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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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Expression levels of TLR2 are inversely correlated with miR-143 in CRC tissues and cell lines. (A) Expression of miR-143 was determined using real-time PCR in 79 CRC tissue samples, and the correlation of TLR2 expression levels with miR-143 in CRC tissues was evaluated using nonparametric tests (R2 = 0.4486; p = 0.000). (B) Expression of miR-143 was determined by northern blotting in 5 normal colon epithelial cells and 4 CRC cell lines (SW480, LS174T, SW620, and HCT116). (C) Expression of miR-143 was determined in 79 CRC tissues (grade1 n = 18; grade2 n = 20; grade3 n = 41) by real-time PCR. The statistical method analysed the correlation between the expression of miR-143 and tumour pathological grade. (D) Expression of miR-143 was determined in 79 CRC tissues (N0 n = 28; N1 n = 31; N2 n = 20) by real-time PCR. The statistical method analysed the correlation between the expression of miR-143 and tumour lymph node metastasis. (E) Higher overall survival of patients with high miR-143 levels (above median; n = 38) compared to patients with low miR-143 levels (below median; n = 41). (***p < 0.001; **p < 0.01; *p < 0.05).
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Figure 3: Expression levels of TLR2 are inversely correlated with miR-143 in CRC tissues and cell lines. (A) Expression of miR-143 was determined using real-time PCR in 79 CRC tissue samples, and the correlation of TLR2 expression levels with miR-143 in CRC tissues was evaluated using nonparametric tests (R2 = 0.4486; p = 0.000). (B) Expression of miR-143 was determined by northern blotting in 5 normal colon epithelial cells and 4 CRC cell lines (SW480, LS174T, SW620, and HCT116). (C) Expression of miR-143 was determined in 79 CRC tissues (grade1 n = 18; grade2 n = 20; grade3 n = 41) by real-time PCR. The statistical method analysed the correlation between the expression of miR-143 and tumour pathological grade. (D) Expression of miR-143 was determined in 79 CRC tissues (N0 n = 28; N1 n = 31; N2 n = 20) by real-time PCR. The statistical method analysed the correlation between the expression of miR-143 and tumour lymph node metastasis. (E) Higher overall survival of patients with high miR-143 levels (above median; n = 38) compared to patients with low miR-143 levels (below median; n = 41). (***p < 0.001; **p < 0.01; *p < 0.05).

Mentions: We detected the expression of TLR2 and mature miR-143 in the same clinical samples using real-time PCR (N = 79). Using nonparametric tests, we found a significant inverse correlation between TLR2 expression and miR-143 expression in CRC tissues (R2 = 0.4486, p = 0.000 for TLR2 and miR-143) (Figure 3A; Table 1). We utilised the same tests to determine whether TLR2 is a putative target of miR-101, miR-154, and miR-340 and found no inverse correlation between TLR2 and these miRNAs in human CRC samples (data not shown). In addition, we examined the endogenous miR-143 expression levels by northern blotting in 5 normal colon epithelial tissues and 4 CRC cells. The expression of miR-143 was significantly lower in 4 CRC cells with high TLR2 expression levels (Figure 3B and Additional file 3: Figure S3A), especially in the poorly differentiated tumour cells SW620 and HCT116, showing that miR-143 levels are inversely correlated with the levels of TLR2 expression. The miR-143 profile of human CRC tissues revealed that low levels of miR-143 are correlated with more advanced pathology grades (1, 2, 3) and lymph node metastasis (N0, N1, N2) (Figures 3C and 3D; ***p < 0.001; **p < 0.01; *p < 0.05), suggesting an association between low miR-143 expression and tumour progression. Additionally, miR-143 levels above the median correlate with a higher overall survival of patients with CRC (p = 0.0018) (Figure 3E).


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)

Expression levels of TLR2 are inversely correlated with miR-143 in CRC tissues and cell lines. (A) Expression of miR-143 was determined using real-time PCR in 79 CRC tissue samples, and the correlation of TLR2 expression levels with miR-143 in CRC tissues was evaluated using nonparametric tests (R2 = 0.4486; p = 0.000). (B) Expression of miR-143 was determined by northern blotting in 5 normal colon epithelial cells and 4 CRC cell lines (SW480, LS174T, SW620, and HCT116). (C) Expression of miR-143 was determined in 79 CRC tissues (grade1 n = 18; grade2 n = 20; grade3 n = 41) by real-time PCR. The statistical method analysed the correlation between the expression of miR-143 and tumour pathological grade. (D) Expression of miR-143 was determined in 79 CRC tissues (N0 n = 28; N1 n = 31; N2 n = 20) by real-time PCR. The statistical method analysed the correlation between the expression of miR-143 and tumour lymph node metastasis. (E) Higher overall survival of patients with high miR-143 levels (above median; n = 38) compared to patients with low miR-143 levels (below median; n = 41). (***p < 0.001; **p < 0.01; *p < 0.05).
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Figure 3: Expression levels of TLR2 are inversely correlated with miR-143 in CRC tissues and cell lines. (A) Expression of miR-143 was determined using real-time PCR in 79 CRC tissue samples, and the correlation of TLR2 expression levels with miR-143 in CRC tissues was evaluated using nonparametric tests (R2 = 0.4486; p = 0.000). (B) Expression of miR-143 was determined by northern blotting in 5 normal colon epithelial cells and 4 CRC cell lines (SW480, LS174T, SW620, and HCT116). (C) Expression of miR-143 was determined in 79 CRC tissues (grade1 n = 18; grade2 n = 20; grade3 n = 41) by real-time PCR. The statistical method analysed the correlation between the expression of miR-143 and tumour pathological grade. (D) Expression of miR-143 was determined in 79 CRC tissues (N0 n = 28; N1 n = 31; N2 n = 20) by real-time PCR. The statistical method analysed the correlation between the expression of miR-143 and tumour lymph node metastasis. (E) Higher overall survival of patients with high miR-143 levels (above median; n = 38) compared to patients with low miR-143 levels (below median; n = 41). (***p < 0.001; **p < 0.01; *p < 0.05).
Mentions: We detected the expression of TLR2 and mature miR-143 in the same clinical samples using real-time PCR (N = 79). Using nonparametric tests, we found a significant inverse correlation between TLR2 expression and miR-143 expression in CRC tissues (R2 = 0.4486, p = 0.000 for TLR2 and miR-143) (Figure 3A; Table 1). We utilised the same tests to determine whether TLR2 is a putative target of miR-101, miR-154, and miR-340 and found no inverse correlation between TLR2 and these miRNAs in human CRC samples (data not shown). In addition, we examined the endogenous miR-143 expression levels by northern blotting in 5 normal colon epithelial tissues and 4 CRC cells. The expression of miR-143 was significantly lower in 4 CRC cells with high TLR2 expression levels (Figure 3B and Additional file 3: Figure S3A), especially in the poorly differentiated tumour cells SW620 and HCT116, showing that miR-143 levels are inversely correlated with the levels of TLR2 expression. The miR-143 profile of human CRC tissues revealed that low levels of miR-143 are correlated with more advanced pathology grades (1, 2, 3) and lymph node metastasis (N0, N1, N2) (Figures 3C and 3D; ***p < 0.001; **p < 0.01; *p < 0.05), suggesting an association between low miR-143 expression and tumour progression. Additionally, miR-143 levels above the median correlate with a higher overall survival of patients with CRC (p = 0.0018) (Figure 3E).

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