Identification of valid endogenous control genes for determining gene expression in C6 glioma cell line treated with conditioned medium from adipose-derived stem cell

Iser, Isabele Cristiana; Campos, Rafael Paschoal de; Bertoni, Ana Paula Santin

doi:10.1016/j.biopha.2015.08.035

Cited by 13 publications

(5 citation statements)

References 76 publications

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“…3G and H). The ACTb and GAPDH genes have been widely used [Iser et al, 2015]. But Huggett et al [2005] published that these genes are inappropriate due to their high variability in biological samples.…”

Section: Discussionmentioning

confidence: 99%

Extracellular Nucleotide Hydrolysis in Dermal and Limbal Mesenchymal Stem Cells: A Source of Adenosine Production

Naasani¹,

Rodrigues²,

Campos³

et al. 2017

J. Cell. Biochem.

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Human Limbal (L-MSCs) and Dermal Mesenchymal Stem Cell (D-MSCs) possess many properties that increase their therapeutic potential in ophthalmology and dermatology. It is known that purinergic signaling plays a role in many aspects of mesenchymal stem cells physiology. They release and respond to purinergic ligands, altering proliferation, migration, differentiation, and apoptosis. Therefore, more information on these processes would be crucial for establishing future clinical applications using their differentiation potential, but without undesirable side effects. This study evaluated and compared the expression of ecto-nucleotidases, the enzymatic activity of degradation of extracellular nucleotides and the metabolism of extracellular ATP in D-MSCs and L-MSCs, isolated from discard tissues of human skin and sclerocorneal rims. The D-MSCs and L-MSCs showed a differentiation potential into osteogenic, adipogenic, and chondrogenic lineages and the expression of markers CD105 , CD44 , CD14 , CD34 , CD45 , as expected. Both cells hydrolyzed low levels of extracellular ATP and high levels of AMP, leading to adenosine accumulation that can regulate inflammation and tissue repair. These cells expressed mRNA for ENTPD1, 2, 3, 5 and 6, and CD73 that corresponded to the observed enzymatic activities. Thus, considering the degradation of ATP and adenosine production, limbal MSCs are very similar to dermal MSCs, indicating that from the aspect of extracellular nucleotide metabolism L-MSCs are very similar to the characterized D-MSCs. J. Cell. Biochem. 118: 2430-2442, 2017. © 2017 Wiley Periodicals, Inc.

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Section: Discussionmentioning

confidence: 99%

Extracellular Nucleotide Hydrolysis in Dermal and Limbal Mesenchymal Stem Cells: A Source of Adenosine Production

Naasani¹,

Rodrigues²,

Campos³

et al. 2017

J. Cell. Biochem.

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“…Five commonly used reference genes were selected: actin beta (ACTB), beta-2-microglobulin (B2M), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), hypoxanthine phosphoribosyltransferase 1 (HPRT1), and TATA-box binding protein (TBP) (Santin et al, 2013;Weber et al, 2014;Iser et al, 2015;Ofinran et al, 2016;Wang et al, 2016a). RT-qPCR was performed using StepOnePlus Real-Time PCR cycler.…”

Section: Rt-qpcrmentioning

confidence: 99%

Cervical cancer stem‐like cells: systematic review and identification of reference genes for gene expression

Campos

Schultz

Mello

et al. 2017

Cell Biology International

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Cervical cancer is the fourth most common cancer affecting women worldwide. Among many factors, the presence of cancer stem cells, a subpopulation of cells inside the tumor, has been associated with a worse prognosis. Considering the importance of gene expression studies to understand the biology of cervical cancer stem cells (CCSC), this work identifies stable reference genes for cervical cancer cell lines SiHa, HeLa, and ME180 as well as their respective cancer stem-like cells. A literature review was performed to identify validated reference genes currently used to normalize RT-qPCR data in cervical cancer cell lines. Then, cell lines were cultured in regular monolayer or in a condition that favors tumor sphere formation. RT-qPCR was performed using five reference genes: ACTB, B2M, GAPDH, HPRT1, and TBP. Stability was assessed to validate the selected genes as suitable reference genes. The evaluation validated B2M, GAPDH, HPRT1, and TBP in these experimental conditions. Among them, GAPDH and TBP presented the lowest variability according to the analysis by Normfinder, Bestkeeper, and ΔC methods, being therefore the most adequate genes to normalize the combination of all samples. These results suggest that B2M, GAPDH, HPRT1, and TBP are suitable reference genes to normalize RT-qPCR data of established cervical cancer cell lines SiHa, HeLa, and ME180 as well as their derived cancer stem-like cells. Indeed, GAPDH and TBP seem to be the most convenient choices for studying gene expression in these cells in monolayers or spheres.

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“…β2-Microglobulin (B2M), a 11 KDa non-glycosylated protein, is encoded by a well-known housekeeping gene 11 – 13 . B2M is expressed by all nucleated cells to form a small invariable light chain subunit of the major histocompatibility complex (MHC) class I antigen on the cell surface 14 .…”

Section: Introductionmentioning

confidence: 99%

Mesenchymal Stromal Cells-Derived β2-Microglobulin Promotes Epithelial–Mesenchymal Transition of Esophageal Squamous Cell Carcinoma Cells

Wang

Yang

Wang

et al. 2018

Sci Rep

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Mesenchymal stromal cells (MSCs) have been considered as one of the pivotal type of cells composing the tumor microenvironment. Although contact-dependent mechanisms and paracrine factors are thought to collaborate in governing the MSCs-based effects on tumors progression, the underlying mechanisms remain largely unknown. In particular, the involvement of MSCs-derived cytokines in the epithelial–mesenchymal transition (EMT) of esophageal squamous cell carcinoma (ESCC) has not been clarified. In this study, we observed that β2-Microglobulin (B2M) is highly expressed in MSCs but scarcely in ESCC cells. Based on the previously described EMT promoting effect of B2M, we investigated the in vitro effect of MSCs-derived B2M on the EMT of ESCC cells, and discovered its subsequent enhancing effects on cell mobility and tumor-initiation. Further xenograft transplantation experiments confirmed the in vivo induction of tumor-initiation by MSCs-derived B2M. Noteworthy, we showed that the B2M expression positively correlated with poor prognosis. The fact that B2M is primarily expressed by the stroma of the ESCC tissue strengthens our hypothesis that in ESCC, MSCs-derived B2M promotes tumor-initiation and invasion via enhancing EMT, resulting in an adverse prognosis for the patients. Our results will be valuable for the prediction of the development and treatment of ESCC.

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Identification of valid endogenous control genes for determining gene expression in C6 glioma cell line treated with conditioned medium from adipose-derived stem cell

Cited by 13 publications

References 76 publications

Extracellular Nucleotide Hydrolysis in Dermal and Limbal Mesenchymal Stem Cells: A Source of Adenosine Production

Extracellular Nucleotide Hydrolysis in Dermal and Limbal Mesenchymal Stem Cells: A Source of Adenosine Production

Cervical cancer stem‐like cells: systematic review and identification of reference genes for gene expression

Mesenchymal Stromal Cells-Derived β2-Microglobulin Promotes Epithelial–Mesenchymal Transition of Esophageal Squamous Cell Carcinoma Cells

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