| The chronic glomerulonephritis is a common disease which is usually lingering , slowly progressive, it' s one of the main cause which leads to chronic renal failure finally. The renal fibrosis is the end-result of all chronic renal disease and the primary cause of the end-stage chronic renal failure. No matter what kind of primary glomerulopathy, diabetes mellitus , or ureter blockage, the impairment of renal fuction correlated closely with the extent of renal fibration. In glomerular focal pathological changes ,the inflammatory cell which get to glomerulus through blood circulation and glomerular inherent cell play the important role in the renal fibration by secreting different cytokine and growth factor. Almost all of renal inherent cell may possess the ability to secrete cytokine , at the same time, they are the target cell which the cytokine produce the biological function, which have been confirmed at present. The cytokine participate in the pathologicalchanges through at least two pathway:the first , the cytokine play the part of aggression inflammatory mediator which trigger or promote the inflammation and impairment in nephridial tissue, like IL—1 β , TNF-α etac;the second, the cytokine as anti-inflammatory factor participate in renal self-defense , promote inflammatory resolution and tissue repair, for example IL-4, IL-10 etac. The end-result of renal glomerular disease which slowly progressive is determined by the comprehensive result between the pre-inflammatory factor and the anti-inflammatory facor. So we study the mechanism of action of Tongmai Oral Solution through observe the cytokine level secreted by the glomerular mesangial cell and the change after the Tongmai Oral Solution intervention, in order to provide the substantial theory and plenitudinous scientic evidence for Tongmai Oral Solution which is composed according to Strengthening-QI and Activating blood method cure chronic glomerularnephritis.In clinical research, we studied the relationship between serum Ⅲ precollagen, Ⅳ collagen and laminin level and syndrome differentiation of TCM in 60 CGN patients. The result illustrated the level of serum PC-Ⅲ, col-Ⅳ and LN in CGN patient is higher than the healthy control group significantly, however the group of chronic renal failure is higher than the group of CGN. In patient of CRF which syndrome differentiation of TCM is Deficiency of Spleen Yang and Kidney Yang , the level of serum Ⅲ precollagen, Ⅳ collagen and laminin rised obviously, but the patient of the Deficiency of QI and Yin have no significant difference. In patient of CGN which syndrome differentiation of TCM is Deficiency of Spleen Qi and Kidney Qi , the level of serum Ⅲ precollagen, Ⅳ collagen and laminin rised obviously, the frequency of accompanied symptoms and signs of Blood Stasis in CGN is the highest, moreover the patient belongs to the Blood Stasis whose serum level of Ⅲ precollagen, Ⅳ collagen and laminin rised obviously.Conclusion: the patient of CGN have different degree fibration in their kidney. With the decrease of renal fuction, the renal fibrosis aggravated gradually. The patient of the Deficiency of QI and Yin or Deficiency of Spleen Yang and Kidney Yang whose serum level of fibrous index rised up significantly. The patient of the Blood Stasis whose serum level of Ⅲ precollagen, Ⅳ collagen and laminin rised obviously. All of this have provided the substantial theory for Tongmai Oral Solution which is composed according to Strengthening-QI and Activating-Blood Method in treatment of chronic glomerulonephritis.Tongmai Oral Solution consist of multiple kind of Traditional Chinese Medicine, for example, Milkvetch Root and Sanchi etac. The previous clinical and experimental study have confirmed that the Tongmai Oral Solution can improve the patient' s clinical symptom and renal function, decrease the proteinuria, adjust the immune function, ameliorate the blood rheology, lessen the renal pathological lesion, defer the glomerular sclerosis. In experiment study, the author under the guidance of the director furtherly probe the mechinesm of Tongmai Oral Solution in treatment of CGN and defer the glomerular sclerosis in cell bilolgy and molecular biology directions, and study the effect of that the different compatibility between Milkvetch Root and Sanchi affect the level of the mesangial cell secrete the cytokine, thus try to find the best compatibility rule and mechanism of action between Milkvetch Rootand Sanchi at last.Objective:To culture the rat glomerular mesangial cell in vitro, we observe the different compatibility of Tongmai Oral Solution have effect on MC proliferation in order to make sure the best compatibility between Milkvetch Root and Sanchi and define the best compatibility of active priciple in Tongmai Oral Solution. We culture the rat MC with drug serum which stimulated by lipopolysaccharide, observe the IL-1 P and IL-10 cytokine level which secreted by MC, and study the effect of IL-1 ￡ mRNA and IL-10mRNA, in order to discover the mechinism of action of Tongmai Oral Solution.Method: To culture the rat MC in vitro with the Tongmai Oral Solution drug serum which consist of different dose-ratio between Milkvetch Root and Sanchi(1. 5:1> 3:K 6:1),we detect the cell proliferation by MTT, thus define the best compatibility between Milkvetch Root and Sanchi. According to the choosed effect ingredient, we design orthogonal experiment to make sure the best compatibility of active priciple in Tongmai Oral Solution. We take advantage of the Elissa technology to analyze the IL-1 ^ and IL-10 level which secreted by MC, and study the the effect of IL-10mRNA and IL-10mRNA with RT-PCR.Result: The Tongmai Oral Solution different dose match between Milkvetch Root and Sanchi can inhibit the MC proliferation.no matter what in normal culture condition or LPS as stimulant. In contrast, the group of 3:1 has the most significant inhibitory action than the group of 6:1 or 1. 5:1. According to the MTT result, we think the complex polycose and the complex saponin are the main active priciple. After orthogonal design, we confirm furtherly that the dose ratio between the complex polycose and the complex saponin is 3:3 which have the most notable inhibitory action on MC proliferation. We culture the rat glomerular mesangial cell with the drug serum of the complex polycose and the complex saponin whose the dose ratio is 3:3, LPS as stimulant, the result we found that the IL-1 3 production increased obviously in LPS group, and arrive at the maximum in 12 hour which has no statistical meaning in contrast to other time. However, the IL-1 0 production of the active priciple group decreased significantly than the LPS group in different time .The IL-10 level have no change in LPS group in different phase. But in the active priciple group the IL-10 level rised up obviously in 6h in contrast to the LPS group. At the same time, the result of RT-PCR stated that the drug serum of the active priciple... |
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