Vitamin C對Quercetin代謝及抗Cisplatin引起的腎毒性之影響

第一部分 Quercetin (Q) 是一種蔬菜、水果及草藥中富含的類黃酮。許多研究顯示Q具有抗氧化、抗發炎及調節基因表現等生物活性。然而，Q的吸收率不佳且進入體內後會在小腸與肝臟中快速經phase II相關酵素進行代謝反應，而使其生物利用率受限並具有變異性。研究顯示有多種因素會影響Q的生物利用率，包括飲食的組成，不過當中vitamin C (C)、eicosapentaenoic acid (EPA) 及bromelain (BM) 的影響情形仍不清楚。因此，本研究首先利用Balb/c小鼠來探討C、EPA及BM與Q合併補充後，是否能夠提高Q的生物利用率。雄性Balb/c小鼠分成六組並介入27週：Control組、Q0.1組、Q0.2組、QC (Q0.1+C) 組、QE (Q0.1+EPA) 組及QB (Q0.1+BM) 組，其中Q是添加於飼料 (0.1%或0.2%；Q0.1及Q0.2) 給予，而C (100 mg/kg B.W.)、EPA (125 mg/kg B.W.) 與BM (10 mg/kg B.W.) 則是每週一次管餵給予。我們在介入期間分析血漿總quercetin與isorhamnetin濃度以評估各測試物對Q生物利用率之影響，結果顯示C相較於EPA及BM更能提升血漿總quercetin濃度，而C、EPA與BM均不影響isorhamnetin濃度。因此，動物犧牲後我們接續比較Q0.1、Q0.2與QC組別中小腸和肝臟代謝酵素uridine 5'-diphospho-glucuronosyltransferase (UGT) 及catechol-O-methyltransferase (COMT) 之活性，結果顯示Q增加小腸及肝臟的UGT和COMT之活性，並且具有劑量效應 (尤其小腸)，而Q合併C補充雖然不影響腸道代謝酵素活性，但是卻能提升肝臟UGT之活性。綜合上述，與單獨補充Q相比，Q合併C補充具有增加血漿總quercetin濃度以及促進肝臟代謝酵素中UGT活性之情形。第二部分 Cispatin (cis-diamminedichloroplatinum II, CDDP) 是一種臨床上廣泛使用的抗癌藥物，然而由於其細胞毒性不具專一性而導致許多種副作用，包含體重流失、淋巴球DNA損傷及腎毒性。我們先前研究發現Q能有效減輕Balb/c小鼠模式中一些CDDP所誘導的傷害，然而，C是否能相加或是相乘性的增強Q的保護效果仍不清楚。因此，本部分研究我們進行一個為期3週的動物實驗，以比較Q及C對CDDP所引發的副作用之單獨或合併保護效果。雄性Balb/c小鼠隨機分成Control組、CDDP組、CDDP+Q組、CDDP+C組及CDDP+QC (Q+C) 組，其中各成份給予方式：CDDP (7.5 mg/kg B.W.) 每週一次經腹腔注射給予；Q是添加於每天的飼料中 (0.1%) 給予；C (100 mg/kg B.W.) 則是每天一次管餵給予。Q與C在CDDP處理前一週開始給予，CDDP介入兩週後，犧牲動物進行各項分析。結果顯示，CDDP顯著減輕小鼠的體重以及脾臟和肝臟的組織相對重量指數 (Tissue somatic index, TSI)，並且增加淋巴球的DNA的損傷，而Q、C及QC則具減輕這些CDDP所造成的傷害之趨勢，不過除了體重之外，合併Q與C之效果並無優於單獨給予。在腎毒性方面，不論CDDP單獨處理或合併給予Q、C或QC對腎臟組織病變及腎功能指標 (血清BUN及creatinine含量) 之影響皆不顯著，不過相較於單獨給予CDDP的組別，Q、C及QC均有降低促發炎細胞激素 (TNF-α及IL-β) 之情況，另外還提升腎臟總抗氧化能力 (TEAC) 及總穀胱甘肽濃度。QC的效果與單獨給予Q或C類似，而C亦不會影響血漿或腎臟中總quercetin和isorhamnetin之濃度。總而言之，合併Q和C能透過其抗氧化及抗發炎的作用來減緩CDDP所引起的體重減輕、臟器毒性、DNA傷害以及腎毒性。然而，Q和C合併使用並無相加或相乘效應。

