Escalated regeneration in sciatic nerve crush injury by the combined therapy of human amniotic fluid mesenchymal stem cells and fermented soybean extracts, Natto

Attenuation of inflammatory cell deposits and associated cytokines prevented the apoptosis of transplanted stem cells in a sciatic nerve crush injury model. Suppression of inflammatory cytokines by fermented soybean extracts (Natto) was also beneficial to nerve regeneration. In this study, the effect of Natto on transplanted human amniotic fluid mesenchymal stem cells (AFS) was evaluated. Peripheral nerve injury was induced in SD rats by crushing a sciatic nerve using a vessel clamp. Animals were categorized into four groups: Group I: no treatment; Group II: fed with Natto (16 mg/day for 7 consecutive days); Group III: AFS embedded in fibrin glue; Group IV: Combination of group II and III therapy. Transplanted AFS and Schwann cell apoptosis, inflammatory cell deposits and associated cytokines, motor function, and nerve regeneration were evaluated 7 or 28 days after injury. The deterioration of neurological function was attenuated by AFS, Natto, or the combined therapy. The combined therapy caused the most significantly beneficial effects. Administration of Natto suppressed the inflammatory responses and correlated with decreased AFS and Schwann cell apoptosis. The decreased AFS apoptosis was in line with neurological improvement such as expression of early regeneration marker of neurofilament and late markers of S-100 and decreased vacuole formation. Administration of either AFS, or Natto, or combined therapy augmented the nerve regeneration. In conclusion, administration of Natto may rescue the AFS and Schwann cells from apoptosis by suppressing the macrophage deposits, associated inflammatory cytokines, and fibrin deposits

CXCL1 Mediated CXCL12/CXCR4 and COX-2/PGE2 Gene Expression Enhances Angiogenesis, Growth and Metastasis

