人工合成抗菌肽Pac525對(duì)種植體周?chē)撞≡种谱饔玫难芯?/H1>
發(fā)布時(shí)間：2018-10-12 08:05
【摘要】：目的：本實(shí)驗(yàn)選用前期實(shí)驗(yàn)已證實(shí)對(duì)牙齦卟啉單胞菌、具核梭桿菌及血鏈球菌這三種種植體周?chē)撞≡胁煌潭纫种谱饔玫娜斯ず铣煽咕腜ac525，通過(guò)活菌計(jì)數(shù)繪制其對(duì)這三種菌的kill-time曲線，以期進(jìn)一步發(fā)現(xiàn)其在不同時(shí)間、不同藥物濃度下Pac525對(duì)這三種菌的抑制作用；建立活菌生物膜，掃描電鏡觀察其生長(zhǎng)狀況；采用結(jié)晶紫法測(cè)定Pac525對(duì)這三種菌完整生物膜形成的藥物最低抑制生物膜濃度(MBIC)和藥物最低清除已形成生物膜濃度(MBRC)。 方法： 一、抗菌肽Pac525對(duì)種植體周?chē)字虏【膋ill-time 依據(jù)Pac525的氨基酸序列，人工合成該抗菌肽。本實(shí)驗(yàn)采用活菌計(jì)數(shù)法研究Pac525對(duì)牙齦卟啉單胞菌、具核梭桿菌及血鏈球菌的kill-time。 二、純鈦表面牙齦卟啉單胞菌、具核梭桿菌及血鏈球菌的菌斑生物膜的建立。 為模擬口腔環(huán)境，用唾液對(duì)光滑鈦片預(yù)處理（37℃，24小時(shí)），使其表面形成獲得性膜。將鈦片放入24孔板中，PBS沖洗，每孔加入1500μl BHI液體培養(yǎng)基和500μl菌液，37℃恒溫培養(yǎng)24~48小時(shí)，脫水干燥，掃描電鏡觀察其生物膜。 三、抗菌肽Pac525對(duì)種植體周?chē)字虏【呱锬さ挠绊?MBIC的測(cè)定：在96孔板上分別建立三種菌的生物膜，加入200μl Pac525藥液培養(yǎng)基和20μl1×107cfu/ml菌液，37℃恒溫培養(yǎng)24~48小時(shí)，PBS沖洗，甲醛固定，結(jié)晶紫染色，無(wú)菌去離子水沖洗，乙醇復(fù)吸，酶標(biāo)儀測(cè)定吸光度。根據(jù)公式計(jì)算其MBIC。使用SPSS17.0統(tǒng)計(jì)軟件進(jìn)行統(tǒng)計(jì)學(xué)分析。 MBRC的測(cè)定：在96孔板上加入200μlBHI液體培養(yǎng)基和20μl1×107CFU/ml菌液37。C恒溫培養(yǎng)24~48小時(shí)，吸出上清和浮游菌，PBS沖洗，加入200μl Pac525藥液培養(yǎng)基24~48小時(shí)，吸出上清和浮游菌，PBS沖洗，甲醛固定，結(jié)晶紫染色，無(wú)菌去離子水沖洗，乙醇復(fù)吸，酶標(biāo)儀測(cè)定吸光度。根據(jù)公式計(jì)算其MBRC。使用SPSS17.0統(tǒng)計(jì)軟件進(jìn)行統(tǒng)計(jì)學(xué)分析。 結(jié)果： 一、抗菌肽Pac525對(duì)種植體周?chē)字虏【膋ill-time 通過(guò)kill-time曲線我們可以看出，隨著時(shí)間的改變，不同濃度的Pac525均顯示出了一定程度的抑菌及殺菌功效。對(duì)于血鏈球菌，0.5h不同濃度的Pac525即顯示出不同程度的抑菌作用，其中2MIC（minimum inhibitory concentration,最小抑菌濃度）和4MIC尤為明顯，1h時(shí)4MIC已殺死全部血鏈球菌,3.5h時(shí)MIC的殺菌作用趨于平穩(wěn)，1h2MIC殺菌作用趨于平穩(wěn)，均保持在較高水平。對(duì)于具核梭桿菌，4h時(shí)不同濃度的Pac525即顯示出不同程度的抑菌作用，8h時(shí)2MIC已殺死全部具核梭桿菌，此時(shí)MIC的殺菌作用也趨于穩(wěn)定。對(duì)于牙齦卟啉單胞菌，4h時(shí)2MIC已殺死全部牙齦卟啉單胞菌，8h時(shí)MIC也已殺死全部牙齦卟啉單胞菌。結(jié)果表明Pac525對(duì)菌的殺死作用隨藥物的濃度和時(shí)間呈正相關(guān)。通過(guò)kill-time曲線可以得出這種作用趨勢(shì)。 二、抗菌肽Pac525對(duì)種植體周?chē)字虏【呱锬ぷ饔媒Y(jié)果 經(jīng)單因素方差分析（P＜0.05）PAC525對(duì)血鏈球菌MBIC50為1mg/ml，為其MIC的8倍。PAC525對(duì)具核梭桿菌MBIC50為0.125mg/ml，為其MIC的16倍。PAC525對(duì)牙齦卟啉單胞菌MBIC500.5mg/ml，，為其MIC的8倍。但是MBRC并未測(cè)出來(lái)。 結(jié)論： 1.抗菌肽Pac525對(duì)種植體周?chē)字虏【例l卟啉單胞菌、具核梭桿菌及血鏈球菌的抑菌作用隨時(shí)間和藥物的濃度呈正相關(guān)改變。 2.抗菌肽Pac525對(duì)三種細(xì)菌的菌斑生物膜均有破壞作用。MBIC為MIC的8~16倍。 3.MBRC50并未測(cè)出，這極有可能與其早期抑菌有關(guān)。
[Abstract]:Objective: In this experiment, we selected the synthetic antimicrobial peptide Pac525 with different degree of inhibition to the pathogenic bacteria in the three implants, such as Pseudomonas sp., Bacillus cereus and Streptococcus sanguis. The kill-time curve of these three kinds of bacteria was plotted by living bacteria count. In order to find out the inhibitory effect of Pac525 on the three kinds of bacteria at different time and different drug concentrations, the growth status of living bacteria biofilm and scanning electron microscope were observed. The lowest inhibitory biofilm concentration (MBIC) and the lowest drug clearance of the drug formed by Pac525 for the three strains of intact biofilm were determined by crystallization violet method (MBRC). Methods: One, antibacterial peptide Pac525 was used to kill pathogenic bacteria in peri-implant