本文選題：腸道菌群 + 短鏈脂肪酸��； 參考：《西南醫(yī)科大學》2017年碩士論文

【摘要】：目的:艾滋病是一種常見的性傳播疾病,當HIV病毒感染后會引起嚴重的細胞免疫缺陷并導致各種機會性感染,嚴重危害人類健康。臨床上絕大多數(shù)發(fā)展至AIDS期的患者會出現(xiàn)慢性腹瀉、腹痛等消化道癥狀,研究證明可能與腸道黏膜損傷、腸道菌群易位、腸道微生態(tài)失衡等病理機制有關(guān),但具體機制尚不明確。正常狀況下,機體與微生物群間存在互利共惠的共生關(guān)系,腸道微生物群及代謝產(chǎn)物在促進營養(yǎng)物質(zhì)消化吸收、維持腸道正常生理功能,調(diào)節(jié)機體免疫等眾多生命活動起重要作用。本實驗的研究對象將選擇尚未發(fā)展至AIDS期的HIV感染者,觀察其腸道微生態(tài)及腸道黏膜損傷情況,以及探究其腸道微生態(tài)與腸黏膜損傷、細胞免疫狀態(tài)是否具有相關(guān)性。本實驗將從以下幾個方面進行研究:1.對比HIV感染者和健康人群腸道菌群分布及其重要代謝產(chǎn)物短鏈脂肪酸的差異;2.對比HIV感染者與健康人群腸粘膜損傷標記物的差異;3.分析腸道菌群分布及不同短鏈脂肪酸與腸粘膜損傷標記物有無相關(guān)性;4.分析腸道菌群分布及不同短鏈脂肪酸與HIV感染者的CD4+、CD8+T細胞數(shù)和CD4/CD8比值有無相關(guān)性;從而為進一步完善HIV感染引起腸道損傷的機制以及腸道微生態(tài)相關(guān)的治療方法用于艾滋病的預(yù)防和治療打下理論基礎(chǔ)。方法:收集來自于傳染病醫(yī)院的符合入選標準的HIV感染者15例以及符合入選標準的健康者10例,所有入選對象均在實驗前簽訂知情同意書,并獲四川省人民醫(yī)院倫理道德委員會批準。采集兩組人群糞便標本后-80℃低溫保存,用高通量測序法檢測兩組人群腸道菌群的分布情況及其差異;用氣相色譜-質(zhì)譜法(gc-ms)聯(lián)合分析兩組人群糞便中各短鏈脂肪酸的含量;采集血液標本,離心后用酶聯(lián)免疫吸附法(elisa)測定兩組人群血清中能夠反映腸粘膜損傷情況的循環(huán)標記腸型脂肪酸結(jié)合蛋白(i-fabp)和d-乳酸的含量;用流式細胞術(shù)(fum)檢測反映hiv感染者細胞免疫狀態(tài)的cd4+、cd8+t細胞數(shù)和cd4/cd8比值。最后使用spss19.0統(tǒng)計軟件,采用mann-whitney檢驗,t檢驗進行分析細菌種屬或者otus組間統(tǒng)計學差異分析,采用person法進行相關(guān)性分析,p0.05提示差異有統(tǒng)計學差義。結(jié)果:1.腸道菌群分布:和健康對照組相比,在門水平上,hiv感染組腸道的擬桿菌門、厚壁菌門、變形菌門、梭菌門、放線菌門不存在顯著差異;在綱水平上,只有negativicutes菌顯著增多;在科水平上,理研菌rikenellaceae和分支桿菌microbacteriaceae菌顯著減少,莫拉菌moraxellaceae顯著增多;在屬水平上,羅氏菌roseburia、毛螺旋菌lachnospiraceae、alistipes菌、瘤胃球菌ruminococcaceae明顯減少;嗜冷桿菌psychrobacter明顯增多。2.各短鏈脂肪酸定量:和健康對照組相比,hiv感染組腸道丁酸和戊酸含量明顯減少,其他短鏈脂肪酸含量不存在差異。3.血清腸道黏膜損傷標記物:hiv感染組和健康對照組相比,血清腸道黏膜損傷標記物i-fabp和d-乳酸值不存在差異。4.細胞免疫:hiv感染者的cd4+t細胞數(shù)目200/μl,cd8+t細胞升高,cd4/cd8比值下降,符合尚未發(fā)展至aids期hiv感染者的細胞免疫狀態(tài)。5.腸道菌群分布與丁酸和戊酸的相關(guān)性分析:在科水平上,hiv感染者腸道丁酸含量的降低主要與理研菌科rikenellaceae、瘤胃球菌科ruminococcaceae呈正相關(guān)。戊酸含量降低主要與理研菌科rikenellaceae、消化球菌科peptococcaceae、瘤胃球菌科ruminococcaceae呈正相關(guān)。在屬水平上,丁酸含量降低主要與理研菌科的alistipes屬;毛螺旋菌科的roseburia屬、blautia屬、lachnospiraceaeucg-005屬、lachnospiraceaefcs020group屬;瘤胃球菌科的ruminococcaceaeucg-003屬、ruminococcus1屬呈正相關(guān)。戊酸含量降低主要與理研菌科的alistipes屬、瘤胃球菌科的eubacteriumcoprostanoligenesgroup屬、ruminococcaceaeucg-002屬、ruminococcaceaeucg-003屬、ruminococcaceaenk4a214group屬、ruminococcaceae1屬;毛螺旋菌科的eubacteriumruminantiumgroup屬;消化球菌科的peptococcus屬呈正相關(guān)。6.腸道菌群與cd4+、cd8+t細胞數(shù)及cd4/cd8比值的相關(guān)性:我們未發(fā)現(xiàn)與cd4+t細胞呈相關(guān)性的菌群。與cd8+t細胞呈正相關(guān)的主要菌群有:變形菌門下的oxalobacer、undibacterium、burkholderia、achromobacter、ramlibacter屬;擬桿菌門下sphingobacteriia科。7.短鏈脂肪酸與CD4+、CD8+T細胞數(shù)及CD4/CD8比值的相關(guān)性:CD4+T細胞、CD4/CD8比值與乙酸呈負相關(guān),CD8+T細胞與戊酸呈正相關(guān)。結(jié)論:1.HIV感染者在尚未進入艾滋病期,發(fā)生消化道癥狀前,腸道微生態(tài)環(huán)境可能已經(jīng)與健康者存在差異;2.HIV感染者的重要腸道粘膜保護因素較健康者降低,如羅氏菌屬Roseburia、毛螺旋菌屬Lachnospiraceae、Alistipes菌屬、瘤胃球菌屬Ruminococcaceae、理研菌科Rikenellaceae等的菌群負荷降低,相關(guān)細菌的代謝產(chǎn)物丁酸和戊酸水平下降;3.HIV感染者腸道粘膜的破壞性因素增加,如變形菌門中的莫拉菌科Moraxellaceae和冷桿菌屬Psychrobacter豐度在腸腔中顯著升高,推測其可能與菌群易位密切相關(guān),從而誘導持續(xù)的炎癥反應(yīng),加重HIV病程的發(fā)展;4.尚未發(fā)展至AIDS期的HIV感染者的細胞免疫水平與腸道菌群分布不一定具有相關(guān)性;5.尚未發(fā)展至AIDS期的HIV感染者的細胞免疫水平與腸道各短鏈脂肪酸SCFAs水平相關(guān)性不明確。
[Abstract]:Objective: AIDS is a common type of sexually transmitted disease. When HIV virus is infected, it can cause serious cellular immune deficiency and cause various opportunistic infections, which seriously harm human health. Most of the patients who develop to AIDS stage may have chronic diarrhea, abdominal pain and other digestive tract symptoms, and the study has proved that it may be associated with intestinal mucosal damage, intestinal mucosa damage, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa injury, intestinal mucosa damage, intestinal mucosa damage, intestinal mucosa damage, intestinal mucosa damage, intestinal mucosa damage, intestinal mucosa damage, intestinal mucosa damage, intestinal mucosa damage, intestinal mucosa damage, intestinal mucosa damage The pathomechanism of the translocation of the flora and the imbalance of the intestinal