胃癌位居全球癌症死亡的第二位。經由臨床病人檢驗檢體分析發現:趨化激素(CXCL1)與胃癌腫瘤癌化進展及轉移有十分密切之關聯性。為了促使有效的治療，探討基因訊息的表現暨釐清調控機轉即更顯重要。在過去一年初步研究成果中發現，CXCL1誘導胃癌細胞媒介物質PGE2及CXCL12導致血管新生腫瘤生長及轉移現象已有令人興奮的結果。我們已經著手研究該主題，並且積極發展或尋求新穎有效、作用在特定分子的抗腫瘤藥物(厚朴酚)，進而應用於臨床上促使抗腫瘤療效更好、副作用反應更小將是理想的策略。CXCL1在血管新生、腫瘤生長及轉移現象扮演重要的角色。由過去研究發現它與趨化激素IL-8序列分析有著極高的相似度，暗示著兩者有類似的功能。然而過去有關CXCL1分子作用機轉卻十分有限，特別是影響與胃癌腫瘤相關的功能性基因，訊息傳遞及治療的可行性。本研究係針對CXCL1誘導胃癌細胞血管新生、腫瘤生長及轉移為腫瘤生物學探討模式主軸，進一步之訊息傳遞及轉錄因子的參與機制。有鑒於腫瘤治療的重要性，我們認為這個研究模型值得深入探討。我們初步的研究發現：1-2. CXCL1誘導胃癌細胞媒介物質PGE2及CXCL12大量生成，導致血管新生腫瘤生長及轉移現象。3-4. CXCL1誘使COX-2及CXCR4蛋白表達顯著增加。5. CXCL1增加轉錄因子CEBPβ之活性。6. CXCL1促使致癌基因Tpl-2活性增加，特別是在致癌基因Tpl-2的絲氨酸的磷酸化而非蘇胺酸的磷酸化。【計畫重點】基於現有的初步研究以及既往之報導發現，我們提出更進一步的研究促使CXCL1對分子機制及腫瘤基因的調控有更深入的瞭解。我們擬以四年時間探討下列重點:1-2.致癌基因Tpl-2是否調控轉錄因子CEBPβ之活性，進而促進COX-2/PGE2 之生成與CXCL12/CXCR4之表達關聯性。3.分別以過度表達轉錄因子CEBPβ及沉默基因CEBPβ深入探討轉錄因子CEBPβ之重要性。4.以體腫瘤異種移植老鼠探討CXCL1促進癌細胞轉移運用。5.純化合物厚朴酚抑制血管新生腫瘤生長及轉移現象之分子機制及腫瘤基因的調控有更深入的瞭解。6.以化學致癌劑MNNG誘導癌化過程中探討CXCL1可能扮演腫瘤癌化生成之角色。我們的初步研究開啟未來四年的研究可行性暨更多的視野，經由探討及瞭解CXCL1 作用在人類胃癌細胞可能扮演的角色及調控蛋白之機制，進而應用於臨床胃癌病人的治療將有莫大的助益。同時本研究的作業平台，將可以更有效地將基礎研究的成果，應用到國人重要癌症篩檢開發，並提供分子藥理開發成為新藥物的靶點，建構出高特異性、作用更精確的藥物，為發展生技製藥奠定基礎暨提升我國生醫科技水準。Gastric cancer is the second most common cause of death from cancer in the world. Clinicalspecimen analysis revealed that increased chemokine CXCL1 expressions correlate with enhancedgastric tumor progression and metastasis. To improve the therapeutic results, the exploration ofexpression gene and elucidation of mechanism of tumor progression are considered to be of greatimportant. Numerously mediators such as prostaglandin E2 (PGE2), CXCL12 promote gastric tumorgrowth by stimulating angiogenesis, cell invasion, and cell growth, and inhibiting apoptosis. We haveinitiated a study to search for cell signaling events involved in human gastric cancer genetic alterations.Molecules that regulate tumor-associated angiogenesis provide promising therapeutic targets fortreatment of gastric cancer as indicated by the recent development of the novel anti-angiogenic agentHonokiol.CXCL1 (melanoma growth-stimulatory activity/growth-regulated protein alpha), plays a majorrole in inflammation, angiogenesis, tumorigenesis, and metastasis. CXCL1 has extensive sequencesimilarity to the IL-8, suggesting functional similarities. In a variety of cancer cell lines, rodent tumormodels and human cancer biopsies, the level of CXCL1 is elevated, correlating with increasedtumorigenicity. Pathological conditions such as tumor growth and malignancy correlate