本文選題：局麻藥　切入點(diǎn)：布比卡因　出處：《南方醫(yī)科大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：局部麻醉藥(簡(jiǎn)稱局麻藥)根據(jù)其化學(xué)結(jié)構(gòu),分為酰胺類和酯類,常用于臨床麻醉。然而,大量研究表明,在多種類型細(xì)胞實(shí)驗(yàn)中,長(zhǎng)時(shí)間或高濃度使用局麻藥,會(huì)產(chǎn)生細(xì)胞毒性。在臨床應(yīng)用,也有關(guān)于局麻藥毒性作用的報(bào)道。盡管局麻藥臨床濃度所致毒性發(fā)生率很低,但一旦發(fā)生,會(huì)產(chǎn)生嚴(yán)重并發(fā)癥。尤其是脊髓麻醉后產(chǎn)生的短暫神經(jīng)綜合征、持續(xù)性腰骶神經(jīng)病變,以及最嚴(yán)重并發(fā)癥馬尾綜合征,多年來(lái)引起廣泛關(guān)注。對(duì)這些并發(fā)癥的發(fā)生,局麻藥的神經(jīng)毒性被認(rèn)為是主要因素。然而,局麻藥神經(jīng)毒性的確切機(jī)制,目前仍未可知。既往已有關(guān)于多種局麻藥神經(jīng)毒性大小的研究,但是對(duì)兩種類型局麻藥(酰胺類-布比卡因,酯類-普魯卡因)所致神經(jīng)毒性機(jī)制是否具有差異的全面性探索,目前國(guó)內(nèi)外仍未見(jiàn)報(bào)道�；诖�,我們?cè)O(shè)計(jì)本課題。目的1.探索布比卡因和普魯卡因?qū)H-SY5Y細(xì)胞活力的影響。2.比較布比卡因和普魯卡因?qū)H-SY5Y細(xì)胞和DRG神經(jīng)元細(xì)胞產(chǎn)生神經(jīng)毒性機(jī)制的異同。方法1.分別用不同濃度布比卡因和普魯卡因處理SH-SY5Y細(xì)胞,隨后使用CCK-8實(shí)驗(yàn)和LDH實(shí)驗(yàn)分別檢測(cè)細(xì)胞活力和細(xì)胞毒性,用曲線擬合計(jì)算兩種藥物L(fēng)D50值,選取該值作為后續(xù)細(xì)胞模型的藥物濃度。2.分別使用布比卡因和普魯卡因LD50值濃度處理SH-SY5Y細(xì)胞。隨后,使用Rhod-2-AM探針檢測(cè)線粒體內(nèi)鈣離子含量,JC-1試劑盒檢測(cè)線粒體膜電位變化,兩者共同反映細(xì)胞內(nèi)線粒體功能;使用DCFH-DA探針檢測(cè)細(xì)胞內(nèi)過(guò)氧化物含量,DHE探針檢測(cè)細(xì)胞內(nèi)超氧化物含量,兩者共同反映細(xì)胞內(nèi)氧化應(yīng)激水平;使用彗星實(shí)驗(yàn)檢測(cè)DNA損傷,Western Blot實(shí)驗(yàn)檢測(cè)DNA損傷標(biāo)志蛋白p-γ-H2AX含量,兩者共同反映DNA損傷;使用TUNEL實(shí)驗(yàn)檢測(cè)細(xì)胞凋亡,Western Blot 實(shí)驗(yàn)檢測(cè)凋亡相關(guān)蛋白 cleaved caspase3、cleaved caspase9 含量,兩者共同反映細(xì)胞凋亡情況。上述檢測(cè)也在DRG神經(jīng)元上進(jìn)行驗(yàn)證。結(jié)果1.布比卡因和普魯卡因均明顯降低SH-SY5Y細(xì)胞活力,并呈劑量依賴性。2.布比卡因和普魯卡因均顯著誘導(dǎo)細(xì)胞線粒體內(nèi)鈣超載、線粒體膜電位下降、氧化應(yīng)激爆發(fā)、DNA損傷以及凋亡產(chǎn)生(P0.05)。在細(xì)胞線粒體損傷和凋亡水平上,兩藥物組間無(wú)顯著差異(P0.05)。但布比卡因比普魯卡因誘導(dǎo)細(xì)胞產(chǎn)生更多超氧化物,早期DNA損傷程度更嚴(yán)重,而普魯卡因比布比卡因誘導(dǎo)細(xì)胞產(chǎn)生更多過(guò)氧化物(P0.05)。結(jié)論布比卡因和普魯卡因產(chǎn)生神經(jīng)毒性的機(jī)制可能涉及細(xì)胞線粒體功能失調(diào)、氧化應(yīng)激爆發(fā)、DNA損傷以及凋亡。兩種藥物誘導(dǎo)產(chǎn)生超氧化物和過(guò)氧化物含量不同,提示我們不同類型局麻藥產(chǎn)生神經(jīng)毒性機(jī)制可能通過(guò)不同途徑。我們可基于各自不同的毒性機(jī)制,實(shí)現(xiàn)個(gè)體化和靶向性治療。
[Abstract]:According to its chemical structure, local anesthetics are divided into amides and esters, which are often used in clinical anaesthesia. However, a large number of studies have shown that in many types of cell experiments, local anesthetics are used for a long time or at a high concentration. In clinical use, there are also reports of the toxic effects of local anesthetics. Although the incidence of toxicity caused by the clinical concentration of local anesthetics is very low, once that happens, Severe complications, especially transient nerve syndrome following spinal anesthesia, persistent lumbosacral neuropathy, and cauda equina syndrome, the most serious complications, have caused widespread concern over the years. The neurotoxicity of local anesthetic drugs is considered to be the main factor. However, the exact mechanism of neurotoxicity of local anesthetic drugs is still unknown. However, whether there are differences in neurotoxic mechanisms between two types of local anesthetic drugs (amide-bupivacaine, ester-procaine) has not been reported at home and abroad. Objective 1. To explore the effects of bupivacaine and procaine on the viability of SH-SY5Y cells. 