多環芳香烴類的化合物是廣泛存在於環境中的污染物，包括工業製程所排放的氣
體、交通運輸排放廢氣、化石燃料的不完全燃燒、拜香和樟腦的使用等，都是環境中多
環芳香烴的污染來源，這些污染物可以經由皮膚接觸、呼吸道或腸胃道吸收進入人體，
由循環系統擴散至各組織，經由細胞色素的代謝活化，引起遺傳物質的突變，或藉由氧
化自由基的生成對細胞產生毒性。血管內皮細胞是位於血管內壁的單層細胞，不僅為血
管壁的屏障，更可以經由EDRF/NO 的釋放，維持血流的順暢，在之前的研究中發現，
短時間暴露低環數多環芳香烴會活化內皮型一氧化氮合成(endothelial nitric oxide
synthase)，合成並釋放NO，引發血管平滑肌的放鬆作用，然而內皮細胞長時間暴露在多
環芳香烴下的影響，卻不十分清楚，因此，本實驗採用人類臍靜脈內皮細胞(human
umbilical vein endothelial cell)來進行更深入的探討。
我們發現高濃度(>50mM)的naphthalene、fluoranthene、fluorene 等多環芳香烴會抑制
細胞生長週期的進展，使細胞週期停頓在G0/G1 時期，而在低濃度(0.1-50mM)時對細胞
生長週期的影響較輕微，也不會造成細胞死亡。此外，多環芳香烴會促進細胞內eNOS
mRNA 的表現，進而增加eNOS 的蛋白轉譯量，並增加eNOS 的活性和NO 的生合成量，
然而，這些多環芳香烴卻不會促進cyclooxygenase-1 的表現，顯示由多環芳香烴所調控
的基因表現具有專一性。在進一步探討多環芳香烴的作用機轉中， 我們發現
phosphatidylinositol 3-kinase 抑制劑wortmannin 和MAP kinase kinase 抑制劑PD98059，均無
法改變多環芳香烴對eNOS mRNA 的正向調節作用。反之當我們利用了鈣離子螯合劑
EGTA 和BAPTA-AM 分別螯合細胞內和外的鈣離子，結果發現在鈣離子被螯合的狀態
下，明顯的抑制由多環芳香烴所誘發的eNOS mRNA 表現，顯示鈣離子參與多環芳香烴
對eNOS mRNA 的調節。相較於低環數的多環芳香烴，高環數benzo(a)pyrene 卻無法促進
eNOS 的表現，因此aryl hydrocarbon receptor 可能不參與以上多環芳香烴對eNOS 基因表
現的影響。
綜合以上的結果，在我們的實驗中發現，長時間暴露多環芳香烴，會增加eNOS 的
蛋白表現，而其作用點主要是在eNOS mRNA 層次，而細胞內的鈣離子則扮演關鍵性的
角色。Polycyclic aromatic hydrocarbons (PAHs) are found in the exhausts of industrial manufacture,
and transportation due to the incomplete combustion of fuel, or the usage of camphor balls
and joss sticks which are common pollutants and wild spread in our environment. These
pollutants can be absorbed into body via respiratory tract, gastrointestinal tract or skin contact,
then distributed to the whole body through blood circulation. In biological system, PAHs have
been found to induce mutagenic or carcinogenic effect after cytochrome P-450 bioactivation.
In addition, PAHs also produced reactive oxygen species during metabolism and resulted in
cytotoxic effects. Endothelial cells localized inside the blood vessels, and function as a
vascular barrier between lumen and vascular smooth muscle. One of the important
pathological and physiological function of endothelial cell is synthesis and releasing of
endothelium-derived relaxating factor, nitric oxide, which can modulate vasotension and
maintain the homeostasis of vascular system. Preliminary studies revealed that endothelial
nitric oxide synthases were activated immediately after low-molecular weight PAHs exposure,
then converted L-arginine to nitric oxide resulted to vasodilation. However, the long term
effect of PAHs on endothelial cell is still unclear. In present research, the human umbilical
vein endothelial cell was used.
We found that several low-molecular weight PAHs such as naphthalene, fluoranthene,
fluorene inhibited cell growth cycle progression at higher concentrations(>50mM), resulting
in cell cycle arrest at G0/G1 phase. However, the effects on cell cycle were absent and no
cytotoxicity was observed in lower concentrations(0.1-50mM). In addition, PAHs will
enhance eNOS mRNA expression in HUVEC, increase eNOS protein content and activity,
and NO production. In contrast, the constitutively expressed cyclooxygenase-1 wasn’t
affected, suggesting that effect of PAHs on eNOS was specific. We also found that the
phosphatidylinositol 3-kinase inhibitor, wortmannin and MAP kinase kinase inhibitor,
PD98059 can not reverse the upregulation of eNOS mRNA levels by PAHs. However, when
the calcium chelators, EGTA and BAPTA-AM were used to chelated extracellular and
intracellular calcium, respectively the effect of PAHs on eNOS mRNA expression was
abolished, suggesting that calcium might be involved. In contrast to low-molecular weight
PAHs, benzo(a)pyrene, a high-molecular weight PAHs didn’t enhance eNOS expression,
implying that the aryl hydrocarbon receptor might not be involved in the previous findings.
In summary, we found that long term exposure of PAHs on HUVEC will upregulate eNOS
mRNA and protein level in a calcium-dependent manner