withneutrophil-activating properties of CXCL1. Evidence shows that CXCL1 correlation with cancerprogression and carcinogenesis. However, little is know about the mechanism and targeted therapyunderlying their molecules action in gastric cancer.Our preliminary data showed that: (1) CXCL1 induces cancer cell growth by in vitro cellproliferation assay, soft agar assay and in vivo tumor growth. (2) CXCL1 promotes angiogenesis by invitro tube formation, CAM assay, in vivo matrigel plug assay, ex vivo aorta ring assay. (3) CXCL1 upregulates cancer cell metastasis by in vitro cancer cell migration, invasion assay, MMP activity and invivo animal study peritoneal metastasis. (4) CXCL1-induced mediators, especially chemokines(CXCL12) and prostaglandins (PGE2) production by ELISA assay. CXCL1-evoked CXCR4 or COX-2mRNA and protein expression in gastric cancer cells and HUVECs. (5) Increased expression ofCXCL1 has been attributed to constitutive activation of transcription factor CEBPβ throughmitogen-activated protein kinase, but not AP-1 and SP-1. The novel pathway is different fromprevious study in the NF-κB activation. (6) CXCL1-induced tumor progression locus 2 (Tpl-2)proto-oncogene kinas activity through serine phosphorylation but not threonine phosphorylation.We propose a four years project to focus on: (1) To elucidate the underlying mechanism thatTpl-2 may trigger CEBPβ activity result in angiogenesis via the COX-2 and PGE2 dependent pathwayin gastric carcinoma. (2) To investigate the underlying mechanism that Tpl-2 may trigger CEBPβactivity result in angiogenesis via the CXCL12/CXCR4 dependent pathway in gastric carcinoma (3)Further to determine involves in CXCL1 enhances angiogenesis and tumor growth signaling cascadeand regulates mechanisms through CEBPβ activation in human gastric cancer. (4) Further to verifyCXCL1 exerts biological activity in cell metastasis and enhances peritoneal dissemination inorthotopic xenograft mouse models. (5) Further to verify the promising drug Honokiol exertsbiological activity in suppression of cancer angiogenesis and metastasis in vivo and in vitro were becarried out. (6) Further to determine the role of CXCL1 in stomach carcinogenesis and tumorigenesisprocess were being achieved.Our results should facilitate the understanding of this expanded role in promoting tumor biologyand provide the therapeutic possibility as a potential chemotherapeutic agent in open new doors totherapeutic intervention