2. To compare the neurotoxic mechanisms of bupivacaine and procaine on SH-SY5Y cells and DRG neurons. Do not treat SH-SY5Y cells with different concentrations of bupivacaine and procaine. CCK-8 test and LDH test were used to detect cell viability and cytotoxicity respectively. The LD50 values of the two drugs were calculated by curve fitting. The concentration of Bupivacaine and procaine LD50 were used to treat SH-SY5Y cells respectively. Then, the intracellular calcium content in mitochondria was detected by Rhod-2-AM probe and the change of mitochondrial membrane potential was detected by JC-1 kit. The DCFH-DA probe was used to detect the content of superoxide in the cells and the level of oxidative stress in the cells was reflected by the two probes. Comet assay was used to detect DNA damage and Western Blot assay was used to detect DNA damage marker protein p- 緯 -H2AX, both of which reflected DNA damage, TUNEL assay was used to detect apoptosis and cleaved caspase3cleaved caspase9 was detected by TUNEL assay. The results showed that: 1. Bupivacaine and procaine significantly decreased the viability of SH-SY5Y cells. In a dose-dependent manner, bupivacaine and procaine could significantly induce intracellular calcium overload, decrease of mitochondrial membrane potential, oxidative stress outbreak of DNA damage and apoptosis. There was no significant difference between the two groups (P 0.05), but bupivacaine induced more superoxide in cells than procaine, and the degree of DNA damage was more serious in the early stage of Bupivacaine than in procaine. Compared with bupivacaine, procaine can induce more peroxides (P0.05). Conclusion the mechanism of neurotoxicity of bupivacaine and procaine may be related to the dysfunction of cell mitochondria. Oxidative stress breaks out DNA damage and apoptosis. The two drugs induce the production of superoxide and peroxide. It is suggested that the neurotoxic mechanisms of different types of local anesthetic drugs may be produced by different ways, and we can realize individualized and targeted therapy based on different toxic mechanisms.
【學(xué)位授予單位】：南方醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R614

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 李樂(lè);盧愛(ài)珠;周樹勤;徐世元;;局麻藥神經(jīng)毒性影響因素及作用機(jī)制研究進(jìn)展[J];實(shí)用藥物與臨床;2012年04期

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本文編號(hào)：1606352