The Experimental Study Of Expression Of CD14 And NF-κBp65 In Septic Immature Rats With Acute Kidney Injury And The Effect Of Ulinastatin On Its Intervention

Objectives:1. To observe changes in renal function(concentration of Scr,BUN),and kidney tissue injury in septic immature rats after the CLP operation by establishing the model of septic immature rats in the method of cecal ligation and puncture(CLP).2. To study the effect of ulinastatin on serum tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) level,and the expression of renal CD14 and NF-κBp65 protein in septic rats with AKI,and to explore its possible protection and mechanism. Methods:1. Grouping:Totally 224 healthy male Sprague-Dawley(SD) rats of clean 4-week-old were randomly divided into four groups:the normal control group(group A,n=32),the sham group(group B,n=32),the CLP group(group C,n=80) and the UTI treatment group(group D,n=80),each group was further randomly divided into four time points at 6h,12 h,24h,48 h.2. Model building and intervention:group A:no celiotomy and other proessing; group B:celiotomy but not ligation cecum and puncture;group C: cecal ligation and puncture;group D:The rats were injected with UTI(50mg/kg) via the tail vein at 0h,24 h after the group D operated and the rats in the other three groups were treated with equal physiological saline.The rats were separately narcotized in batches at 6h,12 h,24h or 48 h after the operation and blood and renal tissues samples were collected.3. Detection index:The concentrations of BUN and Scr were detected with automatic biochemical analyzer;the contents of serum TNF-α and IL-6 were measured by enzyme linked immunosorbent assay;the expression of CD14 m RNA in renal tissue was examined with reverse transcription-polymerase chain reaction(RT-PCR) way;the expression of renal CD14 and NF-κBp65 were investigated by immunohistochemical staining,meanwhile,the pathological changes of renal tissue were observed under optical microscope after HE staining. Results:1. The concentrations of Scr:Compared with the group A and the group B,the concentrations of Scr increased significantly at different time points(except 6 h) in the group C and the group D(P<0.05), it increased at 12 h, and continued to go up until 48 h. And the concentrations of Scr in the group D were lower than those in the group C at each time point(except 6h), and the differences were statistically significant(P<0.01).2. The concentrations of BUN:Compared with the group A and the group B,the concentrations of BUN increased significantly at different time points in the group C and the group D(P<0.01), it increased at 6h, and continued to go up until 48 h. And the concentrations of BUN in the group D were lower than those in the group C at each time point, and the differences were statistically significant(P<0.01).3. The pathological changes of renal tissue:Compared with the group A and the group B,there were a few renal capsule cavities expansion in kidney tissue of group C at 6h, significant expansion of renal capsule cavities could be seen, and significant swelling, degeneration of tubular epithelial cells seen also,part of the glomerulus blood stasis,infiltration of inflammatory cells were visible in renal tissue at12 h,24h,48h; Compared with group C,above pathological changes in renal tissue of group D was significantly reduced at different time points.4. The levels of serum TNF-α and IL-6:Compared with the group A and the group B,the levels of TNF-α and IL-6 increased significantly at different time points in the group C and the group D(P<0.01), it increased at 6h, and continued to go up until 12 h,and it slow fell at 24 h,48h,but were still higher than those in the group A and the group B. And the levels of serum TNF-α and IL-6 in the group D were significantly lower than those in the group C at each time point(P<0.01).5. The expressions of renal tissue NF-κBp65 protein:Compared with the group A and the group B,the expressions of renal tissue NF-κBp65 protein increased significantly at different time points in the group C and the group D(P<0.01),it began to ascend at 6h,and increase gradually at12 h,24h,48 h.And The expressions of renal tissue NF-κBp65 protein in the group D were significantly lower than those in the group C at each time point(P<0.01).6. The expressions of renal tissue CD14 m RNA and CD14 protein:Compared with the group A and the group B,the expressions of renal tissue CD14 m RNA and CD14 protein increased significantly at different time points in the group C and the group D(P<0.01),it began to rise at 6h,reached the peak at 12 h, and it slow dropped off at 24 h,48h,but were still higher than those in the group A and the group B.And the expressions of renal tissue CD14 m RNA and CD14 protein in the group D were lower than those in the group C at each time point(P<0.05). Conclusions:1. The model of septic immature rats is stable and reliable, which was made by cecal ligation and puncture.2. Kidney tissue injury and renal dysfunction can appear early in septic immature rats, to a certain extent.3. Ulinastatin play a protective effect on septic immature rats with acute kidney injury by reducing the expression of renal CD14 and NF-κBp65 in LPS receptor-mediated signal transduction pathway and NF-κB signaling pathway,thereby inhibiting inflammatory factors,such as TNF-α and IL-6,and alleviating inflammation-induced renal tissue injury.