Cleavage of Glucose-Regulated Protein-94 by Honokiol Induces Endoplasmic Reticulum Stress-Mediated Human Gastric Cancer Cell Apoptosis and Tumor Growth in vitro and in vivo

胃癌位居全球癌症死亡的第二位。為了尋求更好的治療效果，探討胃癌細胞內的過度表達基因並進一步釐清其腫瘤生成機轉顯得格外重要。此外，由於目前使用之抗腫瘤藥物仍有許多的缺點及改進空間，所以積極發展或尋求新穎有效、作用在特定分子的抗腫瘤藥物，進而應用於臨床上，促使抗腫瘤療效更好、副作用反應更小將是理想的策略之一。我們已經著手研究人類胃癌細胞內的過度表達基因GRP94, 位居內質網內已知具有抗凋亡的特性。在許多腫瘤細胞株及腫瘤組織病理切片中皆可發現GRP94蛋白大量表達。然而對於其作用機轉、相關訊息傳遞之交互作用，以及將它列為腫瘤標的分子等卻很少有深入的探討。中藥厚樸萃取純化合物厚樸酚(Honokiol)在我們過去的研究中已知具抗氧化、抗發炎的作用。此外之外，也俱備抗血管增生、抗腫瘤特性，然而其分子作用機轉目前尚待研究。特別是影響與胃癌腫瘤相關的功能性基因更須進一步釐清與探討。本研究係針對胃癌腫瘤過量表達蛋白GRP94為標的分子蛋白，以厚樸酚(Honokiol)治療胃癌腫瘤。有鑒於腫瘤治療的重要性，我們認為這個研究模型值得深入探討。我們初步的研究發現：1. GRP94蛋白在胃癌腫瘤細胞株(MKN45, AGS, N87, SCM-1)及胃癌腫瘤組織切片皆有過量表達。2. 在試管內組織培養的環境中，厚樸酚具有促使細胞凋亡效應。3. 厚樸酚誘導胃癌腫瘤細胞株MKN45之GRP94蛋白減少，但GRP78蛋白並不受到影響。【計畫重點】基於現有的初步研究以及既往之報導發現，我們提出更進一步的研究促使厚樸酚對分子機制及腫瘤基因的調控有更深入的瞭解。我們擬以三年時間探討下列重點:1. 探討厚樸酚誘導胃癌腫瘤細胞株(MKN45, AGS, N87, SCM-1)細胞凋亡效應及促使GRP94 蛋白降解之機轉。2. 釐清厚樸酚是否影響細胞膜表面之酪胺酸激.磷酸化之活性或是作用於酪胺酸激.蛋白質本身之表現。3. 分析厚樸酚是否誘導內質網壓力之生成。4. 以動物模式分析厚樸酚是否具有對抗腫瘤生長、增加存活率、降低或降解GRP94 蛋白之表達及可能之藥物毒理之意義。我們的研究將打開更多的視野，暸解人類胃癌細胞GRP94 蛋白之調控機制，進而應用於臨床胃癌病人的治療，將有莫大的助益。Gastric cancer was the second most common cause of death. To improve the therapeuticresults, the exploration of over-expression gene and elucidation of mechanism of tumorprogression are considered to be of great important. Current research in this area is driven bythe need to cover new agents in target specific molecule that will be more effective and havefewer side effects. We have initiated a study to search for cell signaling events involved inhuman gastric cancer genetic alterations.GRP94, is a 94 kDa calcium-binding glycoprotein. GRP94 is a chaperone proteinlocalized in the endoplasmic reticulum with antiapoptotic properties. In a variety of cancercell lines, rodent tumor models and human cancer biopsies, the level of GRP94 is elevated,correlatingwith increased tumorigenicity. Pathological conditions such as tumor growth andmalignancy correlate with cytoprotective protein GRP94 over-expression. Evidence showsthat GRP94 correlation with cancer progression and carcinogenesis. However, little is knowabout the mechanism and targeted therapy underlying their molecules action in gastric cancer.In the search for a more effective single or adjuvant therapy to treat gastric cancer, weinvestigated the effects of the Honokiol, an active component isolated from the herb 現Houpo現(Magnolia officinalis) traditional Chinese herbal medicines, on cell growth and apoptosis ofgastric cancer cells. Honokiol, an activated pure compound, was known to possess potentanti-oxidant, anti-inflammatory, anti-angiogenesis, anti-neoplastic properties. However, themolecular mechanism underlying the anticancer effect of Honokiol is poorly understood.Specifically, whether this pure compound affects the expression of cancer-related genes hasnot been defined. Thus, investigating changes in gene expression profiles, GRP94over-expression, as a result of herbal extract pure compound treatment may help define theunderlying mechanism of action and validate the efficacy of these anticancer reagents.Our preliminary data showed that:1. GRP94 overexpression in gastric cancer in vivo and in vitro.2. Honokiol activates apoptosis.3. Honokiol induces GRP94 protein reduction but not GRP78 protein in gastric cancer celllines.Encouraged by these findings, we decided to investigate whether Honkiol could also beuseful in an oncological context, exerting inhibits tumor growth, induces apoptosis andanti-tumor properties predictive of clinical translation. According to the aforementionedpreliminary data that we propose to study the effect of Honokiol-induced human gastriccancer apoptosis via regulates GRP94 degradation mechanism in in vitro assay and exhibitsantitumor activity in vivo focusing on 4 specific aims as below. Specific aims 1, 2 are to becompleted in the first year, and specific aim 3 in the two years, and specific aim 4 in the thirdyears.We propose a third years project to focus on:Specific Aims 1: Further to determine involves in Honokiol activates apoptosis and GRP94degradation signaling cascade and regulates mechanisms in human gastric cancer.Specific Aims 2: Further to verify Honokiol exerts biological activity in the target cellstyrosine-kinase expression or activity in vitro assay were carried out.Specific Aims 3: To identify whether Honokiol induces endoplasmic reticulum (ER) stress.Specific Aims 4: Further determined involves in Honokiol attenuates tumor GRP94over-expression, tumor burden and survival rate in nude mice.Our results should facilitate the understanding to the regulatory mechanism ofHonokiol-antitumor activity and provide the therapeutic possibility as a potentialchemotherapeutic agent

