Renshen Baidusan Combined with Yurui Enema Treats Intestinal Mucosal Injury in Rat Model of Ulcerative Colitis via PI3K/Akt/NF-κB Pathway / 中国实验方剂学杂志

ABSTRACT

ObjectiveTo explore the mechanisms of internal treatment （Renshen Baidusan）， external treatment （Yurui Enema）， and combination of the two methods in treating intestinal mucosal injury in the rat model of ulcerative colitis （UC） from the changes of phosphatidylinositol-3 kinase （PI3K）/protein kinase B （Akt）/nuclear factor-κB （NF-κB） pathway. MethodFifty SPF-grade SD rats were randomized into blank， model， Renshen Baidusan （15.6 g·kg-1）， Yurui Enema （25 g·kg-1）， and combined treatment （15.6 g·kg-1 Renshen Baidusan + 25 g·kg-1 Yurui Enema） groups （n=10）. The rat model of UC was established in other groups except the blank group by 2，4， 6-trinitrosulfonic acid （TNBS）/ethanol. The rats were administered with corresponding drugs once a day for 14 consecutive days since the 8th day after modeling. The histopathological changes of colon were observed by hematoxylin-eosin （HE） staining. Enzyme-linked immunosorbent assay （ELISA） was employed to measure the levels of tumor necrosis factor （TNF）-α， interferon （IFN）-γ， interleukin （IL）-4， and IL-10 in the colon tissue. The apoptosis of colon epithelial cells was detected by terminal deoxynucleotidyl transferase-mediated nick end labeling （TUNEL）. The location and expression of B-cell lymphoma-2 （Bcl-2）， Bcl-2-associated X protein （Bax）， TNF-α， and IL-6 in the colon tissue were examined by immunohistochemistry. Real-time quantitative fluorescence polymerase chain reaction （Real-time PCR） and Western blot were employed to determine the mRNA and protein levels， respectively， of the proteins in the PI3K/Akt/NF-κB pathway in the colon tissue. ResultIn the model group， HE staining showed a large number of inflammatory cell infiltration in the mucosa and submucosa. Compared with the blank group， the model group showed elevated levels of TNF-α and IFN-γ and lowered levels of IL-4 and IL-10 in the colon tissue， increased apoptosis rate of colon epithelial cells， increased positive expression of Bax， TNF-α， and IL-6， and decreased positive expression of Bcl-2 （P<0.05）. Moreover， the model group showed up-regulated mRNA levels of PI3K， Akt， and NF-κB and protein levels of PI3K， p-PI3K， Akt， p-Akt， p65， p-p65， Bax， and cleaved Caspase-3， increased Bax/Bcl-2 and cleaved Caspase-3/Caspase-3 ratios， and down-regulated protein levels of NF-κB suppressor protein α（IκBα）， Bcl-2， and Caspase-3 in the colon tissue （P<0.05）. Compared with the model group， the internal treatment， the external treatment， and the combination （referred to as the three groups） alleviated the colonic mucosal injury， lowered the levels of TNF-α and IFN-γ and elevated the levels of IL-4 and IL-10 in the colon tissue， decreased the apoptosis rate of colon cells， inhibited the positive expression of Bax， TNF-α， and IL-6， and promoted the positive expression of Bcl-2 （P<0.05）. Furthermore， the combination group down-regulated the mRNA level of PI3K （P<0.05）. The three groups down-regulated the mRNA levels of Akt and NF-κB and the protein levels of p-PI3K， Akt， p-Akt， p65， p-p65， Bax， and cleaved Caspase-3 in the colon tissue， decreased the Bax/Bcl-2 and cleaved Caspase-3/Caspase-3 ratios， and up-regulated the protein levels of IκBα， Bcl-2， and Caspase-3 （P<0.05）. ConclusionRenshen Baidusan， Yurui Enema， and their combination may inhibit the activation of PI3K/Akt/NF-κB signaling pathway and regulate the expression of genes and proteins related to this pathway to achieve anti-inflammatory and anti-apoptotic effects， thus restoring the intestinal mucosal barrier function of UC rats.