評估薑黃水萃取物對海鱺細胞激素與補體之免疫調節作用

薑黃具有抗發炎、抗凝血、抗菌、抗真菌、抗原生動物、抗病毒等的活性，本研究評估薑黃水萃取物對海鱺先天免疫調節作用及對發光桿菌病的抵抗力。第一個實驗中使用水及高壓滅菌方式萃取薑黃粉，離心後收集上清液，將四種不同濃度的上清液噴灑至市售飼料中並餵食海鱺，以0 g.kg-1 (T0)、 1 g.kg-1 (T1)、 5 g.kg-1 (T5) 與 g.kg-1 (T20)，以表示每公斤飼料所含萃取前之薑黃粉克數。於餵食後第1、3、7與14天，測定其血清蛋白濃度(總蛋白、免疫球蛋白)及酵素活性(溶菌酶、過氧化物酶)，同時定量海鱺腸道、前腎與脾中的促炎細胞因子(IFNγ, TNFα, IL-1β, IL6)、抗炎細胞因子(IL10)及補體成分(C1r, C3, C5, MASP1) mRNA。並於餵食14天後，注射發光桿菌至海鱺腹腔，觀察七天的相對存活率。結果表示餵食14天後， T20的總免疫球蛋白、過氧化物酶明顯增加，且腸道中的促炎細胞因子(IFNγ, TNFα, IL-1β, IL6)、抗炎細胞因子(IL10)及補體成分(C1r, C3, C5, MASP1)亦增加。此外，T5前腎中TNFα, IFNγ, IL6, IL10, C1r, C5, 與 MASP1明顯增加，同樣地，餵食後第1天、第7天、第14天後，腸道、前腎中C1r、MASP1表現量亦增加，T1與T20相對存活率(P < 0.05)明顯高於對照組。第二個實驗中餵食海鱺20 g.kg-1薑黃水萃物，並分別於餵食後第0天(0D)、第1天(1D)、第7天(7D)、第14天(14D) 注射發光桿菌至海鱺腹腔。在0D注射PBS (0D+PBS)或 0D+Pdp (0D+Pdp)至海鱺腹腔中，0D+PBS用於檢視PBS的效果，並比較0D+Pdp的補體RNA基因表現。以 0D+Pdp比較1D, 7D與14D的補體mRNA基因表現。於Pdp攻菌前後各一天收集腸道，並分析補體(C3, C5, C1r, MASP1)的mRNA基因表現，同時記錄14天的相對存活率。結果顯示，除了7D攻菌前的補體C1r、C3，其它的補體基因表現皆增加，攻菌後腸道內的C3與C5下降。然而C5在7D和14D明顯增加。在14D相對存活率(37.0%)明顯高於對照組(13 %)。結論為餵食14天後，薑黃水萃取物可提高腸道及前腎的促炎細胞激素(IFNγ, TNFα, IL-1β, IL6)、抗炎細胞激素(IL10)及補體成分(C1r, C3, C5, MASP1)等mRNA的表現，口服薑黃水萃取物可以避免發光桿菌在腸道對C5 mRNA的抑制作用。這也許是發光桿菌攻菌後，存活率增加的其中一項機制，然而薑黃水萃取物能調節細胞激素及補體的mRNA表現，也有可能會增強海鱺對發光桿菌的抵抗力。

關鍵字

薑黃；海鱺；補體系統；細胞激素；發光桿菌病

並列摘要

Turmeric has anti-inflammatory, antioxidant, anticoagulant, antibacterial, antifungal, antiprotozoal, antiviral activities. The present study evaluated the effect of turmeric water extract (TWE) on the innate immune responses and the resistance against Photobacterium damselae subsp. piscicida (Pdp) of cobia. In the first experiment, turmeric powder was extracted in water using autoclave. The supernatant was sprayed to the commercial feed at four concentrations which are indicated as gram of turmeric powder befo The resulted mixture was centrifuged to collect the supernatant. re extracting per kg of feed, e.g., 0 g.kg-1 (T0), 1 g.kg-1 (T1), 5 g.kg-1 (T5) and 20 g.kg-1 (T20). At one, three, seven and fourteen days after feeding, serum protein concentrations (total protein, immunoglobulin) and enzyme activities (lysozyme, peroxidase) were measured. Meanwhile the proinflammatory cytokines (IFNγ, TNFα, IL-1β, IL6), anti-inflammatory cytokine (IL10) and complement components (C1r, C3, C5, MASP1) mRNA were quantified from intestine, head kidney, and spleen. After fourteen days of feeding, cobia was intraperitoneally injected with Pdp, then the relative percentage survival (RPS) was observed for seven days. The results showed that total immunoglobulin and peroxidase were significantly increased after 14 days of feeding in treatment T20. After 14 days of feeding, proinflammatory cytokines (IFNγ, TNFα, IL-1β, IL6), anti-inflammatory cytokines (IL10), complement components (C1r, C3, C5) were up-regulated in the intestine of treatment T20. Besides, TNFα, IFNγ, IL6, IL10, C1r, C5, and MASP1 were significantly upregulated in the head kidney in treatment T5. Also, C1r and MASP1 were all upregulated after one, seven, fourteen days in intestine and head kidney, respectively. The RPS in T1 and T20 was significantly (P < 0.05) higher than the control. In the second experiment, cobia was intraperitoneally injected Pdp after feeding with turmeric at 20 g.kg-1 for 0 day (0D), one day (1D), seven days (7D) and fourteen days (14D). Treatment 0D was injected with PBS (0D+PBS) or with Pdp (0D+Pdp). Treatment 0D+PBS was used to examine the effect of PBS and compare the complements mRNA expression to the treatment 0D+Pdp. The treatment 0D+Pdp was used to compare the complement mRNA expression to the other treatments at1D, 7D, and 14D. The intestine was collected before and one day after Pdp challenge. The mRNA gene expression of the complements (C3, C5, C1r, MASP1) were analyzed. The RPS was recorded for fourteen days. The results showed that complement components were upregulated in the treated groups exception for C1r and C3 in 7D treatment before challenge. After challenge, C3 and C5 were downregulated in the intestine. However, C5 in the treatment 7D and 14D was significantly upregulated. RPS in 14D (37.0%) treatment was significantly higher than the control (13 %). In summary, TWE can upregulate the mRNA gene expression of proinflammatory cytokines (IFNγ, TNFα, IL6), anti-inflammatory cytokine (IL10), and the complement components (C1r, C5) in cobia intestine and head kidney after fourteen days. The oral feeding with TWE can counteract the inhibition of C5 mRNA expression after Pdp challenge in intestine. This may be one of the mechanisms to enhance the survival rate after Pdp challenge. Therefore, Curcuma longa water extract modulates the mRNA expression of cytokines and complements, as well as enhances the resistance against Photobacterium damselae subsp. piscicida on cobia Rachycentron canadum.

並列關鍵字

turmeric ； cobia ； complement ； innate cytokines ； photobacteriosis

參考文獻

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