本文選題：食源性肥胖 + 免疫　； 參考：《天津醫(yī)科大學(xué)》2009年碩士論文

【摘要】： 目的: 1建立食源性肥胖(diet induced obesity,DIO)SD大鼠模型,觀察肥胖大鼠與非肥胖大鼠脾臟CD4~+和CD8~+T淋巴細(xì)胞含量的變化,探究肥胖對大鼠細(xì)胞免疫功能的影響。 2觀察大鼠脾臟淋巴細(xì)胞表面是否存在長型的leptin受體(Ob-Rb)蛋白以及淋巴細(xì)胞內(nèi)是否存在磷酸化的STAT3蛋白。 3觀察肥胖大鼠與非肥胖大鼠脾臟淋巴細(xì)胞Ob-Rb-STAT3信號轉(zhuǎn)導(dǎo)通路活性的差異,從而探究肥胖狀態(tài)下機(jī)體免疫功能的變化是否由leptin通過Ob-Rb-STAT3信號轉(zhuǎn)導(dǎo)通路作用引起。 方法: 1利用高能量高脂肪飲食喂養(yǎng)大鼠14周后,將其與用普通飼料喂養(yǎng)的大鼠的體重進(jìn)行比較,根據(jù)體重分為DIO組以及肥胖抵抗(diet resistant,DR)組。 2分別取對照組、DR組和DIO組大鼠的脾臟,將其制成脾細(xì)胞的單細(xì)胞懸液,再利用流式細(xì)胞術(shù)檢測大鼠脾臟CD4~+和CD8~+T淋巴細(xì)胞在脾細(xì)胞中所占的百分比。 3分別提取對照組、DR組和DIO組大鼠脾細(xì)胞的蛋白,利用Western blot檢測大鼠淋巴細(xì)胞表面的Ob-Rb蛋白以及淋巴細(xì)胞內(nèi)磷酸化的STAT3蛋白的表達(dá)強(qiáng)度。 結(jié)果: 1體重變化的結(jié)果:普通飼料喂養(yǎng)的對照組大鼠與高能量高脂肪飼料喂養(yǎng)的大鼠的初始平均體重相當(dāng),無顯著性差異(P＞0.05)。經(jīng)過14周不同飼料的喂養(yǎng)之后,高能飼料組大鼠的平均體重是對照組大鼠平均體重的122.1%,具有顯著性差異(P＜0.001)。在高能飼料組中,10只體重最高的大鼠設(shè)為DIO組,體重最低的10只大鼠設(shè)為DR組,DIO組大鼠的平均體重是DR組大鼠的136.7%,具有顯著性差異(P＜0.001)。 2三組大鼠脾細(xì)胞中CD4~+和CD8~+T淋巴細(xì)胞含量比較的結(jié)果:通過流式細(xì)胞術(shù)的檢測發(fā)現(xiàn)與對照組和DR組相比較,DIO組大鼠脾細(xì)胞中CD4~+T淋巴細(xì)胞所占百分比變化并不明顯,沒有統(tǒng)計(jì)學(xué)差異(P＞0.05)。與對照組和DR組相比較,DIO組大鼠脾細(xì)胞中CD8~+T淋巴細(xì)胞所占百分比增加(P＜0.001)。上述CD4~+和CD8~+T淋巴細(xì)胞含量的變化導(dǎo)致DIO組大鼠脾臟CD4~+/CD8~+細(xì)胞比值低于DR組大鼠(P＜0.01)以及對照組大鼠(P＜0.001)。 3三組大鼠脾臟淋巴細(xì)胞Ob-Rb蛋白以及磷酸化的STAT3蛋白表達(dá)強(qiáng)度的比較:通過Western blot檢測發(fā)現(xiàn)在各組大鼠脾臟淋巴細(xì)胞所提取的蛋白中,128-kDa以及82-kDa處可見明顯蛋白表達(dá)條帶,與預(yù)期分子量一致,表明Ob-Rb蛋白以及705位酪氨酸磷酸化的STAT3蛋白在大鼠脾臟淋巴細(xì)胞中都有不同程度的表達(dá)。利用UVP凝膠成像系統(tǒng)分析后,DIO組、DR組以及對照組的大鼠脾臟淋巴細(xì)胞表面Ob-Rb蛋白以及細(xì)胞內(nèi)705位酪氨酸磷酸化的STAT3蛋白表達(dá)的密度值并沒有統(tǒng)計(jì)學(xué)的差異(P＞0.05)。 結(jié)論: 1經(jīng)過14周高能量高脂肪的飼料喂養(yǎng),SD大鼠能發(fā)展成為DIO大鼠和DR大鼠兩種體重存在明顯差異動(dòng)物模型,這一結(jié)果的產(chǎn)生是由于DIO大鼠和DR大鼠調(diào)控自身體重的方式不同所造成的。 2流式細(xì)胞術(shù)的實(shí)驗(yàn)結(jié)果表明,與對照組和DR組相比較,DIO組大鼠的脾細(xì)胞中CD4~+T淋巴細(xì)胞所占百分比變化并不明顯,CD8~+T淋巴細(xì)胞所占百分比增加,DIO組大鼠脾臟CD4~+/CD8~+細(xì)胞比值低于DR組以及對照組大鼠。這一實(shí)驗(yàn)結(jié)果表明,肥胖使大鼠處于免疫抑制狀態(tài),其細(xì)胞免疫功能降低,對感染的抵抗力下降,肥胖大鼠的易感性相應(yīng)的增加。 3 Western blot的實(shí)驗(yàn)結(jié)果表明,Ob-Rb蛋白以及磷酸化的STAT3蛋白在大鼠脾臟淋巴細(xì)胞中都有不同程度的表達(dá)。DIO組、DR組以及對照組大鼠脾臟淋巴細(xì)胞表面的Ob-Rb蛋白以及細(xì)胞內(nèi)磷酸化的STAT3蛋白表達(dá)的強(qiáng)度并沒有明顯的差異。這一實(shí)驗(yàn)結(jié)果說明,DIO大鼠細(xì)胞免疫功能的下降與其脾臟淋巴細(xì)胞中的Ob-Rb-STAT3信號轉(zhuǎn)導(dǎo)通路并沒有直接的關(guān)系。
[Abstract]:Objective:
1 the diet induced obesity (DIO) SD rat model was established to observe the changes of CD4~+ and CD8~+T lymphocytes in the spleen of obese rats and non obese rats, and to explore the effect of obesity on the cellular immune function of rats.
2 to observe whether there is long leptin receptor (Ob-Rb) protein on the surface of spleen lymphocytes and whether phosphorylated STAT3 protein exists in lymphocytes.
3 observe the difference in the activity of Ob-Rb-STAT3 signal transduction pathway in the spleen lymphocytes of obese rats and non obese rats, and to explore whether the changes of immune function in the body of obesity are caused by the role of leptin through the Ob-Rb-STAT3 signal transduction pathway.
Method:
1 after 14 weeks of high energy and high fat diet, the rats were compared with the body weight of rats fed with ordinary diet, and the body weight was divided into DIO group and diet resistant (DR) group.
2 the spleen of the control group, the DR group and the DIO group were respectively made into the single cell suspension of the spleen cells, and the percentage of the spleen CD4~+ and the CD8~+T lymphocyte in the spleen cells was detected by flow cytometry.
3 the protein of the spleen cells in the control group, the DR group and the DIO group, and the Western blot were used to detect the Ob-Rb protein on the surface of the rat lymphocyte and the expression intensity of the phosphorylated STAT3 protein in the lymphocyte.
