突變ras基因造成過度表現在骨髓間質幹細胞啟動氧自由基誘導細胞凋亡
特定基因的表現與不同氧自由基的產生,已知會影響細胞的生長和死亡。我\r 個人有興趣利用間質幹細胞體外培養擴充,以為筋骨組織再生的可能應用。因此\r 利用轉植(transfect)突變ras 基因(Glu61Leu)進入骨髓間質細胞株(HS-5 cells),來\r 控制ras 基因表現的高低,進而研究這些間質細胞隨著Ras 蛋白質表現的高低,\r 對氧自由基引導細胞生長與死亡之影響。結果發現ras 基因高度表現的間質細胞\r 生長減緩;相較於原生株平均減少62.4%。進一步研究其生長減少是否與細胞凋\r 亡有關,發現ras 基因高度表現的間質細胞凋亡確實比原生株高22.6%。探究其\r 凋亡原因,發現與caspase-3 有關但和粒腺體功能無關:因為caspase-3 有活化,\r 但是以粒腺體膜電位螢光追蹤劑JC-1 測得的膜電位卻沒有改變。追蹤ras 基因高\r 度表現的間質細胞其細胞內氧自由基的產量,發現Ras 高度表現株其細胞內氧自\r 由基明顯增加。當細胞外加入超氧根轉化酵素(SOD, 500 U/ml)去清除超氧根時,\r 對Ras 高度表現細胞的凋亡沒有影響;但是外加觸化酵素(catalase, 500u/ml)於培\r 養液中,卻可以抑制ras 基因高度表現的間質細胞內caspase-3 活化和細胞凋亡;\r 並且增加細胞生長循環促進分子Cyclin D1 的表現。從這些研究我們歸結出兩點\r 重要新發現: 1) ras 基因突變造成Ras 高度表現時,會促成細胞內特定氧自由基產\r 生,使得細胞生長減緩並進行細胞凋亡,只有特定抗氧化酵素(catalase)才能恢復\r 其異常; 2) Ras 蛋白高度表現而引導氧自由基產生的細胞凋亡與caspase-3 活化\r 有關,但與粒腺體功能無關。根據這些發現,未來我們或許可以朝調節特定氧化\r 還原反應或使用caspase-3 抑制劑去調控間質幹細胞的生長,以供筋骨組織再生\r 的應用。Certain gene’s expression as well as different species of oxygen radicals can\r affect cell growth and apoptosis. We are interested in amplifying mesenchymal\r progenitor cells for the application of musculoskeletal tissue regeneration. Thus, we\r transfected a mutant ras gene (61Glu/Leu) to change Ras protein expression in the\r mesenchymal progenitor cell line (HS-5 cells) and studied how ras expressing levels\r influenced intracellular oxygen radicals, and its relationship to cell growth and\r apoptosis. Results showed that Ras over-expressing HS-5 cells grew slower than those\r with wild type ras HS-5 cells and revealed a higher apoptosis rate. The higher\r apoptosis in Ras over-expressing cells was not related to mitochondrial dysfunction\r since mitochondrial membrane potential was normal as determined by flow\r cytometric analysis of JC-1 fluorescent staining assay. The higher apoptosis was\r related to higher caspase-3 activation. Further studies showed that Ras\r over-expressing HS-5 cells revealed a higher production of intracellular oxygen\r radicals in comparison to those with wild type ras HS-5 cells. Addition of catalase\r (500 u/ml) but not superoxide dismutase (SOD; 500 u/ml) specifically revived the cell\r growth associated with increase of cyclin D1 expression, but decrease of apoptosis\r associated with lower caspase-3 activation. Results from these studies demonstrated\r two important findings: 1) the ras gene over-expressing in a ras-mutant HS-5 cell line\r triggers a higher production of intracellular oxygen radicals resulting in higher cell\r apoptosis; and 2) the higher oxygen radicals related cell apoptosis is mediated by\r caspase-3 but not mitochondrial dysfunction. Based on these findings, we may\r propose to regulate mesenchymal progenitor cell growth for musculoskeletal tissue\r regeneration via modulation of redox reactions or caspase-3 inhibitors in the future.
吸〝氣〞大法-QCM 對有機氣體之吸附與偵測
中文摘要:\r 化學實驗中,常常會使用到各種具有揮發性的有機溶劑,如醇類、酸類、醛類及酮類等。\r 當吸入過量的揮發性有機溶劑時,將會對人體造成嚴重的傷害,故使用時格外需要留意。有\r 鑑於此,我們開始著手研究如何偵測氣相中的有機氣體分子。\r 本研究中,我們利用聚苯胺薄膜吸附氣體分子的特性並結合對質量變化極靈敏的石英晶\r 體微天平,自行設計了一套簡單、藥品用量少且不需昂貴儀器的實驗系統。藉由此系統,我\r 們有效的針對各種揮發性有機溶劑進行偵測,並進一步地探討不同氣體分子對於聚苯胺薄膜\r 的吸附現象。\r \r 英文摘要:\r Students usually use volatile organic solvent in laboratorys such as alcohols, carboxylic acids,\r aldehydes, ketones, etc. It will injure our body when we breathe in too much volatile organic\r solvent. For this purpose, we begin to study how to detect volatile organic compound in air.\r In this work, we combine polyaniline membrane which can adsorb gas molecules and quartz\r crystal microbalance which is highly sensitive to mass changes to design a simple and inexpensive\r system which needs only little chemicals. With this system, we can effectively detect different\r volatile organic compounds. Furthermore, we can also study the adsorption of polyaniline\r membrance for different volatile organic compounds.
