The Mechanism Of Adenosine Signaling Mediated Renal Interstitial Fibrosis

Purpose:To observe the influence of the ADO signal pathway on the renal interstitial fibrosis.Methods:(1) Animals grouping:44male mice were randomly divided into3groups, including sham-operation group (n=12), model group (n=16) and intervention group (n=16). The intervention group was given theophylline and10mg/kg·d intraperitoneal injection (once/day) on the basis of the unilateral ureteral ligation and the model group was given the normal saline intraperitoneal injection based on the UUO. The mice were sacrificed respectively in the1st,3rd,7th and14th days of the experiment, among which three mice were killed in the sham-operation group at each time, and four mice were respectively killed in the model group and the intervention group at each time. The mice were put into the metabolism cage within24hours before being killed to collect the urine so that the content of the urine creatinine and the β2-microglobulin could be determined; the hypoxia probe was injected into the mouse penis vein within1.5hours before being killed. After the mice were killed, the renal tissues of the ligation side were taken for chemical study of the morphology and the immunologic tissue.(2) Observation target:After the renal tissue was stained by HE and Masson, it was observed for the renal tubular injury, the renal interstitial inflammatory cell infiltration and the interstitial fibrosis degree of the each experimental group; the immunohistochemistry technique is used to observe the epithelial cells of the renal tubular injury, proliferation cell nuclear antigen expression of the interstitial cells and the hypoxia degree and distribution of the renal tissues and the HPLC is adopted to detect the adenosine level in the renal tissue.Results:(1) Renal tissues was in the continued hypoxia status after UUO operation and the ADO level in the renal tissues was notably higher than that in the sham-operation group;(2) Broad renal tubular injury occurred after UUO and the8-PT had a protective effect on the renal tubular injury;(3) PCNA positive expression along with the time progress mainly focused on renal interstitial part in the renal interstitial fibrosis process, especially the UUO group, while the renal interstitial cells remained in the high proliferative status;(4) the renal interstitial inflammatory cell infiltration of the model group increased progressively along with the disease course progress and its inflammatory cell infiltration decreased significantly after8-PT intervention;(5) the renal interstitial fibrosis degree of the intervention group was significantly lower than that of the model group in the same period.Conclusion:(1) Hypoxia status of renal tissues exists in the renal interstitial fibrosis process and increases along with ADO level in the tissues;(2) Blocking the ADO signal pathway could restrain inflammatory cell infiltration, renal interstitial cell proliferation and fibrosis, and also has a protective effect on the renal function. Purpose:To observe the relationship between dynamic changes of the fibrogenic cytokines leading to fibrosis and the ADO in the renal interstitial fibrosis process, and to further elucidate the renal interstitial fibrosis mechanism and ADO molecular mechanism leading to the fibrosis.Methods:The RT-PCR is used to dynamically observe the TGF-β1, PAI-1and α1(I) procollagen mRNA expression of the renal tissues in the1st, the3rd, the7th and the14th days after operation; immunohistochemical method is adopted to observe the protein expression and distribution regulation of TGF-β1and a-SMA of renal tissues of each experimental group.Results:(1) The expression of renal tissue cytokines of the mice in the model group raised up and progressively increased along with the extension of the time.(2) The blocking-up of the ADO pathway had a significant inhibition function on the expression of the renal tissue cytokines.(3) The expressions of TGF-β1and a-SMA protein levels of the mouse renal tissue in the model group were significantly up-regulated in various experimental periods; after the ADO pathway was blocked-up, these cytokines’ expressions were obviously inhibited. Conclusion:(1) adenosine signal pathway interferes the progress of the renal interstitial fibrosis by adjusting the cytokines expression.(2) ADO signal pathway could regulate activation and transdifferentiation process of the fibroblast. Purpose:To study the biological behaviour change of the mouse renal fibroblast in different intervention condition.Methods:For the mouse renal fibroblast strain NIH3T3, the TaqMan probe method is used to identify the adenosine receptor type of the fibroblast surface. Four groups were divided:control group; NECA group (NECA); PT group (NECA+8-PT); MRS group (NECA+MRS1754). It was dynamically observed for TGF-β1, α1(Ⅰ)procollagen and α-SMA mRNA expression levels in the cells after the1st, the2nd and the3rd days of the drug intervention. The MMT methods are used to determine the proliferation condition above four groups of cells after being experimented for0,12,24,48and72hours.Results:The main receptor type of the renal fibroblast surface was A2BR;(2) NECA stimulation could up-regulate TGF-β1,α1(Ⅰ)procollagen and a-SMA mRNA expressions, and8-PT and MRS1754could significantly restrain TGF-β1,α1(Ⅰ)procollagen and α-SMA mRNA expressions;(3) NECA could promote the fibroblast proliferation, and8-PT and MRS1754could restrain the fibroblast proliferation.Conclusion:(1) Adenosine up-regulates pro-fibrogenic cytokines and cell proliferation activation in the cells through A2BR inducing on the fibroblast surface, leading to fibrosis occurrence and progress;(2) A2BR blocking agent could effectively restrain proliferation and activation of the fibroblast and the generation of the pro-fibrogenic cytokines.