione reductase、catalase、uinone oxidoreductase 1、superoxide dismutase1、superoxide dismutase2表現，減少肌肉TNF- mRNA表現及紅血球、肝臟、股四頭肌脂質過氧化物含量 (p<0.05)。此外，相較於15週齡年輕小鼠，104週齡老年小鼠腓腸肌MyHC與抗氧化酵素蛋白質表現較低，ALP相關因子LC3Ⅱ、Beclin1和Caspase-3的蛋白質表現較高(p<0.05)。綜合以上結果可知，補充褐藻醣膠可能透過減少肌肉中p38 MAPK訊息傳遞、氧化壓力與發炎反應，減少轉錄因子NF-кB、STAT3及C/EBP-δ，降低UPP及ALP相關分子表現，同時提高抗氧化酵素蛋白的表現，改善低蛋白飲食老年小鼠的肌肉萎縮。

According to World Health Organization definition, Taiwan has become aging society. Aging causes health problems, especially sarcopenia which affects the elderly’s mobility, cognitive function, quality of life, risk of fall, morbidity and mortality. Sarcopenia has been considered a new senile disease, characterized by the gradual and universal loss of skeletal muscle mass and function. The study found that the main cause of muscle loss is dietary protein deficiency and inflammation. Skeletal muscle loss is closely related to ubiquitin-proteasome pathway (UPP) and autophagy-lysosome pathway (ALP) increase protein catabolism. Low molecular weight Fucoidan is a natural sulfated polysaccharide isolated from the brown algae. Fucoidan has multiple health benefits. Previous data have shown that Fucoidan can significantly alleviate muscle loss caused by cancer combined with chemotherapy. The purpose of this experiment is to evaluate the efficacy of Fucoidan in improving sarcopenia in elderly mice feed a low protein diet. 92-week-old C57BL/6J mice (equivalent to 64-year-old human age) were fed chow diet, low protein diet, and a low protein diet combined with garage feeding Fucoidan (300mg/kg, 3 times/week) for 12 weeks. When compared with the chow diet control group, the body weight and muscle weight, as well as cross-sectional area of quadriceps of mice with low protein diet were significantly reduced. Supplementation of Fucoidan significantly improve weight loss and skeletal muscle loss in low protein diet mice. Moreover, compared with control group, myosin heavy chain (MyHC) expression in the gastrocnemius of low protein diet mice was decreased. In addition, UPP-related factors MuRF1 and ALP-related factors microtubule-associated protein 1B light chain 3 (LC3Ⅱ),Beclin1 and Caspase-3, phosphorylated p38 and transcription factor Nuclear factor kappa B (NF-кB), signal transducer and activator of transcription 3 (STAT3) as well as CCAAT/enhancer binding protein-δ (C/EBP-δ) nuclear protein expression was increased in the gastrocnemius of low protein diet, while supplementing with Fucoidan can significantly improve expression of the above protein. Compared with low protein diet mice, supplementation of Fucoidan can increase expression of antioxidant enzymes such as glutathione reductase, catalase, quinone oxidoreductase 1, superoxide dismutase1, superoxide dismutase2, and reduce inflammatory cytokine, TNF-α expression as well as the content of lipid peroxides in red blood cells, liver and quadriceps. In addition, compared with 15-week-old young mice, protein expression of MyHC and antioxidant enzyme in the gastrocnemius of 104-week-old mice was reduced, and protein expression of LC3Ⅱ, Beclin1 and Caspase-3 were increased. In summary, Fucoidan supplementation may through lessened p38 MAPK activation, oxidative stress and inflammation in skeletal muscle reduced NF-кB, STAT3 and C/EBP-δ transcriptional activity which resulted in a decline of the expression of UPP and ALP-related factors and an increase of MyHC expression and muscle weight in aging mice fed with low protein diet.