Immunomodulatory effects of lotus plumule and its extracts on primary immune cells and streptozotocin-induced diabetic mice

Abstract

The aim of this study was to investigate the immunomodulatory effects of lotus (Nelumbo nucifera Gaertn) plumule and its extracts using 40C water, 100C hot water, 50% ethanol and n-hexane on primary immune cells and streptozotocin (STZ) -induced diabetic mice. The chemical compositions including total phenol, sugar, protein and flavonoids of different extracts from lotus plumule were determined. The results showed that differential constitutes existed in the four lotus plumule extracts. Ethanol extracts of lotus plumule consisted of abundant phenolic and sugar contents, however higher protein and flavonoid contents were existed in 40C water extracts of lotus plumule. In an in vitro study using primary immune cells, the immunomodulatory effects of different extracts from lotus plumule were differential. The 40C water extracts demonstrated a pro-inflammatory potential, however the 100C hot water and 50% ethanol extracts exhibited an anti-inflammatory potential in vitro. The immunomodulatory potential of n-hexane extracts was still uncertain. In 28 days consecutive tube feeding study using STZ-induced female BALB/c diabetic mice, lotus plumule and its ethanol extracts were conducted to test the immunomodulatory potential under a diabetic disease stress. The results showed that supplementation with lotus plumule and its ethanol extracts did not significantly affect the mouse body weight compared to the control group. Supplementation with low (1 mg/mouse/day) and high dose (20 mg/mouse/day) crude sample significantly decreased thymus weight, serum glucose and IgG levels. Supplementation with medium dose (5 mg/mouse/day) crude sample significantly decreased serum IgG, IgA and IgM levels. Supplementation with low dose (0.2 mg/mouse/day) ethanol extracts from lotus plumule significantly decreased adipose tissue weight, serum HDL-cholesterol, IgG and IgA levels. Supplementation with medium dose (1 mg/mouse/day) ethanol extracts from lotus plumule significantly decreased thymus weight, triglycerides, HDL-cholesterol, IgG and IgA levels. Supplementation with high dose (4 mg/mouse/day) ethanol extracts significantly decreased serum glucose, HDL-cholesterol and IgG levels. According to the secretion and ratio of Th1 and Th2 cytokines by splenocyte and peritoneal cell cultures from STZ-induced diabetic mice, crude lotus plumule supplementation demonstrated anti-inflammatory effects on peritoneal cells and exhibited Th2-skewed immunodulatory effects on splenocytes. Supplementation with ethanol extracts of lotus plumule also demonstrated anti-inflammatory effects on peritoneal cells and exhibited Th2-skewed immunomodulatory effects on splenocytes, however supplementation with ethanol extracts of lotus plumule seemed to have pro-inflammatory effects on splenocytes. In conclusion, the ethanol extracts of lotus plumule demonstrated an anti-inflammatory effect on peritoneal cells in both in vitro and in vivo studies. However, ethanol extracts of lotus plumule exhibited differential effects on splenocytes in vivo.

本論文之目的在於探討40ºC水、100ºC熱水、50%酒精及正己烷等四種溶劑之蓮子心(蓮Nelumbo nucifera Gaertn)萃取物對免疫細胞及streptozotocin (STZ)誘發之糖尿病BALB/c小鼠免疫調節功能的影響，利用體外(in vitro)及體內(in vivo)等實驗模式，進行抗發炎功效的評估，並對蓮子心的四種溶劑萃取物進行成分分析。 成分分析的結果顯示，蓮子心的四種不同溶劑萃取物成分組成略有差異，總酚及總醣含量是在50%酒精萃取物中較高，而總蛋白及總類黃酮含量則在40ºC水萃取物中較高。 體外免疫細胞實驗的結果顯示，蓮子心四種溶劑萃取物的免疫調節功效略有不同，40ºC水萃取物傾向促發炎趨勢的免疫調節，100ºC熱水及50％酒精萃取物則傾向抗發炎趨勢的免疫調節，正己烷萃取物的免疫調節趨勢則不確定。 以蓮子心及其酒精萃取物連續28天餵食STZ誘發之BALB/c糖尿病小鼠的結果顯示，實驗組及控制組的體重皆無顯著改變，大多呈緩慢增加的趨勢。一般生化指標顯示，餵食低(1mg/mouse/day)及高劑量(20 mg/mouse/day)蓮子心使小鼠胸腺重量下降，並降低血清中葡萄糖及IgG的濃度；餵食中劑量(5 mg/mouse/day)蓮子心，降低IgG、IgA及IgM的濃度。而餵食低劑量(0.2 mg/mouse/day)蓮子心50%酒精萃取物，則使副睪脂肪組織重量下降，並降低HDL、IgG與IgA的濃度；餵食中劑量(1 mg/mouse/day)蓮子心50%酒精萃取物，使胸腺重量下降，並降低TG、HDL、IgG及IgA的濃度；餵食高劑量(4 mg/mouse/day)蓮子心50%酒精萃取物，降低葡萄糖、HDL及IgG的濃度。免疫指標顯示，餵食蓮子心對腹腔細胞具有傾向抗發炎調節趨勢的功效，對脾臟細胞為傾向Th2免疫反應的調節趨勢；餵食蓮子心50%酒精萃取物對腹腔細胞具有傾向抗發炎調節趨勢的功效，但對脾臟細胞則傾向促發炎調節趨勢及傾向Th2免疫反應的調節趨勢。 綜合體外及體內實驗的結果顯示，蓮子心酒精萃取物對腹腔細胞具有抗發炎功效，但對脾臟細胞則呈現不同的功效。