scholarly journals Culture of Mouse Amniotic Fluid-Derived Cells on Irradiated STO Feeders Results in the Generation of Primitive Endoderm Cell Lines Capable of Self-Renewal in vitro

2013 ◽  
Vol 198 (2) ◽  
pp. 111-126 ◽  
Author(s):  
Aleksandar M. Babic ◽  
Sunyoung Jang ◽  
Eugenia Nicolov ◽  
Horatiu Voicu ◽  
Chance J. Luckey
Keyword(s):  
Amniotic Fluid ◽  
Cell Lines ◽  
Primitive Endoderm ◽  
Self Renewal ◽  
Endoderm Cell

The α2δ1 subunit of the voltage-gated calcium channel as a potential candidate for breast cancer tumor initial cells biomarker

2021 ◽  
pp. 1-11
Author(s):  
Meng Li ◽  
Wenmin Zhang ◽  
Xiaodan Yang ◽  
Guo An ◽  
Wei Zhao
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Cell Lines ◽  
Cancer Cell ◽  
Breast Cancer Cell ◽  
Breast Cancer Cells ◽  
Cancer Cell Lines ◽  
Breast Cancer Cell Lines ◽  
Self Renewal

BACKGROUND: The voltage-gated calcium channel subunit alpha 2 delta 1 (α2δ1) is a functional tumor initial cells (TICs) marker for some solid cancer cells. This study aimed to investigate whether α2δ1 can be used as a potential TIC marker for breast cancer cells. METHODS: α2δ1+ and α2δ1- cells were identified and sorted from the breast cancer cell lines MDA-MB-231, MDA-MB-435s and ZR-75-1 by Immunofluorescence (IF) and Fluorescent-activated cell sorting (FACS) analyses. Spheroid formation in vitro and tumorigenesis in NOD/SCID mice were assessed to determine the self-renewal and serial transplantation abilities of these cells. Using a lentivirus infection system for α2δ1 in breast cancer cell lines, we determined the mRNA levels of stemnessassociated genes by quality real-time PCR (qRT-PCR). Boyden chamber and wounding assays were further performed to detect the migration of α2δ1 overexpression cells. Bioinformatics explored the relationship of molecular classification of breast cancer and drug resistance. RESULTS: α2δ1 presents on the cytomembrane of breast cancer cells, with a positive rate of 1.5–3%. The α2δ1+ cells in breast cancer cell lines have a stronger self-renewal ability and tumor initiating properties in vitro and in vivo. Overexpressing α2δ1 successfully enhanced the sphere-forming efficiency, and upregulated the expression of stemness-associated genes, and increased cell migration. However, seldom significant was available between estrogen receptor +/- (ER+/-), progesterone receptor (PR+/-), and Her2+/-. CONCLUSIONS: Breast cancer cells positive for the α2δ1 charactered tumor initiation, and α2δ1 is a potential TIC marker for breast cancer that further promotes the migration.

CHEMOTHERAPEUTIC POTENTIAL OF NOVEL XANTHONE SOURCED FROM SWERTIA CHIRATA AGAINST SKIN CARCINOGENESIS

2020 ◽  
pp. 84-88
Author(s):  
ATISH BARUA ◽  
PRITHA CHOUDHURY ◽  
CHINMAY KUMAR PANDA ◽  
PROSENJIT SAHA
Keyword(s):  
Skin Cancer ◽  
Antitumor Activity ◽  
Cell Line ◽  
Cell Lines ◽  
Cancer Cell ◽  
Cancer Cell Lines ◽  
Self Renewal ◽  
Qrt Pcr ◽  
Swertia Chirata

