嘉義長庚醫院/長庚科大發表論文貴重儀器應用簡述

(2021年6月-9月)

 
項次 論文
1 Yen Ju-Bei (顏如貝醫師), Chen Lee-Wen, Wei Ling-Huei, Hung Chien-Hui, Wang Shie-Shan, Lin Chun-Liang and Chang Pey-Jium*(張沛鈞老師) (2021). "Identification and Characterization of Human Norovirus NTPase Regions Required for Lipid Droplet Localization, Cellular Apoptosis, and Interaction with the Viral P22 Protein." Microbiology Spectrum 9(1): e00422-00421.
NTPase(核苷三磷酸酶)是一種人類諾羅病毒編碼蛋白,將N端區域(1-179)的NTPase融合綠色螢光蛋白(GFP),以流式細胞分析儀檢測人胚胎腎細胞(HEK 293T)轉染NTPase(1-179)-GFP表達質體對其細胞凋亡的影響(分析Annexin V表現)。
共軛焦顯微鏡觀察諾羅病毒蛋白NTPase免疫螢光染色結果,觀察到此病毒蛋白在細胞內會包裹於脂肪滴(Lipid Droplet)外圍,透過表現NTPase不同的刪除片段,找出兩段同時具有親脂性與親水性的LD-targeting motifs (LTM-1及LTM-2),並進一步深入探討其對諾羅病毒複製與致病機轉所扮演的角色。
2 Chang Shun-Fu (張順福博士), Huang Kuo-Chin, Lee Kuan-Han, Chiang Yao-Chang, Lee Wei-Ru, Hsieh Rong-Ze, Su Yu-Ping* and Wu Shun-Chi* (2021). "Effects of Visfatin on Intracellular Mechanics and Catabolism in Human Primary Chondrocytes through Glycogen Synthase Kinase 3β Inactivation." International Journal of Molecular Sciences 22(15): 8107.
為探討內臟脂肪素(visfatin)與骨關節炎(Osteoarthritis, OA)發生的關聯,以共軛焦顯微鏡拍攝螢光小球在人類軟骨細胞內的移動情形(time-lapse imaging),利用軟體依Stokes-Einstein方程式分析螢光小球微觀流變學(microrheology),顯示visfatin處理後會降低人類軟骨細胞的黏彈力,後續由螢光染色共軛焦影像發現visfatin造成軟骨細胞骨架蛋白(microtubule and microfiliment)的表現下降,且其分子機轉為透過P38訊息路徑,經磷酸化而抑制了Glycogen Synthase Kinase 3β(GSK3β),造成細胞骨架蛋白的降解以及發炎因子COX-2的表現增加,這些發現提出抑制GSK3β或許可作為OA治療的方向。
3 Lin Martin Hsiu-Chu (林修竹醫師), Lai Ping-Shan, Chang Li-Ching, Huang Wei-Chao, Lee Ming-Hsueh, Chen Kuo-Tai, Chung Chiu-Yen*(鐘秋燕博士) and Yang Jen-Tsung*(楊仁宗醫師) (2021). "Characterization and Optimization of Chitosan-Coated Polybutylcyanoacrylate Nanoparticles for the Transfection-Guided Neural Differentiation of Mouse Induced Pluripotent Stem Cells." International Journal of Molecular Sciences 22(16): 8741.
以高分子量殼聚醣包覆聚山梨醇酯80聚氰基丙烯酸丁酯奈米顆粒(HMWC-PS80 PBCA NP)作為基因轉染載體用於小鼠誘導性多能幹細胞(iPSC)的轉染引導神經分化後,利用流式細胞分析儀檢測神經元特異性β III微管蛋白(beta III tubulin)的表達水平。
共軛焦顯微鏡拍攝小鼠誘導多能幹細胞(mouse induced pluripotent stem cells, miPSC)攝入奈米粒子的情形,評估Chitosan表面修飾對於提升PS80 PBCA奈米粒子被miPSC攝入的效果,此奈米粒子攜帶BDNF(brain-derived neurotrophic factor)基因的質體DNA,進入miPSC後可使其表現BDNF蛋白,並誘導miPSC分化為神經細胞。利用共軛焦顯微鏡拍攝miPSC免疫螢光染色結果,評估miPSC攝入奈米粒子後,表現BDNF、TrkB及神經細胞分化標記NF-H、Nestin蛋白,結果顯示Chitosan表面修飾可提升PS80 PBCA奈米粒子對於miPSC的轉染效率。
4 Huang Yen-Lin, Chen Po-Ru, Lai Ying-Ju*(賴盈如老師) and Hsu Hsao-Hsun* (2021). "Sirolimus Suppresses Phosphorylation of Cofilin and Reduces Interstitial Septal Thickness in Sporadic Lymphangioleiomyomatosis." International Journal of Molecular Sciences 22(16): 8564.
