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The regulation of Toll-like receptor 2 by miR-143 suppresses the invasion and migration of a subset of human colorectal carcinoma cells.

Guo H, Chen Y, Hu X, Qian G, Ge S, Zhang J - Mol. Cancer (2013)

Bottom Line: We also found that miR-143, a putative tumour suppressor that is down-regulated in CRC tissues, reduces the invasion and migration of CRC cells primarily via TLR2.Utilising a xenograft mouse model, we demonstrated that re-expression of miR-143 inhibits CRC cell colonisation in vivo. miR-143 blocks the TLR2 signalling pathway in human CRC cells.This knowledge may pave the way for new clinical applications utilising miR-143 mimics in the treatment of patients with CRC.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Clinical Laboratory, Shanghai Third People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai 200019, PR China. sxguohaiyan@126.com

ABSTRACT

Background: The Toll-like receptor 2 (TLR2)-driven tissue response may promote neoangiogenesis and tumour growth by mechanisms that are poorly understood.

Methods: We investigated the expression levels of TLR2 and associated-miRNAs in colorectal carcinoma (CRC) tissues and cell lines using real-time PCR, northern blotting and western blotting. Survival curver was generated by Log-Rank test and the role of TLR2 signalling in tumour invasion and migration was determined by transwell analysis kits.

Results: We observed that the tissues from CRC patients express relatively high levels of TLR2. Targeting TLR2 markedly reduces the invasion and migration of CRC cells. We also found that miR-143, a putative tumour suppressor that is down-regulated in CRC tissues, reduces the invasion and migration of CRC cells primarily via TLR2. Utilising a xenograft mouse model, we demonstrated that re-expression of miR-143 inhibits CRC cell colonisation in vivo.

Conclusion: miR-143 blocks the TLR2 signalling pathway in human CRC cells. This knowledge may pave the way for new clinical applications utilising miR-143 mimics in the treatment of patients with CRC.

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TLR2 modulation accounts for the antimetastatic effect of miR-143. (A) Western blot showing TLR2 expression in SW620 cells 48 h after transfection with the artificial TLR2 expression vector (TLR2). (B) The effects of miR-143, TLR2, and miR-143 combined with TLR2 on the migration and invasion of SW620 cells. (C) Western blot showing TLR2 translation in HCT116 cells 48 hours after transfection with TLR2. (D) The effects of miR-143, TLR2, and miR-143 combined with TLR2 on the migration and invasion of HCT116 cells. (E) Aliquots of 1 ×106 stable miR-143 or miR-NC over-expressing SW620 cells were injected into immunodeficient mice and evaluated for lung colonisation capacity in tail-vein assays. Lung colonisation was measured using bioluminescence at 1, 2, 3, 4, 5 and 6 weeks after injection. (F) The bar graph represents 24 hour time-point measurements of the normalised photon flux from animals injected with SW620 cells. Representative images are shown. n = 5 for each group; error bars indicate SD.
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Figure 5: TLR2 modulation accounts for the antimetastatic effect of miR-143. (A) Western blot showing TLR2 expression in SW620 cells 48 h after transfection with the artificial TLR2 expression vector (TLR2). (B) The effects of miR-143, TLR2, and miR-143 combined with TLR2 on the migration and invasion of SW620 cells. (C) Western blot showing TLR2 translation in HCT116 cells 48 hours after transfection with TLR2. (D) The effects of miR-143, TLR2, and miR-143 combined with TLR2 on the migration and invasion of HCT116 cells. (E) Aliquots of 1 ×106 stable miR-143 or miR-NC over-expressing SW620 cells were injected into immunodeficient mice and evaluated for lung colonisation capacity in tail-vein assays. Lung colonisation was measured using bioluminescence at 1, 2, 3, 4, 5 and 6 weeks after injection. (F) The bar graph represents 24 hour time-point measurements of the normalised photon flux from animals injected with SW620 cells. Representative images are shown. n = 5 for each group; error bars indicate SD.

Mentions: To determine whether TLR2 is the critical mediator of the role of miR-143 in cellular invasion and migration, we constructed a TLR2 transcript with a 3′UTR deletion mutation (TLR2). First, we performed a western blot analysis 48 h after transfecting the TLR2 expression vector with the 3′UTR deletion mutation (TLR2) into SW620 (Figure 5A and Additional file 5: Figure S5A) and HCT116 cells (Figures 5C and Additional file 5: Figure S5B). As shown in the figures, compared to the negative control group (pcDNA3.0), ectopic expression of TLR2 significantly increased the expression of TLR2.


The regulation of Toll-like receptor 2 by miR-143 suppresses the invasion and migration of a subset of human colorectal carcinoma cells.

