先-腦內啡-黑色素激素-腎上腺皮質素激素(POMC)是許多神經胜肽的先驅物，其中包括adrenocorticotropin(ACTH)，而ACTH有調節腎上腺皮質功能的作用。本實驗將在培養細胞以及動物模型，研究傳送POMC基因對腎上腺皮質細胞所產生的影響。實驗材料包括人類腎上腺皮質H295R細胞，比較POMC基因刺激cortisol分泌以及細胞增生的作用。此外也將32隻SD rats分成三組，由尾部靜脈分別注射下列三種物質作比較：1) 注射生理食鹽水 (n=8)；2) 攜帶GFP的腺病毒(Ad-GFP) (n=12)；3) 攜帶POMC的腺病毒(Ad-POMC) (n=12)。注射後定期秤老鼠體重，且取出腎上腺秤重量比較。並將腎上腺檢體切片，用抗增生細胞核抗原及抗MC2-R抗原染色。以及 測量老鼠血清cortisol及VEGF的濃度。結果發現在人類腎上腺皮質H295R細胞，經攜帶POMC基因之腺病毒傳送後ACTH上升50至100倍，而cortisol僅上升20至100%。此外，在H295R細胞，POMC重組腺病毒明顯抑制細胞的增生，而更高比例的apoptotic (pre-G0)細胞也因POMC基因傳送而產生。此外，MC2-R蛋白的表現也因POMC基因傳送而減少。於前2週，傳送Ad-POMC的大鼠所增加的體重，少於控制組和傳送Ad-GFP的大鼠，但是整個實驗過程，傳送Ad-POMC和傳送Ad-GFP這兩組體重並無明顯差異。傳送Ad-POMC的大鼠，腎上腺的重量於前4週並無明顯變化，但第6週時顯著增加而大於第2週和第4週。至第8週時，Ad-POMC大鼠的腎上腺重量則高於Ad-GFP的大鼠。傳送Ad-POMC的大鼠，第8週的VEGF濃度也明顯高於Ad-GFP的大鼠。傳送Ad-POMC的大鼠，於第八週的腎上腺皮質切片，表現出多數的抗增生細胞核抗原染色細胞，而傳送Ad-GFP的大鼠腎上腺皮質切片，則未見明顯的染色細胞，顯示腎上腺皮質細胞受POMC的刺激而增生。但是傳送Ad-POMC的大鼠，於第八週的腎上腺皮質切片，所表現出的抗MC2-R抗原染色細胞則明顯少於傳送Ad-GFP的大鼠。Ad-POMC組的大鼠，血清的cortisol濃度大於Ad-GFP組，但是卻沒有達到統計上的意義。由以上實驗顯示POMC基因傳送可以調控腎上腺皮質的型態及功能。POMC基因經由造成MC2-R down-regulation以及產生apoptosis而抑制H295R細胞增生。但POMC基因傳送對大鼠的腎上腺，則產生雙相式的刺激作用，後期的快速生長則和VEGF有關。而生理調節cortisol的合成遠比ACTH的產生更為嚴格。

Pro-opiomelanocortin (POMC) is the precursor of many neuropeptides which includ adrenocorticotropin (ACTH). ACTH has a biological activity in regulating adrenocortical function. In the present study, we will investigate the effect of POMC gene transfer on adrenal cortex cells in cell cultures and animal models. The study included adrenal cortical H295R cells for adenovirus-mediated gene delivery. The effects of POMC gene on H295R cell steroidogenesis and cell proliferation were investigated. In addition, there were 32 SD rats dividing into three groups. 1) Control, injected with normal saline via tail vein (n = 8); 2) Ad-GFP, injected with adenovirus containing GFP (n=12); 3) Ad-POMC, injected with adenovirus containing recombinant POMC gene (n=12). Body weight (BW) was measured. Adrenals were collected, fixed and a series of sections were cut for stains for PCNA and MC2-R. The plasma cortisol and VEGF levels of rats were measured. The results showed that Ad-POMC delivery significantly increased the ACTH and cortisol levels by 50-100 fold and 20-100% in H295R cells, respectively. In addition, Ad-POMC delivery significantly inhibited the cell proliferation and increased the apoptotic cells. The expression of MC2-R protein of H295R cells was also suppressed after Ad-POMC delivery. In the study of SD rats, the Ad-POMC-treated rats exhibited reduced weight gain compared with other groups in the first 2 weeks; however, there was no significant change in BW between Ad-POMC and Ad-GFP groups during the experimental period. The weight of adrenal in Ad-POMC-treated rats was significantly higher than Ad-GFP group in the 8th week. Comparing the sequential adrenal weights in Ad-POMC group, those in 6th week were significantly higher than in 2nd and 4th weeks. The plasma VEFG levels of Ad-POMC-treated rats were higher than Ad-GFP group in the 8th week. The adrenal sections showed that Ad-POMC treated rats had moreanti-PCNA stained cells than Ad-GFP treated rats in 8th week. However, less anti-MC2R stained cells were found in Ad-POMC treated rats in 8th week. Ad-POMC treated rats had higher plasma cortisol levels than those in Ad-GFP treated rats, however, there were no statistical significances. In conclusion, POMC gene transfer modulates the morphology and function of the adrenal cortex. POMC gene inhibits the H295R cells proliferation by inducing MC2-R down-regulation and cells apoptosis. In SD rat adrenal, however, it stimulates adrenal cortex in biphasic pattern. The rapid growing pattern noted in the later phase may be due to the effect of VEGF. Besides, the physical regulation of cortisol synthesis is much stricter than that of ACTH.

目錄 頁次
中文摘要 6
英文摘要 8
縮寫 10
前言 11
材料和方法 17
結果 21
討論 26
參考文獻 33

參考文獻
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