Result:
1 the results of body weight change: the average weight of the rats fed with the normal diet was equal to the rats fed with high energy and high fat feed, and there was no significant difference (P > 0.05). After 14 weeks of feeding, the average weight of the rats in the high energy diet group was 122.1% of the control group (P 0.001). In the high energy diet group, 10 rats with the highest weight were set as DIO group, and the 10 rats with the lowest weight were set as group DR, and the average weight of group DIO was 136.7% in group DR (P < 0.001).
2 Comparison of the content of CD4~+ and CD8~+T lymphocytes in the splenocytes of the three groups: compared with the control group and the DR group by flow cytometry, the percentage of CD4~+T lymphocyte in the spleen cells of the DIO group was not significantly different (P > 0.05). Compared with the control group and the DR group, the spleen cells of the DIO group rats were compared with the control group and the DR group. The percentage of CD8~+T lymphocytes increased (P < 0.001). The changes in the content of CD4~+ and CD8~+T lymphocytes in the DIO group were lower than that of the DR group (P < 0.01) and the control group (P < 0.001).
3 Comparison of the expression intensity of Ob-Rb protein and phosphorylated STAT3 protein in the spleen lymphocytes of the three groups of rats: through Western blot detection, it was found that in the protein extracted from the spleen lymphocytes of the rats of each group, the obvious protein bands were found at 128-kDa and 82-kDa, which was consistent with the expected molecular weight, indicating the Ob-Rb protein and 705 tyrosine phosphorus. The acidified STAT3 protein had different degrees of expression in the spleen lymphocytes of rats. After the analysis of the UVP gel imaging system, there was no statistical difference in the density of Ob-Rb protein on the surface of spleen lymphocyte and the expression of 705 tyrosine phosphorylation of STAT3 protein in the rats of DIO, DR and control groups (P > 0.05).
Conclusion:
1 after 14 weeks of high energy and high fat feed, SD rats were able to develop into DIO rats and DR rats with significant differences in animal models. This result was caused by different ways of regulating the body weight of DIO rats and DR rats.
The results of 2 flow cytometry showed that compared with the control group and the DR group, the percentage of CD4~+T lymphocyte in the spleen cells of the DIO group was not obvious, the percentage of CD8~+T lymphocytes increased, and the ratio of CD4~+/CD8~+ cells in the spleen of the DIO group was lower than that of the DR group and the control group. Mice are in an immunosuppressive state, their cellular immune function is reduced, their resistance to infection is reduced, and the susceptibility of obese rats increases correspondingly.
The experimental results of 3 Western blot showed that the Ob-Rb protein and the phosphorylated STAT3 protein had different levels of.DIO in the spleen lymphocytes of rats. There was no significant difference between the Ob-Rb protein on the surface of the spleen lymphocyte and the intensity of the phosphorylated STAT3 protein expression in the spleen lymphocytes of the DR group and the control group. The results showed that the decrease of cellular immune function in DIO rats was not directly related to the Ob-Rb-STAT3 signal transduction pathway in splenic lymphocytes.
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2009
【分類號】：R392.12

【共引文獻(xiàn)】

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