Objective: Swertia chirata forms a rich source of bio-active compounds, among which xanthones form an important part. Among the xanthones present in it, 1,5,8 Tri-hydroxy-3-methoxy xanthone (TMX) was found to be the most active. The present study aims to evaluate the chemotherapeutic potential of it against metastatic skin cancer cell lines. Methods: In this study, the antitumor activity of TMX (the active component of chirata plant) was evaluated in A431, SKMEL-5, and A375 cell line by using in-vitro assays such as cell viability assay, cell cycle analysis, caspase 3 activity assay, intracellular reactive oxygen species (ROS) level determination by dichlorofluorescein diacetate, and quantitative real-time polymerase chain reaction (qRT-PCR). Results: In vitro studies showed that TMX from S. chirata exhibited significant antitumor activity by inducing apoptosis and restricting proliferation in both melanoma and non-melanoma skin cancer cell lines, but no such activity was seen in normal skin cancer cell line WS1. The qRT-PCR analysis revealed that in both the melanoma ad non-melanoma cell lines, TMX could exert its antitumor activity by downregulating c-Myc, cyclin-D1, and β-catenin and up-regulating Wnt antagonist gsk-3β, thereby suppressing wnt self-renewal pathway, but such regulation was absent in normal cell line. Conclusions: TMX from chirata could effectively inhibit the proliferation of metastatic skin cancer (both melanoma and non-melanoma) cell lines while being non-toxic to normal cell lines. The chemotherapeutic potential of TMX against metastatic skin cancer cell lines was achieved by downregulating several key regulatory genes enabling the suppression of the self-renewal pathway, the chief reason behind the invasiveness of cancer cells.

scholarly journals SOX17 Is Not Required for the Derivation and Maintenance of Mouse Extraembryonic Endoderm Stem Cell Lines

2022 ◽  
Author(s):  
Xudong Dong ◽  
Ailing Ding ◽  
Jiangwei Lin
Keyword(s):  
Stem Cell ◽  
Cell Lines ◽  
Embryonic Stem ◽  
Cell Lineage ◽  
Preimplantation Embryos ◽  
Primitive Endoderm ◽  
Extraembryonic Endoderm ◽  
Gene Expression Analyses ◽  
Stem Cell Lines

Extraembryonic endoderm stem (XEN) cell lines can be derived and maintained in vitro and reflect the primitive endoderm cell lineage. SOX17 is thought to be required for the derivation and maintenance of mouse XEN cell lines. Here we have re-evaluated this requirement for SOX17. We derived multiple SOX17-deficient XEN cell lines from preimplantation embryos of a SOX17-Cre knockout strain and chemically converted multiple SOX17-deficient embryonic stem cell lines into XEN cell lines by transient culturing with retinoic acid and Activin A. We confirmed the XEN profile of SOX17-deficient cell lines by immunofluorescence with various markers, by NanoString gene expression analyses, and by their contribution to the extraembryonic endoderm of chimeric embryos produced by injecting these cells into blastocysts. Thus, SOX17 is not required for the derivation and maintenance of XEN cell lines.

lncRNA, PVT-1, controls tumorigenesis and cancer stem-like phenotypes in osteosarcoma through PI3K/TSC2.

2020 ◽  
Vol 38 (15_suppl) ◽  
pp. e23512-e23512
Author(s):  
Susan Tsang ◽  
Nino Carlo Rainusso ◽  
Jason Todd Yustein
Keyword(s):  
Stem Cell ◽  
Cancer Stem Cell ◽  
Cell Lines ◽  
Bone Cancer ◽  
High Rate ◽  
Osteosarcoma Cell ◽  
Self Renewal ◽  
Cell Phenotypes

e23512 Background: Osteosarcoma is the most common pediatric bone cancer and a key genetic characteristic of this particular malignancy is its complex karyotype. Specifically it has been reported that 40% of osteosarcoma patients’ present with 8q24 amplification. The presence of this specific amplification has been previously associated with a high rate of relapse and poor prognosis for osteosarcoma patients. Within this amplicon resides, a long non-coding RNA gene, PVT-1. Prior studies indicates that PVT-1 has pro-oncogenic properties however the function of PVT-1 in osteosarcoma is not well characterized. Methods: To understand PVT-1 copy number, Fluorescent In Situ Hybridization was performed on both osteosarcoma cell lines and osteosarcoma patient-derived xenografts. In addition the PVT-1 RNA level is elevated in a majority of osteosarcoma samples compared to normal bone. To test PVT-1 pro-oncogenic role in osteosarcoma, several functional assays were performed. Results: Our studies demonstrated that overexpression of PVT-1 in osteosarcoma cell lines promotes multiple tumorigenic behaviors including enhanced proliferation, migration, invasion and chemotherapeutic resistance to cisplatin. PVT-1’s ability to mediate metastasis and contribute to chemotherapeutic sensitivity is a shared phenotype of cancer stem cells. Based on this observation, we hypothesize targeting PVT-1 will reduce cancer stem-cell properties. Osteosarcoma lines with increased levels of PVT-1 exhibited higher expression of cancer stem cell genes: Nanog, SOX2, c-Myc, and Oct4 at both the transcriptomic and proteomic level. In Vitro and In Vivo self-renewal capacity studies showed enhanced osteosarcoma cell self-renewal in the PVT-1 overexpression cohort. Additional molecular studies were performed in order to gain additional insights into potential mechanism of action for PVT-1 including Reverse Phase Protein Array. Initial analysis suggest a role for PVT-1 in regulating the PI3K-AKT-TSC2 pathway. Conclusions: This suggests a potential oncogenic pathway in which PVT-1 enhances cancer stem cell phenotypes. On-going investigations are addressing potential PI3K/TSC2 pathway inhibitors, BEZ-2335 and LY3023414, which could be utilized to regulate PVT-1 mediated tumorigenic roles and cancer stem-like properties.

