| PrefaceAristolochi Acid Nephropathy ( AAN) is a kind of tubulointerstitial injury caused by Aristolochi Acid ( AA) , which can be expressed as acute renal failure, chronic renal failure and tubules acidosis. The pathologic characteristics of AAN is extensive renal interstitial fibrosis, damage and shelling of renal tubular epithelial cells. It has been widely accepted that the extensive renal interstitial fibrosis stems from fibroblasts recruitments or activation, and transformation polarization of renal tubular epithelial cells, while few reports about the important role of peri - tubular microcirculation injury and microcirculation dysfunction on mechanism of AAN can be found. The microthrombosis in renal tissue plays an important role in the formation and development of pathological changes in vassal disease. Endothelial cells damage results in the function unintegrity of vassal wall. The enhancement of substance for vasoconstrict shrink blood vessels, slow blood stream, resulting in local anoxemia, as well as the releasing enhancement of procoagulant factor. The reducing anticoagulant factor strength high coagulation of blood, resulting in formation of thrombus easily, which in turn strength the anoxia of endothelial cells and form a vicious circle. The purpose of this work is to find out the significance of capillary vessel disease in AAN by measuring the role of Endothelin - 1 and Von willebrand factor in kidney of Wistar rats with immunochemical technique. At the same time, a treatment with GBE was attempted to study the effect of medication for improving microcirculation inAAN treatment.Materials and Methods1. Materials1. 1 Animals: 50 female Wistar rats were divided into three groups random-ly.1. 2 Major reagent and physics: Manchurian Dutchmanspipe, Ginkgo Bilola Extract, ET - 1 test kits, VWF test kits.1. 3 Apparatus: spectrophotometer, auto - biochemical - analyzer, centrifugal machine, microcamera, microimage analyzer, optical microscope.2 Methods2. 1 Manchruian Dutchmanspipe solution was made with intensity of 2g/ml. 2. 2 The division of animal: (1) control group ( n = 10) : give water with10ml/kg · d semidiurnal at first 8 weeks, and 7. 5ml/Kg · d once per day at the following 8 weeks. (2) AAN group (n =20) : give Manchurian Dutchmanspipe solution with 10 ml / Kg · d semidiurnal at first 8 weeks , and give water with 7. 5ml/Kg · d once per day at the following 8 weeks. (3 ) AAN therapy group ( n =20) : same as the AAN group at first 8 weeks; give GBE with 7.5ml/Kg · d once per day at the following 8 weeks.2. 3 Biochemical detectionThe rats were killed at 12 and 16 weeks respectively. Sulfosalicylic acid -natrium sulfuricum turbidimetry was used to detect 24 hours urine protein; hemoglobin was detected by light - electricity chromatometry; BUN and Scr was detected with auto - biochemical - instrument.2.4 Pathology detectionThe paraffin sample cutted into 3μm slide were stained with HE method.2. 5 Immunohisto - chemical staining was performed by stretavidin - biotin technique to detect ET - 1 and VWF on renal tissue.2. 6 Statistical analysisAll data appear as x ± s. SPSS 11.0 software was employed to analyze the variance with T - test between the different groups. The difference is significantwhen p<0.05.ResultsAfter twelve and sixteen weeks, mean weight level was found significantly lower in AAN group than in control group and AAN therepy group ( P <0. 01) ; Mean hemoglobin level was found significantly lower in AAN group than the other groups ( P <0. 01 ) ; Mean 24 hours urine protein excretion level was found significantly higher in AAN group ( P < 0. 05 ) , mean BUN and Scr level was found significantly higher in non - therepy group than the other two groups ( P < 0. 05).The damage of renal pathology is more serious in AANgroup than AAN therapy group; the integrated optical density of ET - 1 and VWF with Immuno-histo -chemical staining is higher than control and AAN therapy group( P <0. 01).DiscussionIn the last decade, AAN has been paid more and more attention by neph-rologists and urologists. It is popular in domestic reports that most of renal biopsies results are local interstitial fibrosis, accompanied with degradation, necrosis and shrinkage of uriniferous tubules. Recently, the role of renal peri - tubular microcirculation injury in the formation and development of AAN starts to be valued. Choi etc. reported that human chronic interstitial diseases, regardless of the initial cause, all exist peri - tubular microcirculation injury. Not only the local distributed vein lesion happens at the same part in kidney with tubulointersti-tial lesion, but also its loss is connected with renal interstitial fibrosis degree. The capillary around uriniferous tubules is the main location for microcirculation impairment in kidney, which plays a very important role in maintaining the normal construction and supplying oxygen and nourishment, and penetrating urine of uriniferous tubules.ET distributes primarily in vas endothelial cell and has four kinds of iso-mer. ET - 1 is the strongest polyuncleotide for vasconstrinct and can causing the afferent constringency. The renal vas contract and flux descent can resulting hypoxia , impairmention, activation or proliferation of tubulointerstitial cells, inducing cell factor sythesize , accelerating hypoxia factor formation and increasing the expression of Fas, which in turn cause the tubulointerstitial injury. ET - 1 can damage the vas smooth muscle cells and uriniferous tubules endothelial cells; ET - 1 can also stimulate the creation of transforming growth factor - β, and promote interstitial fibrosis, which causing vascular wall thicken and aggravating the renal anoxemia. VWF is a kind of glycoprotein with molecular weight of 2700 thousands Dalton synthesized and excreted by endothelial cells, which mainly stored in Weibel - Palade granules in endothelial cells and plays an important role in thrombosis. It can be regarded as a standardization to measure function status and damage degree of endothelial cells. The mechanism of VWF to accelerate blood coagulation and thrombosis can be understood in two ways: (1) In biological circulation VWF does not integrate with stable platelet. VWF is released into blood and integrate with endothelioid tissue and shift its structure when endothelial cells in vasals were damaged, therefore, it integrates with collagen by the way of " GPIb-VWF-Microfibrin" , the latter further integrate with activated platelet CD41 acceptor, resulting in broad platelet collection and thrombosis irreversibly. (2) VWF and VM as a compound take part in thrombosis. So the expression of VWF in interstitial endothelial cells can cause thrombosis and microcirculation obstacle easily. All these effects can influence transportation of oxygen and nourishment , resulte chronic anoxemia, and impairment in uriniferous tubules and interstitium finally. In this work, it was found that the expression of ET -1 and VWF in renal interstitium in AAN group is higher than in control group, which indicates the impairment of vas endothelial cell and microcirculation dysfunction. So we deduced that there maybe exist medicial vas endothelial injury in AAN.GBE ( Ginkgo Bilola Extract) can reduce the penetration of capillary, expand vessels, increase blood flux, improve microcirculation, eliminate oxygen -derived free radical, protect vas endothelial cells and restrain the release of inflammation. It can also compete acceptor with PAF( platelet activating factor)...
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