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Endothelial cells promote stem-like phenotype of glioma cells through activating the Hedgehog pathway.

Yan GN, Yang L, Lv YF, Shi Y, Shen LL, Yao XH, Guo QN, Zhang P, Cui YH, Zhang X, Bian XW, Guo DY - J. Pathol. (2014)

Bottom Line: Microenvironmental regulation of cancer stem cells (CSCs) strongly influences the onset and spread of cancer.We observed that CD133(+) GSCs were located closely to Shh(+) endothelial cells in specimens of human glioblastoma multiforme (GBM).Knockdown of Smo in glioma cells led to a significant reduction of their CSC-like phenotype formation in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pathology and Southwest Cancer Centre, Southwest Hospital, Third Military Medical University and Key Laboratory of Tumour Immunopathology, Ministry of Education of China, Chongqing, 400038, People's Republic of China.

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Endothelial cells promote tumour sphere formation and CD133 expression by glioma cells in vitro and tumour growth in vivo. (A) Tumour spheres formed by GL261 cells cultured with b.END3 cells (left; scale bar = 100 µm) or by U251 cells cultured with HUVECs (right; scale bar = 300 µm) were larger in size than those formed by GL261, or U251 cells alone, respectively. (B) Efficiency of tumour sphere formation by GL261 cells cultured with b.END3 cells or by U251 cells cultured with HUVECs was higher than that by GL261 or U251 cells alone, respectively (*p < 0.05). (C) The numbers of CD133+ GL261 (left) or U251 cells (right) were significantly increased after co-culture with b.END3 cells or HUVECs, respectively (*p < 0.05). (D) Survival times of mice that were orthotopically co-injected with GL261 and b.END3 cells were shorter than those of mice orthotopically injected with GL261 cells alone (*p < 0.05). (E) Tumour volume of orthotopic allografts generated by co-injection of GL261 with b.END3 cells was significantly larger than that of orthotopic allografts generated by injection of GL261 cells alone (*p < 0.05).
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fig02: Endothelial cells promote tumour sphere formation and CD133 expression by glioma cells in vitro and tumour growth in vivo. (A) Tumour spheres formed by GL261 cells cultured with b.END3 cells (left; scale bar = 100 µm) or by U251 cells cultured with HUVECs (right; scale bar = 300 µm) were larger in size than those formed by GL261, or U251 cells alone, respectively. (B) Efficiency of tumour sphere formation by GL261 cells cultured with b.END3 cells or by U251 cells cultured with HUVECs was higher than that by GL261 or U251 cells alone, respectively (*p < 0.05). (C) The numbers of CD133+ GL261 (left) or U251 cells (right) were significantly increased after co-culture with b.END3 cells or HUVECs, respectively (*p < 0.05). (D) Survival times of mice that were orthotopically co-injected with GL261 and b.END3 cells were shorter than those of mice orthotopically injected with GL261 cells alone (*p < 0.05). (E) Tumour volume of orthotopic allografts generated by co-injection of GL261 with b.END3 cells was significantly larger than that of orthotopic allografts generated by injection of GL261 cells alone (*p < 0.05).

Mentions: To explore the mechanism underlying the interaction between GSCs and ECs, we utilized transwell co-culture. Both GL261 and U251 cells were tested for their ability to form tumour spheres after being cultured with corresponding ECs. As shown in Figure 2, tumour spheres generated from both cells cultured with ECs were much larger than those generated from both cells in the absence of ECs (Figure 2A). Meanwhile, proliferation of GL261 cells was not affected by their interaction with b.END3 cells (see supplementary material, Figure S2A). A limiting dilution assay revealed that the sphere-forming capacity of GL261 and U251 cells precultured with ECs was higher than that of glioma cells alone (Figure 2B). FACS analysis demonstrated that CD133+ glioma cells were significantly enriched after being co-cultured with corresponding ECs (Figure 2C). In order to verify the effect of ECs on glioma cells in vivo, we performed grafted tumour experiments, using both subcutaneous and orthotopic transplantation. Our orthotopic transplanted tumour model revealed that co-injection of GL261 and b.END3 cells led to earlier deaths of mice (Figure 2D) with larger allografts (Figure 2E) than GL261 cells alone, which was also confirmed by subcutaneous transplantation experiments (see supplementary material, Figure S2B). Further investigation revealed that allografts generated by co-injection of GL261 and b.END3 cells contained more CD133+ GSCs (see supplementary material, Figure S2C), higher microvessel density (MVD; see supplementary material, Figure S2D) and Ki-67 index (see supplementary material, Figure S2E), as compared to the allografts generated from GL261 cells alone. Although the cell cycle of GL261 cells was not changed after co-culture with b.END3 cells, which is inconsistent with previous findings by others 25,26, the higher Ki67 index indicated that endothelial cells could enhance not only the CSC phenotype but also proliferation in glioma cells. Our data suggest that ECs enhance tumour initiation and growth, possibly through promoting CSC-like phenotype of glioma cells.


