請用此 Handle URI 來引用此文件：
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/18236完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 李君男(Chun-Nan Lee) | |
| dc.contributor.author | Pei-Yun Hung | en |
| dc.contributor.author | 洪珮芸 | zh_TW |
| dc.date.accessioned | 2021-06-08T00:56:00Z | - |
| dc.date.copyright | 2015-03-12 | |
| dc.date.issued | 2015 | |
| dc.date.submitted | 2015-02-12 | |
| dc.identifier.citation | References
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/18236 | - |
| dc.description.abstract | 第一型單純皰疹病毒(Herpes simplex virus type 1, HSV-1)是一種普遍存在的病毒，會潛伏在人體內並可能造成嚴重疾病。現行治療單純皰疹病毒感染之藥物Acyclovir使用量不斷增加，因而產生具有抗藥性之病毒，故急需發展新的藥物以治療單純皰疹病毒之感染。魚腥草為一天然中草藥，已有文獻指出魚腥草可經由作用於第二波NF-κB來活化抑制第一型單純皰疹病毒。然而對於更詳細的抑制機轉則還不清楚。本研究結果顯示魚腥草水萃取物抑制HSV-1 、HSV-2以及具有Acyclovir抗性的HSV-1(HSV-AR)其作用主要是透過抑制病毒和細胞結合以及抑制病毒進入宿主細胞。除此之外，魚腥草水萃取物也可以經由抑制第一波NF-κB活化來減少病毒複製。進一步分析魚腥草水萃取物之六個主要成分，顯示quercetin可以同時抑制病毒進入細胞以及NF-κB活化，而isoquercitrin可以抑制NF-κB活化。上述結果顯示魚腥草水溶液萃取物可以經由不同機制來抑制單純皰疹病毒感染，並且可能對於未來抗單純皰疹病毒藥物的發展有所幫助。
另外，微核醣核酸(microRNAs)最近也被用來研究作為抗病毒的標的。微核醣核酸是近來發現的一類小片段、非蛋白質編碼的核醣核酸，可藉由內源性RNA干擾方式來調控廣泛的生物功能，其中包括宿主與病毒的相互作用。目前對於微核醣核酸在單純皰疹病毒感染過程中的角色仍不全然清楚。研究細胞生成的微核醣核酸如何調控病毒感染有助於了解病毒和宿主細胞之間的交互作用。本研究使用單核球細胞株THP1，並利用即時反轉錄聚合酶連鎖反應分析平台，比較第一型單純皰疹病毒感染THP1後微核醣核酸表現量的差異。MicroRNA-27a*為 第一型單純皰疹病毒感染細胞後下降最顯著的微核醣核酸。用相關電腦軟體分析，指出第二型成對類免疫球蛋白受體α (paired immunoglobulin-like type 2 receptor α, PILRα)為其可能作用之標的基因。PILRα是第一型單純皰疹病毒感染時的受體，除此之外，在它的細胞質端區域具有免疫受體酪氨酸抑制模體(ITIM)可以傳遞抑制的訊號。在本研究中，我們假設第一型單純皰疹病毒感染後會抑制 miR-27a*，因而增加PILRα的表現並進而幫助抑制宿主細胞抗病毒的先天免疫反應。進一步的實驗顯示病毒在高度表現miR-27a*的細胞中複製會被抑制高達20倍。除此之外，細胞激素TNF-α，IFN-β以及 IL-6的表現也有增加的現象。如果在細胞中送入對PILRα有特異性的siRNA也可以觀察到相似的結果。綜合以上實驗顯示miR-27a*對於經由抑制 PILRα表現來調控細胞抗病毒的先天免疫扮演著重要角色。 | zh_TW |
| dc.description.abstract | Herpes simplex virus (HSV), a common latent virus in humans, causes certain severe diseases. Extensive use of acyclovir (ACV) results in the development of drug-resistant HSV strains, hence, there is an urgent need to develop new drugs to treat HSV infection. Houttuynia cordata (H. cordata), a natural herbal medicine, has been reported to exhibit anti-HSV effect which is partly NF-κB-dependent. However, the molecular mechanisms by which H. cordata inhibits HSV infection are not elucidated thoroughly. In this study, we found that H. cordata water extract (HCWE) inhibited the infection of HSV-1, HSV-2, and acyclovir-resistant HSV-1, mainly via blocking viral binding and penetration in the beginning of infection. HCWE also suppressed HSV replication. Furthermore, HCWE attenuated the first-wave of NF-κB activation, which is essential for viral gene expression. Further analysis of six compounds identified in H. cordata revealed that quercetin and isoquercitrin inhibit NF-κB activation and additionally, quercetin also has an inhibitory effect on viral entry. These results indicate that H. cordata could inhibit HSV infection through multiple mechanisms and could be a potential lead for development of new drugs for treating HSV.
