發(fā)布時(shí)間：2018-05-21 14:58

  本文選題：鹽酸去氧腎上腺素 + 絲裂霉素��； 參考：《東南大學(xué)》2005年碩士論文

【摘要】： 【目的】觀察鹽酸去氧腎上腺素和絲裂霉素(MMC)對(duì)體外培養(yǎng)家兔角膜內(nèi)皮的作用,探討臨床應(yīng)用這兩種藥物的可能毒性作用。 【方法】顯微分離兔角膜后彈力層-內(nèi)皮細(xì)胞層,聯(lián)合貼壁法和(或)消化法培養(yǎng)角膜內(nèi)皮細(xì)胞,利用含小牛血清的高糖DMEM作為培養(yǎng)液,不添加生長(zhǎng)因子。培養(yǎng)的角膜內(nèi)皮細(xì)胞,傳代接種于96孔板,鹽酸去氧腎上腺素設(shè)定為4個(gè)實(shí)驗(yàn)組0.625mg/ml、1.25mg/ml、2.5mg/ml、5mg/ml,4個(gè)相應(yīng)陰性對(duì)照組;MMC分為6個(gè)實(shí)驗(yàn)組1×10~(-6)mg/ml、1×10~(-5)mg/ml、1×10~(-4)mg/ml、1×10~(-3)mg/ml、1×10~(-2)mg/ml、1×10~(-1)mg/ml,均設(shè)空白對(duì)照2組。不同時(shí)相相差倒置顯微鏡下觀察細(xì)胞的形態(tài)變化和增殖情況,去氧腎上腺素組于加藥72h后,MMC組于加藥22h后MTT法檢測(cè)各組細(xì)胞增殖情況,對(duì)比不同濃度組細(xì)胞吸光度并進(jìn)行統(tǒng)計(jì)學(xué)處理。去氧腎上腺素組再次設(shè)定4個(gè)實(shí)驗(yàn)組0.625mg/ml、1.25mg/ml、2.5mg/ml、5mg/ml,在加藥后5min、10min、30min、1h、1h30min更換實(shí)驗(yàn)組的培養(yǎng)液,進(jìn)一步觀察細(xì)胞的空泡變性能否逆轉(zhuǎn)。 【結(jié)果】培養(yǎng)的兔角膜內(nèi)皮細(xì)胞呈單層密集生長(zhǎng),形態(tài)為六角形或多邊形,具有活體細(xì)胞的形態(tài)特征。去氧腎上腺素5mg/ml實(shí)驗(yàn)組加藥4h后細(xì)胞出現(xiàn)空泡,2.5mg/ml組加藥5h后細(xì)胞出現(xiàn)空泡,1.25mg/ml組加藥后10h出現(xiàn)明顯空泡,加藥12h后3個(gè)實(shí)驗(yàn)組均顯示明顯空泡,30h后2.5mg/ml、5mg/ml實(shí)驗(yàn)組細(xì)胞部分死亡,60h后1.25mg/ml、2.5mg/ml實(shí)驗(yàn)組原空泡變細(xì)胞大部分死亡,72h后3個(gè)濃度組幾乎全部死亡。0.625mg/ml濃度組和4個(gè)陰性對(duì)照組均未出現(xiàn)形態(tài)學(xué)變化。1.25mg/ml、2.5mg/ml、5mg/ml實(shí)驗(yàn)組細(xì)胞吸光度值與空白及陰性對(duì)照的差異經(jīng)統(tǒng)計(jì)學(xué)分析有極顯著意義(P0.001);0.625mg/ml試驗(yàn)組、4個(gè)陰性對(duì)照組與空白對(duì)照組相比較,差異無(wú)統(tǒng)計(jì)學(xué)意義(P0.05)。去氧腎上腺素接觸內(nèi)皮細(xì)胞后,5mg/ml、2.5mg/ml、1.25mg/ml濃度組5min以內(nèi)換液,細(xì)胞均未出現(xiàn)空泡變,加藥后6h再次換液,已出現(xiàn)的細(xì)胞空泡變性減輕,所有換液復(fù)孔細(xì)胞均未死亡。MMC組:加藥后1h、2h、3h、5h各組細(xì)胞形態(tài)變化不明顯,1×10~(-2)mg/ml、1×10~(-1)mg/ml實(shí)驗(yàn)組8h后細(xì)胞部分變圓,20h后細(xì)胞幾乎全部變圓死亡,其余各組變化不明顯。1×10~(-6)mg/ml、1×10~(-5)mg/ml組和空白對(duì)照組經(jīng)統(tǒng)計(jì)學(xué)分析差異無(wú)顯著性(P0.05),1×10~(-4)組和空白對(duì)照組比較差異有顯著性(P0.05),1×10~(-3)mg/ml、1×10~(-2)mg/ml、1×10~(-1)mg/ml組和空白對(duì)照組比較有極顯著差異(P0.001)。 【結(jié)論】鹽酸去氧腎上腺素和絲裂霉素對(duì)體外培養(yǎng)兔角膜內(nèi)皮細(xì)胞有損傷作用,并有時(shí)間和濃度相關(guān)性。盡早和及時(shí)的換液可避免或減輕鹽酸去氧腎上腺素對(duì)角膜內(nèi)皮細(xì)胞的毒性作
[Abstract]:[objective] to observe the effects of noradrenaline hydrochloride and mitomycin MMCon on rabbit corneal endothelium in vitro and to explore the possible toxic effects of these two drugs in clinical application. [methods] the rabbit corneal posterior elastic layer and endothelial cell layer were separated microscopically. Corneal endothelial cells were cultured with adherent method and / or digestion method. High glucose DMEM containing calf serum was used as culture medium without adding growth factor. Cultured corneal endothelial cells were cultured on 96 well plates. Noradrenaline hydrochloride was set at 0.625 mg / ml 1.25 mg / ml ~ (2.5 mg / ml). The four corresponding negative control groups were divided into 6 groups: 1 脳 10 ~ (-1) -6 mg / ml -1 脳 10 ~ (-5) mg / ml -1 脳 10 ~ (-4) mg / ml -1 脳 10 ~ (-3) mg / ml -1 脳 10 ~ (-1) mg / ml, respectively. The morphologic changes and proliferation of the cells were observed