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Knockdown of CD44 inhibits the invasion and metastasis of hepatocellular carcinoma both in vitro and in vivo by reversing epithelial-mesenchymal transition.

Gao Y, Ruan B, Liu W, Wang J, Yang X, Zhang Z, Li X, Duan J, Zhang F, Ding R, Tao K, Dou K - Oncotarget (2015)

Bottom Line: First, we noticed that CD44 expression was associated with the mesenchymal phenotype in HCC cell lines, and knocking down CD44 with lentivirus-mediated shRNA in HCC cell lines resulted in the mesenchymal-epithelial-transition (MET) and the subsequent impaired migration and invasion in vitro.Moreover, in a metastatic mice model established by tail vein injection of luciferase labelled MHCC97-H cells, we confirmed that CD44 knockdown resulted in the decreased metastasis of HCC cells.Furthermore, we found that the induction of MET by CD44 inhibition might be achieved, at least in part, by repressing the ERK/Snail pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Hepato-Biliary and Pancreto-Splenic Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an, China.

ABSTRACT
Mounting evidence has shown that induction of epithelial-mesenchymal transition (EMT) contributes to the the expression of CSC (cancer stem cell) markers. However, whether and how CSC markers could be involved in regulating EMT has rarely been reported. CD44, being one of the most commonly used CSC markers in hepatocellular carcinoma (HCC), has been demonstrated to act as a multidomain, transmembrane platform that serves to integrate a wide variety of extracellular signals. Therefore, we determined to seek whether CD44 is necessary for the EMT process in HCC. First, we noticed that CD44 expression was associated with the mesenchymal phenotype in HCC cell lines, and knocking down CD44 with lentivirus-mediated shRNA in HCC cell lines resulted in the mesenchymal-epithelial-transition (MET) and the subsequent impaired migration and invasion in vitro. Moreover, in a metastatic mice model established by tail vein injection of luciferase labelled MHCC97-H cells, we confirmed that CD44 knockdown resulted in the decreased metastasis of HCC cells. Furthermore, we found that the induction of MET by CD44 inhibition might be achieved, at least in part, by repressing the ERK/Snail pathway.

No MeSH data available.


Related in: MedlinePlus

CD44 knockdown in luciferase-labeled MHCC97-H cells inhibited lung metastasis generated by tail vein injection in nude mice(A) qRT-PCR and (B) Western-blot analysis of CD44 expression in luciferase-labeled MHCC97-H cells that were transfected with shRNA of CD44 (KD) or NC. (C) The 10 nude mice in each group were luminescently imaged 45 days after injection (Supplemental Figure 1), and representative images for each group are shown. (D) There was a higher incidence of lung metastases in the CD44 knockdown group than in the normal control group. (E) To quantitively compare the mass of the metastasis, the photon counts per second in each group are shown (*** means p<0.001 by t-test). (F) The lungs of the nude mice from each group were removed and sectioned for evaluation lung metastasis after H&E staining (G).
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Figure 4: CD44 knockdown in luciferase-labeled MHCC97-H cells inhibited lung metastasis generated by tail vein injection in nude mice(A) qRT-PCR and (B) Western-blot analysis of CD44 expression in luciferase-labeled MHCC97-H cells that were transfected with shRNA of CD44 (KD) or NC. (C) The 10 nude mice in each group were luminescently imaged 45 days after injection (Supplemental Figure 1), and representative images for each group are shown. (D) There was a higher incidence of lung metastases in the CD44 knockdown group than in the normal control group. (E) To quantitively compare the mass of the metastasis, the photon counts per second in each group are shown (*** means p<0.001 by t-test). (F) The lungs of the nude mice from each group were removed and sectioned for evaluation lung metastasis after H&E staining (G).

Mentions: We transfected the luciferase-labeled MHCC97-H cells with lentivirus containing shRNA of CD44 and normal control as for SMMC-7721 and MHCC97-H cells. The knockdown efficiency of CD44 mRNA was 84.03% (Fig. 4A), which was verified by Western blot (Fig. 4B). The bioluminescent imaging of nude mice 45 days after injection showed that the incidence of lung metastasis was lower in mice injected with MHCC97-H-luc-KD compared with MHCC97-H-luc-NC (Fig. 4D) and that the bioluminescence from MHCC97-H-luc-NC metastases was stronger than that of MHCC97-H-luc-KD (Fig. 4C, E), revealing that the knockdown of CD44 in MHCC97-H-luc cells inhibited lung metastasis in nude mice.