Honokiol Activates PPAR-Gamma Activity in vitro and Exhibits Antitumor Activity in vivo

Peroxisome Proliferator-Activated Receptor gamma (PPAR )是轉錄因子也是細胞核之接受器，與retinoid X receptor (RXR)形成異二元體。當轉錄因子PPAR 接受刺激活化後會與RXR 結合進入細胞核與DNA 結合、進一步啟動下游標的基因。PPAR 在許多腫瘤細胞皆有大量過度表現，胃癌腫瘤細胞也是如此。胃癌是常見的消化道惡性腫瘤，但目前的治療成效卻十分有限。所以發展有效抗腫瘤、降低副作用的新藥物將是重要的治療策略之一。我們已經初步著手探討研發新的轉錄因子PPAR 之活化劑-厚樸酚(Honokiol)，具活性之純化合物-厚樸酚、可由中藥厚樸萃取出。厚樸酚(Honokiol)已廣泛使用於傳統醫藥之應用，目前已知具有抗發炎、抗血管增生之效用。然而抗腫瘤生長及其分子機轉至今仍然不十分清楚。目前就已知的文獻中，厚樸酚(Honokiol)對胃癌腫瘤PPAR 之活性作用、影響腫瘤之侵襲能力、對抗腫瘤生長作用等等皆尚未釐清其功用暨調控機制。我們目前之初步研究發現:1. 厚樸酚(Honokiol)可以有效誘導胃癌腫瘤細胞之凋亡.、增加凋亡.活性、進一步造成細胞凋亡。2. PPAR 之拮抗劑GW9662，可以有效抑制細胞凋亡.活性、有效抑制細胞凋亡。3. PPAR 之配體、促進劑(15d-PGJ2)，可以有效增加胃癌腫瘤細胞之細胞凋亡.活性。4. 厚樸酚(Honokiol)活化轉錄因子PPAR ，促使PPAR 與DNA 結合活性增加。基於既往之報導以及現有之實驗結果，我們提出更進一步的研究促使厚樸酚(Honokiol)對PPAR 之分子機制、影響腫瘤侵襲之能力、對實驗動物之抗腫瘤生長能力有更深入的瞭解。我們擬以兩年時間探討下列重點:1. 更進一步探討厚樸酚(Honokiol)對轉錄因子PPAR 之作用基因轉錄活性。2. 比較分析厚樸酚(Honokiol)與不同的PPAR 配體、促進劑在胃癌腫瘤細胞之活性效用。3. 分析厚樸酚(Honokiol)對抗腫瘤細胞之侵襲能力。4. 探討可能之藥物毒理意義--誘導實驗動物胃癌腫瘤形成之動物模式，分析厚樸酚(Honokiol)對抗腫瘤生長之效應及存活率

