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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 陳玉怜(Yuh-Lien Chen) | |
| dc.contributor.author | Wen-Ching Shen | en |
| dc.contributor.author | 沈紋君 | zh_TW |
| dc.date.accessioned | 2021-06-16T13:41:26Z | - |
| dc.date.available | 2013-09-24 | |
| dc.date.copyright | 2013-09-24 | |
| dc.date.issued | 2013 | |
| dc.date.submitted | 2013-07-12 | |
| dc.identifier.citation | Anzalone R, Lo Iacono M, Corrao S, Magno F, Loria T, Cappello F, Zummo G, Farina F, La Rocca G. New emerging potentials for human Wharton's jelly mesenchymal stem cells: immunological features and hepatocyte-like differentiative capacity. Stem Cells and Development. 2010; 19: 423-438.
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/62328 | - |
| dc.description.abstract | 由於逐漸攀高的糖尿病盛行率及其防治與治療已被視為非常重要的課題。嚴重糖尿病可引起諸多併發症。其中，病患的足部血管病變造成微循環不良，更是容易導致糖尿病患者下肢傷口不易癒合及其相關感染的產生，嚴重時常會導致下肢截肢。近幾年來研究發現骨髓衍生之內皮前驅細胞(EPC)可以降低動物組織缺血性傷害，促使血管新生並提高存活率。隨著其他器官幹細胞的發現以及對幹細胞特性，包括如何幫助受損器官修補之潛能的瞭解，運用幹細胞於治療缺血性傷害是一新的策略。有別於單一藥物的治療，幹細胞可於受傷組織分泌細胞激素、改善血管功能或直接分化為需要的細胞來幫助血管再生。本篇研究之主要目的想要藉由臍帶間質幹細胞所分化之內皮前驅細胞(WJ-EPC)，植入至糖尿病鼠之缺血下肢，探討EPC影響糖尿病小鼠之缺血性傷害及其相關機轉。首先，藉由股動脈截斷術造成下肢缺血，然後將EPC植入至糖尿病鼠下肢缺血的肌肉內，藉由下肢血流、外觀等觀察來驗證植入EPC的治療效果。實驗結果發現，在缺血性下肢糖尿病鼠的血流會降低、血管密度減少以及增加骨骼肌細胞凋亡。而這些組織傷害可藉由植入EPC而被明顯減緩。除此之外，以免疫組 織化學染色法鑑別，發現植入骨骼肌之EPC在缺氧的微環境下有表現缺氧誘導因子(Hypoxia-inducible factor-1a，HIF-1α)及介白素8 ( interleukin-8，IL-8)。在體外實驗方面，我們將EPC培養在缺氧環境下，發現EPC會大量表現HIF-1α並且可在細胞培養液中測得大量的IL-8。此外，我們以在缺氧所獲得EPC培養液(CM-H)與骨骼肌細胞株(NOR)共同培養在缺氧環境中，結果發現CM-H可以降低骨骼肌細胞之促細胞凋亡蛋白質的表現量、增加骨骼肌細胞抗細胞凋亡蛋白質的表現量以及促進EPC的移動性及脈管形成。上述這些現象在轉染HIF-1α siRNA 或IL-8 siRNA至EPC後，其缺氧所獲得EPC培養液不再具有保護骨骼肌細胞免於細胞凋亡的功效，也失去促進EPC的移動性及脈管形成的能力。綜合本研究結果顯示，WJ可當為EPC的主要來源，同時在糖尿病缺血性組織疾病的臨床治療，WJ-EPC的治療可視為一新穎性策略療法。在本研究中，植入EPC至糖尿病鼠的缺血性下肢可經由HIF-1α/IL-8訊號路徑減緩傷害，所以EPC及其訊號機轉的保護作用扮演一非常重要角色。 | zh_TW |
| dc.description.abstract | Peripheral arterial diseases, the major complication of diabetes, can result in lower limb amputation. As endothelial progenitor cells (EPCs) are involved in neovascularization, the study was to examine whether EPCs isolated from Wharton’s jelly (WJ-EPCs) of the umbilical cord, a rich source of mesenchymal stem cells, could reduce ischemia-induced hind limb injury in diabetic mice. We evaluated the effects of WJ-EPC transplantation on hind limb injury caused by femoral artery ligation in mice with streptozotocin (STZ)-induced diabetes. We found that the ischemic hind limb in mice with streptozotocin-induced diabetes showed decreased blood flow and capillary density and increased cell apoptosis and that these effects were significantly inhibited by injection of WJ-EPCs. In addition, HIF-1α and IL-8 were highly expressed in transplanted WJ-EPCs in the ischemic skeletal tissues and were present at high levels in hypoxia-treated cultured WJ-EPCs. Moreover, incubation of the NOR skeletal muscle cell line under hypoxic conditions in conditioned medium from EPCs cultured for 16 h under hypoxic conditions resulted in decreased expression of pro-apoptotic proteins and increased expression of anti-apoptotic proteins. The inhibition of HIF-1α or IL-8 expression by EPCs using HIF-1α siRNA or IL-8 siRNA, respectively, prevented this change in expression of apoptotic-related proteins. Wharton’s jelly in the umbilical cord is a valuable source of EPCs and transplantation of these EPCs represents an innovative therapeutic strategy for treating diabetic ischemic tissues. The HIF-1α/IL-8 signalling pathway plays a critical role in the protective effects of EPCs in the ischemic hind limb of diabetic mice. