本文選題：缺血再灌注損傷 + 皮瓣��； 參考：《第二軍醫(yī)大學(xué)》2017年博士論文

【摘要】：第一部分:脂氧素A4對大鼠帶蒂皮瓣缺血再灌注損傷保護作用研究目的:研究脂氧素A4對大鼠帶蒂皮瓣缺血再灌注損傷的保護作用。方法:建立大鼠腹壁上動脈帶蒂皮瓣缺血再灌注損傷模型,靜脈注射脂氧素A4,在術(shù)后72小時,通過大體形態(tài)計算皮瓣成活面積,激光多普勒血流成像檢測皮瓣血流灌注。HE染色觀察皮瓣組織學(xué)改變,ELISA檢測皮瓣中IL-1β,IL-6,TNF-α炎癥因子含量。同時檢測皮瓣中MDA含量及SOD活性。TUNEL染色觀察皮瓣細(xì)胞凋亡比例,通過western blotting檢測抗氧化應(yīng)激相關(guān)蛋白Nrf2、HO-1及凋亡相關(guān)蛋白Bax,Bcl2含量變化。通過免疫組織化學(xué)染色和western blotting檢測LC3蛋白含量。結(jié)果:LA4顯著增加缺血再灌注損傷皮瓣成活面積,增加皮瓣血流灌注量。LA4減輕缺血再灌注導(dǎo)致的組織損傷,降低缺血再灌注損傷皮瓣中MDA含量,升高SOD活性,同時抑制炎癥因子IL-1β,IL-6,TNF-α釋放。LA4抑制皮瓣缺血再灌注損傷所致細(xì)胞凋亡,同時降低Bax表達,促進Bcl2表達。LA4促進Nrf2向細(xì)胞核內(nèi)轉(zhuǎn)位并促進HO-1表達。LA4還促進缺血再灌注損傷皮瓣組織中LC3Ⅰ向LC3Ⅱ轉(zhuǎn)變,提高細(xì)胞自噬水平。結(jié)論:LA4通過抑制氧化應(yīng)激、炎癥反應(yīng)、凋亡,調(diào)節(jié)自噬水平保護皮瓣組織缺血再灌注損傷。第二部分:脂氧素A4對大鼠骨骼肌缺血再灌注損傷保護作用研究目的:研究LA4對大鼠骨骼肌缺血再灌注損傷的保護作用及相關(guān)機制。方法:建立大鼠下肢肌肉缺血再灌注損傷模型,通過靜脈注射不同劑量的LA4。再灌注后3小時,觀察下肢肌肉組織水腫,免疫熒光染色法標(biāo)記中性粒細(xì)胞浸潤,HE染色觀察肌肉組織損傷狀態(tài),ELISA檢測肌肉組織中炎癥因子TNF-α、IL-Iβ及IL-6含量。同時檢測抗氧化應(yīng)激相關(guān)酶SOD、CAT、GSH-Px活性,TUNEL染色觀察肌細(xì)胞凋亡比例。Western blotting檢測抗氧化應(yīng)激相關(guān)蛋白Nrf2、HO-1和凋亡相關(guān)蛋白Bax、Bcl2表達變化。透射電鏡觀察肌肉組織自噬體數(shù)量。免疫熒光染色和western blotting檢測自噬相關(guān)蛋白LC3和Beclin1含量變化。結(jié)果:大鼠肌肉缺血再灌注損傷導(dǎo)致肌肉水腫,中性粒細(xì)胞浸潤、肌肉組織學(xué)損傷,炎癥因子水平增高,抗氧化應(yīng)激相關(guān)酶活性下降,細(xì)胞凋亡比例增高。LA4能減輕肌肉組織水腫,減少中性粒細(xì)胞浸潤。LA4還能減輕肌肉組織學(xué)損傷程度。LA4降低缺血再灌注組織中炎癥因子含量,增加抗氧化應(yīng)激相關(guān)酶的活性。LA4還增加抗凋亡蛋白bcl2表達并降低促凋亡蛋白bax表達。la4對肌肉缺血再灌注損傷的保護作用與其激活nrf2/ho-1信號通路相關(guān),當(dāng)加入ho-1抑制劑(znppⅨ)時,la4對肌肉組織缺血再灌注損傷的保護作用明顯被抑制。la4還促進缺血再灌注損傷肌肉組織中自噬體的形成,促進lc3Ⅰ向lc3Ⅱ轉(zhuǎn)變,增加beclin1表達。結(jié)論:la4通過抑制炎癥反應(yīng)、氧化應(yīng)激、細(xì)胞凋亡、調(diào)節(jié)自噬保護肌肉組織缺血再灌注損傷,其保護作用與激活nrf2/ho-1信號通路相關(guān)。第三部分:脂氧素a4對骨骼肌細(xì)胞氧化應(yīng)激損傷的保護作用研究目的:通過體外培養(yǎng)的骨骼肌細(xì)胞氧化應(yīng)激損傷模型,觀察la4對骨骼肌細(xì)胞氧化應(yīng)激損傷的保護作用。方法:使用不同濃度的h2o2(0,25,50,100,200,400μmol/l)刺激大鼠骨骼肌成肌細(xì)胞(l6細(xì)胞),模擬骨骼肌缺血再灌注損傷,確定h2o2損傷骨骼肌細(xì)胞的最佳時間和濃度。在h2o2損傷細(xì)胞前,不同濃度(0,0.1,1,10,100nmol/l)la4預(yù)處理細(xì)胞12小時,確定la4對l6細(xì)胞氧化應(yīng)激損傷的最佳保護濃度。mtt實驗觀察細(xì)胞活性變化,檢測ros含量判斷細(xì)胞氧化應(yīng)激損傷程度。光鏡下形態(tài)學(xué)觀察細(xì)胞氧化應(yīng)激損傷所致細(xì)胞形態(tài)學(xué)改變,并檢測培養(yǎng)液中ck、ldh活性判斷細(xì)胞損傷嚴(yán)重程度。測定細(xì)胞內(nèi)gsh含量判斷細(xì)胞抗氧化應(yīng)激能力。tunel染色觀察骨骼肌細(xì)胞凋亡比例,westernblotting檢測凋亡相關(guān)蛋白bcl2,bax含量變化。結(jié)果:h2o2氧化應(yīng)激損傷骨骼肌細(xì)胞的最佳濃度為200μmol/l,最佳作用時間為4小時,可造成細(xì)胞活性下降至對照組55%。la4(10nmol/l)顯著增加h2o2損傷的骨骼肌細(xì)胞活力。la4還能夠抑制h2o2損傷所致ros的生成,抑制ck、ldh向培養(yǎng)液中釋放,促進細(xì)胞內(nèi)gsh的消耗。la4促進bcl2表達,抑制bax表達,降低h2o2損傷所致細(xì)胞凋亡比例。結(jié)論:la4在體外培養(yǎng)的骨骼肌細(xì)胞氧化應(yīng)激損傷中,顯著增加細(xì)胞活力、抑制ros生成、增加細(xì)胞抗氧化應(yīng)激能力,抑制氧化應(yīng)激損傷所致細(xì)胞凋亡,對骨骼肌氧化應(yīng)激損傷有保護作用。第四部分:脂氧素a4對骨骼肌細(xì)胞氧化應(yīng)激損傷保護機制研究目的:在骨骼肌細(xì)胞氧化應(yīng)激損傷模型中,研究la4保護作用的相關(guān)機制。方法:在體外骨骼肌細(xì)胞氧化應(yīng)激損傷模型中,通過免疫熒光染色,westernblotting檢測la4對nrf2/ho-1信號通路及自噬水平的影響。同時通過westernblotting檢測la4對erk、p38、jnk、pi3k/akt信號通路的作用,并研究erk、p38、jnk、pi3k/akt信號通路與nrf2/ho-1信號通路、自噬的關(guān)系。結(jié)果:LA4在體外骨骼肌細(xì)胞氧化應(yīng)激損傷時激活Nrf2/HO-1信號通路,促進Nrf2向細(xì)胞核內(nèi)轉(zhuǎn)移,同時增加HO-1蛋白的表達。LA4提高骨骼肌細(xì)胞氧化應(yīng)激損傷時自噬水平。LA4通過激活ERK通路而激活Nrf2/HO-1信號通路。LA4對P38和JNK、PI3K/AKT信號通路無明顯調(diào)節(jié)作用。LA4通過激活ERK/Nrf2信號通路而提高骨骼肌氧化應(yīng)激損傷時自噬水平,抑制ERK/Nrf2信號通路后,自噬水平降低。結(jié)論:LA4通過激活ERK/Nrf2信號通路提高氧化應(yīng)激損傷的骨骼肌細(xì)胞自噬水平,增強抗氧化應(yīng)激蛋白HO-1的表達,從而保護骨骼肌細(xì)胞氧化應(yīng)激損傷。