共軛焦顯微鏡搭配免疫螢光染色法觀察偶發性淋巴管平滑肌增生症(Sporadic lymphangioleiomyomatosis, S-LAM)病人肺部組織切片中HMB-45、Desmin及p-Cofilin蛋白的表現,探討Sirolimus此藥物對於治療S-LAM的作用機制。
5 Tsai Ming-Shao (蔡明劭醫師), Yang Yao-Hsu, Lin Yu-Shih, Chang Geng-He, Hsu Cheng-Ming, Yeh Reming-Albert, Shu Li-Hsin, Cheng Yu-Ching, Liu Hung-Te, Wu Yu-Huei, Wu Yu-Heng, Shen Rou-Chen and Wu Ching-Yuan*(吳清源醫師) (2021). "GB-2 blocking the interaction between ACE2 and wild type and mutation of spike protein of SARS-CoV-2." Biomedicine & Pharmacotherapy 142: 112011.
人胚胎腎細胞(HEK 293T)轉染pCEP4-MYC-ACE2表達質體後以不同濃度的抗SARS-CoV-2預防配方GB-2進行預先處理,然後加入來自293T細胞轉染pcDNA3-SARS-CoV-2-S-RBD-sfGFP表達質體後收集的RBD-sfGFP條件培養基及MYC抗體進行培養後,利用流式細胞分析儀檢測MYC及GFP表現,以評估GB-2對人體細胞表面的血管收縮素轉化酵素2(ACE2)及SARS-CoV-2表面刺蛋白(Spike Protein)的受體結合區域(RBD)之間結合的影響。
6 Cheng Kung-Chuan, Kuo Hsing-Chun, Hsieh Meng-Chiao, Huang Cheng-Yi, Teng Chih-Chuan, Tung Shui-Yi, Shen Chien-Heng, Lee Kam-Fai, Yang Ya-Ling and Lee Ko-Chao* (2021). "Identification of Two Novel CIL-102 Upregulations of ERP29 and FUMH to Inhibit the Migration and Invasiveness of Colorectal Cancer Cells by Using the Proteomic Approach." Biomolecules 11(9): 1280.
流式細胞分析儀檢測抗菌藥物CIL-102對大腸癌細胞(DLD-1)的影響,包含分析細胞凋亡Annexin V表現、活性氧ROS濃度(H2DCFDA標定)。
7 Chen Miao-Fen*(陳妙芬醫師), Hsieh Ching-Chuan, Chen Ping-Tsung and Lu Ming-Shian (2021). "Role of Nutritional Status in the Treatment Outcome for Esophageal Squamous Cell Carcinoma." Nutrients 13(9): 2997.
小鼠以氧化型致突變劑4-nitroquinoline-1-oxide(4NQO)誘導變成食道癌模型,治療過程中分成對照組(標準飲食)及營養組(額外餵食營養補充劑)進行飼養,再利用流式細胞分析儀檢測小鼠脾臟中骨髓源性抑制細胞MDSC(CD11b+Gr1+)的比例。
8 Hsu Cheng-Ming (徐正明醫師), Yang Ming-Yu, Tsai Ming-Shao, Chang Geng-He, Yang Yao-Hsu, Tsai Yao-Te, Wu Ching-Yuan*(吳清源醫師) and Chang Shun-Fu*(張順福博士) (2021). "Dihydroisotanshinone I as a Treatment Option for Head and Neck Squamous Cell Carcinomas." International Journal of Molecular Sciences 22(16): 8881.