Knockdown of CD44 inhibits the invasion and metastasis of hepatocellular carcinoma both in vitro and in vivo by reversing epithelial-mesenchymal transition.

Gao Y, Ruan B, Liu W, Wang J, Yang X, Zhang Z, Li X, Duan J, Zhang F, Ding R, Tao K, Dou K - Oncotarget (2015)

CD44 knockdown in luciferase-labeled MHCC97-H cells inhibited lung metastasis generated by tail vein injection in nude mice(A) qRT-PCR and (B) Western-blot analysis of CD44 expression in luciferase-labeled MHCC97-H cells that were transfected with shRNA of CD44 (KD) or NC. (C) The 10 nude mice in each group were luminescently imaged 45 days after injection (Supplemental Figure 1), and representative images for each group are shown. (D) There was a higher incidence of lung metastases in the CD44 knockdown group than in the normal control group. (E) To quantitively compare the mass of the metastasis, the photon counts per second in each group are shown (*** means p<0.001 by t-test). (F) The lungs of the nude mice from each group were removed and sectioned for evaluation lung metastasis after H&E staining (G).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4480719&req=5

Figure 4: CD44 knockdown in luciferase-labeled MHCC97-H cells inhibited lung metastasis generated by tail vein injection in nude mice(A) qRT-PCR and (B) Western-blot analysis of CD44 expression in luciferase-labeled MHCC97-H cells that were transfected with shRNA of CD44 (KD) or NC. (C) The 10 nude mice in each group were luminescently imaged 45 days after injection (Supplemental Figure 1), and representative images for each group are shown. (D) There was a higher incidence of lung metastases in the CD44 knockdown group than in the normal control group. (E) To quantitively compare the mass of the metastasis, the photon counts per second in each group are shown (*** means p<0.001 by t-test). (F) The lungs of the nude mice from each group were removed and sectioned for evaluation lung metastasis after H&E staining (G).
Mentions: We transfected the luciferase-labeled MHCC97-H cells with lentivirus containing shRNA of CD44 and normal control as for SMMC-7721 and MHCC97-H cells. The knockdown efficiency of CD44 mRNA was 84.03% (Fig. 4A), which was verified by Western blot (Fig. 4B). The bioluminescent imaging of nude mice 45 days after injection showed that the incidence of lung metastasis was lower in mice injected with MHCC97-H-luc-KD compared with MHCC97-H-luc-NC (Fig. 4D) and that the bioluminescence from MHCC97-H-luc-NC metastases was stronger than that of MHCC97-H-luc-KD (Fig. 4C, E), revealing that the knockdown of CD44 in MHCC97-H-luc cells inhibited lung metastasis in nude mice.

Bottom Line: First, we noticed that CD44 expression was associated with the mesenchymal phenotype in HCC cell lines, and knocking down CD44 with lentivirus-mediated shRNA in HCC cell lines resulted in the mesenchymal-epithelial-transition (MET) and the subsequent impaired migration and invasion in vitro.Moreover, in a metastatic mice model established by tail vein injection of luciferase labelled MHCC97-H cells, we confirmed that CD44 knockdown resulted in the decreased metastasis of HCC cells.Furthermore, we found that the induction of MET by CD44 inhibition might be achieved, at least in part, by repressing the ERK/Snail pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Hepato-Biliary and Pancreto-Splenic Surgery, Xijing Hospital, The Fourth Military Medical University, Xi'an, China.

ABSTRACT
Mounting evidence has shown that induction of epithelial-mesenchymal transition (EMT) contributes to the the expression of CSC (cancer stem cell) markers. However, whether and how CSC markers could be involved in regulating EMT has rarely been reported. CD44, being one of the most commonly used CSC markers in hepatocellular carcinoma (HCC), has been demonstrated to act as a multidomain, transmembrane platform that serves to integrate a wide variety of extracellular signals. Therefore, we determined to seek whether CD44 is necessary for the EMT process in HCC. First, we noticed that CD44 expression was associated with the mesenchymal phenotype in HCC cell lines, and knocking down CD44 with lentivirus-mediated shRNA in HCC cell lines resulted in the mesenchymal-epithelial-transition (MET) and the subsequent impaired migration and invasion in vitro. Moreover, in a metastatic mice model established by tail vein injection of luciferase labelled MHCC97-H cells, we confirmed that CD44 knockdown resulted in the decreased metastasis of HCC cells. Furthermore, we found that the induction of MET by CD44 inhibition might be achieved, at least in part, by repressing the ERK/Snail pathway.

No MeSH data available.


Related in: MedlinePlus