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  本文選題：急性鼻-鼻竇炎　切入點：肺炎鏈球菌　出處：《南昌大學》2011年碩士論文　論文類型：學位論文

【摘要】：第一部分大鼠急性鼻一鼻竇炎模型的建立 目的：探討運用Merocel止血棉和肺炎鏈球菌制作大鼠急性鼻—鼻竇炎模型。 方法：正常SD大鼠50只,隨機分為實驗組(40只)和對照組(10只)。實驗組大鼠右側(cè)鼻腔均置入條棒狀Merocel止血棉,同時滴加0.1m1肺炎鏈球菌菌液于止血棉上；對照組大鼠右側(cè)鼻腔單純滴入等量生理鹽水。實驗組大鼠分別于接種細菌后1、2、3、4周處死10只；對照組大鼠均于第1周處死。麻醉、斷頭、剔除鼻面部軟組織,取包含鼻腔鼻竇的組織,4%多聚甲醛固定、10%EDTA脫鈣、酒精梯度脫水、石蠟包埋、切片、HE染色觀察鼻腔鼻竇炎性反應(yīng)情況。 結(jié)果：實驗組大鼠鼻腔鼻竇黏膜均有不同程度的炎癥細胞侵潤及黏膜上皮變薄,第1周炎癥反應(yīng)出現(xiàn),第2周炎癥反應(yīng)最重,第3周炎癥較前減輕,第4周最輕,基本恢復(fù)正常,對照組大鼠鼻腔鼻竇黏膜沒有炎癥反應(yīng)。 結(jié)論：運用Merocel止血棉作為肺炎鏈球菌的載體可建立較其他方法更為穩(wěn)定便捷的大鼠急性鼻—鼻竇炎模型,為鼻—鼻竇炎的研究提供了可靠的實驗基礎(chǔ)。 第二部分嗅覺功能的檢測 目的：探討運用埋藏食物小球?qū)嶒?buffed food pellet test, BFPT)檢測大鼠嗅覺功能的可行性,并分析嗅覺功能的變化情況。 方法：正常SD大鼠100只,隨機分為實驗組(80只)和對照組(20只)。在建立上述大鼠急性鼻—鼻竇炎模型的基礎(chǔ)上,采用Nathan等提出的行為學方法一埋藏食物小球法(buffed food pellet test, BFPT)檢測各組大鼠的嗅覺功能,并以林靜等提出的超過300s未找到食物小球作為大鼠嗅覺障礙的參考標準。實驗組大鼠分別于造模后的第1、2、3、4周檢測嗅覺功能,每次檢測20只(檢測之前抽出大鼠右側(cè)鼻腔內(nèi)填塞的Merocel止血棉),對照組大鼠于第1周檢測。記錄每只大鼠從放入籠內(nèi)到找到食物小球(雙前肢抱住或牙齒咬住食物小球)的時間。每只大鼠測試兩次,分別在受試當天的上午和下午進行測試,間隔6個小時。兩次結(jié)果取平均值,并進行統(tǒng)計學分析。 結(jié)果：各實驗組大鼠尋找到食物小球的時間均較對照組為長,以第2周最長。并且前三周均超過300s,即均伴有不同程度的嗅覺功能障礙。第4周盡管低于300s,但仍較對照組長。差別均具統(tǒng)計學意義(P0.01)。 結(jié)論：埋藏食物小球法與病理組織學檢查相結(jié)合,增加了實驗的客觀性,可作為評估動物嗅覺功能的一種方法。 第三部分成熟嗅感覺神經(jīng)元變化的實驗觀察 目的：觀察大鼠急性鼻一鼻竇炎模型中成熟嗅感覺神經(jīng)元的變化情況。 方法：在上述建立大鼠急性鼻一鼻竇炎動物模型的基礎(chǔ)上,用免疫熒光標記法觀察嗅黏膜上皮中嗅感覺神經(jīng)元的變化情況。實驗組大鼠分別于接種細菌后1、2、3、4周處死20只；對照組大鼠均于第1周處死。麻醉、斷頭、剔除鼻面部軟組織,于4%多聚甲醛中取包含鼻中隔黏膜的組織,依次進行固定、脫水、包埋、制作冰凍切片、順序標號。采用抗嗅覺蛋白受體(OMP,標記成熟神經(jīng)元)觀察成熟嗅感覺神經(jīng)元的變化。 結(jié)果：對照組大鼠嗅黏膜中下層可見大量排列整齊呈圓形或橢圓形的成熟嗅感覺神經(jīng)元,固有層亦有散在分布。實驗中觀察到造模后第1周,成熟嗅感覺神經(jīng)元出現(xiàn)減少。至第2周炎癥反應(yīng)高峰期,成熟嗅感覺神經(jīng)元減少,尤為顯著。造模后第3周,成熟嗅感覺神經(jīng)元開始增加。至第4周,成熟嗅感覺神經(jīng)元較前增多,但是尚未恢復(fù)正常神經(jīng)元的總數(shù)。 結(jié)論：鼻一鼻竇炎導(dǎo)致成熟嗅感覺神經(jīng)元凋亡增加,并能夠誘導(dǎo)嗅感覺神經(jīng)元再生,但炎性環(huán)境下新生嗅感覺神經(jīng)元的成熟障礙。 第四部分嗅鞘細胞變化的實驗觀察 目的：觀察大鼠急性鼻一鼻竇炎模型中嗅鞘細胞的變化情況,以進一步探索嗅鞘細胞在鼻—鼻竇炎嗅覺功能恢復(fù)中發(fā)揮的作用。 方法：在上述建立大鼠急性鼻一鼻竇炎動物模型的基礎(chǔ)上,應(yīng)用免疫熒光雙標法觀察嗅黏膜上皮中嗅鞘細胞的變化情況。在第三部分所制冰凍切片中,選取兩張,采用p75NTR抗體和GFAP抗體觀察嗅鞘細胞的變化情況。 結(jié)果：正常嗅黏膜固有層可見大量排列整齊的呈梭形或圓形的嗅鞘細胞。造模后第1周,嗅上皮變薄,固有層嗅鞘細明顯減少,同時嗅上皮偶可見OECs生長。造模后第2周,嗅上皮顯著變薄,固有層OECs較上周增多,嗅上皮可見明顯增多的OECs生長,多為GFAP單陽性未成熟扁圓形,群集生長。造模后第3周,嗅上皮厚度增加,固有層OECs增多,部分形成集落。嗅上皮仍可見少量OECs生長。造模后第4周,嗅上皮厚度基本恢復(fù)正常,固有層嗅鞘細胞基本恢復(fù)正常,嗅上皮偶可見OECs生長。 結(jié)論：,急性鼻—鼻竇炎不僅可導(dǎo)致ORNs凋亡增加,同時OECs亦會出現(xiàn)短時間的減少。但隨之OECs出現(xiàn)應(yīng)激性再生增加,尤以第2周炎癥高峰期最為明顯,直至炎癥消退。提示OECs在急性鼻—鼻竇炎嗅覺障礙的恢復(fù)中可能發(fā)揮了不可忽視的作用。
[Abstract]:The first part of the rat model of acute rhinosinusitis
Objective: To explore the model of acute rhinosinusitis in rats by using Merocel hemostat and Streptococcus pneumoniae.
Methods: 50 SD rats were randomly divided into experimental group (40 rats) and control group (10 rats). Rats in the experimental group were implanted into the right nasal rod dropping the Merocel sponge, 0.1m1 of Streptococcus pneumoniae bacteria on hemostatic cotton; the control group on the right side of rat nasal drops the same amount of pure life saline. Experimental group rats were killed 10 weeks after inoculation of bacteria 1,2,3,4; the control group rats were killed in first weeks. Anesthesia, decapitated, removing nasal facial soft tissue, the nasal tissue, 4% paraformaldehyde, 10%EDTA decalcified, dehydrated with gradient ethanol, paraffin embedded, sliced. HE staining was used to observe the nasal inflammation reaction.