under different phase contrast microscope, and the proliferation of the cells in the noradrenaline group was detected by MTT assay after 22 hours of administration after 72 h of administration of noradrenaline. The cell absorbance of different concentration groups was compared and treated statistically. In the noradrenaline group, 0.625 mg / ml 1.25 mg / ml ~ (2.5 mg / ml) 2.5 mg / ml ~ (-1) mg 路ml ~ (-1) 路ml ~ (-1)? [results] the cultured rabbit corneal endothelial cells showed monolayer dense growth, with hexagonal or polygonal morphology, with the morphological characteristics of living cells. Vacuoles appeared in the cells of the 5mg/ml group treated with noradrenaline 2.5 mg / ml for 4 h, and the vacuoles appeared in the cells of the 1.25 mg / ml group 10 h after the addition of the drug for 5 h, and the vacuoles appeared in the cells in the group of 2.5 mg / ml of noradrenaline at 10 h after addition of the drug. After 12 h of administration, all the three experimental groups showed significant vacuolation 30 h, 2.5 mg / ml 5 mg / ml after partial cell death 60 h later, 1.25 mg / ml 2.5 mg / ml after most of the original vacuolar cells died 72 h later, almost all of the 3 groups died. 0.625 mg / ml and 4 negative control groups. There were no morphological changes. 1.25 mg / ml 2.5 mg / ml 5 mg / ml cell absorbance value of the experimental group was significantly different from the blank and negative control groups. There was significant difference between the four negative control groups and the blank control group, compared with the blank control group, the four negative control groups were compared with the blank control group. The difference was not statistically significant (P 0.05). After noradrenaline was exposed to endothelial cells, 5 mg / ml of 2.5 mg / ml ~ (-1) mg 路ml ~ (-1) mg / ml of 5min did not change the vacuolation of the cells, 6 h after adding the drug, the vacuolar denaturation of the cells was alleviated. There was no death in all the cells in the fluid exchange. MMC group: the morphological changes of the cells in each group were not obvious 1 脳 10 ~ (-2) mg / ml ~ (-1) 10~(-1)mg/ml after 8 h, and almost all the cells became round and died after 20 h. There was no significant difference between the other groups (1 脳 10 ~ (-6) mg 路ml ~ (-1) 10~(-5)mg/ml) and the blank control group (P ~ (0.05) 1 脳 10 ~ (-4) and the blank control group (P ~ (0.05) ~ (1 脳 10 ~ (-1) mg 路ml ~ (-1) 路ml ~ (-1) 脳 10 ~ (-1) mg 路ml ~ (-1) 路ml ~ (-1) 10~(-1)mg/ml and the control group (P ~ (0.001). [conclusion] noradrenaline hydrochloride and mitomycin can injure rabbit corneal endothelial cells in vitro and have time and concentration correlation. Early and timely fluid exchange can avoid or reduce the toxicity of noradrenaline hydrochloride to corneal endothelial cells.
【學(xué)位授予單位】：東南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2005
【分類號(hào)】：R329

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本文編號(hào)：1919693