中国医药
中國醫藥
중국의약
CHINA MEDICINE
2014年
9期
1388-1392
,共5页
胡阳黔%李静%刘坚%王文峰%宋杰
鬍暘黔%李靜%劉堅%王文峰%宋傑
호양검%리정%류견%왕문봉%송걸
黄芪多糖%游离脂肪酸%腺苷酸活化蛋白激酶%脂肪酸结合蛋白4%巨噬细胞%脂质沉积
黃芪多糖%遊離脂肪痠%腺苷痠活化蛋白激酶%脂肪痠結閤蛋白4%巨噬細胞%脂質沉積
황기다당%유리지방산%선감산활화단백격매%지방산결합단백4%거서세포%지질침적
Astragalus polysaccharides%Free fatty acid%Adenosine monophosphate activated protein kinase%Fatty acid binding protein 4%Macophage%Intracellular lipid accumulation
目的：探讨黄芪多糖对高游离脂肪酸导致巨噬细胞内脂质沉积的作用及机制。方法将培养的单核源性巨噬细胞（ THP-1）分为对照组（加入与游离脂肪酸组等量的无水乙醇及牛血清清蛋白）、黄芪多糖组（培养液中加入黄芪多糖200 mg/L）、游离脂肪酸组（培养液中加入长链游离脂肪酸0．25 mmol/L）、游离脂肪酸加黄芪多糖组（培养液中加入黄芪多糖200 mg/L及长链游离脂肪酸0．25 mmol/L）及游离脂肪酸加黄芪多糖及腺苷酸活化蛋白激酶（AMPK）阻断剂（compound C）组（培养液中加入黄芪多糖200 mg/L、长链游离脂肪酸0．25 mmol/L及compound C 50μmol/L）。采用游离脂肪酸检测试剂盒和三酰甘油试剂盒分别检测游离脂肪酸及三酰甘油含量，油红O染色观察细胞内脂质蓄积，蛋白质印迹法检测细胞内AMPK、磷酸化腺苷酸活化蛋白激酶（p-AMPK）、脂肪酸结合蛋白4（FABP4）的蛋白表达情况。结果①游离脂肪酸组三酰甘油及游离脂肪酸含量[（3．35±0．79）mmol/L、（29．3±1．7）μmol/L]明显高于对照组[（0．81±0．19）mmol/L、（15．7±1．3）μmol/L]和游离脂肪酸加黄芪多糖组[（1．46±0．25）mmol/L、（15．5±2．1）μmol/L]，差异均有统计学意义（均P＜0．01）。②油红O染色结果：游离脂肪酸组脂质含量较对照组增多，游离脂肪酸加黄芪多糖组脂质含量较游离脂肪酸组减少，提示游离脂肪酸增加细胞内脂质蓄积，而黄芪多糖减少细胞内脂质蓄积。③蛋白质印迹法检测AMPK、p-AMPK、FABP4的蛋白表达情况发现，与对照组相比，黄芪多糖组各种蛋白表达无明显变化，游离脂肪酸组p-AMPK表达明显少于对照组和游离脂肪酸加黄芪多糖组[（54．3±3．0）％比（100．0±1．4）％、（96．5±3．9）％]，FABP4表达明显高于对照组和游离脂肪酸加黄芪多糖组[（138．3±2．3）％比（100．0±3．1）％、（72．3±2．6）％]；与游离脂肪酸加黄芪多糖组比较，游离脂肪酸加黄芪多糖及compound C组FABP4表达明显增多[（136．9±4．3）％比（72．3±2．6）％]，差异均有统计学意义（均P＜0．01）。结论黄芪多糖可以通过AMPK-FABP4途径减少高游离脂肪酸导致的巨噬细胞内脂质蓄积。
目的：探討黃芪多糖對高遊離脂肪痠導緻巨噬細胞內脂質沉積的作用及機製。方法將培養的單覈源性巨噬細胞（ THP-1）分為對照組（加入與遊離脂肪痠組等量的無水乙醇及牛血清清蛋白）、黃芪多糖組（培養液中加入黃芪多糖200 mg/L）、遊離脂肪痠組（培養液中加入長鏈遊離脂肪痠0．25 mmol/L）、遊離脂肪痠加黃芪多糖組（培養液中加入黃芪多糖200 mg/L及長鏈遊離脂肪痠0．25 mmol/L）及遊離脂肪痠加黃芪多糖及腺苷痠活化蛋白激酶（AMPK）阻斷劑（compound C）組（培養液中加入黃芪多糖200 mg/L、長鏈遊離脂肪痠0．25 mmol/L及compound C 50μmol/L）。採用遊離脂肪痠檢測試劑盒和三酰甘油試劑盒分彆檢測遊離脂肪痠及三酰甘油含量，油紅O染色觀察細胞內脂質蓄積，蛋白質印跡法檢測細胞內AMPK、燐痠化腺苷痠活化蛋白激酶（p-AMPK）、脂肪痠結閤蛋白4（FABP4）的蛋白錶達情況。結果①遊離脂肪痠組三酰甘油及遊離脂肪痠含量[（3．35±0．79）mmol/L、（29．3±1．7）μmol/L]明顯高于對照組[（0．81±0．19）mmol/L、（15．7±1．3）μmol/L]和遊離脂肪痠加黃芪多糖組[（1．46±0．