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MicroRNA-21 regulates hTERT via PTEN in hypertrophic scar fibroblasts.

Zhu HY, Li C, Bai WD, Su LL, Liu JQ, Li Y, Shi JH, Cai WX, Bai XZ, Jia YH, Zhao B, Wu X, Li J, Hu DH - PLoS ONE (2014)

Bottom Line: As an important oncogenic miRNA, microRNA-21 (miR-21) is associated with various malignant diseases.Introduction of PTEN cDNA led to a remarkable depletion of hTERT and PI3K/AKT at the protein level as well as inhibition of miR-21-induced proliferation.In addition, Western-blot and qRT-PCR analyses confirmed that hTERT was the downstream target of PTEN.

View Article: PubMed Central - PubMed

Affiliation: Department of Burns and Cutaneous Surgery, Xijing Hospital, Fourth Military Medical University, Xi'an, Shaanxi, People's Republic of China.

ABSTRACT

Background: As an important oncogenic miRNA, microRNA-21 (miR-21) is associated with various malignant diseases. However, the precise biological function of miR-21 and its molecular mechanism in hypertrophic scar fibroblast cells has not been fully elucidated.

Methodology/principal findings: Quantitative Real-Time PCR (qRT-PCR) analysis revealed significant upregulation of miR-21 in hypertrophic scar fibroblast cells compared with that in normal skin fibroblast cells. The effects of miR-21 were then assessed in MTT and apoptosis assays through in vitro transfection with a miR-21 mimic or inhibitor. Next, PTEN (phosphatase and tensin homologue deleted on chromosome ten) was identified as a target gene of miR-21 in hypertrophic scar fibroblast cells. Furthermore, Western-blot and qRT-PCR analyses revealed that miR-21 increased the expression of human telomerase reverse transcriptase (hTERT) via the PTEN/PI3K/AKT pathway. Introduction of PTEN cDNA led to a remarkable depletion of hTERT and PI3K/AKT at the protein level as well as inhibition of miR-21-induced proliferation. In addition, Western-blot and qRT-PCR analyses confirmed that hTERT was the downstream target of PTEN. Finally, miR-21 and PTEN RNA expression levels in hypertrophic scar tissue samples were examined. Immunohistochemistry assays revealed an inverse correlation between PTEN and hTERT levels in high miR-21 RNA expressing-hypertrophic scar tissues.

Conclusions/significance: These data indicate that miR-21 regulates hTERT expression via the PTEN/PI3K/AKT signaling pathway by directly targeting PTEN, therefore controlling hypertrophic scar fibroblast cell growth. MiR-21 may be a potential novel molecular target for the treatment of hypertrophic scarring.

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RNAi-mediated hTERT knockdown inhibits HSFBs growth and promotes apoptosis.A, hTERT mRNA expression; B, hTERT protein expression; C, proliferation; D, E, apoptosis were downregulated following RNAi-mediated hTERT knockdown.
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pone-0097114-g005: RNAi-mediated hTERT knockdown inhibits HSFBs growth and promotes apoptosis.A, hTERT mRNA expression; B, hTERT protein expression; C, proliferation; D, E, apoptosis were downregulated following RNAi-mediated hTERT knockdown.

Mentions: Previous studies have indicated that the hTERT overexpression correlates closely with drug sensitivity and tumor growth in carcinoma-associated fibroblasts; however, the mechanism of hTERT activity in HSFBs has not been characterized [40]. To determine whether the biological effect of hTERT in HSFBs is consistent with the proposed functions of miR-21 and PTEN/P13K/AKT signaling, we established cell lines that expressed low levels of hTERT using hTERT-specific RNAi method (Figure 5A and 5B). MTT analysis indicated that hTERT knockdown reduced HSFBs proliferation and increased apoptosis (Figure 5C, 5D and 5E). This was consistent with the effects observed in HSFBs transfected with miR-21, which further confirmed that the downregulation of hTERT is essential for miR-21-induced and PTEN/P13K/AKT-mediated cell proliferation.


