維生素D治療大鼠結(jié)腸炎模型療效觀察及免疫調(diào)節(jié)機制的初探
發(fā)布時間：2019-05-28 16:12
【摘要】：研究背景 炎癥性腸病是一組發(fā)病原因尚不明確的亞急性和慢性腸道炎癥性疾病,其臨床類型主要包括UC和CD�，F(xiàn)在普遍認為CD是Thl細胞異常激活引起的疾病。維生素D近年來被發(fā)現(xiàn)在多種自身免疫性疾病中有重要的免疫調(diào)節(jié)作用,尤其是Thl細胞介導的自身免疫疾病。維生素D對免疫的調(diào)控可以通過多個途徑達到,其中之一就是調(diào)控APC上的TLRs家族的表達及功能。通過TLRs信號通路可影響初始T細胞的分化,改變Thl細胞和Treg細胞在組織中的比例。目前有國外研究小組在動物實驗上使用1,25(OH)2D3干預,發(fā)現(xiàn)1,25(OH)2D3有抑制炎癥性腸病腸道炎癥的作用。但尚無使用維生素D3及其不同劑量對于IBD療效比較的研究。 研究目的 探索維生素D3和1,25(OH)2D3對IBD動物模型的療效及其可能的免疫機制。 研究方法 54只SD大鼠隨機分為9組,每組6只,分為：空白對照組,TNBS組,5-ASA (0.4g/kg/d)組,活性VD(1,25(OH)2D30.2ug/kg/d)組,活性VD(1,25(OH)2D30.2ug/kg/d)+5-ASA(0.4g/kg/d)組,大劑量維生素D3(7500IU/d)組,大劑量維生素D3(7500IU/d)+5-ASA(0.4g/kg/d)組,小劑量維生素D3(1800IU once)組,小劑量維生素D3(1800IU once)+5-ASA(0.4g/kg/d)組。治療9天后于第10天處死取材。采用DAI評分,大體炎癥評分,組織病理評分和結(jié)腸組織MPO測定來進行炎癥程度的評估。采用免疫組化的方法評估TLR4、TLR9、Foxp3在各組大鼠結(jié)腸局部中的表達。并用RT-PCR法測定大鼠結(jié)腸組織中TLR9mRNA的含量。 實驗結(jié)果 1、各TNBS處理組DAI評分、大體評分、組織病理學評分和MPO測定均較空白對照組明顯升高(P0.01)。各治療組結(jié)腸組織MPO值均顯著低于TNBS組(P0.01),但仍高于空白對照組。各治療組DAI評分、大體評分和組織病理學評分較TNBS組無明顯下降(P0.05)。各治療組之間療效無明顯差異(P0.05)。大劑量VD組和大劑量VD+5-ASA組血清鈣顯著高于其余各組(P0.01)。大劑量VD組血清肌酐高于其余各組(P0.05)。 2、空白對照組結(jié)腸固有層和粘膜下層炎性細胞胞膜和胞質(zhì)僅有少量TLR4和TLR9表達。TNBS造模組可見大量TLR4和TLR9表達,較空白組明顯增多(P0.01)。在空白對照組中僅有散在的Foxp3表達,主要不均勻分布于粘膜固有層淋巴細胞的細胞核內(nèi)。TNBS組中Foxp3表達較空白組明顯增多(P0.01)。單純5-ASA治療組TLR4、TLR9和Foxp3表達較TNBS組弱(P0.05)。各維生素D干預組TLR9和Foxp3表達較TNBS組弱(P0.05),而TLR4表達較TNBS組無明顯差異(P0.05)。各維生素D干預組之間TLR4、TLR9和Foxp3表達無明顯差異(P0.05)。TLR9mRNA在空白對照組中表達較少。各組之間條帶灰度比值無差異(P0.05)。 實驗結(jié)論 1、5-ASA、維生素D3和1,25(OH)2D能緩解大鼠結(jié)腸炎癥,聯(lián)合使用維生素D3和5-ASA及聯(lián)合使用1,25(OH)2D和5-ASA療效無增強。維生素D3抑制炎癥的作用和劑量無關。 2、正常大鼠結(jié)腸中TLR4、TLR9和Foxp3的表達較少。 3、TNBS誘導的大鼠結(jié)腸組織中TLR4、TLR9和Foxp3表達增多。與TNBS組比較,5-ASA干預后上述三者表達均可被下調(diào)；1,25(OH)2D3或維生素D3干預后TLR9和Foxp3表達減少,TLR4表達無明顯差異；不同劑量維生素D3干預后TLR4、TLR9和Foxp3的表達無明顯差異。聯(lián)合使用1,25(OH)2D3和5-ASA、維生素D3和5-ASA較單獨使用5-ASA結(jié)腸TLR4、TLR9和Foxp3的表達無明顯差異。各組TLR9在mRNA層面表達無明顯差異。
[Abstract]:Study Background Inflammatory bowel disease is a group of sub-acute and chronic intestinal inflammatory diseases that are not well-defined in a group, and its clinical types mainly include UC and C D. It is widely believed that CD is a disease caused by the abnormal activation of Thl cells Vitamin D has been found to be an important immunomodulatory activity in a variety of autoimmune diseases, especially Thl cell-mediated autoimmune diseases. The regulation of vitamin D to immunity can be achieved through multiple pathways, one of which is to regulate the expression and work of the TLRs on the APC Energy. The differentiation of the initial T cells can be affected by the TLRs signaling pathway, and the ratio of the Thl cells and Treg cells to the tissue is changed The results of this study were as follows:1,25 (OH) 2D3 was used to control the intestinal inflammation of inflammatory bowel disease. A study on the efficacy of vitamin D3 and its different doses in the treatment of IBD To study. Objective To explore the effect of vitamin D3 and 1,25 (OH) 2D3 on the animal model of IBD and its possible effect Immunologic machine Methods 54 SD rats were randomly divided into 9 groups, each group was divided into two groups: the blank control group, the TNBS group, the 5-ASA (0.4 g/ kg/ day) group, the active VD (1,25 (OH) 2D30.2 ug/ kg/ day) group, the active VD (1,25 (OH) 2D30.2 ug/ kg/ day) + 5-ASA (0.4 g/ kg/ day) group, the large-dose vitamin D3 (7500 IU/ d) group, the large-dose vitamin D3 (7500 IU/ d) + 5-ASA (0.4 g/ kg/ day) group, the low-dose vitamin D3 (180 0 IU on), small dose of vitamin D3 (1800 IU on) + 5-ASA (0. 