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Study of common Chinese herb on anti-cancer ability The effect of andrographolide on ultraviolet B radiation induced apoptosis in human keratinocytes HaCaT cell Part II：The study of mediate for effect of oxidative stress by osmanthus fragrans flower extra
No full text誌謝 iii
中文摘要 - 3 -
Abstract - 5 -
第一部分：穿心蓮內酯對紫外線輻射B誘導人類角質層細胞之影響
- 1 -
第一章 前言 - 2 -
一、常用中草藥植物 - 2 -
二、穿心蓮（ Andrographis paniculata ( Burm. F. ) Nees ） - 3 -
三、穿心蓮內酯 ( Andrographolide；Andro ) - 6 -
四、紫外線（Ultraviolet radiation ; UVR）簡介 - 9 -
五、細胞凋亡 - 11 -
六、研究目的 - 14 -
七、研究架構 - 15 -
第二章 材料與方法 - 16 -
第三章 結果 - 23 -
參考文獻 - 30 -
圖表 - 36 -
第二部分：桂花萃取物調控人類肝癌細胞株氧化壓力之影響 - 45 -
第一章 前言 - 46 -
一、氧化壓力 - 46 -
二、肝癌 - 47 -
三、天然藥草植物 - 49 -
四、桂花 - 50 -
五、目的 - 53 -
六、研究架構 - 54 -
第二章 材料與方法 - 55 -
第 四 章 結果 - 61 -
第五章 討論 - 63 -
圖表 - 69 -
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"Scavenging effects of phenylpropanoid glycosides from Pedicularis
on superoxide anion and hydroxyl radical by the spin trapping method" Biochemical Pharmacology. pp. 687-691.[[abstract]]我們將本論文分做兩個部分進行探討。
第一部分：紫外線的輻射是造成許多皮膚病變的原因之一，長時間在紫外線的照射下會造成角質細胞的損傷。而紫外線輻射的中波UVB所造成細胞的損害尤其嚴重，可能造成DNA或蛋白質等分子的損害，當這些損傷過於嚴重的時候會使得角質細胞趨向於癌化。因此，如何抑制已損傷的角質細胞走向癌化並減緩皮膚癌的發生是本文研究的動機。先前國內外之研究證實穿心蓮內酯能夠發揮抗發炎及抗癌的效用，我們進一步探討穿心蓮內酯對於紫外線所造成的人類角質層細胞（HaCaT）的影響極其造成的死亡機轉。實驗中以不同濃度的穿心蓮內酯(0-50 μM)與UVB（1.5 mJ/cm2）的照射分別處理下，結果顯示在低劑量的穿心蓮內酯（1 μM ~ 5 μM）對HaCaT細胞是不具細胞毒性的，以及在UVB照射的試驗則發現當細胞單獨照射UVB 1.5 mJ/cm2的劑量會造成約4%的細胞死亡。而在穿心蓮內酯濃度(2.5 μM和5 μM)與1.5 mJ/cm2的UVB共同作用細胞後，可更加促進細胞走向凋亡。由此我們推測此傳統中草藥「穿心蓮內酯」具有預防紫外線照射後角質細胞癌化之潛力，對於抑制或減緩人類皮膚癌應有相當的裨益。
第二部分：活性氧分子（Reactive oxygen species; ROS）是造成氧化壓力的原因之ㄧ，氧化壓力的增加可能對細胞造成傷害，使細胞可能借由細胞凋亡或是細胞壞死的途徑而走向死亡。在近年來的研究也得知利用調控ROS的生成可抑制癌細胞的生長； 桂花（Osmanthus fragrans flower）含有許多的抗氧化物質，其萃取物具有增加人類肝癌細胞HepG2 cell氧化壓力的效果。在MTT及LDH的分析結果發現對HepG2具有生長抑制的效果。然而在本實驗中並未發現Osmanthus fragrans flower萃取物會造成HepG2凋亡的表現。此外，我們發現桂花萃取物也對於正常人類肝細胞Chang liver cell則有促進生長且增加細胞模的完整性的功能。因此，桂花萃取物可能應用在護肝上及肝癌相關的治療是具有相當潛力的。
We will devide it into two parts to introduction in the study.
In part one： Ultraviolet radiation（UV） is the cause of many skin diseases, and being exposing under ultraviolet radiation on long-time bases could cause the injury of keratinocyte cell. The UVB of Ultraviolet radiation could caused the most serious injury to cells, and the UVB could caused of injury in DNA or protein and lading to promoter tumor produced. Therefore, Is been interested by how to suppressed the cell injury. Function of andrographolide could to anti-cancer and anti- immunity. In our study, we used different concentrate of andrographolide ( 0 μM-50 μM )on effect of low dose UVB（ 1.5 mJ/cm2 ） induced apoptosis in human keratinocyte cell. In the result, low dose andrographolide （1 μM、2.5 μM、5 μM）is non toxic effect in HaCaT cell. Induced 4 % death by 1.5 mJ/cm2 of UVB in HaCaT cell, and the 2.5 and 5μM andrographolide combined UVB could enhance apoptosis landing to cell death. In future, can application use the andrographolide in inhibit UVB promoter tumor by Chinese medical herb in human keratinocyte cell, and reduce occurrence of skin cancer.
In part two：Oxidative stress is caused by exposure to reactive oxygen species, which can affect the damage effect of proteins and cell membranes, even lead to cell death via necrosis or apoptosis. So, the cancer cell inhibited by regulating intracellular ROS has been described in recently studies.
