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Piwil2-transfected human fibroblasts are cancer stem cell-like and genetically unstable.

ABSTRACT: Uncontrolled cell proliferation and inhibition of apoptosis are considered to be vital for cancer initiation, maintenance, infiltration, metastasis and recurrence after anti-cancer therapy. Here we report the generation of a novel cell line by reprogramming child foreskin fibroblast with the full length apoptosis inhibitor gene PIWIL2. The fibroblasts transfected with PIWIL2 expressed the stem cell markers OCT-4, NANOG, SOX-2, KLF-4 and C-MYC; endoderm marker AFP and GATA6; mesoderm markers ACTA2 and BRACHYURY; and ectoderm markers NESTIN and TUBB3. The karyotype was found to be hyperdiploid. The PIWIL2 transfected fibroblast cells grew into tumorous masses within 5 weeks of subcutaneous injection into adult nude mice. Although the injected cell expressed markers for all three germlines, ectoderm, mesoderm, and endoderm, they did not form teratomas in vivo. This study indicates that the PIWIL2 gene could play a key role in cancer induction and maintenance. This method for generating induced tumorigenic cells (ITGC) provides a new research tool to study oncogenesis that in turn may lead to a better understanding of cancer etiology and the development of novel anti-cancer therapies.

SUBMITTER: Zhang D

PROVIDER: S-EPMC5355342 | biostudies-literature | 2017 Feb

REPOSITORIES: biostudies-literature

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Piwil2-transfected human fibroblasts are cancer stem cell-like and genetically unstable.

Zhang Deying D Wu Xin X Liu Xing X Cai Chunhong C Zeng Guangping G Rohozinski Jan J Zhang Yuanyuan Y Wei Guanghui G He Dawei D

Oncotarget 20170201 7

Uncontrolled cell proliferation and inhibition of apoptosis are considered to be vital for cancer initiation, maintenance, infiltration, metastasis and recurrence after anti-cancer therapy. Here we report the generation of a novel cell line by reprogramming child foreskin fibroblast with the full length apoptosis inhibitor gene PIWIL2. The fibroblasts transfected with PIWIL2 expressed the stem cell markers OCT-4, NANOG, SOX-2, KLF-4 and C-MYC; endoderm marker AFP and GATA6; mesoderm markers ACTA ...[more]

PMID: 28103575

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Project description:Recently, several studies have demonstrated that cancer cell?derived exosomes can facilitate tumor development and metastasis formation. However, the detailed function of exosomes released by cancer stem cells (CSCs) requires further investigation. The aim of the present study was to investigate the role of CSC?derived exosomes in tumor development. For this purpose, Piwil2?induced cancer stem cells (Piwil2?iCSCs) were used as exosome?generating cells, while fibroblasts (FBs) served as recipient cells. Exosomes were isolated by the ultracentrifugation of Piwil2?iCSC?conditioned medium and identified by transmission electron microscopy, nanoparticle tracking analysis and western blot analysis. To evaluate the effects of the exosomes on cell proliferation, migration and invasion, cell counting assay (CCK?8), a wound healing assay and a Transwell assay were performed. Protein expression [matrix metalloproteinase (MMP)2, MMP9, ??smooth muscle actin (??SMA) and vimentin and fibroblast?activating protein (FAP)] was examined in FBs by western blot analysis. It was found that the Piwil2?iCSC?derived exosomes (Piwil2?iCSC?Exo) were oval or spherical, membrane?coated vesicles with a uniform size (30?100 nm in diameter). They are characterized by the surface expression of CD9, CD63, Hsp70 and Piwil2 proteins. Additional results from functional analyses revealed that Piwil2?iCSC?Exo enhanced the proliferative, migratory and invasive abilities of FBs, accompanied by the upregulated expression of MMP2 and MMP9. In addition, the increased expression of ??SMA (P<0.05), vimentin (P<0.01 vs. control group, P<0.05 vs. PBS group) and FAP (P<0.001 vs. control group, P<0.01 vs. PBS group) following exposure to Piwil2?iCSC?Exo suggested that the exosomes induced FB transformation into cancer?associated fibroblasts (CAFs). On the whole, the findings of this study demonstrate that Piwil2?iCSC?Exo induce the cancer?associated phenotype in fibroblasts in vitro, suggesting that CSCs can promote tumor development through the modulation of the tumor microenvironment.

Project description:Reprogramming of differentiated somatic cells into induced pluripotent stem (iPS) cells has potential for derivation of patient-specific cells for therapy as well as for development of models with which to study disease progression. Derivation of iPS cells from human somatic cells has been achieved by viral transduction of human fibroblasts with early developmental genes. Because forced expression of these genes by viral transduction results in transgene integration with unknown and unpredictable potential mutagenic effects, identification of cell culture conditions that can induce endogenous expression of these genes is desirable. Here we show that primary adult human fibroblasts have basal expression of mRNA for OCT4, SOX2, and NANOG. However, translation of these messages into detectable proteins and their subcellular localization depends on cell culture conditions. Manipulation of oxygen concentration and FGF2 supplementation can modulate expression of some pluripotency related genes at the transcriptional, translational, and cellular localization level. Changing cell culture condition parameters led to expression of REX1, potentiation of expression of LIN28, translation of OCT4, SOX2, and NANOG, and translocation of these transcription factors to the cell nucleus. We also show that culture conditions affect the in vitro lifespan of dermal fibroblasts, nearly doubling the number of population doublings before the cells reach replicative senescence. Our results suggest that it is possible to induce and manipulate endogenous expression of stem cell genes in somatic cells without genetic manipulation, but this short-term induction may not be sufficient for acquisition of true pluripotency. Further investigation of the factors involved in inducing this response could lead to discovery of defined culture conditions capable of altering cell fate in vitro. This would alleviate the need for forced expression by transgenesis, thus eliminating the risk of mutagenic effects due to genetic manipulation.

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Piwil2-transfected human fibroblasts are cancer stem cell-like and genetically unstable.

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Piwil2-transfected human fibroblasts are cancer stem cell-like and genetically unstable.

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