關鍵字

槲皮素；維生素C ；代謝酵素；順鉑；腎毒性

並列摘要

Part I Quercetin is a flavonoid widely present in fruits, vegetables, and herbs. Growing evidence shows that quercetin possesses several bioactivities including anti-oxidation, anti-inflammation, and regulation of gene expressions. However, quercetin has a poor and variable bioavailability because of its poor absorption and quick phase II metabolism in the small intestine and liver. Studies suggest that several factors may affect the bioavailability of quercetin, including food components. However, the precise effects of vitamin C (C), eicosapentaenoic acid (EPA), and bromelain (BM) remain unclear. Thus, in the present study, we first investigated whether the presentation of C, EPA, or BM increased the bioavailability in Balb/c mice. The male Balb/c mice were randomly assigned to the following six groups for 27 weeks: control, Q0.1, Q0.2, QC (Q0.1+C), or QE (Q0.1+EPA), QB (Q0.1+BM). Quercetin was given by a diet containing 0.1% or 0.2% quercetin (Q0.1 and Q0.2, respectively); while C (100 mg/kg B.W.), EPA (125 mg/kg B.W.), and BM (10 mg/kg B.W.) were given by gavage once per week. We examined the total plasma quercetin and isorhamnetin levels to determine the effects of C, EPA, and BM on the bioavailability of quercetin during the experiment. The results showed that C rather than EPA and BM increased the total serum quercetin level. None of C, EPA, and BM affected the level of isorhamnetin. Thus, after sacrificed we compared small intestinal and hepatic enzyme activities of uridine 5'-diphospho-glucuronosyltransferase (UGT) and catechol-O- methyltransferase (COMT) among Q0.1, Q0.2, and QC groups. The results show that Q increased UGT and COMT activities in the intestine and liver (especially in the intestine) in a dose-dependent manner. C did not affect intestinal enzyme activities but enhanced the UGT activity in the liver. In summary, Q in combination with C tends to increase the total plasma quercetin level and hepatic UGT activities compared with Q alone. Part II Cisplatin (CDDP) is a common chemotherapeutic drug for several cancers. However, the non-specific cytotoxicity of CDDP induces several side effects including body weight loss, lymphocyte DNA damage, and nephrotoxicity. Our previous study found that Q decreased some cisplatin-induced harmful effects in Balb/c mice. However, whether C additively or synergistically enhances the preventive effect of Q is unclear. Thus, in this part of the study, we performed a 3-week animal study to compare the individual and combined preventive effect of Q, C, and QC on the toxic effects of cisplatin (CDDP). The Balb/c mice were randomly assigned to the control, CDDP, CDDP+Q, CDDP+C, or CDDP+QC groups. The animals were treated Q or/and C 1 week before the CDDP treatment; Q was given by a diet containing 0.1% quercetin, while C (100 mg/ kg B.W.) was given by gavage every day. CDDP (7.5 mg/kg B.W.) was given by i.p. injection once a week for 2 weeks. The results showed that CDDP significantly decreased the body weight, and tissue somatic index (TSI) of spleen and liver but increased the lymphocyte DNA damage. Q, C, and QC tended to decrease the effects of CDDP similarly. Except for the body weight, the efficiency of QC did not better than that of Q or C. Regarding nephrotoxicity, although CDDP alone or in combination with C, Q, or QC did not affect the degree of lesions in renal tissues and serum markers of renal functions; C, Q, and QC tended decreased the levels of proinflammatory cytokines (TNF-α and IL-β) compared with CDDP alone group. C, Q, and QC also increased the renal levels of Trolox equivalent antioxidant capacity (TEAC) and total glutathione compared with CDDP alone group. The combined effects of QC were similar to that of C or Q alone. The presentation of C also did not affect total quercetin or isorhamnetin levels in serum or kidney. Taken together, the combination of Q and C attenuated CDDP-induced body weight loss, change in TSI, DNA damage, and nephrotoxicity by the antioxidant and anti-inflammatory properties. However, in general, the combined effects of Q and C were not additive or synergistic.