scholarly journals Mutated p53 Promotes the Symmetric Self-Renewal of Cisplatin-Resistant Lung Cancer Stem-Like Cells and Inhibits the Recruitment of Macrophages

2019 ◽  
Vol 2019 ◽  
pp. 1-9
Author(s):  
Yu Xu ◽  
Zhi Xu ◽  
Qi Li ◽  
Liang Guo ◽  
Yao Wang ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Cell Lines ◽  
Cancer Cell ◽  
Cancer Recurrence ◽  
Cancer Cell Lines ◽  
The Self ◽  
Mutant P53 ◽  
Lung Cancer Cell ◽  
Self Renewal

It has been proposed that mutant p53 is correlated with the recurrence of lung cancer. Recently, a small population of cells with asymmetric or symmetric self-renewal potential has been identified in lung cancer, which was termed as cancer stem-like cells (CSCs) and was speculated to be the reason for cancer recurrence after chemotherapy. In this study, we used lung cancer cell lines with different TP53 backgrounds to elucidate the potential role of mutant p53 in regulating lung CSC self-renewal and on lung cancer recurrence. Cisplatin-resistant lung cancer cells with different TP53 backgrounds were generated in vitro by exposing A549, H460, and H661 lung cancer cell lines repeatedly to cisplatin. CD44+/CD90+ stem-like cells were identified in above cisplatin-resistant lung cancers (termed as cisplatin-resistant lung cancer stem-like cells, (Cr-LCSCs)) and stained with PKH26 dye which was used to define the self-renewal pattern. The proportion of symmetric divisions was significantly higher in Cr-LCSCs with mutant (mt) p53 compared with Cr-LCSCs with wild-type (wt) p53, and forced expression of mt p53 promoted the symmetric division of Cr-LCSCs. Furthermore, fewer macrophages accumulated in subcutaneously implanted xenografts consisting of mt p53 Cr-LCSCs compared with wt p53 Cr-LCSCs. These results indicated that mt p53 might accelerate the recurrence of lung cancer by regulating the self-renewal kinetics of Cr-LCSCs as well as the recruitment of macrophages.

scholarly journals Skipping Exon-v6 from CD44v6-Containing Isoforms Influences Chemotherapy Response and Self-Renewal Capacity of Gastric Cancer Cells

Cancers ◽  
2020 ◽  
Vol 12 (9) ◽  
pp. 2378
Author(s):  
Silvana Lobo ◽  
Carla Pereira ◽  
Carla Oliveira ◽  
Gabriela M. Almeida
Keyword(s):  
Gastric Cancer ◽  
Cell Lines ◽  
De Novo ◽  
Chemotherapy Response ◽  
Gastric Cancer Cells ◽  
Self Renewal ◽  
Cd44 Isoforms ◽  
Reading Frame ◽  
Phosphorodiamidate Morpholino Oligomers

