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Tumorsphere Derivation and Treatment from Primary Tumor Cells Isolated from Mouse Rhabdomyosarcomas
Published on: September 13, 2019
Mouse fibroblasts lacking RB1 function form spheres and undergo reprogramming to a cancer stem cell phenotype
Yongqing Liu1, Brian Clem, Ewa K Zuba-Surma
1Molecular Targets Program, Brown Cancer Center, University of Louisville Health Sciences Center, Louisville, KY 40202, USA.
Abstract:
Activation of the RB1 pathway triggers the cell-cycle arrest that mediates cell-cell contact inhibition. Accordingly, mutation of all three RB1 family members leads to loss of contact inhibition and outgrowth of fibroblasts into spheres where cell-cell contacts predominate. We present evidence that such outgrowth triggers reprogramming to generate cells with properties of cancer stem cells. Fibroblasts with only a single RB1 mutation remain contact inhibited; however, if this contact inhibition is bypassed by forcing the RB1(-/-) cells to form spheres in suspension, cells with properties of cancer stem cells are also generated. These cells not only form tumors in nude mice but also generate differentiated cells. We propose that contact inhibition imposed by the RB1 pathway performs an unexpected tumor suppressor function by preventing cell outgrowth into structures where cells with properties of cancer stem cells can be generated from differentiated somatic cells in advancing cancers.
Insights
Loss of RB1 pathway function causes cell outgrowth, generating cancer stem cells. This suggests RB1
Area of Science:
- Cell Biology
- Cancer Research
- Molecular Biology
Background:
- The RB1 pathway is crucial for cell-cycle arrest and contact inhibition.
- Mutations in RB1 family members disrupt these processes, leading to uncontrolled cell growth.
- Contact inhibition normally prevents excessive cell proliferation in differentiated somatic cells.
Purpose of the Study:
- To investigate the role of RB1 pathway-mediated contact inhibition in preventing cancer stem cell generation.
- To explore how bypassing contact inhibition in RB1-mutated cells affects cellular reprogramming.
- To understand the tumor suppressor function of RB1 in the context of cancer stem cell formation.
Main Methods:
- Fibroblast cell culture and manipulation of RB1 gene status.
- Induction of cell-cell contact and sphere formation in suspension culture.
- Tumorigenicity assays in nude mice.
- Analysis of cell differentiation and stem cell properties.
Main Results:
- Complete RB1 mutation led to loss of contact inhibition and fibroblast outgrowth into spheres.
- These outgrowing cells exhibited cancer stem cell properties, forming tumors and differentiating.
- Even with partial RB1 mutation, forcing sphere formation bypassed contact inhibition and generated cancer stem cells.
Conclusions:
- RB1 pathway-mediated contact inhibition acts as a tumor suppressor by preventing the generation of cancer stem cells from differentiated somatic cells.
- Loss of RB1 function and subsequent cell outgrowth create conditions conducive to cancer stem cell formation.
- Maintaining contact inhibition is critical for preventing the acquisition of cancer stem cell characteristics in advancing cancers.
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