乳酸菌Lactobacillus paracasei 菌體及發酵菌液對免疫功效之評估

Immunomodulation Function of Lactobacillus paracasei and its Fermentation Broth

黃建彰(Jian-Jhang Huang)

指導教授 ： 葉怡玲

國立虎尾科技大學/文理學院/生物科技研究所/碩士(2012年)

摘要

近年來乳酸菌的研究發現其具有免疫調節及改善過敏、氣喘的效果，也具有預防腸道疾病發生的能力，但並非所有的乳酸菌皆具有免疫調節的功效，由先前本實驗室研究成果發現，Lactobacillus paracasei ( L.paracasei )菌體具有改善氣喘發炎現象，但機制尚不明確，因此本研究將以 L.paracasei 菌體及其培養發酵菌液做為研究對象，評估本菌株對氣喘小鼠動物模式是否具有預防的功效。將菌體餵食由 ovalbumin ( OVA ) 所誘發的過敏小鼠，實驗結果發現其體內過敏特異性抗體 IgE 有劑量依賴性的減少趨勢，且經流式細胞儀分析後發現，餵食L.paracasei 菌體小鼠之脾臟細胞中 CD4+CD25+ T 細胞有劑量依賴性的增加現象，但對於 OVA 特異性抗體 IgG1、IgG2a、IgE 表現及肺沖洗液發炎細胞並無顯著改善的差異。
進一步我們觀察發酵菌液中是否具有可調節免疫反應之蛋白，因此使用了蛋白膠體電泳進行分析。由結果得知其培養後的菌液中存在大小介於 170 KDa - 35 KDa 中間的蛋白，而根據蛋白大小不同，將之分為 100-35 KDa ( A分層)，55 - 35 KDa ( B分層)，35 KDa ( C分層)，將三分層菌液蛋白及未分離之菌液蛋白與人類腸道上皮細胞株 HT-29 細胞共培養後觀察出C 分層具有明顯細胞增生活性且可抑制interleukin-8 ( IL-8 ) 的分泌。而後將 C 分層與小鼠骨髓樹突狀細胞共培養，並同時給予 OVA 或lipopolysaccharides ( LPS )，結果發現細胞表面分子 CD11c、CD80、CD86 以及 MHC II 的表現有受到抑制的現象，且 IL-10 有明顯 ( P < 0.05 ) 的劑量依賴性的增加。
由實驗結果發現本實驗所使用之菌株 L.paracasei 及其發酵菌液蛋白 ( 特別是 C 分層 ) 具有免疫調節能力，可以做為未來預防發炎疾病發生的候選成分開發應用。

關鍵字

乳酸菌 ； 免疫調節 ； 過敏氣喘 ； 發酵菌液蛋白

並列摘要

In recent years, lactic acid bacteria have been reported that bearing immunomodulatory activity for many inflammatory diseases. In our study, oral feeding with Lactobacillus paracasei (L. paracasei) could reduce ovalbumin ( OVA ) specific antibody IgE secretion and CD4+CD25+ regulatory T cells population of splenocytes has dose-dependently increased in OVA induced allergy mice. We found that L. paracasei could alleviate allergy inflammation and increase the population of regulatory T cells.
Using Tricine gel electrophoresis, the soluble proteins in culture medium from L. paracasei were from 170 KDa to 35 KDa. Three fractions were separated by Amicon. The fraction A protein size was from 100-35 KDa, fraction B protein size was from 55-35 KDa, and the protein fraction C was majorly in 35 KDa. After respectively co-culturing 3 fractions with HT-29 cell line for 48 hr, the Fraction C has the ability to promote cell proliferation and to inhibit IL-8 secretion. Furthermore, the expression of cell surface molecule CD11c、CD80、CD86 and MHC II was suppressed and IL-10 secretion was induced by mouse bone marrow dendritic cells ( BMDCs ) after stimulated with Fraction C and OVA or lipopolysaccharides ( LPS ).
The soluble proteins of L. paracasei (particularly Fraction C ) have the immunomodulatory ability and might be potential candidates for the prevention and/or treatment for inflammatory diseases.

並列關鍵字

allergy asthma ； immunmodulation ； L .paracasei ； soluble proteins

參考文獻

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