The extract of Osmanthus fragrans flower is rich with antioxidant compounds and enhances ROS produced in Hep G2 cell line. MTT and LDH assays showed that the extract of Osmanthus fragrans flower reduced cell viability in Hep G2 cell, but not showed apoptosis. Furthermore, no toxicity was found in the Chang liver cell treated with Osmanthus fragrans flower extract and must be maintain the integrity of the cell membrane. These results suggest that Osmanthus fragrans flower extract may be valuable for a hepatoprotective agent and possible to development of the therapeutic management of human hepatoma
In charge of the paoduct and enclose the tactics to reduce the blood and dialysis patients blood than the volume rate-Take one regional hospital in the south as an example
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Nutrient Control of the Cellular Growth in the Prothoracic Glands of the Silkworm, Bombyx Mori
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Analysis of variants biological function of recombinant human SCF 1-165 and SCF1-141 produced by the yeast expression system
Get PDF[[abstract]]中文摘要
人類幹細胞生長因子(human Stem cell factor; hSCF)，為一種醣蛋白，可廣泛運用在基礎、臨床研究、生物科技與工業製程之發展。SCF藉由結合KIT胞外的部份，導致酪氨酸激酶的活化，進而起始了很多細胞內的反應，包括造血、器官生成、黑色素生成、細胞增生、分化、遷移、生存等功能。為了進一步探討SCF結構與功能的關係，本研究乃利用酵母菌表現系統誘導表現重組SCF1-165(wild type)與SCF1-141(truncated)，並分析兩者間生物活性與熱穩定性差異。SCF1-141為SCF1-165之接受器結合功能區(receptor binding domain)，是SCF1-165在C-terminus處短缺一段序列(STLSPEKDSRVSVTKKPFMLPPVA)，兩者皆屬可溶式幹細胞生長因子(soluble stem cell factor；sSCF)。結果顯示，利用酵母菌表現系統可成功地大量表現rhSCF1-165或rhSCF1-141。在純化重組SCF蛋白質的過程中，我們選擇疏水性交互作用層析法(Hydrophobic interaction chromatography)及陰離子交換樹脂層析法(Anion exchange chromatography)。利用Alamar blue測定TF-1細胞存活率實驗中，發現在加入不同濃度的重組SCF1-141或SCF1-165 具有dose-dependent增生的效果。在固定濃度(100 ng/ml)、固定時間(十分鐘)及不同溫度(25，50，70，90，110℃)條件下各加熱十分鐘，可觀察到重組SCF1-165生物活性均優於SCF1-141。再者，固定濃度(100 ng/ml)、固定溫度(90℃)，在加熱不同時間(0、2、10、30、60、150分鐘)，則觀察到重組SCF1-165分別在90℃加熱0分鐘、2分鐘、10分鐘與30分鐘，生物活性都高於SCF1-141。若測定SCF1-141和SCF1-165二者在不同溫度(25、50、70、90℃)及不同濃度(500、100、10 ng/ml)下加熱十分鐘後細胞存活率之分析，結果顯示，SCF1-141 及SCF1-165兩者在高濃度(500、100 ng/ml)及不同溫度(25、50、70、90℃)下，生物活性雖有下降趨勢，但均較控制組高。
Abstract
Human stem cell factor (hSCF), as a glycoprotein, can be widely used in basic and clinical research, biotechnology and the development of industrial processes. It has been shown that SCF initiated diverse cellular responses including hematopoiesis, organgenesis, melanin pigmentation, and development of proliferation, differentiation, migration and survival of cells. To further explore the relationship between structure and function of SCF, we produced rhSCF1-165 (wild type) or rhSCF1-141 (truncated) by yeast expression system and compared their differences between their biological activity and thermal stability. Recombinant hSCF1-141, which is truncated form of rhSCF1-165 in its C-terminus (STLSPEKDSRVSVTKKPFMLPPVA), is the receptor binding domain of rhSCF1-165. Both of them are soluble-type stem cell factor (sSCF). In this study, we showed that overexpression of rhSCF1-165 or rhSCF1-141 can be successfully performed by yeast expression systems in a large amount. In the purification processes of rhSCF, we use hydrophobic interaction chromatography and anion exchange resin chromatography. The biological function study of rhSCF1-165 or rhSCF1-141 was measured by using Alamar blue in TF-1 cell survival assay. We found that it has a dose-dependent proliferation manner at the different concentrations of rhSCF1-141 or rhSCF1-165. At the concentration of 100 ng/ml and different temperatures (25, 50, 70, 90, 110 ℃) under the conditions of the heat 10 minutes, we can observed that the biological activity of rhSCF1-165 are better than rhSCF1-141. Moreover, we found that the biological activity of rhSCF1-165 is higher than rhSCF1-141 when they were heated at 90 ℃ for 0, 2, 10 or 30 minutes at the concentration of 100 ng/ml under the different heating time conditions for 0, 2, 10, 30, 60, or 150 minutes. If the determination of rhSCF1-141 or rhSCF1-165 both at different temperatures (25, 50, 70, 90 ℃) and different concentrations (500, 100, 10 ng/ml) after heating 10 minutes, it showed that both has decline in their biological activity but still be higher than control group in high concentration (500, 100 ng/ml) and under different temperature condition(25, 50, 70, 90 ℃)