1,25-（OH）₂D₃on The Interstitial Infiltration Of Mononuclear/m Acrophages In Renal Interstitial Fibrosis Of UUO Rats

Objective: We choosed the unilateral ureteral obstruction of rats asthe model of renal interstitial fibrosis（RIF）, and detected the effect of1,25-（OH）₂D₃on interstitial infiltration of mononuclear/macrophages（ED-1positive cells）、the serum TNF-α Concentration and the mRNA and protein expression of monocyte chemoattractant protein-1（MCP-1）, matrixmetalloproteinase-9（MMP-9）, Tissue-Inhibitor of Metalloproteinase-1（TIMP-1） in rats＇ RIF after UUO. Therefore this study is to explore whether1,25-（OH）₂D₃exerted a role to mononuclear/macrophages in vivo and provide a new method and academic foundation for blocking RIF.Methods:1Fifty male Sprague-Dawley rats were randomly assigned to fivegroups, each group had10animals:â‘ UUO+high dose1,25-（OH）₂D₃,â‘¡UUO+low dose1,25-（OH）₂D₃â‘¢UUO+Benazopril groupâ‘£UUO operation group⑤sham operated group.1,25-（OH）₂D₃was administered daily at a dose6ng/100g、3ng/100g dissolved in olive oil1ml in high1,25-（OH）2D3and low treatment groups, benazopril was administered daily ata dose0.17mg/100g dissolved in NS1ml after UUO and daily for4weeks, Meanwhile, the group①＆②were lavaged daily in1ml NS and the groupâ‘¢was lavaged daily in1ml olive oil; the group④＆⑤wereadministered daily in1ml olive oil and NS.2Then the rats were killed and the sample blood were collected to detected the concentration of the serum TNF-α Concentration、the obstructed kidney was harvested to observe typeⅠ、Ⅲ collagen of renal interstitial by Sirius red staining（SRS） and the infiltration of Mononuclear/macrophages by immunohistochemistry（IHC） and the expression of MCP-1、MMP-9、TIMP-1by RT-PCR and western blot, the enumeration dat a was recorded asχ±SD.Results:1The Sirius red staining analysis of CVF in different groups of rats.There is little collagen in renal interstitial of the sham operated group, the CVF in UUO operation group was significantly more than the sham operated group（P＜0.05）. In the UUO+1,25-（OH）₂D₃treatment group,the CVF was significantly less than the UUO operation group. Therewas no significant difference betweenUUO+1,25-（OH）₂D₃treatment group and UUO+Benazopril group（P＞0.05）.2The renal infiltration of mononuclear/macrophages（ED-1positive cells）in different groups of rats.There are few ED-1positive cells in renal interstitial of the sham operated group, the number of ED-1positive cells in UUO operation group was significantly more than the sham operated group（P＜0.05）, which was mainly seen in the renal interstitial with focal or diffuse distribution. In the UUO+1,25-（OH）₂D₃treatment group the number of ED-1positive cells was significantly less than the UUO operation group. Therewas no significant difference betweenUUO+1,25-（OH）₂D₃treatment group and UUO+Benazopril group（P＞0.05）.3The serum TNF-α concentration in different groups of rats.The serum TNF-α concentration in UUO operation group was significantly more than the sham operated group（P＜0.05）. In the all medicine treatment group the serum TNF-α concentration was significantly lessthan the UUO operation group（P＜0.05）. There were no significant difference between UUO+high dose1,25-（OH）₂D₃treatment group and UUO+Benazopril group（P＞0.05）.4The expression of MCP-1、MMP-9、TIMP-1mRNA in different groups rats4.1MCP-1、TIMP-1: The expression of MCP-1、TIMP-1mRNA inUUO operation group significantly increased what compared with shamoperated group（P＜0.05）, and also significantly increased what compared with UUO+1,25-（OH）₂D₃/Benazopril treatment group（P＜0.05）.4.2MMP-9: The expression of MMP-9mRNA in UUO+1,25-（OH）2D3/Benazopril treatment group significantly increased what compared with UUO operation group, the differences of MMP-9mRNA between UUO operation group and sham operated group have statistically significant（P＜0.05）.There were no significant difference in the expression of MCP-1、MMP-9、TIMP-1mRNA between UUO+high dose1,25-（OH）₂D₃treatment group and UUO+Benazopril group（P＞0.05）, and were basal expression of MCP-1、MMP-9、TIMP-1mRNA in sham operated group.5The protein expression of MCP-1、MMP-9、TIMP-1in different groups rats5.1MCP-1、TIMP-1: The protein expression of MCP-1、TIMP-1inUUO operation group significantly increased what compared with shamoperated group（P＜0.05）, and also significantly increased what compared with UUO+1,25-（OH）₂D₃/Benazopril treatment group（P＜0.05）.5.2MMP-9: The protein expression of MMP-9in UUO operationgroup was significantly higher than what in sham operated group（P＜0.05）, the UUO+1,25-（OH）₂D₃/Benazopril treatment group significantly increased what compared with UUO operation group（P＜0.05）.There were no significant difference in the protein expression of MCP-1、MMP-9、TIMP-1between UUO+high dose1,25-（OH）₂D₃treatment group and UUO+Benazopril group（P＞0.05）, and were basal proteinexpression of MCP-1、MMP-9、TIMP-1in sham operated group.Conclusions:11,25-（OH）₂D₃can inhibit the infiltration and activation of mononuclear/macrophages in rat renal interstitial after UUO by reducing the expression of MCP-1in kidney.21,25-（OH）₂D₃can protect to the rats＇ kideny through lowering the serum TNF-α concentration of rats after UUO, and then reduce stimulating effect of TNF-α on intrinsic renal cells and inflammatory cells. 31,25-（OH）₂D₃can alleviate the imbalance of MMP-9/TIMP-1in rats’ kidney after UUO by escalating the expression of MMP-9and by lowering TIMP-1of the kidney, then promote the degradation of ECM and reduce renal interstitial fibrosis.4The effects of1,25-（OH）₂D₃on alleviating the pathological changes of renal interstitial fibrosis in UUO rats were similar to Benazepril.