Recently, the microRNA also can be the antiviral target. MicroRNA is a class of small non-protein-coding RNAs that may act via endogenous RNA interference. However, few studies have explored the role of microRNA in HSV infection. In this study, we used the monocytic cell line THP1 and found that the expression of miR-27a* in THP1 was reduced after HSV-1 infection. Potential target genes of miR-27a* were predicted by the target prediction program, miRanda. One of the predicted targets is paired immunoglobulin-like type 2 receptor α (PILRα), the co-receptor of HSV-1, which contains an immunoreceptor tyrosine-based inhibition motif (ITIM) and can deliver inhibitory signals in immune cells. We hypothesized that HSV-1 infection could down-regulate the expression of miR-27a* and increase the expression of PILRα and then inhibit the antiviral immunity. Further studies showed that overexpression of miR-27a* in cells could significantly inhibit virus replication and also could increase the expression of TNF-α, IFN-β, and IL-6. Similar results were also observed by delivering PILRα-specific siRNA. Collectively, miR-27a* had important role in regulating innate antiviral immunity by inhibiting the expression of PILRα. Further studies will be required to elucidate the role of PILRα in HSV-1 infection. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-08T00:56:00Z (GMT). No. of bitstreams: 1 ntu-104-F95424014-1.pdf: 8221607 bytes, checksum: c1640e59b547b28b95348204afb926a7 (MD5) Previous issue date: 2015 | en |
| dc.description.tableofcontents | 1.5 Biosynthesis and functions of microRNA …………..………………….…………29
1.6 Host microRNA and HSV………………………………………………………….32 1.7 Virus-encoded microRNAs……………………………………………………….33 1.8 Specific aims…………………………………….………………...……………….35 Chapter 2 Materials and Methods……………..………………...…….…….…….37 2.1 Virus and cell culture……………..…………………………………..…………….37 2.2 HCWE preparation………………………….…………………….……………….37 2.3 Plaque reduction assay……………….……………………………….……………38 2.4 Cytotoxicity assay……………….…………………………………………………38 2.5 Cell pre-treatment and virus pre-treatment assays…………………………………38 2.6 Binding and penetration assays……………….……………………………………39 2.7 Time-of-addition assay……………….……………………………………….……40 2.8 Western blot assay……………….………………………………………………41 2.9 Immunofluorescence assay…………………………………………..…………..41 2.10 Plasmid construction for ICP0 promoter……………….…………………………42 2.11 Production of recombinant glycoprotein D……………….………………………43 2.12 Promoter luciferase assay……………….……………………………...…………43 2.13 HPLC instrumentation and conditions……………….…………………...………44 2.14 Compounds preparation……………….…………………………………..………44 2.15 RNA extraction and microRNA profiling………………..………...…………44 2.16 Quantification of miR-27a*……………….……...……………………….………45 2.17 Plasmid constructions of PILRα 3’UTR……………………………....…….……45 2.18 Luciferase assay……………….…………………………………………..………46 2.19 siPILRα transfections……………….…………………………………….………46 2.20 Statistical Analysis……………….……………………………………….………47 Chapter 3 Results…….……….……………………………….…..………….………48 3.1 HCWE inhibited HSV replications……………..…………………….…….....……48 3.2 HCWE inhibited HSV infections by targeting virus particles directly….…………49 3.3 HCWE inhibited viral entry……………………………………………….…..……50 3.4 HCWE blocked recombinant gD binding to host cells…………………………..52 3.5 HCWE suppressed HSV replication after viral entry………………...…………….53 3.6 HCWE suppressed viral gene expression via inhibiting HSV-inducedNF-κB activation……………………………..…..…..……………………………….…..53 3.7 HCWE universally inhibited HSV replication in different cell lines…………….56 3.8 HPLC analysis of the HCWE collected from Taiwan…………………….………..57 3.9 Major compounds of HCWE possess anti-HSV activity…………………………..57 3.10 The expression of microRNA-27a* is decreased by HSV-1 infection……..……..59 3.11 PILRα is a target of miR-27a*………………………………….……..…………..59 3.12 HSV can infect and replicate in immune cell………………………....…..…61 3.13 MicroRNA-27a* involved in host immune responses after HSV-1 infection….....62 3.14 Overexpression of miR-27a* reduces virus production…………………………..63 3.15 UV-inactivated HSV-1 could inhibit the expression of miR-27a*…………...…..64 Chapter 4 Discussion………………………..…………….…………….…....……..66 Chapter 5 Conclusions………………………………………………………………..76 References………………………………..……………………..……………..………77 Tables………..……………………...………………………….…………………....101 Figures…..…………………………..…………….…………….……………...….…104 | |
| dc.language.iso | en | |
| dc.title | 單純皰疹病毒感染中魚腥草抗病毒作用之機制及miR-27a*所扮演之角色 | zh_TW |
| dc.title | Mechanism of Anti-virus Activity of Houttuynia Cordata and The Role of miR-27a* in Herpes Simplex Virus Infection | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 103-1 | |
| dc.description.degree | 博士 | |
| dc.contributor.oralexamcommittee | 高全良(Chuan-Liang Kao),張淑媛(Sui-Yuan Chang),劉旻禕(Min-yi Liu),陳基旺(Ji-Wang Chern) | |
| dc.subject.keyword | 單純皰疹病毒,魚腥草,微核醣核酸, | zh_TW |
| dc.subject.keyword | Herpes simplex virus,Houttuynia cordata,microRNA, | en |
| dc.relation.page | 146 | |
| dc.rights.note | 未授權 | |
| dc.date.accepted | 2015-02-12 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 醫學檢驗暨生物技術學研究所 | zh_TW |
| 顯示於系所單位： | 醫學檢驗暨生物技術學系 | |
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