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-16T13:41:26Z (GMT). No. of bitstreams: 1 ntu-102-D98446006-1.pdf: 2706929 bytes, checksum: bfbd0e2708242c37697a90fe55fe64d7 (MD5) Previous issue date: 2013 | en |
| dc.description.tableofcontents | 中文摘要------------------------------------------------- I
英文摘要------------------------------------------------- III 英文縮寫與全文對照----------------------------------------- V 第一部份 文獻回顧------------------------------------------------- 1 一 引言------------------------------------------------- 1 二 糖尿病及下肢缺血之成因----------------------------------- 2 三 前驅細胞之來源種類及功能應用------------------------------ 3 四 Hypoxia-inducible factor-1α ( HIF-1α)與缺血性下肢之關聯- 6 五 Interleukin-8 (IL-8)與下肢缺血及血管新生之關聯------------ 8 研究動機------------------------------------------------- 10 儀器設備------------------------------------------------- 11 實驗材料與試劑-------------------------------------------- 11 實驗用溶液配方-------------------------------------------- 15 材料與方法----------------------------------------------- 17 一 內皮前驅細胞之純化與培養--------------------------------- 17 二 內皮前驅細胞之鑑定-------------------------------------- 18 DiI-acetylated LDL uptake------------------------------ 18 免疫細胞螢光染色------------------------------------------ 18 流式細胞儀分析-------------------------------------------- 19 三 糖尿病鼠模式之建立-------------------------------------- 19 四 雷射都卜勒影像分析內皮前驅細胞植入缺血下肢之血流分布---------- 20 五 下肢功能性之評分--------------------------------------- 21 六 組織切片之收集，形態分析和免疫組織化學染色------------------ 22 七 電子顯微鏡術------------------------------------------ 23 八 TUNEL染色------------------------------------------- 23 九 分析細胞與骨骼肌組織之蛋白質表現-------------------------- 24 十 條件式培養液(condition medium, CM)的製備及其對NOR細胞的影響 --------------------------------------------------------25 十一 多重免疫分析與酵素免疫分析法檢測細胞培養液中IL-8之含量------ 26 十二 以電穿孔法轉染siRNA---------------------------------- 26 十三 細胞移行分析----------------------------------------- 27 十四 脈管形成分析----------------------------------------- 27 十五 數據統計分析----------------------------------------- 28 結果--------------------------------------------------- 29 一 臍帶間質細胞具有分化成內皮前驅細胞(WJ-EPC)之能力----------- 29 二 植入內皮前驅幹細胞(EPC)可有效恢復糖尿病鼠下肢股動脈截斷後之血流及功能 性恢復--------------------------------------------------- 29 三 植入EPC可有效改善糖尿病鼠下肢缺血後之肌肉組織形態------------ 30 四 植入EPC在糖尿病鼠下肢缺血後之血管新生情形------------------ 31 五 植入EPC減少下肢缺血性組織細胞之凋亡及來自缺氧性EPC所獲得培養液(CM-H)增加缺氧性骨骼肌細胞株(NOR cells)抗凋亡蛋白質表現情形-------- 32 六 植入EPC經由HIF-1α/IL-8訊號路徑減少糖尿病鼠缺血性下肢之組織傷害 ------------------------------------------------33 七 HIF-1α及IL-8 可以提高EPC移動性和脈管形成能力-------------- 35 討論---------------------------------------------------- 36 結論---------------------------------------------------- 43 參考文獻------------------------------------------------- 44 附圖---------------------------------------------------- 52 圖一 臍帶內皮前驅細胞 (WJ-EPC)之特徵------------------------ 52 圖二 STZ誘導ICR小鼠高血糖---------------------------------- 