Studies on the cellular signaling pathways in hyperglycemia-induced cell proliferation and apoptosis

糖尿病為一常見的內分泌代謝疾病，目前已經成為公共衛生問題的一部份。在世界各地隨著肥胖人數日驅增加，糖尿病的人數也隨之顯著提高。而糖尿病對身體造成最大的問題是對實質性組織器官的傷害；尤其是心血管系統以及腎臟的影響。也由於在世界各地糖尿病的病人愈來愈多，其導致的複雜變化以及合併病發症更值得我們去重視。 糖尿病病人體內血糖過高的訊息是不可逆的並且最後導致組織器官的功能障礙或損傷。由過去的文獻報導以及臨床病人報告中已知：糖尿病的病理過程可引起血管內皮細胞功能缺失，造成的血管功能障礙、加速冠狀動脈硬化並對預後不佳。然而進一步地導致血管內皮細胞凋亡的機制至今仍不十分清楚。探討此一異常訊息是如何造成人類血管內皮細胞凋亡？其機制為何？進而如何避免糖尿病在人類心血管系統功能障礙及合併症的產生，以及病程的惡化是十分重要的課題。於本論文第一部分的研究主要為探討高血糖的狀態下誘導人類血管內皮細胞凋亡之機制，以及PI3K所調控之環氧化蛋白酶表現在其中的角色。本實驗發現暴露於高血糖的環境中皆可測得早期血管內皮細胞凋亡訊號。在細胞凋亡的形態學上以Hoechst染色觀察細胞形態以及以Annexin V/Propidium Iodide偵測早期細胞凋亡訊號。高血糖狀態也會誘導環氧化蛋白酶的表現增加，同時也誘導環氧化蛋白酶的下游產物PGE2生成，進而促使凋亡酶-3 (caspase-3)活性增強而導致細胞走向凋亡。出乎意外地，給予PI3K抑制劑皆可有效地抑制高血糖狀態之環氧化蛋白酶蛋白表現、環氧化蛋白酶的下游產物PGE2生成、凋亡酶-3活性和細胞凋亡。高血糖誘導PI3K活化也依序促使Akt的磷酸化。更進一步地，高血糖引發氧化自由基的生成和NFDiabetes has become a public health crisis. With the incidence of obesity rising in the world, the number of diabetics will grow considerably. Of greatest concern is the impact this trend will have on damage to the kidneys and cardiovascular disease. The incidence of diabetes is increasing worldwide, with subsequent increase in the incidence of diabetic complication. The hyperglycemic signal exchange occurs ubiquitously and irreversibly in patients with diabetes mellitus, and its consequences are especially relevant to organ dysfunctions. In type-2 diabetes, a greater proportion of patients have overt nephropathy at shortly and vascular complications after diagnosis of diabetes. In this thesis, the studies are divided into two parts for description. Diabetes has been demonstrated to accelerate vascular dysfunction, coronary atherosclerosis, and the prognosis were worse following cardiac events. Therefore, the part I will investigate the regulation of abnormal signalings that promote human umbilical vein endothelial cells apoptosis under high glucose condition and relevant in understanding the intracellular signaling associated with vascular disease and preventing the development of vascular complication in diabetics, principally the evolution of this practice from its beginning until cell death. Diabetes mellitus causes endothelial dysfunction. The precise molecular mechanisms by which hyperglycemia causes apoptosis in endothelial cells are not yet well understood. The aim of this study was to explore the role of cyclooxygenases-2 (COX-2) and the possible involvement of phosphoinositide 3-kinase (PI3K) signaling in high glucose (HG)-induced apoptosis in human umbilical vein endothelial cells (HUVECs). For detection of apoptosis, the morphological Hoechst staining and Annexin V/Propidium Iodide staining were used. Glucose up-regulated COX-2 protein expression, which was associated with the induction of prostaglandin E2 (PGE2), caspase-3 activity and apoptosis. Unexpectedly, we found that PI3K inhibitors could suppress COX-2 expression, PGE2 production, caspase-3 activity, and the subsequent apoptosis under HG condition. Glucose-induced activation of PI3K resulted in the down stream effector Akt phosphorylation. PI3K inhibitors effectively attenuated the intracellular reactive oxygen species (ROS) generation and NF-TABLE OF CONTENTS 中文摘要 1 Abstract 5 List of abbreviations 9 PART I CHAPTER 1 Introduction 1.1 Diabetes "Basics" 11 1.2 Classification 11 1.3 Signs and complications 14 1.4 Endothelium cellS in cardiovascular system 16 1.5 The role of transcription factors in human umbilical vein endothelial cells pathophysiology of diabetes 17 1.6 The role of cyclooxygenase-2 (COX-2) in human umbilical vein endothelial cells pathophysiology of diabetes 18 1.7 The functions of glomerular mesangial cell 20 1.8 The role of glomerular mesangial cell in pathophysiology of diabetes 20 1.9 The role of COX-2 in glomerular mesangial cell 22 1.10 The role of NF- B in glomerular mesangial cell 24 1.11 The role of PI3K in glomerular mesangial cell 25 1.12 Purpose and rationales of present research 27 CHAPTER 2 Materials and Methods 28 2.1 Primary culture HUVECs 29 2.2 Apoptosis and Caspase-3/cpp32 activity 29 2.3 Phosphoinositide 3-kinase (PI3K) activity 30 2.4 Immunoblotting 30 2.5 Electrophoretic mobility shift assay (EMSA) 31 2.6 Reactive oxygen species production 31 2.7 PGE2 production 32 2.8 Mesangial cell culture 32 2.9 Cell proliferation assay MTS Assay 33 [3H]Thymidine incorporation 33 2.10 RT-PCR for COX-2 expression 34 2.11 Preparation of nuclear extracts 35 2.12 Transient transfection with dominant-negative vectors-DN-P85 and DN-Akt 35 2.13 Immunofluorescence staining for NF- B 36 2.14 Statistical analyses 36 CHAPTER 3 Results in endothelial cells 38 3.1 COX-2 protein expression and involvement in high glucose (HG)-induced apoptosis 38 3.2 PI3K activity and involvement in HG-induced COX-2 protein expression and apoptosis 38 3.3 Effects of PI3K inhibitors on HG-induced NF

Inhibition of Nadph Oxidase-Related Oxidative Stress-Triggered Signaling by Honokiol Suppresses High Glucose-Induced Human Endothelial Cell Apoptosis

