| Objective To investigate the expression of CHI3L1 in gastric cancer tissues and patients' serum,and to analyze the correlation between the expression of CHI3L1 and the clinicopathological parameters of patients,and the influence on the prognosis of patients.Methods Western Blot(WB)was used to detect the expression of CHI3L1 in gastric cancer tissues and paracancerous tissues from 5 patients.Then,the expression of CHI3L1 in gastric cancer tissue microarray was detected by immunohistochemistry(IHC),and the expression of CHI3L1 in serum of patients with gastric cancer and normal controls was detected by Enzyme-Linked Immunosorbent Assay(ELISA).Results We found that in 5 cases of gastric cancer,the expression of CHI3L1 in the cancer tissues was significantly higher than that in the adjacent tissues.Consistent with WB results,immunohistochemistry results showed that the expression level of CHI3L1 in cancer tissues was higher than that in adjacent tissues,and CHI3L1 was significantly higher in cancer tissues in metastases.We further analyzed the relationship between clinicopathological parameters and the expression of CHI3L1 in 100 cases of gastric cancer and found that the expression level of CHI3L1 was related to the depth of tumor invasion,lymph node metastasis,and tumor stage.Prognostic analysis showed that patients with high expression of CHI3L1 had poorer prognosis than patients with low expression.Stratified analysis showed that in patients with T3-T4,N2-N3,and III-IV,the overall survival was significantly shorter in patients with high CHI3L1 expression.Serological results showed that the expression of CHI3L1 in gastric cancer patients was higher than that in the normal control group,and the serum CHI3L1 level was positively correlated with the degree of malignancy of the tumor.Conclusion The expression of CHI3L1 in patients with gastric cancer is significantly related to the clinicopathological parameters of patients,and its expression has an impact on the prognosis of patients.Objective To investigate which receptors CHI3L1 exerts its biological functions,and to further investigate whether IL-13R?2 regulates its downstream signaling pathways through co-receptors with CD44.Methods The co-immunoprecipitation(Co-IP)method was used to detect whether CHI3L1 interacted with CD44.Then immunofluorescence co-localization analysis was used to detect the cell membrane of AGS and MGC803 cells.CHI3L1 Co-localization with CD44.To confirm the above findings,we further applied immunofluorescence colocalization to analyze whether CHI3L1 interacts with CD44 in gastric cancer tissues.Similarly,we used Co-IP and immunofluorescence co-localization methods to detect whether IL-13R? interacts with CD44.In order to verify whether the CHI3L1-IL-13R?2/CD44 signaling axis can regulate the proliferation and metastasis of tumor cells,we designed a signal-axis-enhanced cell function experiment for this signaling axis to enhance its signaling pathway and inhibit its signal transduction.The CHI3L1-IL-13R?2/CD44 signaling axis was confirmed to function in vitro.RESULTS: In gastric cancer AGS and MGC803 cells,we found that CHI3L1 was able to pull down CD44 by co-immunoprecipitation.Corresponding to this,CHI3L1 can also be pulled down by CD44 antibody,suggesting that there is an interaction between CHI3L1 and CD44 in gastric cancer cells.To confirm the above findings,immunofluorescence co-localization analysis showed that CHI3L1 and CD44 can co-localize on the surface of tumor cell membranes in AGS and MGC803 cells.The immunofluorescence co-localization of gastric cancer tissues showed that there was also a co-localization between the two.Similarly,by co-immunoprecipitation and immunofluorescence colocalization analysis,we also confirmed that IL-13R?2 and CD44 can interact to form a co-receptor.The proliferation assay(CCK8)showed that CHI3L1 stimulated cell proliferation in a concentration-dependent manner.We have found that the use of CD44 neutralizing antibodies or CD44 KO can significantly inhibit CHI3L1-induced tumor proliferation and invasion.CD44 neutralizing antibodies are also capable of inhibiting epithelial-mesenchymal transition(EMT)of CHI3L1-induced tumor cells.Conclusion Our study revealed for the first time that a new receptor for CHI3L1 is CD44,and CHI3L1 exerts its biological function via CD44 receptor.On the other hand,IL-13R?2 can form a co-receptor with CD44.The CHI3L1-IL-13R?2/CD44 signaling axis can promote the proliferation and metastasis of gastric cancer cells.Objective To investigate whether the CHI3L1-IL-13R?2/CD44 axis regulates which signals play a role in promoting tumor proliferation and metastasis,and further confirms that CHI3L1 or IL-13R?2 interacts with that CD44-cleaved body.To investigate whether the CHI3L1-IL-13R?2/CD44 signal axis functions in vivo.Methods The use of CD44 neutralizing antibodies in gastric cancer cells can block the activation of PKB/AKT signaling pathway,MAPK/ERK signaling pathway,and Wnt/?-catenin signaling pathway induced by CHI3L1.Whether sh RNA knocks down CD44 of gastric cancer cells can inhibit CHI3L1-induced signal pathway activation.In CD44 KO mice,mouse bone marrow-derived macrophages(BMDM)were stimulated with mouse CHI3L1 to observe the activation of the PKB/AKT signaling pathway and the MAPK/ERK signaling pathway.Biolayer Interferometry(BLI)was used to detect the affinity between CHI3L1 and CD44.And the affinity between IL-13R?2 and CD44.Results Our study showed that CD44 neutralizing antibodies can significantly inhibit PK3/AKT-induced PKB/AKT signaling,MAPK signaling,and Wnt/?-catenin signaling pathways.Similarly,knockdown of CD44 by sh RNA also inhibits PKB/ AKT signaling pathway,MAPK signaling pathway.Similarly,in CD44 knockout mice,CHI3L1 could not induce the activation of PKB/AKT signaling pathway and MAPK signaling pathway in BMDM cells.Using Biolayer Interferometry,we found that CHI3L1 binds to CD44v3 and IL-13R?2 also binds to CD44v3.In the mouse xenograft tumor model,CHI3L1 was knocked out from the tumor cells.Compared with the control group,knockout of CHI3L1 significantly inhibited tumor proliferation.In a mouse metastatic tumor model,we found that knocking out CD44 on the surface of tumor cells can reduce lung metastases in gastric cancer.In the melanoma metastasis model,we found that compared with the control mice,the number of lung metastases was significantly reduced in the CD44 KO group,and the total TGF-?1 and activated TGF-?1 levels in the alveolar lavage fluid were lower than those in the control group.Conclusion The results of this study suggest that CHI3L1 interacts with CD44v3.On the other hand,IL-13R?2 also forms a co-receptor with CD44v3.The CHI3L1-IL-13R?2/CD44 signaling axis also controls the proliferation and metastasis of gastric cancer cells in vivo. |
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