[Abstract]:The first part: the protective effect of lipoxin A4 on ischemic reperfusion injury in rats with pedicle flap: To study the protective effect of lipoxygenin A4 on ischemia reperfusion injury in rats with pedicle flap. Method: establish the model of ischemia reperfusion injury in the upper abdominal wall flap of the abdominal wall of rats, intravenous lipoxygenase A4, through the large body shape after 72 hours of operation. The survival area of the skin flap was calculated. The skin flap was stained by laser Doppler flow imaging to observe the histological changes of the skin flap. The content of IL-1 beta, IL-6, and TNF- alpha in the skin flap was detected by ELISA. The MDA content in the flap and the proportion of the apoptosis of the skin flap were detected by SOD activity.TUNEL, and the proportion of apoptosis in the skin flap was observed by SOD activity.TUNEL, and the antioxidant should be detected by Western blotting. Changes in the content of Nrf2, HO-1 and apoptosis related protein Bax and Bcl2. The content of LC3 protein was detected by immunohistochemical staining and Western blotting. Results: LA4 significantly increased the survival area of the skin flap with ischemia-reperfusion injury, increased the volume of blood flow perfusion of the flap to reduce the tissue damage caused by ischemia-reperfusion, and reduced the ischemia-reperfusion injury. The content of MDA in the wound skin flap, the increase of SOD activity, and the inhibition of the inflammatory factor IL-1 beta, IL-6, and TNF- alpha release.LA4 inhibit the cell apoptosis induced by the ischemia-reperfusion injury of the flap, and reduce the expression of Bax, promote the expression of.LA4 to promote the transposition of Nrf2 to the nucleus and promote HO-1 expression.LA4 to promote the ischemia-reperfusion injury in the skin flap. Change, improve the level of autophagy. Conclusion: LA4 protects the ischemic reperfusion injury of skin flap by inhibiting oxidative stress, inflammatory response, apoptosis and regulating autophagy level. The second part: the protective effect of lipoxygenin A4 on skeletal muscle ischemia reperfusion injury in rats: To study the protective effect of LA4 on skeletal muscle ischemia reperfusion injury in rats and the protective effect of lipoxygenin on the protection of ischemia reperfusion injury in rat skeletal muscle and Methods: to establish a rat model of lower limb muscle ischemia reperfusion injury, the edema of the muscle tissue of the lower extremities was observed 3 hours after intravenous injection of different doses