4 g/ kg/ day).9 days after treatment The samples were sacrificed on Day 10. The DAI score, the gross inflammatory score, the tissue pathology score, and the colon tissue MPO assay were used. The assessment of the degree of inflammation was carried out. TLR4, TLR9 and Foxp3 were assessed by immunohistochemistry. Expression of the local part of the colon of the rat, and the TL-PCR method was used to determine the TL in the colon tissue of the rat. R9mR The results were as follows:1. The scores of DAI, the general score, the histopathological score and the MPO in the TNBS treatment group were all blank. The number of MPO in the colon of each treatment group was significantly lower than that of the TNBS group (P0.01). 01), but still higher than the blank control group. The DAI score, general score and histopathological score of each treatment group were lower than that of the TNBS There was no significant decrease in the treatment group (P0.05). There was no significant difference between the large-dose VD group and the large-dose VD + 5-ASA group. The serum of the large-dose VD group was higher than that of the other groups (P0.01). It was higher than that of the other groups (P0.05). The cells and the cytoplasm of the lamina propria and the submucosal inflammatory cells in the blank control group were higher than that of the other groups (P0.05). There is only a small amount of TLR4 and TLR9 expression. A large number of TLR4 and TLR9 expression is visible in the TNBS fabrication module, In the blank group, only the expression of Foxp3 in the blank group was significantly increased (P0.01). The expression of Foxp3 in TNBS group The number of TLR4, TLR9 and Foxp3 in the 5-ASA treatment group was significantly higher than that in the blank group (P0.01). The expression of TLR9 and Foxp3 in each vitamin D group was weaker than that of the TNBS group (P0.05), while the expression of TLR4 was higher than that of the TNBS group (P0.05). There was no significant difference in the expression of TLR4, TLR9 and Foxp3 among the groups of vitamin D (P0.05). TLR9 MRNA was expressed in the blank control group. band-to-gray ratio The results suggest that 1,5-ASA, vitamin D3 and 1,25 (OH) 2D can relieve the inflammation of the colon of the rat, and the combination of vitamin D3 and 5-ASA and the combination of 1, 25 (OH) 2D and 5-ASA with no increase in efficacy strong. Vitamin D3 inhibits the role and dose of inflammation. Normal rat colon The expression of TLR4, TLR9 and Foxp3 was less. The expression of TLR4, TLR9 and Foxp3 in intestinal tissue was increased. Compared with TNBS group, the expression of TLR4, TLR9 and Foxp3 could be reduced after 5-ASA intervention. The expression of TLR9 and Foxp3 decreased after the intervention of 1,25 (OH) 2D3 or vitamin D3, and the expression of TLR4 was not significant. There was no significant difference in the expression of TLR4, TLR9 and Foxp3. The combined use of 1,25 (OH) 2D3 and 5-ASA, vitamin D3, and 5-ASA used alone in the 5-ASA colon The expression of TLR4, TLR9 and Foxp3 was not obvious.
