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系統識別號 U0026-0208201716185700
論文名稱(中文) 陰道鞭毛蟲與人類類中性白血球共培養下IL-1及IL-8的反應
論文名稱(英文) Production of IL-1 and IL-8 by neutrophil-like cell (HL-60) stimulated with Trichomonas vaginalis
校院名稱 成功大學
系所名稱(中) 微生物及免疫學研究所
系所名稱(英) Department of Microbiology & Immunology
學年度 105
學期 2
出版年 106
研究生(中文) 白明旭
研究生(英文) Sina Taheri Baghmisheh
學號 S46047012
學位類別 碩士
語文別 英文
論文頁數 57頁
口試委員 指導教授-辛致煒
口試委員-黃一修
口試委員-林威辰
口試委員-黃國洋
中文關鍵字 none 
英文關鍵字 Trichomonas vaginalis  dHL-60 cells  inflammation  cytokines 
學科別分類
中文摘要 陰道毛滴蟲是一種厭氧原蟲,是人類最常見的非病毒性傳播疾病之一。陰道炎的傳播途徑通常藉由性交,如口交或肛交感染個體。此外,陰道炎也會在出生時藉由母親傳染胎兒。世界衛生組織估計,2011年全球共有2.49億例感染病例,而在2015年,美國發病率達至300萬例。陰道炎約有70%的感染者沒有任何症狀,但感染陰道毛滴蟲的婦女可能會產生痛癢、生殖器灼熱、發紅或酸痛,排尿不適或過度不適。在本實驗中,我們在相同的培養條件下創建了嗜中性白血球(dHL-60)細胞和陰道毛滴蟲的模擬條件,我們的培養基(DMEM-YIS)支持兩種嗜中性白血球(dHL-60)細胞和陰道毛滴蟲。此外,在我們的合成培養基中,嗜中性白血球(dHL-60)細胞和陰道毛滴蟲進行反應,這意味著陰道毛滴蟲刺激嗜中性白血球(dHL-60)細胞在我們的合成培養基中產生細胞因子反應,並且在共同培養期間,陰道毛滴蟲產生SOD6。
在這項研究中,我們已經證明我們的合成培養基可以支持兩種細胞的存活,也可以使陰道毛滴蟲能刺激嗜中性白血球細胞產生細胞因子如IL-1和IL-8 。
英文摘要 Trichomonas vaginalis, an anaerobic protozoa, is one of the most common non-viral sexually transmitted diseases in human. The transmission way for T. vaginalis is typically transmitted through vaginal, oral, or anal sex with an infected individual. It can also be passed from a mother to her baby at birth. The WHO has estimated that 248 million cases of infection was acquired worldwide in 2011, and there are 3 million new cases in the US in 2015. About 70% of infected people do not have any signs or symptoms, but women with trichomoniasis may notice itching, burning, redness or soreness of the genitals, discomfort with urination, or excessive discharge. In this study we created the mimic condition for neutrophil-like (dHL-60) cells and T. vaginalis in a same culture condition, which our medium (DMEM-YIS) supported the two microorganism which are neutrophil (dHL-60) cells and T.vaginalis. Furthermore, in our synthetic medium neutrophil-like (dHL-60) cells and T. vaginalis went to a reaction, which means that T. vaginalis stimulated the neutrophil-like (dHL-60) cells to produce the cytokines responses in our synthetic medium, and also, T. vaginalis produced the SOD6 during the co-incubation.
In this study we have been showed that our synthetic medium which is DMEM-YIS could support the two microorganism for surviving, and also, T. vaginalis could stimulate the neutrophil-like cells to produce the cytokines such as IL-1 and IL-8.
論文目次 T.O.C.
Chinese abstract .................………………….…………………….…… I
English abstract ...............….……………….………………………….. II
Summary.............................................. III
Chapter I. Introduction................................ 1
Chapter II. Materials and Methods ................………… 7
2.1 Experimental Design........................... 7
2.2 Trichomonas vaginalis culture ……………………..………………….. 7
2.2.1 Tirichomonas vaginalis growth medium preparation ……. 8
2.2.2 Trichomonas vaginalis hawing ................ 8
2.2.3 Trichomonas vaginalis subculture................... 8
2.2.4 Trichomonas vaginalis preservation…............... 9
2.3 Trichomonas vaginalis growth curve and doubling time counting ……….. 9
2.4 Domestication of Trichomonas vaginalis .............. 10
2.5 HL-60 cell culture ………………………………………. 10
2.5.1 HL-60 cell growth medium preparation………………………………. 10
2.5.2 HL-60 cells thawing .............................. 10
2.5.3 HL-60 cells subculture ….................... 11
2.5.4 HL-60 cell preservation........................... 11
2.6 HL-60 cells growth curve………………… 12
2.7 HL-60 cells domestication.......................... 12
2.8 HL-60 cell differentiation culture…………………………………… 12
2.9 HL-60 cell differentiation analyzing by NBT reduction assay................ 12
2.10 Co-incubation of Trichomonas vaginalis with HL-60 cells…………. 13
2.11 PCR reaction………........................ 14
2.11.1 RNA purification.......................... 14
2.11.2 cDNA synthesis ............................ 15
2.11.3 PCR nucleic acid electrophoresis................ 15
2.12 Reverse Transcriptase PCR (RT-PCR) ............... 16
2.12.1 RT-PCR reaction steps ...................... 16
2.12.2 RT-PCR reaction conditions .................... 16
2.12.3 PCR nucleic acid product electrophoresis ........ 17
Chapter III. Results ........................... 18
3.1 Domestication of Trichomonas vaginalis …………………… 18
3.2 Domestication of HL-60 cells ………………………………….. 18
3.3 HL-60 cells differentiation …………………………………………. 19
3.4 Establishing of co-incubation system .............. 19
3.5 Production of SOD6 by Trichomonas vaginalis ………………….. 20
3.6 The effect of Trichomonas vaginalis on HL-60 cells to produce the cytokines response...................... 20
Chapter IV. Conclusion …………………………………. 22
Chapter V. Discussion ……………………............... 23
Chapter VI. Future’s works …………………………………… 29
REFERENCES ..................................... 31
Appendix 1: INSTRUMENTS AND MATERIALS .................. 53
Appendix 2: Oligo GEArray Human Common Cytokines Microarray... 56
Appendix 3: Oxygen scavenging system of Trichomonas vaginalis..................... 57
Table 1. The list of primitives used in this paper……… 41
Figure directory
Figure 1.Growth curve of HL-60……………………….. 42
Figure 2. Process of HL-60 cells domestication…………...... 43
Figure 3. Growth curve HL-60 cells after domestication ………….. 44
Figure 4. Growth curve of T. vaginalis………………... 45
Figure 5. Process of T. vaginalis domestication........ 46
Figure 6. Growth curve of T. vaginalis after domestication......... 47
Figure 7. process of HL-60 cells differentiation to neutrophil-like (dHL-60) cells …………. 48
Figure 8. Differentiation curve of HL-60 cells …........ 49
Figure 9. HL-60 cells before and after differentiation…. 50
Figure 10. Production of SOD6 by Trichomonas vaginalis. 51
Figure 11. Cytokines production...................... 52
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