of LA4.. Immunofluorescence staining was used to mark neutrophils infiltration, HE staining was used to observe the injury state of the muscle tissue. ELISA was used to detect the inflammatory factor TNF- a, IL-I beta in the muscle tissue. And IL-6 content. Detection of antioxidant stress related enzymes SOD, CAT, GSH-Px activity. TUNEL staining was used to observe the apoptosis ratio of muscle cells,.Western blotting was used to detect antioxidant stress related protein Nrf2, HO-1 and apoptosis related proteins Bax, Bcl2 expression. Transmission electron microscopy was used to observe the number of autophago in muscle tissue. Immunofluorescence staining and Western assay Changes in the content of autophagy related proteins LC3 and Beclin1. Results: muscle edema, neutrophil infiltration, muscle tissue injury, increased inflammatory factors, decreased activity of antioxidant stress related enzymes, and increased apoptosis ratio.LA4 could reduce muscle edema and decrease neutrophils infiltration.LA4 in rats. .LA4 can reduce the degree of muscle tissue injury, reduce the content of inflammatory factors in ischemia reperfusion tissue, increase the activity of antioxidant stress related enzymes, and increase the expression of anti apoptotic protein BCL2 and reduce the protective effect of.La4 on myocardial ischemia reperfusion injury, which is related to the activation of nrf2/ho-1 signaling pathway, when ho is added to ho. When -1 inhibitor (ZnPP IX), the protective effect of LA4 on ischemia-reperfusion injury of muscle tissue is obviously inhibited by.La4, which also promotes the formation of autophagic in the muscle tissue of ischemia reperfusion injury, promotes the transformation of LC3 I to LC3 II, and increases the expression of Beclin1. Conclusion: LA4 can regulate autophagy by inhibiting the inflammatory response, oxidative stress, and apoptosis. The protective effect of ischemic reperfusion injury is related to the activation of nrf2/ho-1 signaling pathway. Third part: the protective effect of lipoxygenin A4 on oxidative stress injury in skeletal muscle cells: the protective effect of LA4 on oxidative stress damage in skeletal muscle cells is observed by the oxidative stress damage model of skeletal muscle cells in vitro. The rat skeletal myoblasts (L6 cells) were stimulated with different concentrations of H2O2 (0,25,50100200400 mol/l) to simulate the injury of skeletal muscle ischemia-reperfusion injury, and the optimal time and concentration of H2O2 damaged skeletal muscle cells were determined. Before H2O2 damaged cells, 0,0.1,1,10100nmol/l LA4 pretreated cells for 12 hours to determine LA4 on the oxidation of L6 cells. The best protective concentration of stress injury.Mtt test observed cell activity changes, detect ROS content to judge the degree of oxidative stress injury. Morphological observation of cell morphology changes caused