Guo H, Chen Y, Hu X, Qian G, Ge S, Zhang J - Mol. Cancer (2013)

TLR2 modulation accounts for the antimetastatic effect of miR-143. (A) Western blot showing TLR2 expression in SW620 cells 48 h after transfection with the artificial TLR2 expression vector (TLR2). (B) The effects of miR-143, TLR2, and miR-143 combined with TLR2 on the migration and invasion of SW620 cells. (C) Western blot showing TLR2 translation in HCT116 cells 48 hours after transfection with TLR2. (D) The effects of miR-143, TLR2, and miR-143 combined with TLR2 on the migration and invasion of HCT116 cells. (E) Aliquots of 1 ×106 stable miR-143 or miR-NC over-expressing SW620 cells were injected into immunodeficient mice and evaluated for lung colonisation capacity in tail-vein assays. Lung colonisation was measured using bioluminescence at 1, 2, 3, 4, 5 and 6 weeks after injection. (F) The bar graph represents 24 hour time-point measurements of the normalised photon flux from animals injected with SW620 cells. Representative images are shown. n = 5 for each group; error bars indicate SD.
© Copyright Policy - open-access
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Figure 5: TLR2 modulation accounts for the antimetastatic effect of miR-143. (A) Western blot showing TLR2 expression in SW620 cells 48 h after transfection with the artificial TLR2 expression vector (TLR2). (B) The effects of miR-143, TLR2, and miR-143 combined with TLR2 on the migration and invasion of SW620 cells. (C) Western blot showing TLR2 translation in HCT116 cells 48 hours after transfection with TLR2. (D) The effects of miR-143, TLR2, and miR-143 combined with TLR2 on the migration and invasion of HCT116 cells. (E) Aliquots of 1 ×106 stable miR-143 or miR-NC over-expressing SW620 cells were injected into immunodeficient mice and evaluated for lung colonisation capacity in tail-vein assays. Lung colonisation was measured using bioluminescence at 1, 2, 3, 4, 5 and 6 weeks after injection. (F) The bar graph represents 24 hour time-point measurements of the normalised photon flux from animals injected with SW620 cells. Representative images are shown. n = 5 for each group; error bars indicate SD.
Mentions: To determine whether TLR2 is the critical mediator of the role of miR-143 in cellular invasion and migration, we constructed a TLR2 transcript with a 3′UTR deletion mutation (TLR2). First, we performed a western blot analysis 48 h after transfecting the TLR2 expression vector with the 3′UTR deletion mutation (TLR2) into SW620 (Figure 5A and Additional file 5: Figure S5A) and HCT116 cells (Figures 5C and Additional file 5: Figure S5B). As shown in the figures, compared to the negative control group (pcDNA3.0), ectopic expression of TLR2 significantly increased the expression of TLR2.

Bottom Line: We also found that miR-143, a putative tumour suppressor that is down-regulated in CRC tissues, reduces the invasion and migration of CRC cells primarily via TLR2.Utilising a xenograft mouse model, we demonstrated that re-expression of miR-143 inhibits CRC cell colonisation in vivo. miR-143 blocks the TLR2 signalling pathway in human CRC cells.This knowledge may pave the way for new clinical applications utilising miR-143 mimics in the treatment of patients with CRC.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Clinical Laboratory, Shanghai Third People's Hospital, School of medicine, Shanghai Jiao Tong University, Shanghai 200019, PR China. sxguohaiyan@126.com

ABSTRACT

Background: The Toll-like receptor 2 (TLR2)-driven tissue response may promote neoangiogenesis and tumour growth by mechanisms that are poorly understood.

Methods: We investigated the expression levels of TLR2 and associated-miRNAs in colorectal carcinoma (CRC) tissues and cell lines using real-time PCR, northern blotting and western blotting. Survival curver was generated by Log-Rank test and the role of TLR2 signalling in tumour invasion and migration was determined by transwell analysis kits.

Results: We observed that the tissues from CRC patients express relatively high levels of TLR2. Targeting TLR2 markedly reduces the invasion and migration of CRC cells. We also found that miR-143, a putative tumour suppressor that is down-regulated in CRC tissues, reduces the invasion and migration of CRC cells primarily via TLR2. Utilising a xenograft mouse model, we demonstrated that re-expression of miR-143 inhibits CRC cell colonisation in vivo.

Conclusion: miR-143 blocks the TLR2 signalling pathway in human CRC cells. This knowledge may pave the way for new clinical applications utilising miR-143 mimics in the treatment of patients with CRC.

Show MeSH
Related in: MedlinePlus