25）mmol/L、（15．5±2．1）μmol/L]，差異均有統計學意義（均P＜0．01）。②油紅O染色結果：遊離脂肪痠組脂質含量較對照組增多，遊離脂肪痠加黃芪多糖組脂質含量較遊離脂肪痠組減少，提示遊離脂肪痠增加細胞內脂質蓄積，而黃芪多糖減少細胞內脂質蓄積。③蛋白質印跡法檢測AMPK、p-AMPK、FABP4的蛋白錶達情況髮現，與對照組相比，黃芪多糖組各種蛋白錶達無明顯變化，遊離脂肪痠組p-AMPK錶達明顯少于對照組和遊離脂肪痠加黃芪多糖組[（54．3±3．0）％比（100．0±1．4）％、（96．5±3．9）％]，FABP4錶達明顯高于對照組和遊離脂肪痠加黃芪多糖組[（138．3±2．3）％比（100．0±3．1）％、（72．3±2．6）％]；與遊離脂肪痠加黃芪多糖組比較，遊離脂肪痠加黃芪多糖及compound C組FABP4錶達明顯增多[（136．9±4．3）％比（72．3±2．6）％]，差異均有統計學意義（均P＜0．01）。結論黃芪多糖可以通過AMPK-FABP4途徑減少高遊離脂肪痠導緻的巨噬細胞內脂質蓄積。
목적：탐토황기다당대고유리지방산도치거서세포내지질침적적작용급궤제。방법장배양적단핵원성거서세포（ THP-1）분위대조조（가입여유리지방산조등량적무수을순급우혈청청단백）、황기다당조（배양액중가입황기다당200 mg/L）、유리지방산조（배양액중가입장련유리지방산0．25 mmol/L）、유리지방산가황기다당조（배양액중가입황기다당200 mg/L급장련유리지방산0．25 mmol/L）급유리지방산가황기다당급선감산활화단백격매（AMPK）조단제（compound C）조（배양액중가입황기다당200 mg/L、장련유리지방산0．25 mmol/L급compound C 50μmol/L）。채용유리지방산검측시제합화삼선감유시제합분별검측유리지방산급삼선감유함량，유홍O염색관찰세포내지질축적，단백질인적법검측세포내AMPK、린산화선감산활화단백격매（p-AMPK）、지방산결합단백4（FABP4）적단백표체정황。결과①유리지방산조삼선감유급유리지방산함량[（3．35±0．79）mmol/L、（29．3±1．7）μmol/L]명현고우대조조[（0．81±0．19）mmol/L、（15．7±1．3）μmol/L]화유리지방산가황기다당조[（1．46±0．25）mmol/L、（15．5±2．1）μmol/L]，차이균유통계학의의（균P＜0．01）。②유홍O염색결과：유리지방산조지질함량교대조조증다，유리지방산가황기다당조지질함량교유리지방산조감소，제시유리지방산증가세포내지질축적，이황기다당감소세포내지질축적。③단백질인적법검측AMPK、p-AMPK、FABP4적단백표체정황발현，여대조조상비，황기다당조각충단백표체무명현변화，유리지방산조p-AMPK표체명현소우대조조화유리지방산가황기다당조[（54．3±3．0）％비（100．0±1．4）％、（96．5±3．9）％]，FABP4표체명현고우대조조화유리지방산가황기다당조[（138．3±2．3）％비（100．0±3．1）％、（72．3±2．6）％]；여유리지방산가황기다당조비교，유리지방산가황기다당급compound C조FABP4표체명현증다[（136．9±4．3）％비（72．3±2．6）％]，차이균유통계학의의（균P＜0．01）。결론황기다당가이통과AMPK-FABP4도경감소고유리지방산도치적거서세포내지질축적。
Objective To investigate the effects of astragalus polysaccharide ( APS) on lipid accumula-tion in macrophages induced by free fatty acids ( FFA) and mechanism.Methods The cultured derived THP-1 monocyte macrophages were divided into 5 groups: the control group ( ethanol and bovine serum albumin , equal with FFA group), APS group (APS 200 mg/L), FFA group (long chain FFA 0.25 mmol/L), FFA and APS group (APS 200 mg/L+long chain FFA 0.25 mmol/L), FFA +APS +amp activated protein kinase(AMPK) inhibitor (compound C) group (APS 200 mg/L+long chain FFA 0.25 mmol/L+compound C 50μmol/L).The FFA detection kit and triacylglycerol ( TG) kit were used to detect FFA and TG .The intracellular lipid accumula-tion was analyzed by oil red O staining , and protein expressions of AMPK , phosphorylated AMPK ( p-AMPK) and fattyacidbindingprotein4(FABP4)wereanalyzedbywesternblotting.Results ①ThecontentsofTGandFFA were significantly higher in FFA group than in the control group [(3.35 ±0.79) mmol/L vs (0.81 ±0.19) mmol/L, (29.3 ±1.7) μmol/L vs (15.7 ±1.3) μmol/L]; the contents of TG and FFA were significantly decreased in FFA and APS group than those in FFA group [(1.46 ±0.25) mmol/L vs (3.35 ±0.79) mmol/L, (15.5 ± 2.1) μmol/L vs (29.3 ±1.7) μmol/L], the differences were statistically significant (all P<0.01).②The oil red O staining results indicated that lipid content was significantly increased in FFA group than in the control group;lipid content was significantly lower in FFA and APS group than in FFA group , which suggested that APS could reduce lipid accumulation induced by FFA .③The expression of AMPK , p-AMPK and FABP4 protein were no difference between APS group and control group .Compared with the control group , the FFA group decreased p-AMPK expression, and increased FABP4 expression [(54.3 ±3.0)%vs (100.0 ±1.4)%,(138.3 ±2.3)%vs (100.0 ±3.1)%];compared with the FFA group , the FFA and APS group increased p-AMPK expression, and decreased FABP4 expression [(96.5 ±3.9)% vs (54.3 ±3.0)%,(72.3 ±2.6)% vs (138.3 ±2.3)%];compared with the FFA and APS group , the FFA +APS +compound C group increased FABP 4 expression [(136.9 ±4.3) vs (72.3 ±2.6)%], the differences were statistically significant (all P<0.01).Conclusion PS can reduce the lipid accumulation in macrophage , which induced by high level of free fatty acid , which is through the AMPK-FABP4 pathway .