De novo expressed CD44 isoforms containing exon-v6 are frequently associated with gastric cancer (GC) aggressiveness, and may predict chemotherapy response in vitro. Whether exon-v6 itself is responsible for conferring these properties to CD44v6-containing isoforms remains to be elucidated. CRISPR/Cas9 and Phosphorodiamidate Morpholino oligomers (PMOs) were used to induce specific exon-v6 skipping, maintaining the CD44 reading frame, in two GC cell lines endogenously expressing CD44v6. Cisplatin and 5-fluorouracil treatment response, and self-renewal ability was compared between CRISPR/Cas9-edited, CD44v6 knockdown and mock cells. We obtained homozygous genome-edited cell lines with exon-v6 deletion. Edited cells transcribed CD44v isoforms presenting in frame v5–v7 splicing, mimicking exon-v6 skipping. Results showed that removing specifically exon-v6 sensitizes cells to cisplatin and impairs cells’ self-renewal ability, similarly to CD44v6 knockdown. In parallel, we also tested a clinically feasible approach for transient exon-v6 skipping with a PMO-based strategy. We demonstrate that exon-v6 specific removal from CD44v isoforms increases cell sensitivity to cisplatin and impairs GC cells self-renewal. We trust that a PMO approach designed towards CD44v6 overexpressing GC cells may be a suitable approach to sensitize tumor cells for conventional therapy.

scholarly journals Inhibiting Oncogenic RAS in Multiple Myeloma By Targeting Scaffold-ERK Interactions

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 2089-2089
Author(s):  
Christian B. Gocke ◽  
Qiuju Wang ◽  
Ross McMillan ◽  
Vesselin Penchev ◽  
Julien Sage ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Disease Progression ◽  
Cell Lines ◽  
Mapk Signaling ◽  
Ras Signaling ◽  
Self Renewal ◽  
Oncogenic Ras ◽  
Clonogenic Growth ◽  
Tumor Regrowth

Abstract The introduction of novel therapeutic agents over the past decade has significantly improved the outcomes of patients with newly diagnosed multiple myeloma (MM). Despite these advances, virtually all patients relapse and develop drug-resistant disease that carries a dismal prognosis. In addition to chemoresistance, relapse and tumor regrowth are dependent on self-renewal to maintain clonogenic growth potential over time. A better understanding of the factors responsible for tumor self-renewal and regrowth may lead to novel therapies and improved survival rates. RAS mutations are common in MM and have been demonstrated to increase in frequency with relapse and disease progression. These findings implicate RAS in chemoresistance, self-renewal, and tumor regrowth upon relapse. We have begun to directly examine the functional role of mutant RAS signaling in MM by stably transducing KRASmut MM cells with a tetracycline-inducible shRNA and found that KRAS knockdown decreases MM clonogenic growth in vitro. Although inhibiting oncogenic RAS may be beneficial for treating relapsed disease it remains an elusive drug target. Recently, our collaborators have developed a novel small peptide RAS signaling inhibitor that mimics the WW domain of IQGAP1 which competes for ERK binding to scaffolds. The WW peptide specifically targets solid tumors containing RAS (or BRAF) mutations and is well tolerated in mouse models. Thus, by targeting only tumorigenic cells it may demonstrate decreased toxicity compared to existing small molecule inhibitors of MAPK signaling. We analyzed gene expression profiling data from the University of Arkansas (GSE2658) and found that elevated IQGAP1 expression levels (top 12.5%) are associated with increased disease-related mortality in MM (log-rank p = 0.0002, HR 4.3, 95% CI 2.0-9.1). This suggests that IQGAP1 may play an important biological role in MM disease progression and that inhibition of IQGAP1 and RAS signaling with the WW peptide may have efficacy and specificity in MM patients. We have treated RAS mutated MM cell lines with the WW peptide or a scrambled (SCR) control peptide and found that the WW peptide effectively decreases pERK levels after 24 hours at doses as low as 20 nM. The WW peptide also decreases cell growth in RAS mutated but not wild-type MM cells by inducing a cell cycle arrest (by BrdU/7-AAD staining). Importantly, treatment of RAS mutated MM cell lines with the WW peptide significantly decreased colony formation in vitro and led to a reduction in the frequency of highly clonogenic CD138neg MM cells. Finally, an immunodeficient mouse xenograft model was used to determine the effect of the WW peptide on engraftment potential. We treated NCI-H929NRAS(G13D) cells with SCR or WW peptide and injected them into NOD/SCID/IL2γchainKO (NSG) mice. Engraftment as detected by kappa light chain ELISA was significantly reduced in the WW peptide treated group. These data further implicate RAS signaling as well as MAPK signaling in MM self-renewal and identify a potential novel therapy for treatment of relapsed/refractory MM. Disclosures No relevant conflicts of interest to declare.

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