微陣列基因分析法探討心肌細胞在機械性展延下的基因表現
高血壓所誘發的機械性展延是造成心肌肥大的基本因子,本實驗即藉由微陣列基因分析法同時大量的分析機械性展延所造成心肌細胞的基因表現。將新生鼠的心肌細胞施以 20﹪的機械性展延,抽取其 mRNA製作成 cDNA 探針與現成的 cDNA 晶片進行雜漬反應 (此晶片上包含了 480個如訊息傳遞、控制細胞生長週期、細胞骨架等的已知基因),在眾多有因為機械性展延而造成基因表現差異的基因中,我們選擇了 eNOS 基因(內皮細胞 NO合成?)進行西方墨點法及 NOS活性和 NO 產生量測定的實驗,進一步證實 eNOS 的基因表現量的確是增加的,此一結果與微陣列基因分析所得之結果不謀而合。 Mechanical stretch induced by high blood pressure is an initial factor laeding to cardiac\r hypertrophy. The use of cDNA microarrays has made it possible to simultaneously analyze\r stretch-induced gene expression in cardiomyocytes. Neonatal rat cardiomyocytes were cultured on\r malleable silicone dishes and were stretched by 20%. We compared the transcript profiles of\r cardiomyocytes under mechanical stretch for 60 minutes by hybridization of cell-derived cDNA to\r DNA probes immobilized on microarrays. The microarrays contained probes for 480 known genes\r including signal transduction, cell cycle regulators, cytoskeleton and cell motility, and so on. Eighteen\r genes were indentified that showed significantly differential expression in response to mechanical\r stretch in cardiomyocytes. Of the represented genes expressed, endothelial nitric oxide synthase (eNOS)\r genes was the most interesting one. Northern blot and western blot analysis further quantified the\r expression of eNOS gene. Mechanical stretch also increased constitutive NOS activity and NO\r production. Our results indicate that mechanical stretch induces eNOS gene expression thus increases\r constitutive NOS activity and NO production in cardiomyocytes.
光子晶體合成、特性與應用
We report an investigation on the synthesis, characterization, and application of photonic crystal. In the study of the synthesis of SiO? nanoparticles for the building blocks of photonic crystal, it is found that by changing the concentration of NH3 solution, we are able to control the size of SiO? nanoparticles. After trying several different methods, we discover that the vertical substrate method is the best way to arrange nanoparticles into a periodic structure. From scanning electron microscope, we confirm that SiO? nanoparticles can form a three dimensional hexagonal photonic crystal. From transmission experiment, we find that the wavelength of the minimum transmission is proportional to the size of nanoparticles. This result implies that using photonic crystals we can control the behavior of electromagnetic wave. Finally, we fabricate CdS nanoparticles on the top of photonic crystals with different diameter of SiO? nanoparticles. Using photoluminescence measurements, we show that by controlling the lattice constant of a photonic crystal the luminescent efficiency of CdS nanoparticles can be substantially enhanced. Out results, therefore demonstrate that photonic crystals are very important for the application of light emitting devices. 本研究主要是著重於探討光子晶體合成、特性分析及其應用。在有關合成光子晶體之奈米二氧化矽顆粒方面, 發現在合成過程中利用氨水的溶量可以控制顆粒的大小。在將奈米顆粒排列成光子晶體的研究中, 嘗試了多種方法後, 發現垂直基座法為最快速有效的方法。從掃瞄電子顯微鏡的觀測, 證實奈米顆粒是以六角對稱排列成整齊的光子晶體。在光子晶體的特性分析中, 利用光穿透實驗, 發現電磁波穿透率最小的波長與奈米顆粒成正比關係, 這顯示出可以利用光子晶體來控制光的行為。最後,本研究將光子晶體與硫化鎘奈米顆粒結合,經由光激螢光譜, 證明光子晶體確實可以增進物體之發光效率,這對發光元件的應用, 將有很大的幫助, 可以節省大量的能源
雞胰臟去氧核醣核酸水解?多型性之探討
去氧核糖核酸水解?,(deoxyribonuclease,簡稱DNase)為一種核酸內切?。目前研究得最透徹的去氧核糖核酸水解?為牛胰臟的DNase I,有四種以上的同功?存在。而牛和雞的DNA序列有極高的相似度,由之前的研究中在等電點焦集電泳法顯示雞胰臟DNase I 具A、B 兩種同功?,本實驗則希望能進一步了解兩種同功?的差異原因。在抽取RNA 後,以RT-PCR 方法合成cDNA,將其以限制?切割,再以大腸桿菌作為載體進行轉殖及繁殖培養,最後將其DNA 定序後進行分析比較。目前已完成第一次定序,但因引子接合過程中有部分位置錯亂造成殖體無法進行表現。另外在此次所得之序列中有發現一個胺基酸的轉變,而其是否為多型性之表現則有待進一步的實驗證實。The most advanced research on deoxyribonuclease in current states is on the bovine pancreatic DNase I, more than four of which isoforms have been found. It is shown that the DNA sequences of chicken and bovine have relatively high similarity. In addition, according to the result of isoelectric focusing from previous researches, chicken pancreatic DNaseI has two isoforms (A and B.) In this research, the author expects to establish more knowledge on the differences on the isoforms and the causes. After receiving the RNA, the RT-PCR is preformed to incorporate the cDNA, which is later cleaved by restriction enzyme and inserted into the plasmid DNA of the E. coli host cell to be cloned. So far a polynucleotide sequence has been deduced from clones of the cDNA, but it cannot be expressed successfully in E.coli due to few random mistaken splicing. There is an alternation of one amino acid, and whether it is the actual state of isoform or not still requires further proof.