【學位授予單位】：北京協(xié)和醫(yī)學院
【學位級別】：博士
【學位授予年份】：2012
【分類號】：R-332;R574.62
本文編號：2487190
[Abstract]:Study Background Inflammatory bowel disease is a group of sub-acute and chronic intestinal inflammatory diseases that are not well-defined in a group, and its clinical types mainly include UC and C D. It is widely believed that CD is a disease caused by the abnormal activation of Thl cells Vitamin D has been found to be an important immunomodulatory activity in a variety of autoimmune diseases, especially Thl cell-mediated autoimmune diseases. The regulation of vitamin D to immunity can be achieved through multiple pathways, one of which is to regulate the expression and work of the TLRs on the APC Energy. The differentiation of the initial T cells can be affected by the TLRs signaling pathway, and the ratio of the Thl cells and Treg cells to the tissue is changed The results of this study were as follows:1,25 (OH) 2D3 was used to control the intestinal inflammation of inflammatory bowel disease. A study on the efficacy of vitamin D3 and its different doses in the treatment of IBD To study. Objective To explore the effect of vitamin D3 and 1,25 (OH) 2D3 on the animal model of IBD and its possible effect Immunologic machine Methods 54 SD rats were randomly divided into 9 groups, each group was divided into two groups: the blank control group, the TNBS group, the 5-ASA (0.4 g/ kg/ day) group, the active VD (1,25 (OH) 2D30.2 ug/ kg/ day) group, the active VD (1,25 (OH) 2D30.2 ug/ kg/ day) + 5-ASA (0.4 g/ kg/ day) group, the large-dose vitamin D3 (7500 IU/ d) group, the large-dose vitamin D3 (7500 IU/ d) + 5-ASA (0.4 g/ kg/ day) group, the low-dose vitamin D3 (180 0 IU on), small dose of vitamin D3 (1800 IU on) + 5-ASA (0. 4 g/ kg/ day).9 days after treatment The samples were sacrificed on Day 10. The DAI score, the gross inflammatory score, the tissue pathology score, and the colon tissue MPO assay were used. The assessment of the degree of inflammation was carried out. TLR4, TLR9 and Foxp3 were assessed by immunohistochemistry. Expression of the local part of the colon of the rat, and the TL-PCR method was used to determine the TL in the colon tissue of the rat. R9mR The results were as follows:1. The scores of DAI, the general score, the histopathological score and the MPO in the TNBS treatment group were all blank. The number of MPO in the colon of each treatment group was significantly lower than that of the TNBS group (P0.01). 01), but still higher than the blank control group. The DAI score, general score and histopathological score of each treatment group were lower than that of the TNBS There was no significant decrease in the treatment group (P0.05). There was no significant difference between the large-dose VD group and the large-dose VD + 5-ASA group. The serum of the large-dose VD group was higher than that of the other groups (P0.01). It was higher than that of the other groups (P0.05). The cells and the cytoplasm of the lamina propria and the submucosal inflammatory cells in the blank control group were higher than that of the other groups (P0.05). There is only a small amount of TLR4 and TLR9 expression. A large number of TLR4 and TLR9 expression is visible in the TNBS fabrication module, In the blank group, only the expression of Foxp3 in the blank group was significantly increased (P0.01). The expression of Foxp3 in TNBS group The number of TLR4, TLR9 and Foxp3 in the 5-ASA treatment group was significantly higher than that in the blank group (P0.01). The expression of TLR9 and Foxp3 in each vitamin D group was weaker than that of the TNBS group (P0.05), while the expression of TLR4 was higher than that of the TNBS group (P0.05). There was no significant difference in the expression of TLR4, TLR9 and Foxp3 among the groups of vitamin D (P0.05). TLR9 MRNA was expressed in the blank control group. band-to-gray ratio The results suggest that 1,5-ASA, vitamin D3 and 1,25 (OH) 2D can relieve the inflammation of the colon of the rat, and the combination of vitamin D3 and 5-ASA and the combination of 1, 25 (OH) 2D and 5-ASA with no increase in efficacy strong. Vitamin D3 inhibits the role and dose of inflammation. Normal rat colon The expression of TLR4, TLR9 and Foxp3 was less. The expression of TLR4, TLR9 and Foxp3 in intestinal tissue was increased. Compared with TNBS group, the expression of TLR4, TLR9 and Foxp3 could be reduced after 5-ASA intervention. The expression of TLR9 and Foxp3 decreased after the intervention of 1,25 (OH) 2D3 or vitamin D3, and the expression of TLR4 was not significant. There was no significant difference in the expression of TLR4, TLR9 and Foxp3. The combined use of 1,25 (OH) 2D3 and 5-ASA, vitamin D3, and 5-ASA used alone in the 5-ASA colon The expression of TLR4, TLR9 and Foxp3 was not obvious.
【學位授予單位】：北京協(xié)和醫(yī)學院
【學位級別】：博士
【學位授予年份】：2012
【分類號】：R-332;R574.62
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