The Effect Of Pantoprazole On Skeletal Muscle Loss In A Mouse Model Of Cancer Cachexia

Objective To investigate the effect of pantoprazole on skeletal muscle wasting in cancer cachexia and the possible mechanism.Methods 48 male BALB/c mice were randomly divided into 6 groups,including normal control group(NC group),cancer cachexia group(CC group),lower dose pantoprazole-treated group(CL group),medium-dose pantoprazole-treated group(CM),higher dose pantoprazole-treated group(CH),and saline group(CS).The mouse colon adenocarcinoma cell line C26 was inoculated in the right forelimb of male BALB/C mice to establish a cancer cachexia model.The animals were treated with or without different concentrations of pantoprazole orally,and the body weight,tumor growth,spontaneous activity,and muscle functions were determined at various time points.Two weeks later,the levels of serum IL-6 and TNF-?,the m RNA levels of gastrocnemius JAK2 and STAT3,and the expression levels of p-JAK2,p-STAT3,Fbx32,and Mu RF1 were examined with ELISA assay,q RT-PCR assay,and Western blotting,respectively.Further studies were performed to assess the levels of Fbx32 and Mu RF1 expression,as well as morphological changes with immunohistochemistry and H&E staining.SPSS17.0 software was used to analyze the data.Results 1 Cancer cachexia model was successfully established After 5 days of tumor cell implantation,visible tumors were found in mice implanted with C26 cells.At 12 days post-implantation,the body weight of the mice implanted with the C26 cells were significant decreased compared with that of the mice in the normal control group(p<0.05).Spontaneous activity was also decreased in the C26 cell-implanted mice.Furthermore,other cachexia phenotypes,including matted hair,luster loss,and mental weakness,were observed in these groups.2 The body weight and change of the gastrocnemius muscle in mice After 5 days of tumor cell implantation,visible tumors were found in mice implanted with C26 cells.However,the differences of body weight in each group were undetectable.At 12 days post-implantation,the body weight of the mice implanted with the C26 cells were significant decreased compared with that of the mice in the normal control group(p<0.05).The body weight of the mice in the CC group was significantly lower than that of the NC group(p<0.05).The body weight of the other four groups of pantoprazole-treated mice was dose-dependent(CS<CL<CM<CH).The weight of the gastrocnemius muscle as well as the cross-sectional area of muscle fiber were substantially decreased in the CC group compared with those in the NC group(p<0.05).In the pantoprazole-treated groups,including the CL,CM,and CH group,the weight of gastrocnemius muscle and the cross-sectional area of muscle fiber were significantly increased in a dose-dependent manner.Compared with the CS group,the sectional areas of these three groups(CL,CM,and CH groups)were increased by 13.9%,39%,and 64.9%,respectively(Table 2).3 The levels of serum inflammatory factors in mice The levels of serum IL-6 and TNF-? were significantly up-regulated in the CC group compared with the NC group.Additionally,the levels of serum IL-6 and TNF-? were significantly lower in all of the pantoprazole-treated groups(CL,CM,and CH groups)than the CS groups in a dose-dependent manner.The levels of serum IL-6 were reduced by 34.1%,50.6%,and 67.6%,whereas the levels of serum TNF-? were reduced by 23.2%,41.2%,and 56.3% in the CL,CM,and CH groups,respectively 4 The change of inflammatory pathways JAK2 / STAT3 expression in mice The m RNA levels of JAK2 and STAT3 in the CC group were substantially higher than those in the NC group(p<0.05).Moreover,pantoprazole treatment significantly inhibited the m RNA levels of JAK2 and STAT3 in three pantoprazole-treated groups in a dose-dependent manner in the CL,CM,and CH groups.Notably,the expression levels of p-JAK2 and p-STAT3 were substantially higher in the CC group than those in the NC group,whereas the expression levels of p-JAK2 and p-STAT3 were remarkably lower in the three pantoprazole-treated groups than the CS group.The expression levels of p-JAK2 and p-STAT3 in the CH group were substantially lower than those in the other groups(p<0.01)5 The change of ubiquitin proteasome system in mice skeletal muscle The expression levels of Mu RF1 and Fbx32 in the gastrocnemius muscle tissue of CC group were substantially higher than those of the NC group.Similar results were also found by immunohistochemistry assay(p<0.01).Furthermore,western blot analysis and immunohistochemistry demonstrated that pantoprazole treatment can down-regulate the expression of these two proteins in the CL,CM,and CH groups in a dose-dependent mannerConclusion We successfully established a model of cancer cachexia with C26 cells.Pantoprazole can significantly alleviate cancer cachexia-induced body weight reduction and inhibit skeletal muscle wasting in a dose-dependent manner.Pantoprazole treatment can also decrease the levels of serum IL-6 and TNF-? and inhibit the activation of the JAK2/STAT3 signaling pathway in a cancer cachexia mice model.Moreover,the expression of Mu RF1 and Fbx32,which are ubiquitin proteasome-related factors,were also suppressed after pantoprazole treatment.