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來源資料
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題名 | 抑制人類多瘤性病毒,BK 病毒,感染之中藥篩檢=Screening of Chinese Herbal Medicine to Inhibit Human Polyomaviurs, BK Virus, Infection |
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作者姓名(中文) | 張德卿; | 書刊名 | 中醫藥年報 |
卷期 | 29:8 2011.09[民100.09] |
頁次 | 頁515-540 |
專輯 | 中醫藥基因體相關研究 |
分類號 | 414.52 |
關鍵詞 | BK病毒; 多瘤性病毒腎病變; 中藥抗病毒; BK virus; Polyomavirus-associated nephrolpathy; Herbal anti-virus; |
語文 | 中文(Chinese) |
中文摘要 | 人類多瘤性病毒,BK 病毒(BKV),屬於DNA 病毒。BKV 於腎臟組織再活化,常導致腎臟移植病人腎衰竭,即為多瘤性病毒相關腎病變(Polyomavirusassociated nephropathy, PVN)。截至目前為止,仍缺乏有效抗BKV 的藥物,本研究計畫的目的即在於尋找有效抑制BKV 活性的中藥,以提供治療PVN的新藥物。實驗先以抑制BKV VP1 類病毒殼體血液凝集,篩選可能具有抑制BK 病毒與細胞受體結合能力之中藥,再以免疫螢光染色確認中藥處理後,BKV 感染細胞後病毒蛋白質表現。同時以溶劑梯度沖提分析中藥之圖譜,確認分離中藥的較佳條件;接著收集不同部分之中藥分離物,測定其血液凝集抑制活性,以作為後續分離鑑定中藥之依據。結果顯示紅景天、枸杞、白芍、黃芩及山楂,可抑制BK VP1類病毒殼體血液凝集活性。經由免疫螢光染色,確認紅景天及山楂,具有明顯抑制病毒蛋白質表現之活性。為了探討鑑定紅景天及山楂之有效成分,以不同極性溶劑沖提分離其成分物質,HAI 結果顯示無論以不同濃度甲醇、或乙酸乙酯及正丁醇萃取,紅景天及山楂之有效成分均位於水萃取層;再以5-50%乙晴分離此水萃取層中紅景天及山楂之有效成分,結果顯示此部分純化之20%乙晴萃取物具有最高之HAI 活性,且濃度分別增加了128 及2048 倍HAI。未來我們將繼續以不同溶劑及管柱分離純化紅景天及山楂之20%乙晴萃取物中的有效成分,並搭配NMR,2D-NMR,IR,UV,ESI-Mass,Element Analysis 等方法決定其構造式。接著以LiChrospher 100 RP-18e 層析管柱,移動相TFA-CH3CN 的條件,以不同比率之甲醇濃度沖提分離紅景天及山楂萃取物,繼續以抑制血液凝集試驗分析。結果顯示80:20 之甲醇濃度時有較佳的血液凝集抑制活性。因此紅景天及山楂抑制BKV 與細胞受體結合的成分可能位於此分離物中。我們將繼續分離鑑定此物質,以其找到有效抑制BKV 感染的中藥,作為臨床治療PVN 用藥之新選擇。 |
英文摘要 | Human polyomvirus, BK virus (BKV), is a small DNA virus. Reactivation of BKV with high level viral replication results in lytic disruption of kidney cells. BKV reactivation may cause renal dysfunction and graft loss in renal transplant patients. This is polyomavirus-associated nephropathy (PVN). There are no effective drugs available for anti-BKV infection. The purpose of this study is to identify effective compounds for anti-BKV infection from Chinese herbal extracts. Herbal extacts were then analyzed for the inhibition of BK VLP hemagglutination activity (HAI). Immunofluoescence assay was performed to analyze viral protein expression in BKV infected cells after herbal treatment. Candidate herbal extracts were separated by chromatography using gradient solvents. The elunts from herbal extacts were then screened for HAI activity. Five examined herbal extracts, Rhodiola rosea L, Paeonia albiflora Palla. Lycium chinense Mill, Scutellaria. Baicalensis Georg, and Crataegus pinnatifida Bunge, were able to show HAI activity. In vivo immunofluorescence assay (IFA) demonstrated that only Rhodiola rosea L. and Crataegus pinnatifida Bunge exhibited a viral infection inhibition effect. Fifty-100% methanols, or butanol and ethylacetate solvent were employed to extract the effective compounds from Rhodiola rosea L. and Crataegus pinnatifida Bunge. The buffer layer was demonstrated to exhibit HAI acitivity. Five-50% acetonitril were then followed to extract the effective compunds from the buffer layer. HAI activity was concentrated to reach 128 and 2048 fold in the partial purified 20% acetonitrile extracts. Further solvent extraction and chromatography separation combined with NMR, IR, ESI-Mass will be employed to identify the structure of the effective compounds from Rhodiola rosea L. and Crataegus pinnatifida Bunge. |
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