MicroRNA-21 regulates hTERT via PTEN in hypertrophic scar fibroblasts.

Zhu HY, Li C, Bai WD, Su LL, Liu JQ, Li Y, Shi JH, Cai WX, Bai XZ, Jia YH, Zhao B, Wu X, Li J, Hu DH - PLoS ONE (2014)

RNAi-mediated hTERT knockdown inhibits HSFBs growth and promotes apoptosis.A, hTERT mRNA expression; B, hTERT protein expression; C, proliferation; D, E, apoptosis were downregulated following RNAi-mediated hTERT knockdown.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4016251&req=5

pone-0097114-g005: RNAi-mediated hTERT knockdown inhibits HSFBs growth and promotes apoptosis.A, hTERT mRNA expression; B, hTERT protein expression; C, proliferation; D, E, apoptosis were downregulated following RNAi-mediated hTERT knockdown.
Mentions: Previous studies have indicated that the hTERT overexpression correlates closely with drug sensitivity and tumor growth in carcinoma-associated fibroblasts; however, the mechanism of hTERT activity in HSFBs has not been characterized [40]. To determine whether the biological effect of hTERT in HSFBs is consistent with the proposed functions of miR-21 and PTEN/P13K/AKT signaling, we established cell lines that expressed low levels of hTERT using hTERT-specific RNAi method (Figure 5A and 5B). MTT analysis indicated that hTERT knockdown reduced HSFBs proliferation and increased apoptosis (Figure 5C, 5D and 5E). This was consistent with the effects observed in HSFBs transfected with miR-21, which further confirmed that the downregulation of hTERT is essential for miR-21-induced and PTEN/P13K/AKT-mediated cell proliferation.

Bottom Line: As an important oncogenic miRNA, microRNA-21 (miR-21) is associated with various malignant diseases.Introduction of PTEN cDNA led to a remarkable depletion of hTERT and PI3K/AKT at the protein level as well as inhibition of miR-21-induced proliferation.In addition, Western-blot and qRT-PCR analyses confirmed that hTERT was the downstream target of PTEN.

View Article: PubMed Central - PubMed

Affiliation: Department of Burns and Cutaneous Surgery, Xijing Hospital, Fourth Military Medical University, Xi'an, Shaanxi, People's Republic of China.

ABSTRACT

Background: As an important oncogenic miRNA, microRNA-21 (miR-21) is associated with various malignant diseases. However, the precise biological function of miR-21 and its molecular mechanism in hypertrophic scar fibroblast cells has not been fully elucidated.

Methodology/principal findings: Quantitative Real-Time PCR (qRT-PCR) analysis revealed significant upregulation of miR-21 in hypertrophic scar fibroblast cells compared with that in normal skin fibroblast cells. The effects of miR-21 were then assessed in MTT and apoptosis assays through in vitro transfection with a miR-21 mimic or inhibitor. Next, PTEN (phosphatase and tensin homologue deleted on chromosome ten) was identified as a target gene of miR-21 in hypertrophic scar fibroblast cells. Furthermore, Western-blot and qRT-PCR analyses revealed that miR-21 increased the expression of human telomerase reverse transcriptase (hTERT) via the PTEN/PI3K/AKT pathway. Introduction of PTEN cDNA led to a remarkable depletion of hTERT and PI3K/AKT at the protein level as well as inhibition of miR-21-induced proliferation. In addition, Western-blot and qRT-PCR analyses confirmed that hTERT was the downstream target of PTEN. Finally, miR-21 and PTEN RNA expression levels in hypertrophic scar tissue samples were examined. Immunohistochemistry assays revealed an inverse correlation between PTEN and hTERT levels in high miR-21 RNA expressing-hypertrophic scar tissues.

Conclusions/significance: These data indicate that miR-21 regulates hTERT expression via the PTEN/PI3K/AKT signaling pathway by directly targeting PTEN, therefore controlling hypertrophic scar fibroblast cell growth. MiR-21 may be a potential novel molecular target for the treatment of hypertrophic scarring.

Show MeSH
Related in: MedlinePlus