by oxidative stress injury under light microscope, and detection of CK, LDH activity to judge the severity of cell damage in the culture medium. The determination of GSH content in cells to determine cell resistance The apoptosis ratio of skeletal muscle cells was observed by oxidative stress.Tunel staining, and westernblotting was used to detect the changes of BCL2 and Bax content of apoptosis related proteins. Results: the optimum concentration of skeletal muscle cells by H2O2 oxidative stress was 200 mu mol/l and the best time of action was 4 hours, which could cause the cell viability to decrease to 55%.la4 (10nmol/l) in the control group. The damage of skeletal muscle cell vitality.La4 can also inhibit the formation of ROS caused by H2O2 damage, inhibit the release of CK, LDH into the culture medium, promote the consumption of GSH in the cell.La4 to promote BCL2 expression, inhibit Bax expression and reduce the percentage of apoptosis induced by H2O2 damage. Conclusion: LA4 in the oxidative stress injury of skeletal muscle cells cultured in vitro, significantly increased Cell vitality, inhibiting ROS formation, increasing cell antioxidant stress ability, inhibiting apoptosis induced by oxidative stress injury and protecting skeletal muscle oxidative stress damage. The fourth part: Study on the protective mechanism of lipoxygenin A4 on oxidative stress injury in skeletal muscle cells: the study of LA4 protection in the oxidative stress damage model of skeletal muscle cells Methods: in the model of oxidative stress injury in skeletal muscle cells in vitro, the effects of LA4 on nrf2/ho-1 signaling pathway and autophagy were detected by immunofluorescence staining, and the effects of LA4 on ERK, p38, JNK, pi3k/akt signaling pathway were detected by westernblotting, and ERK, p38, JNK, and signaling pathways were studied. The relationship with autophagy with nrf2/ho-1 signaling pathway. Results: LA4 activates Nrf2/HO-1 signaling pathway during oxidative stress injury in skeletal muscle cells in vitro, promotes the transfer of Nrf2 into the nucleus, and increases the expression of HO-1 protein expression.LA4 to increase the autophagy level in skeletal muscle cell oxidative stress,.LA4 activates the Nrf2/HO-1 signal through the activation of the ERK pathway. Road.LA4 has no obvious regulating effect on P38 and JNK, PI3K/AKT signaling pathway..LA4 increases autophagy level in oxidative stress injury of skeletal muscle by activating ERK/Nrf2 signaling pathway and reduces autophagy level after ERK/Nrf2 signaling pathway. Conclusion: LA4 enhances autophagy level in skeletal muscle cells damaged by oxidative stress by activating ERK/Nrf2 signaling pathway. The expression of strong antioxidant stress protein HO-1 can protect skeletal muscle cells from oxidative stress injury.
【學(xué)位授予單位】：第二軍醫(yī)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：R658