microflora is not clear. Under normal conditions, there is a symbiotic relationship between the organism and the microorganism. The intestinal microbes and the metabolites can promote the digestion and absorption of nutrients, maintain the normal physiological function of the intestinal tract, and regulate the many life activities of the body. The subjects of this study will choose HIV infected people who have not developed to AIDS stage, observe the intestinal microflora and intestinal mucosal damage, and explore whether the intestinal microflora and intestinal mucosa damage, and whether the cell immune state is related. This experiment will be studied from the following aspects: 1. comparison of HIV infected persons and The distribution of intestinal microflora and the difference between the important metabolites of short chain fatty acids in the healthy population; 2. compared the difference of intestinal mucosal damage markers between HIV infected people and healthy people; 3. analysis of the distribution of intestinal flora and the correlation between different short chain fatty acids and intestinal mucosal damage markers; 4. analysis of the distribution of intestinal microflora and different short chain fatty acids and HIV There is no correlation between the number of CD4+, CD8+T cells and the ratio of CD4/CD8 in the infected people, which provides a theoretical basis for further improving the mechanism of intestinal damage caused by HIV infection and the treatment of the intestinal microecology related to the prevention and treatment of AIDS. Methods: 15 cases of HIV infected with the criteria of admission from infectious diseases hospitals were collected. And 10 healthy persons who met the standard of admission, all the selected subjects signed informed consent before the experiment, and approved by the ethics committee of Sichuan Provincial People's Hospital. Two groups of fecal specimens were collected at -80 C and preserved at low temperature. The distribution and difference of intestinal microflora of two groups of people were detected by high flux sequencing method, and gas chromatography-quality was used. The content of the short chain fatty acids in the feces of two groups of people was analyzed by GC-MS. Blood samples were collected, and after centrifugation, the contents of circulating labeled intestinal fatty acid binding protein (I-FABP) and d- lactic acid in the serum of two groups were measured by enzyme linked immunosorbent assay (ELISA); the flow cytometry (fum) was used to detect the content of the intestinal mucosa injury. Cd4+, cd8+t cell number and cd4/cd8 ratio in the cell immune state of HIV infected people. Finally, using the spss19.0 statistics software, using Mann-Whitney test, t test to analyze the statistical difference between the bacteria species and the OTUs group, and using person method to analyze the correlation. The P0.05 suggests the difference is statistically difference. Results: 1. intestinal flora distribution: Compared with the healthy control group, there was no significant difference in the intestinal bacteriobacteria, the thicker door, the deformable bacteria gate, the Clostridium, and the actinomycetes in the HIV infection group. At the level of the class, only the negativicutes bacteria increased significantly; at the level of the family, the rikenellaceae and the Mycobacterium microbacteriaceae were significantly reduced, Molarand moraxell Aceae was significantly increased; at the level of the genus Roche Roseburia, spirulae lachnospiraceae, alistipes, and rumen cocci ruminococcaceae decreased significantly; Psychrobacter significantly increased.2. short chain fatty acids in.2.: compared with the healthy control group, the content of butyric acid and valeric acid in the intestinal tract of HIV infection group decreased significantly, and