Results: the nasal mucosa of rats in experimental group were significantly different degrees of infiltration of inflammatory cells and epithelial thinning, inflammatory reaction appeared first weeks second weeks, the most important inflammatory response, third weeks before the fourth week of inflammation is reduced, the light returned to normal, the control group of nasal mucosa in rats without inflammation.
Conclusion: the use of Merocel hemostatic cotton as carrier of Streptococcus pneumoniae can establish a more stable and convenient model of acute rhinosinusitis in rats than other methods. It provides a reliable experimental basis for the research of rhinosinusitis.
Detection of olfactory function in the second part
Objective: To explore the feasibility of buffed food pellet test (BFPT) in detecting olfactory function in rats and analyze the change of olfactory function.
Methods: 100 SD rats were randomly divided into experimental group (80 rats) and control group (20 rats). Based on the establishment of the rat acute nose - sinusitis model, proposed by Nathan and the behavior of a buried food pellet method methodology (buffed food pellet test, BFPT) were measured the rat olfactory function, and more than to put forward by Lin Jing 300s did not find a food pellet as olfactory dysfunction in rats. The rats of the experimental group reference standard respectively after modeling 1,2,3,4 weeks of detection of olfactory function, each test 20 (before testing out the rat right nasal packing Merocel hemostatic cotton). The rats in the control group in the first week test. Records of each rat from into the cage to find food pellets (double teeth or hold forelimb food pellet) time. Each rat was tested two times respectively in the subjects on the day of the morning and afternoon test interval of 6 hours. The average value of the two results was taken and the statistical analysis was carried out.
Results: the experimental rats to find the food pellet time were higher than those in control group, second weeks and three weeks before the longest. More than 300s, which were accompanied by varying degrees of olfactory dysfunction. Fourth weeks despite lower than 300s, but still than in the control group. The difference was statistically significant (P0.01).
Conclusion: the combination of buried food ball method and histopathological examination can increase the objectivity of the experiment, and can be used as a method to evaluate the olfactory function of animals.
Experimental observation on the changes of the third part of the mature olfactory sensory neurons
Objective: To observe the changes of the mature olfactory sensory neurons in the rat model of acute rhinosinusitis.
Methods: in the basis of acute rat nasal sinusitis animal model, to observe the changes of olfactory epithelium of olfactory sensory neurons by immunofluorescence method. The experimental group rats were killed 20 weeks after inoculation of bacteria 1,2,3,4; the rats in control group were dead at first weeks. Anesthesia, decapitation, culling nasal and facial soft tissue in 4% paraformaldehyde in the nasal mucosa tissue were fixed, dehydrated, embedded, frozen sections were made, sequence labeling. The anti olfactory receptor protein (OMP, markers of mature neurons) observed in mature olfactory sensory neurons.
