| Objective: The common complications of diabetes mellitus(DM), which mainly occur in the small and large blood vessels, such as diabetic nephropathy, retinopathy, coronary heart disease, are the main causes that induced injury or disability. Currently, the pathogenesis of complications of, meanwhile the role of chronic inflammation in complications has been increasingly recognized.The activation of inflammatory is promoted by Toll.1ike receptor 4(TLR4), as the pattern recognition receptors existing in kinds of immune cells, may be associated with the inflammation of complications in DM. As we known, TLR4 is expressed on the surfaces of macrophages and other known innate immune cells, once diabetes attack, the endogenous ligand such as heat shock protein 60(HSP60), high mobility group protein 1(HMGBI) will activate TLR4 and induce inflammations. Furthermore our previous clinical and experimental studies confirmed the expression of TLR4 in various kinds of cells under high glucose condition significantly increased as well as the expression of related cytokines IL-1 β and TNF-α, however it has not been fully understand that the expression of TLR4 in other organs and The mechanism of the interaction with related inflammatory factors. Therefore, we intend to construct a model of DM rat and detect the expression of TLR4, IL-1 β and TNF-α in organs of the liver, the heart and the kidney,investigate the extent of inflammatory infiltration in tissues of rats, learn of the inflammatory characteristics of organs of DM rats. Then we explore the mechanism of complications of DM and provide the experimental basis for the anti-inflammatory treatment of it.Methods:1 The construction of the model of diabetic rats: STZ(45mg / kg) have been injected from Abdominal cavity to male SD rats, blood glucose was measured after 72 h from tail vein blood. When two detection concentration was higher than 16.7mmol / L, the model is constructed successfully. There are 10 DM rats in the experimental group and normal rats in the control group each other. 2 Experimental methods 2.1 The detection to expression of m RNA from TLR4 and inflammatory factors in different organs:12 weeks later, we extract tissues in rats kidney, heart and liver, and detect the expression level of TLR-4, TNF-α and IL-1β by the Real-time PCR method. 2.2 The detection to the infiltration of Inflamed tissue from different organs: we construct the kidney, heart and liver tissue sections, and detect the inflammatory cells of inflammation tissues by means of HE staining. 3 Statistical methods We analyze the data by using Excel and SPSS13.0 statistical software,indicate the data on the mean and standard deviation and adopt single independent samples t test for the mean for the difference between the groups, the signal of statistical significance is P<0.05.Results:1 To construct the model of type II diabetic rats successfully. 2 The expression of TLR4 m RNA TLR4 expression of Real-time PCR results showed, the rat heart, liver and kidney is as follows, the expression level of TLR4 m RNA in the heart in the healthy control group was 1.104 ± 0.189 and that of DM group was 1.079 ± 0.503, there was no significant difference between control group and DM group; the expression level of TLR4 m RNA in the liver of the healthy control group was 0.933 ± 0.275 and that of DM group was 8.899 ± 1.769, which was significantly higher than the control group; the expression of TLR4 m RNA in kidney of healthy control group was 1.032 ± 0.125 and that of DM group was 2.766 ± 0.663, which has increased significantly compared to the healthy control group. 3 The expression of IL-1β m RNAReal-time PCR results showed that the expression of IL-1β m RNA in heart of the healthy control group was 1.025 ± 0.263 and that of DM group was 1.120 ± 0.311, there was no significant difference between them; the expression of IL-1β m RNA in the liver of healthy control group was 1.175 ± 0.275 and that of DM group was 3.940 ± 0.958, which was significantly higher than that of the control group; the expression of IL-1β m RNA in the kidney of the healthy control group was 1.125 ± 0.222 and that of DM group was 6.900 ± 1.700, which was significantly higher than that of the control group. 4 The expression of TNF-α m RNA The expression of TNF-α m RNA in the heart of the healthy control group was 1.075 ± 0.359 and that of DM group was 1.080 ± 0.449. There was no significant difference between them; the expression of TNF-α m RNA in the liver of the healthy control group was 1.150 ± 0.311 and that of DM group was 4.060 ± 1.097, which was significantly higher than that of healthy control group; the expression of TNF-α m RNA in the kidney of the healthy control group was 1.050 ± 0.208 and that of DM group was 10.38 ± 1.753, which was significantly higher than that of the control group. 5 HE staining results HE staining results in the heart, liver, kidney tissue showed that: 12 week later, compared with healthy control group, cardiac myocyte hypertrophy, interstitial edema, vascular inflammatory cells appeared in DM group; the infiltration of fatty inflammatory cells in liver is higher than that of the control group; there are larges of infiltrations in intercellular substance of the kidney of DM group.Conclusions:1 The high expression of TLR4 m RNA in the liver, kidney of DM groups exists indeed. 2 the expression of IL-1β and TNF-α m RNA in the liver and kidney of the DM group were significantly increased, while no significant changes took place in the heart.3 Infiltration has been shown in the inflammatory cells of heart, liver and kidney of DM group. |
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