【相似文獻】

相關(guān)期刊論文 前10條

1 吳升華;脂氧素研究進展[J];國外醫(yī)學(xué).臨床生物化學(xué)與檢驗學(xué)分冊;2003年05期

2 周曉燕;張力;吳萍;熊維;李詠生;葉篤筠;;脂氧素對粒細(xì)胞集落刺激因子表達的影響及其機制初探[J];細(xì)胞生物學(xué)雜志;2007年03期

3 倪鐫;黃引平;;脂氧素在妊娠期高血壓疾病中的作用[J];國外醫(yī)學(xué)(婦產(chǎn)科學(xué)分冊);2007年05期

4 張力;萬敬員;;脂氧素受體及其介導(dǎo)的促炎癥消退信號[J];生命的化學(xué);2007年06期

5 代先坤;萬敬員;張力;龔霞;倪安紅;周岐新;;脂氧素對脂多糖誘導(dǎo)巨噬細(xì)胞炎癥相關(guān)因子的影響及機制[J];第四軍醫(yī)大學(xué)學(xué)報;2009年14期

6 馬利;黃中華;梁寧;;炎癥調(diào)控介質(zhì)-脂氧素的研究進展[J];中國臨床新醫(yī)學(xué);2010年01期

7 胡珊;毛應(yīng)啟梁;王彥青;;脂氧素在炎癥中作用的研究進展[J];國際藥學(xué)研究雜志;2011年02期

8 張彥亮;史會連;陶臻;;脂氧素在疾病中的作用[J];國際病理科學(xué)與臨床雜志;2011年04期

9 吳真;孫劍;;脂氧素與炎癥的消退[J];現(xiàn)代生物醫(yī)學(xué)進展;2012年31期

10 張力,萬敬員,葉篤筠;脂氧素與炎癥消退[J];生命的化學(xué);2004年01期

相關(guān)會議論文 前10條

1 易攀;龐花艷;黃引平;;脂氧素保護臍靜脈內(nèi)皮細(xì)胞氧化損傷的機制研究[A];中國病理生理學(xué)會第九屆全國代表大會及學(xué)術(shù)會議論文摘要[C];2010年

2 劉松君;黃引平;李詠生;周曉燕;蔡蕾;葉篤筠;;脂氧素對缺氧損傷臍靜脈內(nèi)皮細(xì)胞的保護及其機制研究[A];浙江省醫(yī)學(xué)會圍產(chǎn)醫(yī)學(xué)分會成立大會暨“圍產(chǎn)醫(yī)學(xué)熱點問題”學(xué)術(shù)會議論文匯編[C];2008年

3 龔建明;黃引平;湯彪;陳金霞;周潔;黃艷君;;脂氧素對滋養(yǎng)細(xì)胞氧化損傷的保護作用及機制的研究[A];2011年浙江省婦產(chǎn)科學(xué)學(xué)術(shù)年會暨“婦產(chǎn)科常見疾病的臨床研究新進展”學(xué)習(xí)班論文匯編[C];2011年

4 周曉燕;王紅梅;李詠生;吳萍;蔡震宇;徐方云;葉篤筠;;脂氧素對HepG2肝癌細(xì)胞侵襲能力的影響及機制研究[A];2009醫(yī)學(xué)前沿論壇暨第十一屆全國腫瘤藥理與化療學(xué)術(shù)會議論文集[C];2009年

5 何文靜;彭思思;郝華;蔡蕾;;脂氧素受體激動劑BML-111在巨噬細(xì)胞表型轉(zhuǎn)變中的作用[A];中國病理生理學(xué)會第九屆全國代表大會及學(xué)術(shù)會議論文摘要[C];2010年

6 胡珊;張輝;李倩;王小微;毛應(yīng)啟梁;米文麗;吳根誠;王彥青;;脂氧素及阿司匹林誘導(dǎo)的脂氧素對脛骨癌痛大鼠機械性痛覺超敏的影響[A];第十屆全國針刺麻醉針刺鎮(zhèn)痛及針刺調(diào)整效應(yīng)學(xué)術(shù)研討會論文集[C];2010年

7 郝華;劉淼;吳萍;葉篤筠;;脂氧素A4通過重塑腫瘤微環(huán)境抑制前列腺癌[A];中國病理生理學(xué)會第九屆全國代表大會及學(xué)術(shù)會議論文摘要[C];2010年

