Differential marker expression by cultures rich in mesenchymal stem cells

Wetzig, Andrew Roberts; Alaiya, Ayodele; Al‐Alwan, Monther; Pradez, Christian Benedict; Pulicat, Manogaran S; Al-Mazrou, Amer; Shinwari, Zakia; Sleiman, Ghida Majed; Ghebeh, Hazem; Al-Humaidan, Hind; Gaafar, Ameera; Kanaan, Imaduddin; Adra, Chaker N.

doi:10.1186/1471-2121-14-54

Cited by 36 publications

(31 citation statements)

References 45 publications

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“…Immunofluorescence assay was conducted as described previously [28]. Briefly, MCF-7 cells treated with the test compounds for the indicated time intervals were plated on glass slides at a density of 20000 cells/cm 2 and cultured for 7 days then fixed in 4% formaldehyde.…”

Section: Methodsmentioning

confidence: 99%

Aryl hydrocarbon receptor/cytochrome P450 1A1 pathway mediates breast cancer stem cells expansion through PTEN inhibition and β-Catenin and Akt activation

2017

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BackgroundBreast cancer stem cells (CSCs) are small sub-type of the whole cancer cells that drive tumor initiation, progression and metastasis. Recent studies have demonstrated a role for the aryl hydrocarbon receptor (AhR)/cytochrome P4501A1 pathway in CSCs expansion. However, the exact molecular mechanisms remain unclear.MethodsThe current study was designed to a) determine the effect of AhR activation and inhibition on breast CSCs development, maintenance, self-renewal, and chemoresistance at the in vitro and in vivo levels and b) explore the role of β-Catenin, PI3K/Akt, and PTEN signaling pathways. To test this hypothesis, CSC characteristics of five human breast cancer cells; SKBR-3, MCF-7, and MDA-MB231, HS587T, and T47D treated with AhR activators or inhibitor were determined using Aldefluor assay, side population, and mammosphere formation. The mRNA, protein expression, cellular content and localization of the target genes were determined by RT-PCR, Western blot analysis, and Immunofluorescence, respectively. At the in vivo level, female Balb/c mice were treated with AhR/CYP1A1 inducer and histopathology changes and Immunohistochemistry examination for target proteins were determined.ResultsThe constitutive mRNA expression and cellular content of CYP1A1 and CYP1B1, AhR-regulated genes, were markedly higher in CSCs more than differentiating non-CSCs of five different human breast cancer cells. Activation of AhR/CYP1A1 in MCF-7 cells by TCDD and DMBA, strong AhR activators, significantly increased CSC-specific markers, mammosphere formation, aldehyde dehydrogenase (ALDH) activity, and percentage of side population (SP) cells, whereas inactivation of AhR/CYP1A1 using chemical inhibitor, α-naphthoflavone (α-NF), or by genetic shRNA knockdown, significantly inhibited the upregulation of ALDH activity and SP cells. Importantly, inactivation of the AhR/CYP1A1 significantly increased sensitization of CSCs to the chemotherapeutic agent doxorubicin. Mechanistically, Induction of AhR/CYP1A1 by TCDD and DMBA was associated with significant increase in β-Catenin mRNA and protein expression, nuclear translocation and its downstream target Cyclin D1, whereas AhR or CYP1A1 knockdown using shRNA dramatically inhibited β-Catenin cellular content and nuclear translocation. This was associated with significant inhibition of PTEN and induction of total and phosphorylated Akt protein expressions. Importantly, inhibition of PI3K/Akt pathway by LY294002 completely blocked the TCDD-induced SP cells expansion. In vivo, IHC staining of mammary gland structures of untreated and DMBA (30 mg/kg, IP)- treated mice, showed tremendous inhibition of PTEN expression accompanied with an increase in the expression p-Akt, β-Catenin and stem cells marker ALDH1.ConclusionsThe present study provides the first evidence that AhR/CYP1A1 signaling pathway is controlling breast CSCs proliferation, development, self-renewal and chemoresistance through inhibition of the PTEN and activation of β-Catenin and Akt pathways.

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Section: Methodsmentioning

confidence: 99%

Aryl hydrocarbon receptor/cytochrome P450 1A1 pathway mediates breast cancer stem cells expansion through PTEN inhibition and β-Catenin and Akt activation

2017

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“…In other experiments the expression of CD90 with a value above 80% in AdMSC populations of different species was verified (Al-Nbaheen et al, 2013;Carvalho et al, 2013;Maia et al, 2012). However, as MSC markers from bone marrow and adipose tissue in humans, they are the same in dermis fibroblast cells and olfactory epithelium, and the development of a standardized technique for the isolation of different populations in clinical practice (Wetzig et al, 2013) is required. In humans, MSCs from bone marrow, adipose tissue, umbilical cord and neural origin showed common expression for a set of markers, the difficulty to standardize the reported immunophenotypic profile in other studies being attributed to the absence of evaluation of the set of specific marker variations in the protocols used and the lack of characterization of several obtaining sites (New et al, 2015) Despite the minimum criteria proposed for human MSCs (Dominici et al, 2006), CD markers are quite controversial.…”

Section: Discussionmentioning

confidence: 99%

Immunophenotyping, plasticity tests and nanotagging of stem cells derived from adipose tissue of wild rodent agouti (Dasyprocta prymnolopha)

Rocha

Leite

Silva

et al. 2019

Arq. Bras. Med. Vet. Zootec.