inflammation. l-time according to the amino acid sequence of Pac525, The antibacterial peptide was artificially synthesized. The experiment was carried out by using live bacteria counting method. ill-time. di, pure titanium surface gum base, pseudomonas, Bacillus cereus and blood chain Establishment of a plaque biofilm for cocci. To simulate the oral environment, pre-treat a smooth titanium sheet with saliva (37.degree. C., 24 (h) forming an acquired film on the surface thereof, placing the titanium sheet in a 24-well plate, washing with PBS, adding 1500 & mu; l of BHI liquid culture medium and 500 & mu; l of bacteria liquid per hole, culturing at 37 & deg; C for 24-48 hr, and removing The biofilm was observed by water drying and scanning electron microscope. The antibacterial peptide Pac525 The effect of MBIC on dental plaque biofilm was determined: three kinds of biofilm were established on 96-well plates, 200 ul Pac525 solution and 20. mu. l/ 107cfu/ ml of bacteria liquid were added, and 37 鈩
本文編號(hào)：2265392
本文鏈接：http://sikaile.net/yixuelunwen/kouq/2265392.html
[Abstract]:Objective: In this experiment, we selected the synthetic antimicrobial peptide Pac525 with different degree of inhibition to the pathogenic bacteria in the three implants, such as Pseudomonas sp., Bacillus cereus and Streptococcus sanguis. The kill-time curve of these three kinds of bacteria was plotted by living bacteria count. In order to find out the inhibitory effect of Pac525 on the three kinds of bacteria at different time and different drug concentrations, the growth status of living bacteria biofilm and scanning electron microscope were observed. The lowest inhibitory biofilm concentration (MBIC) and the lowest drug clearance of the drug formed by Pac525 for the three strains of intact biofilm were determined by crystallization violet method (MBRC). Methods: One, antibacterial peptide Pac525 was used to kill pathogenic bacteria in peri-implant inflammation. l-time according to the amino acid sequence of Pac525, The antibacterial peptide was artificially synthesized. The experiment was carried out by using live bacteria counting method. ill-time. di, pure titanium surface gum base, pseudomonas, Bacillus cereus and blood chain Establishment of a plaque biofilm for cocci. To simulate the oral environment, pre-treat a smooth titanium sheet with saliva (37.degree. C., 24 (h) forming an acquired film on the surface thereof, placing the titanium sheet in a 24-well plate, washing with PBS, adding 1500 & mu; l of BHI liquid culture medium and 500 & mu; l of bacteria liquid per hole, culturing at 37 & deg; C for 24-48 hr, and removing The biofilm was observed by water drying and scanning electron microscope. The antibacterial peptide Pac525 The effect of MBIC on dental plaque biofilm was determined: three kinds of biofilm were established on 96-well plates, 200 ul Pac525 solution and 20. mu. l/ 107cfu/ ml of bacteria liquid were added, and 37 鈩
本文編號(hào)：2265392
本文鏈接：http://sikaile.net/yixuelunwen/kouq/2265392.html
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