8 艾鳳庭;高莉莉;關(guān)鳳軍;董晨;趙彤;辛顯芳;;脂氧素A4對幼年肥胖鼠T淋巴細(xì)胞免疫失衡的作用研究[A];2012年江浙滬兒科學(xué)術(shù)年會暨浙江省醫(yī)學(xué)會兒科學(xué)分會學(xué)術(shù)年會、兒內(nèi)科疾病診治新進展國家級學(xué)習(xí)班論文匯編[C];2012年

9 吳升華;張永梅;陶慧嫻;董玲;;脂氧素A4抑制CTGF誘導(dǎo)的人腎小管上皮細(xì)胞轉(zhuǎn)分化[A];中華醫(yī)學(xué)會第十五次全國兒科學(xué)術(shù)大會論文匯編（上冊）[C];2010年

10 艾鳳庭;高莉莉;關(guān)鳳軍;董晨;趙彤;辛顯芳;;脂氧素A4對幼年肥胖鼠T淋巴細(xì)胞免疫失衡的作用研究[A];中華醫(yī)學(xué)會第十七次全國兒科學(xué)術(shù)大會論文匯編（下冊）[C];2012年

相關(guān)博士學(xué)位論文 前8條

1 宗海洋;脂氧素A4對大鼠皮瓣和肌肉組織缺血再灌注損傷保護作用及機制研究[D];第二軍醫(yī)大學(xué);2017年

2 金勝威;促炎癥消退介質(zhì)脂氧素對內(nèi)毒素性肺損傷的保護作用及機制[D];華中科技大學(xué);2007年

3 陳志橋;脂氧素對心臟驟停后綜合征心功能不全的作用及機制研究[D];武漢大學(xué);2013年

4 吳樂;脂氧素A_4抑制5-脂氧酶和白三烯參與其抗腦缺血再灌注損傷的機制研究[D];華中科技大學(xué);2012年

5 熊晶;脂氧素A_4對小鼠胚胎著床的干預(yù)作用及機制研究[D];華中科技大學(xué);2013年

6 紀(jì)宇東;脂氧素受體激動劑BML-111抑制小鼠肺纖維化及相關(guān)機制研究[D];華中科技大學(xué);2013年

7 葉習(xí)紅;甲酯化脂氧素A_4對局灶性腦缺血再灌注損傷的保護作用及其機制研究[D];華中科技大學(xué);2010年

8 郝華;脂氧素A_4抗肝癌作用及機制研究[D];華中科技大學(xué);2011年

相關(guān)碩士學(xué)位論文 前10條

1 苗貴申;脂氧素A_4在椎間盤突出所致大鼠根性神經(jīng)痛中的作用及機制研究[D];山東大學(xué);2015年

2 朱旭利;脂氧素A_4及脂氧素A_4受體在慢性鼻—鼻竇炎上皮—間質(zhì)轉(zhuǎn)化中的作用[D];福建醫(yī)科大學(xué);2016年

3 代先坤;脂氧素對脂多糖誘導(dǎo)巨噬細(xì)胞炎癥相關(guān)因子的影響及機制的研究[D];重慶醫(yī)科大學(xué);2009年

4 柯有力;脂氧素對小鼠異位移植心存活的影響[D];華中科技大學(xué);2007年

5 易攀;脂氧素A_4調(diào)節(jié)人臍靜脈內(nèi)皮細(xì)胞通透性的機制初探[D];華中科技大學(xué);2010年

6 翟恒;15-差向異構(gòu)體—脂氧素A_4對脂多糖誘導(dǎo)小膠質(zhì)細(xì)胞內(nèi)活性氧產(chǎn)生的影響及其機制研究[D];華中科技大學(xué);2011年

7 易婷婷;脂氧素A_4對尿酸誘導(dǎo)的人臍靜脈內(nèi)皮細(xì)胞炎癥反應(yīng)的影響[D];川北醫(yī)學(xué)院;2014年

8 陳碩;脂氧素抑制子宮內(nèi)膜異位癥的分子機制[D];廈門大學(xué);2014年

9 黃利興;脂氧素對鼠源性巨噬細(xì)胞內(nèi)TLR4/NF-κB信號通路的影響及其作用機制[D];南昌大學(xué);2014年

10 唐艷榮;脂氧素A4對心肌缺血再灌注損傷的保護作用及其機制研究[D];南京醫(yī)科大學(xué);2012年

，

本文編號：2041625