53 圖三 植入的EPC能有效的改善糖尿病鼠因下肢股動脈截斷手術後所引起的下肢缺血血流---------------------------------------------------- 54 圖四 動脈截除手術後立即植入EPC，在第7天後，植入的細胞能有效的改善肌肉組織形態-------------------------------------------------- 56 圖五 植入的EPC，在第7天後，糖尿病鼠缺血性下肢肌肉組織之微血管密度表現情形------------------------------------------------------ 57 圖六 植入的EPC，在第7天後，糖尿病鼠缺血性下肢肌肉組織之細胞凋亡情形，以及在缺氧環境中NOR cells與CM-H在缺氧環境共同培養下其凋亡或抗凋亡蛋白質之表現情形----------------------------------------------- 58 圖七 HIF-1α及IL-8在缺血性下肢肌肉組織之表現以及來自EPC之HIF-1α及IL-8對NOR cells在缺氧環境中影響相關凋亡蛋白質之表現---------------- 61 圖八 CM-H對EPC移動能力和脈管形成能力之影響-------------------- 64 圖九 HIF-1α及IL-8訊號路徑在活體內及活體外參與調控缺血性下肢與缺氧環境中NOR cells之血管新生及凋亡作用------------------------------ 66 第二部份 中文摘要------------------------------------------------- 67 英文摘要------------------------------------------------- 69 文獻回顧------------------------------------------------- 71 引言---------------------------------------------------- 71 一 急性腎損傷與IS及致炎性細胞激素之關聯----------------------- 73 二 IS (indoxyl sulfate, IS)與黏附蛋白之關係 ----------------74 進行中之初步結果------------------------------------------ 77 一 探討IS是否會增強IL-1β所誘導之HUVEC表現E-selectin黏附子----- 77 1. IS處理對 HUVEC不具毒性--------------------------------- 77 2. IL1-β刺激HUVEC表現E selectin 之情形-------------------- 77 3. IS對IL1β處理HUVEC表現E- selectin之影響----------------- 78 4. OAT抑制劑對IS刺激IL1-β處理HUVEC表現E-selectin之影響------ 78 二 探討IS對IL1β處理HUVEC是否經由MAPKs訊號參與調控E-selectin之表現 ----------------------- --------------------------------79 1. IS刺激IL1-β處理HUVEC之MAPKs的磷酸化表現情形---------------79 2. MAPKs抑制劑對IS刺激IL1-β處理HUVEC表現 E-selectin之影響--- 79 未來實驗研究方向-------------------------------------------81 參考文獻------------------------------------------------- 83 附圖---------------------------------------------------- 86 圖一 IS對HUVEC細胞活性之影響------------------------------- 86 圖二 IS對IL-1β所處理之HUVEC表現E-selectin黏附因子之影響------ 87 圖三 IS加強IL-1β所處理之HUVEC表現E-selectin黏附因子之MAPK的影響 --------------------------------------------------------89 Appendix----------------------------------------------- 91 | |
| dc.language.iso | zh-TW | |
| dc.subject | 內皮前驅細胞 | zh_TW |
| dc.subject | 介白素8 | zh_TW |
| dc.subject | 缺氧誘導因子 | zh_TW |
| dc.subject | endothelial progenitor cells | en |
| dc.subject | interleukin-8 | en |
| dc.subject | Hypoxia-inducible factor | en |
| dc.title | 臍帶間質組織衍生內皮前驅細胞經由HIF-1α/IL-8
減少糖尿病鼠下肢缺血性傷害 | zh_TW |
| dc.title | Endothelial progenitor cells derived from Wharton’s jelly of the umbilical cord reduces ischemia-induced hind limb injury in diabetic mice by inducing HIF-1α/IL-8 expression | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 101-2 | |
| dc.description.degree | 博士 | |
| dc.contributor.oralexamcommittee | 歐陽品(Pin Ouyang),王懷詩(Hwai-Shi Wang),吳建春(Jiahn-Chun Wu),林水龍(SHUEI-LIONG LIN),王淑慧(Shu-Huei Wang) | |
| dc.subject.keyword | 內皮前驅細胞,缺氧誘導因子,介白素8, | zh_TW |
| dc.subject.keyword | endothelial progenitor cells,Hypoxia-inducible factor,interleukin-8, | en |
| dc.relation.page | 91 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2013-07-12 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 解剖學暨細胞生物學研究所 | zh_TW |
| 顯示於系所單位： | 解剖學暨細胞生物學科所 | |
文件中的檔案：
| 檔案 | 大小 | 格式 | |
|---|---|---|---|
| ntu-102-1.pdf 未授權公開取用 | 2.64 MB | Adobe PDF |
系統中的文件，除了特別指名其著作權條款之外，均受到著作權保護，並且保留所有的權利。