Angiopathy is a major complication of diabetes. Abnormally high blood glucose is a crucial risk factor for endothelial cell damage. Nuclear factor-kappaB (NF-kappaB) has been demonstrated as a mediated signaling in hyperglycemia or oxidative stress-triggered apoptosis of endothelial cells. Here we explored the efficacy of honokiol, a small molecular weight natural product, on NADPH oxidase-related oxidative stress-mediated NF- kappaB-regulated signaling and apoptosis in human umbilical vein endothelial cells (HUVECs) under hyperglycemic conditions. The methods of morphological Hoechst staining and annexin V/propidium iodide staining were used to detect apoptosis. Submicromolar concentrations of honokiol suppressed the increases of NADPH oxidase activity, Rac-1 phosphorylation, p22(phox) protein expression, and reactive oxygen species production in high glucose (HG)-stimulated HUVECs. The degradation of IkappaBalpha and increase of NF-kappaB activity were inhibited by honokiol in HG-treated HUVECs. Moreover, honokiol (0.125-1 microM) also suppressed HG-induced cyclooxygenase (COX)-2 upregulation and prostaglandin E(2) production in HUVECs. Honokiol could reduce increased caspase-3 activity and the subsequent apoptosis and cell death triggered by HG. These results imply that inhibition of NADPH oxidase-related oxidative stress by honokiol suppresses the HG-induced NF-kappaB-regulated COX-2 upregulation, apoptosis, and cell death in HUVECs, which has the potential to be developed as a therapeutic agent to prevent hyperglycemia-induced endothelial damage

A study of the relationship among entrepreneurial attitude, entrepreneurial perception and entrepreneurial behavior

近年來，隨著全球經濟不景氣及金融海嘯的影響，國內失業人口亦逐漸攀高，面臨經濟低迷的衝擊，創業實為促進經濟成長與增加就業機會的途徑之一。創業活動的蓬勃發展除能替個人創造財富，亦能降低失業率與通貨膨脹，為國家帶來經濟效益，可見創業活動與國家經濟發展關係密切。因此了解臺灣民眾的創業行為狀況及其影響因素已成為值得關注的課題。 過去曾有學者從內在因素與外在因素來探討創業行為之影響因素，其中內在因素包含人格特質、個人價值觀、創業態度、創業認知等，外在因素包含政府政策、創業教育、社會資本、組織資源等。本研究之主要目的為藉由實證研究，探討臺灣成年民眾在創業態度、創業觀感上的認知與創業行為間之關聯性，並試圖找出影響創業行為（包含創業傾向、創業活動）的關鍵因素。 研究結果顯示，有創業傾向的比例約占29%左右，參與投入創業活動的比例約占12%左右。影響創業傾向與創業活動之因素並不完全相同，創業傾向除了受到創業態度的影響之外，還與創業觀感有所關聯，而是否參與創業活動則只與創業態度有關，其中以認為自己擁有足夠創業能力的人，以及認識的人當中有創業經驗的人，其有參與創業活動及未來有創業傾向的可能性較高

High Glucose Induces Human Endothelial Cell Apoptosis through a Phosphoinositide 3-Kinase-Regulated Cyclooxygenase-2 Pathway

Objectives-: Diabetes mellitus causes endothelial dysfunction. The precise molecular mechanisms by which hyperglycemia causes apoptosis in endothelial cells are not yet well understood. The aim of this study was to explore the role of cyclooxygenase-2 (COX-2) and the possible involvement of phosphoinositide 3-kinase (PI3K) signaling in high glucose (HG)-induced apoptosis in human umbilical vein endothelial cells (HUVECs). Methods and Results-: For detection of apoptosis, the morphological Hoechst staining and Annexin V/propidium iodide staining were used. Glucose upregulated COX-2 protein expression, which was associated with the induction of prostaglandin (PG) E2 (PGE2), caspase- 3 activity, and apoptosis. Unexpectedly, we found that PI3K inhibitors could suppress COX- 2 expression, PGE2 production , caspase-3 activity, and the subsequent apoptosis under HG condition. Glucose-induced activation of PI3K resulted in the downstream effector Akt phosphorylation. PI3K inhibitors effectively attenuated the intracellular reactive oxygen species (ROS) generation and nuclear factor [kappa]B (NF-[ kappa]B) activation. Blocking the PI3K and Akt activities with the dominant-negative vectors greatly diminished the HG -triggered NF-[kappa]B activation and COX-2 expression and apoptosis. Conclusions-: These results suggest that HG, via PI3K/Akt signaling, induces NF-[kappa]B-related upregulation of COX-2, which in turn triggers the caspase-3 activity that facilitates HUVEC apoptosis. Also, HG may cause ROS generation in HUVECs through a PI3K/Akt-dependent pathway