Endothelial cells promote stem-like phenotype of glioma cells through activating the Hedgehog pathway.

Yan GN, Yang L, Lv YF, Shi Y, Shen LL, Yao XH, Guo QN, Zhang P, Cui YH, Zhang X, Bian XW, Guo DY - J. Pathol. (2014)

Endothelial cells promote tumour sphere formation and CD133 expression by glioma cells in vitro and tumour growth in vivo. (A) Tumour spheres formed by GL261 cells cultured with b.END3 cells (left; scale bar = 100 µm) or by U251 cells cultured with HUVECs (right; scale bar = 300 µm) were larger in size than those formed by GL261, or U251 cells alone, respectively. (B) Efficiency of tumour sphere formation by GL261 cells cultured with b.END3 cells or by U251 cells cultured with HUVECs was higher than that by GL261 or U251 cells alone, respectively (*p < 0.05). (C) The numbers of CD133+ GL261 (left) or U251 cells (right) were significantly increased after co-culture with b.END3 cells or HUVECs, respectively (*p < 0.05). (D) Survival times of mice that were orthotopically co-injected with GL261 and b.END3 cells were shorter than those of mice orthotopically injected with GL261 cells alone (*p < 0.05). (E) Tumour volume of orthotopic allografts generated by co-injection of GL261 with b.END3 cells was significantly larger than that of orthotopic allografts generated by injection of GL261 cells alone (*p < 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig02: Endothelial cells promote tumour sphere formation and CD133 expression by glioma cells in vitro and tumour growth in vivo. (A) Tumour spheres formed by GL261 cells cultured with b.END3 cells (left; scale bar = 100 µm) or by U251 cells cultured with HUVECs (right; scale bar = 300 µm) were larger in size than those formed by GL261, or U251 cells alone, respectively. (B) Efficiency of tumour sphere formation by GL261 cells cultured with b.END3 cells or by U251 cells cultured with HUVECs was higher than that by GL261 or U251 cells alone, respectively (*p < 0.05). (C) The numbers of CD133+ GL261 (left) or U251 cells (right) were significantly increased after co-culture with b.END3 cells or HUVECs, respectively (*p < 0.05). (D) Survival times of mice that were orthotopically co-injected with GL261 and b.END3 cells were shorter than those of mice orthotopically injected with GL261 cells alone (*p < 0.05). (E) Tumour volume of orthotopic allografts generated by co-injection of GL261 with b.END3 cells was significantly larger than that of orthotopic allografts generated by injection of GL261 cells alone (*p < 0.05).
Mentions: To explore the mechanism underlying the interaction between GSCs and ECs, we utilized transwell co-culture. Both GL261 and U251 cells were tested for their ability to form tumour spheres after being cultured with corresponding ECs. As shown in Figure 2, tumour spheres generated from both cells cultured with ECs were much larger than those generated from both cells in the absence of ECs (Figure 2A). Meanwhile, proliferation of GL261 cells was not affected by their interaction with b.END3 cells (see supplementary material, Figure S2A). A limiting dilution assay revealed that the sphere-forming capacity of GL261 and U251 cells precultured with ECs was higher than that of glioma cells alone (Figure 2B). FACS analysis demonstrated that CD133+ glioma cells were significantly enriched after being co-cultured with corresponding ECs (Figure 2C). In order to verify the effect of ECs on glioma cells in vivo, we performed grafted tumour experiments, using both subcutaneous and orthotopic transplantation. Our orthotopic transplanted tumour model revealed that co-injection of GL261 and b.END3 cells led to earlier deaths of mice (Figure 2D) with larger allografts (Figure 2E) than GL261 cells alone, which was also confirmed by subcutaneous transplantation experiments (see supplementary material, Figure S2B). Further investigation revealed that allografts generated by co-injection of GL261 and b.END3 cells contained more CD133+ GSCs (see supplementary material, Figure S2C), higher microvessel density (MVD; see supplementary material, Figure S2D) and Ki-67 index (see supplementary material, Figure S2E), as compared to the allografts generated from GL261 cells alone. Although the cell cycle of GL261 cells was not changed after co-culture with b.END3 cells, which is inconsistent with previous findings by others 25,26, the higher Ki67 index indicated that endothelial cells could enhance not only the CSC phenotype but also proliferation in glioma cells. Our data suggest that ECs enhance tumour initiation and growth, possibly through promoting CSC-like phenotype of glioma cells.

Bottom Line: Microenvironmental regulation of cancer stem cells (CSCs) strongly influences the onset and spread of cancer.We observed that CD133(+) GSCs were located closely to Shh(+) endothelial cells in specimens of human glioblastoma multiforme (GBM).Knockdown of Smo in glioma cells led to a significant reduction of their CSC-like phenotype formation in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pathology and Southwest Cancer Centre, Southwest Hospital, Third Military Medical University and Key Laboratory of Tumour Immunopathology, Ministry of Education of China, Chongqing, 400038, People's Republic of China.

Show MeSH
Related in: MedlinePlus