南美蟛蜞菊相剋作用之探討
Few plants can grow in the place where Wedelia trilobata grows. In the following experiment, the leaves ans stalks(of the plants from Wedelia trilobata) are extracted for experimental purpose .The data and records of the experiment have shown that Wedelia trilo ata indeed refain the growth of plants such as mung bean, teeding seed, yeast’s respiration and the photosynthesis. We use the function of dieing the polyphenols to know that the Wedelia trilobata can produce Allelochemicals called as the polyphemols The mesophycell produce the polyphemols from chloroplast of cortex,the polyphemols transport to root and release from root exudation. The plant will absorb polyphemols of the soil, and will affect the growth of the plant. If we can separate and purify the Allelochemicals, we can make it as natural herbicideb. 南美蟛蜞菊所在之處,少有其他植物,本實驗採南美蟛蜞菊植栽區之土壤與南美蟛蜞菊莖或葉之萃取液進行相剋作用之研究,結果顯示南美蟛蜞菊確實會抑制小麥與綠豆種子萌發及幼苗生長,並抑制酵母菌的呼吸作用及植物光合作用中之光反應。 利用多酚類染色方法得知南美蟛蜞菊產生的相剋化合物為多酚類,且該多酚類由葉肉細胞與莖內皮層中具葉綠體之細胞產生,經韌皮部輸送至根,再由根經泌濾作用釋出。土壤中的多酚類會被植物吸收,進而影響植物的生長與發育。 若能將該相剋化合物---多酚類分離與純化,或可作為具選擇性之天然除草劑。
「渦」藏「聖」機--以渦流脫離重新詮釋聖嬰發生的原因
If we place a block in continuous, steadily flowing water, some periodic eddy currents will appear behind it. In my report, I would like to introduce a new way to explain how the periodical movement of eddy currents would help to triggering El Nino. According to my experiment, these eddy currents have a certain life cycle, and the eddy current give a force, which drag water form the area they just went by. It will cause water level became lowest for a period of time. These phenomena are also shown in the real data of “Sea Surface Height Anomaly”, it is a quiescence period, and then changes into the highest one it is interesting that highest one always follows by the starting of El Nino. Another important result from my experiment is that, in the same fluid, the frequency of eddy currents is controlled by 2 variables: Size of the block and velocity of the flow, From this result, then get the frequency of eddy current-occur every 3.2 years. It is just correspond with frequency of historical data the El Nino from the 70’s to the present.
本報告嘗試以南極繞極環流在南美洲南端產生週期性渦流脫離的現象來解釋聖嬰的發生根據水槽實驗的結果,水流在通過障礙物後渦流脫離瞬間,會形成一水位低的空區,之後在周圍的水大量湧入,水位突然升高,並有一段時間的振盪。另外,實驗結果也顯示,渦流形成和脫離頻率大致上和注水流速成正比,且和障礙物的大小成反比。根據此結果,對照實驗結果,我認為1993 年3-4 月,1996 年1-6 月及2001 年7-10 月三個「海平面高度異常」的最低值密集帶,此極可能是渦流脫離的時段。且對照「海平面高度異常」實際資料後,發現「海平面高度異常」的「最低值密集帶」後均伴隨水面「寧靜期」,接著就出現周圍暖水流大量湧入的「最高值密集帶」,此時南美洲東南側海水溫度驟增,接著馬上接續著聖嬰的到來。「海平面高度異常」分析的結果,指示渦流脫離的「最低值密集帶」和顯示著聖嬰到來的「最高密值密集帶」間約間隔半年左右。我們推算水的史托爾數約為1,以這個值代入實際南極繞極環流通過南美洲南端,得到渦流脫離的頻率約每3.2 年一次,和長期聖嬰發生的平均頻率非常接近。