流式細胞分析儀檢測中藥丹蔘萃取物二氫異丹蔘酮 I (DT)對人類頭頸部鱗狀細胞癌細胞(Detroit 562、SCC-4、SCC-25)的影響,以Annexin V分析細胞凋亡表現。
9 Peng Kuo-Ti*(彭國狄醫師), Chen Jiun-Liang, Kuo Liang-Tseng, Yu Pei-An, Hsu Wei-Hsiu, Lee Chiang-Wen, Chang Pey-Jium and Huang Tsung-Yu*(黃琮輿醫師) (2021). "GMI, an Immunomodulatory Peptide from Ganoderma microsporum, Restrains Periprosthetic Joint Infections via Modulating the Functions of Myeloid-Derived Suppressor Cells and Effector T Cells." International Journal of Molecular Sciences 22(13): 6854.
收集小鼠股骨中的骨髓細胞(bone marrow cell),分別處理金黃色葡萄球菌生物膜(S. aureus biofilms)或不同濃度的小孢子靈芝免疫調節蛋白質(Ganoderma microsporum immunomodulatory protein, GMI)培養後,再利用流式細胞分選儀篩選出骨髓源性抑制細胞MDSC(CD11b+Gr1+)進行實驗。利用流式細胞分析儀檢測細胞培養基或小鼠血清中IL-6、TNF-α等發炎細胞激素蛋白濃度(搭配Cytokine Bead Array, CBA試劑組),以及檢測處理GMI對MDSC釋放活性氧ROS濃度(DCFDA標定)及抑制T細胞增殖proliferation(CFSE標定)的影響。
10 Chen Cing-Yu, Wu Cheng-Hsun, Wu King-Chuen, Shiao Lian-Ru, Chuang Chin-Min, Leung Yuk-Man* and Chow Louis Wc (2021). "A basal level of γ-linolenic acid depletes Ca2+ stores and induces endoplasmic reticulum and oxidative stresses to cause death of breast cancer BT-474 cells." Chinese Journal of Physiology 64(4): 202.
流式細胞分析儀檢測γ-亞麻酸(GLA)對乳腺癌細胞(BT-474)的影響,包含分析細胞凋亡Annexin V表現、活性氧ROS濃度(DCFH2-DA標定)、粒線體Ca2+濃度(Rhod 2標定)。
11 Chen Yu-Chen, Sung Hsin-Ching*, Chuang Tzu-Yi*, Lai Tsai-Chun, Lee Tzu-Lin, Lee Chiang-Wen, Lee I-Ta and Chen Yuh-Lien* (2021). "Vitamin D3 decreases TNF-α-induced inflammation in lung epithelial cells through a reduction in mitochondrial fission and mitophagy." Cell Biology and Toxicology: 1-24.
經Vitamin D3預先處理的肺癌細胞(A549)再利用腫瘤壞死因子α(TNF-α)誘導其炎症,以流式細胞分析儀檢測Vitamin D3對炎症的影響,包含分析粒線體活性氧ROS濃度(MitoSOX Red標定)、粒線體膜電位變化(JC-1標定)。
12 Chuang Min-Chun(莊閔鈞醫師), Lung Jrhau, Chen Yi-Chuan, Lin Yu-Ching, Li Ya-Chin and Hung Ming-Szu*(洪明賜醫師) (2021). "The Association of Annexin A1 and Chemosensitivity to Osimertinib in Lung Cancer Cells." Cancers 13(16): 4106.
肺癌細胞(H1975、H1650)經基因轉染對照載體(Ctrl)或膜聯蛋白A1(ANXA1)沉默載體(SiANXA1)後,以流式細胞分析儀檢測有無沉默ANXA1對奧希替尼(Osimertinib)化療藥物治療肺癌之細胞凋亡的影響(分析Annexin V表現)。
13 Yu Chih-Chia, Chan Michael W.Y., Lin Hon-Yi, Chiou Wen-Yen, Lin Ru-Inn, Chen Chien-An, Lee Moon-Sing, Chi Chen-Lin, Chen Liang-Cheng, Huang Li-Wen, Chew Chia-Hui, Hsu Feng-Chun, Yang Hsuan-Ju and Hung Shih-Kai* (2021). "IRAK2, an IL1R/TLR Immune Mediator, Enhances Radiosensitivity via Modulating Caspase 8/3-Mediated Apoptosis in Oral Squamous Cell Carcinoma." Frontiers in Oncology 11(2361).
口腔癌細胞原始株OML1及抗輻射株OML1-R經基因轉染介白素1受體相關激酶2 (IRAK2)後,以流式細胞分析儀檢測放射線治療(RT)的電離輻射(IR)對過度表達IRAK2的口腔癌細胞凋亡的影響(分析Annexin V表現)。




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