Results: in the control group in the rat olfactory mucosa layer shows a large number of rows in the mature olfactory sensory neurons round or oval, are scattered in the lamina propria. Observed first weeks after the experiment, the mature olfactory sensory neurons decreased to second weeks. The inflammatory response peak, mature olfactory sensory neurons decreased significantly at third weeks after modeling, the mature olfactory sensory neurons began to increase. At fourth weeks, the mature olfactory sensory neurons was increased, but the total number of neurons have not yet returned to normal.
Conclusion: rhinosinusitis can increase the apoptosis of mature olfactory sensory neurons and induce the regeneration of olfactory sensory neurons, but the maturation of new olfactory sensory neurons in inflammatory environment.
Experimental observation on the changes of olfactory ensheathing cells in the fourth part
Objective: To observe the changes of olfactory ensheathing cells in acute rhinosinusitis model in rats, so as to further explore the role of olfactory ensheathing cells in the recovery of olfactory function in rhinosinusitis.
Methods: in the basis of acute rat nasal sinusitis animal model, double immunofluorescence method was used to observe the changes of olfactory ensheathing cells of the olfactory epithelium. In the third part of the frozen section, select two, using p75NTR and GFAP antibody to observe the changes of olfactory ensheathing cells.
Results: the normal olfactory mucosa lamina propria in large fusiform or round arrangement of olfactory ensheathing cells were neat. First weeks after modeling, the thinning of the olfactory epithelium, lamina propria of olfactory ensheathing fine was reduced, while the olfactory epithelium even visible growth of OECs. Second weeks after modeling, significant thinning of the olfactory epithelium, lamina propria OECs last week increased, the olfactory epithelium increased significantly OECs visible growth for GFAP single positive immature oblate, cluster growth. Third weeks after modeling, the increase of the thickness of the olfactory epithelium, lamina propria OECs increased, part of the colony formation. The olfactory epithelium was still a small growth of OECs. Fourth weeks after modeling, the thickness of the olfactory epithelium basic return to normal, the lamina propria of olfactory ensheathing cells returned to normal, the olfactory epithelium showed occasional OECs growth.
Conclusion: acute rhinosinusitis can not only lead to the apoptosis of ORNs, while OECs will also appear to reduce short time. But with OECs stress of regeneration increased, especially in the second week peak of inflammation was most obvious, until the inflammation subsided. It suggested that OECs might play a noticeable role in olfactory dysfunction in acute rhinosinusitis the recovery.

【學位授予單位】：南昌大學
【學位級別】：碩士
【學位授予年份】：2011
【分類號】：R765.21

【引證文獻】

相關(guān)博士學位論文 前1條

1 賈旭錦;鼻竇炎口服液對急性鼻—鼻竇炎大鼠鼻黏膜β防御素影響的實驗研究[D];成都中醫(yī)藥大學;2012年

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本文編號：1558291