The Possible Effect And Mechanism Of Transforming Growth Factor Beta Activator Kinase 1 Inhibitor In Diabetic Db/db Mice

Objective: To investigate the possible effect and mechanism of transforming growth factor beta activator kinase 1(TAK1) inhibitor 5Z-7-oxozeaenol(OZ) in diabetic db/db mice,the changes of general indicators in mice were detected. PAS stain and transmission electron microscope were used to observe the pathological changes of renal tissues. In addition, immunohistochemistry、 Western blot were used to detect the expression of inflammation related indicators in renal tissues. Methods:12 healthy male WT mice and 24 healthy male db/db mice were randomly divided into three groups: 1.the control group(the WT group), comprising of 12 WT mice; 2.the model group(the db/db group), comprising of 12 db/db mice; 3.the inhibitor group( the db/db+ OZ group), comprising of 12 db/db mice, OZ 2mg/kg was administrated by intraperitoneal injection every other day. After 8 and 12 weeks of 5Z-7-oxozeaenol treatment, blood glucose, body weight, kidney weight and 24 h urinary albumin excretion rate of mice in each group were measured. Light microscopy and electron microscopy were used to observe the pathological changes of kidney tissues. Immunohistochemistry was used to detect the expression of inflammation related factors, such as NF-κBp65, MCP-1, TNF-α and TGF-β1. Western blot was used to detect the expression of p-TAK1, TAB1 and IL-1β. Results 1. The db/db mice gradually appeared the diabetic symptoms of polydipsia, polyphagia, polyuria and loss of body weight, while the symptoms of mice in db/db+OZ group reduced compared with the db/db mice group. 2. Compared with the WT mice, the levels of blood glucose, body weight, kidney weight and UAER were much higher in db/db mice group, and the difference was statistically significant(P<0.01). Meanwhile, the body weight, kidney weight and UAER levels were significantly decreased in db/db+OZ group compared with the db/db mice and the difference was statistically significant as well(P<0.05, P<0.01). 3. PAS staining shows that the strong positive range in week 8 and 12 WT mice was relatively less, while the PAS strong positive range of db/db mice was dramatically increased compared with the control group. In addition, the PAS strong positive range of db/db+OZ mice was reduced compared with the db/db mice. 4. Electron microscopic showed that glomerular basement membrane in week 8 and 12 WT mice was homogeneous and there was no electron dense deposits, and the podocyticprocess distributed homogeneous with no fusion. In the 8th week, the glomerular mesangial cell and mesangial matrix of db/db mice were increased, the glomerular basement membrane thickened with local high upwarping, the adjacent podocyticprocess was fusion or loss. These pathological lesions were aggravated in the 12 th week. On the other hand, the pathological lesions in kidney tissues were positively improved by TAK1 inhibitor. 5. Immunohistochemistry showed that NF-κB p65, MCP-1, TNF-α,TGF-β1 expression significantly increased in the kidney tissues of week 8 and 12 db/db mice compared with WT mice, and the difference was statistically significant(P<0.05). While the expression significantly reduced in db/db+OZ mice compared with db/db mice, the difference was statistically significant(P<0.05). 6. Western blot showed that p-TAK1, TAB1 and IL-1β expression levels were higher in week 8 and 12 db/db mice compared with WT mice(P<0.05) and lower in db/db+OZ group compared with db/db mice(P<0.05). Conclusions 1. After TAK1 inhibitor injection,the expression of NF-κB p65, p-TAK1, TAB1 was down regulated and the expression of MCP-1, TGF-β1, TNF-α, IL-1β significantly decreased in the kidney tissues. 2. TAK1 inhibitors could down regulate the NF-κB signaling pathway by inhibit the expression of TAK1. Then the expression of inflammatory cytokines reduced, the inflammatory reaction was inhibited and the kidney damage was improved.