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There is a growing interest in the study of unspecialized mesenchymal stem cells, for there are still some discussions about their in vitro behavior. Regenerative medicine is a science undergoing improvement which develops treatments as cell therapy using somatic stem cells. In several studies, adipose tissue is presented as a source of multipotent adult cells that has several advantages over other tissue sources. This study aimed to characterize and evaluate the tagging of mesenchymal stem cells from the agoutis adipose tissue (Dasyprocta prymonolopha), with fluorescent intracytoplasmic nanocrystals. Fibroblast cells were observed, plastic adherent, with extended self-renewal, ability to form colonies, multipotency by differentiation into three lineages, population CD90 + and CD45 - expression, which issued high red fluorescence after the tagging with fluorescent nanocrystals by different paths and cryopreserved for future use. It is possible to conclude that mesenchymal stem cells from agouti adipose tissue have biological characteristics and in vitro behavior that demonstrate its potential for use in clinical tests.

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“…Equal amount of proteins was taken from each sample to generate a pool of patients as one group, a pool of carriers, and a third pool of controls. For each analysis sample group, 200 μg complex protein mixtures was subjected to in-solution digestion for mass spectrometry analysis as previously described [ 22 , 23 ]. We have used the 1-Dimensional Nano Acquity liquid chromatography coupled with tandem mass spectrometry on Synapt G2 (Waters, Manchester, UK) to generate expression proteomics data based on quantitative protein changes between the three sample groups.…”

Section: Methodsmentioning

confidence: 99%

The molecular significance of methylated BRCA1 promoter in white blood cells of cancer-free females

et al. 2014

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BackgroundBRCA1 promoter methylation has been detected in DNA from peripheral blood cells of both breast cancer patients and cancer-free females. However, the pathological significance of this epigenetic change in white blood cells (WBC) remains an open question. In this study, we hypothesized that if constitutional BRCA1 methylation reflects an elevated risk for developing breast cancer (BC), WBC that harbor methylated BRCA1 in both cancer-free females and BC patients should exhibit similar molecular changes.MethodsBRCA1 promoter methylation was examined by methylation-specific PCR in WBC from 155 breast cancer patients and 143 cancer-free females. The Human Breast Cancer EpiTect Methyl II Signature PCR Array and The Human Breast Cancer RT2 Profiler™ PCR Array were used to study the methylation status and the expression profile of several breast cancer-related genes, respectively. In addition, we used label-free MS-based technique to study protein expression in plasma.ResultsWe have shown that 14.2% of BC patients and 9.1% of cancer-free females (carriers) harbored methylated BRCA1 promoter in their WBC. Interestingly, 66.7% of patients harbored methylated BRCA1 promoter in both WBC and tumors. Importantly, we have shown the presence of epigenetic changes in 9 other BC-related genes in WBC of both patients and carriers. Additionally, BRCA1 and 15 other important cancer –related genes were found to be differentially expressed in WBC from patients and carriers as compared to controls. Furthermore, we have shown that the carriers exhibited a unique plasma protein pattern different from those of BC patients and controls, with 10 proteins similarly differentially expressed in patients and carriers as compared to controls.ConclusionsThe present results suggest the presence of a strong link between aberrant methylation of the BRCA1 promoter in WBC and breast cancer –related molecular changes, which indicate the potential predisposition of the carriers for developing breast cancer. This informs the potential use of the aberrant methylation of BRCA1 promoter in WBC as a powerful non-invasive molecular marker for detecting predisposed individuals at a very early age.

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Differential marker expression by cultures rich in mesenchymal stem cells

Cited by 36 publications

References 45 publications

Aryl hydrocarbon receptor/cytochrome P450 1A1 pathway mediates breast cancer stem cells expansion through PTEN inhibition and β-Catenin and Akt activation

Aryl hydrocarbon receptor/cytochrome P450 1A1 pathway mediates breast cancer stem cells expansion through PTEN inhibition and β-Catenin and Akt activation

Immunophenotyping, plasticity tests and nanotagging of stem cells derived from adipose tissue of wild rodent agouti (Dasyprocta prymnolopha)

The molecular significance of methylated BRCA1 promoter in white blood cells of cancer-free females

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