the other short chain fatty acids were found in the HIV infection group. There was no difference in.3. serum intestinal mucosa damage markers: the value of serum intestinal mucosal damage markers I-FABP and d- lactate in HIV infection group and healthy control group did not differ from.4. cell immunity: the number of cd4+t cells in HIV infected persons was 200/ U L, cd8+t cells increased and cd4/cd8 ratio decreased, which was in line with those who had not developed to AIDS stage. The relationship between the distribution of.5. intestinal flora and the relationship between butyric acid and valerate: at the level of the family, the decrease of butyric acid content in the intestinal tract of HIV infected people is mainly related to the rikenellaceae of the family of the family of LAD and the ruminococcaceae of the family of rumen, and the decrease of the content of valerate is mainly with the rikenellaceae in the family of science and medicine, the peptococcaceae of the family of digestive bacteria, and the tumor. A positive correlation is found in the ruminococcaceae of the family of stomach cocci. At the level of the genus, the decrease of butyric acid content is mainly with the alistipes genus of the family of the family, Roseburia, blautia, lachnospiraceaeucg-005, lachnospiraceaefcs020group, ruminococcaceaeucg-003, ruminococcus1, and valeric acid in the family of the family of the family of the family rumen. The genus alistipes, eubacteriumcoprostanoligenesgroup, ruminococcaceaeucg-002, ruminococcaceaeucg-003, ruminococcaceaenk4a214group, ruminococcaceae1, eubacteriumruminantiumgroup, and eubacteriumruminantiumgroup genera of the family helioris; peptococcus genera in the family of digestistate families are positively related to.6. intestines. The correlation between the number of cd4+, cd8+t cells and the ratio of cd4/cd8: we did not find the bacteria associated with the cd4+t cells. The main groups that were positively related to the cd8+t cells were: oxalobacer, undibacterium, Burkholderia, Achromobacter, ramlibacter genera under the Proteus, sphingobacteriia family under the bacteriobacteria and the.7. short chain fatty acids. The correlation between CD8+T cell number and CD4/CD8 ratio: CD4+T cells, CD4/CD8 ratio is negatively correlated with acetic acid, CD8+T cells are positively correlated with valerate. Conclusion: 1.HIV infected people may have different intestinal microecological environment before the onset of AIDS, and the intestinal microecological environment may have been different from those of healthy people; the important intestinal mucosa protection of 2.HIV infected people The factors such as Roseburia of Roche, Lachnospiraceae, Alistipes, Ruminococcaceae of rumen, and Rikenellaceae in the family of Rikenellaceae and so on were reduced, and the levels of butyric acid and valeric acid in the metabolites of the related bacteria decreased, and the destructive factors of intestinal mucosa in the 3.HIV infected people were increased, such as the deformable bacteria. The abundance of Moraxellaceae and Psychrobacter in the genus Mora and colibacilli was significantly increased in the intestinal cavity. It is suggested that it may be closely related to the bacterial translocation, thus inducing sustained inflammatory response and aggravating the development of the HIV disease course; 4. the cellular immune level of the HIV infected persons who have not yet developed to the AIDS stage is not necessarily related to the distribution of intestinal flora; 5 The correlation between the level of cellular immunity and the short chain fatty acid SCFAs level of intestinal tract in HIV infected persons who have not yet developed to AIDS stage is not clear.
【學位授予單位】：西南醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R512.91

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