Bmi1-positive cells in the lingual epithelium could serve as cancer stem cells in tongue cancer.
ABSTRACT: We recently reported that the polycomb complex protein Bmi1 is a marker for lingual epithelial stem cells (LESCs), which are involved in the long-term maintenance of lingual epithelial tissue in the physiological state. However, the precise role of LESCs in generating tongue tumors and Bmi1-positive cell lineage dynamics in tongue cancers are unclear. Here, using a mouse model of chemically (4-nitroquinoline-1-oxide: 4-NQO) induced tongue cancer and the multicolor lineage tracing method, we found that each unit of the tumor was generated by a single cell and that the assembly of such cells formed a polyclonal tumor. Although many Bmi1-positive cells within the tongue cancer specimens failed to proliferate, some proliferated continuously and supplied tumor cells to the surrounding area. This process eventually led to the formation of areas derived from single cells after 1-3 months, as determined using the multicolor lineage tracing method, indicating that such cells could serve as cancer stem cells. These results indicate that LESCs could serve as the origin for tongue cancer and that cancer stem cells are present in tongue tumors.
Project description:Despite the strong need for the establishment of a lingual epithelial cell culture system, a simple and convenient culture method has not yet been established. Here, we report the establishment of a novel lingual epithelium organoid culture system using a three-dimensional matrix and growth factors. Histological analyses showed that the generated organoids had both a stratified squamous epithelial cell layer and a stratum corneum. Very recently, we showed via a multicolor lineage tracing method that Bmi1-positive stem cells exist at the base of the epithelial basal layer in the interpapillary pit. Using our new culture system, we found that organoids could be generated by single Bmi1-positive stem cells and that in the established organoids, multiple Bmi1-positive stem cells were generated at the outermost layer. Moreover, we observed that organoids harvested at an early point in culture could be engrafted and maturate in the tongue of recipient mice and that the organoids generated from carcinogen-treated mice had an abnormal morphology. Thus, this culture system presents valuable settings for studying not only the regulatory mechanisms of lingual epithelium but also lingual regeneration and carcinogenesis.
Project description:Although the existence of cancer stem cells in intestine tumors has been suggested, direct evidence has not been yet provided. Here, we showed, using the multicolor lineage-tracing method and mouse models of intestinal adenocarcinoma and adenoma that Bmi1- or Lgr5- positive tumorigenic cells clonally expanded in proliferating tumors. At tumor initiation and during tumor propagation in the colon, the descendants of Lgr5-positive cells clonally proliferated to form clusters. Clonal analysis using ubiquitous multicolor lineage tracing revealed that colon tumors derived from Lgr5-positive cells were monoclonal in origin but eventually merged with neighboring tumors, producing polyclonal tumors at the later stage. In contrast, the origin of small intestine tumors was likely polyclonal, and during cancer progression some clones were eliminated, resulting in the formation of monoclonal tumors, which could merge similar to colon tumors. These results suggest that in proliferating intestinal neoplasms, Bmi1- or Lgr5-positive cells represent a population of cancer stem cells, whereas Lgr5-positive cells also function as cells-of-origin for intestinal tumors.
Project description:Taste bud cells arise from local epithelial stem cells in the oral cavity and are continuously replaced by newborn cells throughout an animal's life. However, little is known about the molecular and cellular mechanisms of taste cell turnover. Recently, it has been demonstrated that SOX2, a transcription factor expressed in epithelial stem/progenitor cells of the oral cavity, regulates turnover of anterior tongue epithelium including gustatory and non-gustatory papillae. Yet, the role of SOX2 in regulating taste cell turnover in the posterior tongue is unclear. Prompted by the fact that there are regional differences in the cellular and molecular composition of taste buds and stem/progenitor cells in the anterior and posterior portions of tongue, which are derived from distinct embryonic origins, we set out to determine the role of SOX2 in epithelial tissue homeostasis in the posterior tongue. Here we report the differential requirement of SOX2 in the stem/progenitor cells for the normal turnover of lingual epithelial cells in the posterior tongue. Sox2 deletion in the stem/progenitor cells neither induced active caspase 3-mediated apoptotic cell death nor altered stem/progenitor cell population in the posterior tongue. Nevertheless, morphology and molecular feature of non-gustatory epithelial cells were impaired in the circumvallate papilla but not in the filiform papillae. Remarkably, taste buds became thinner, collapsed, and undetectable over time. Lineage tracing of Sox2-deleted stem/progenitor cells demonstrated an almost complete lack of newly generated basal precursor cells in the taste buds, suggesting mechanistically that Sox2 is involved in determining stem/progenitor cells to differentiate to gustatory lineage cells. Together, these results demonstrate that SOX2 plays key roles in regulating epithelial tissue homeostasis in the posterior tongue, similar but not identical to its function in the anterior tongue.
Project description:The use of mice that are mosaic for reporter gene expression underlies many lineage-tracing studies in stem cell biology. For example, using mosaic LacZ reporter mice, it was shown that limbal epithelial stem cells (LESCs) around the periphery of the cornea maintain radial sectors of the corneal epithelium and that radial stripe numbers declined with age. Originally, the corneal results were interpreted as progressive, age-related loss or irreversible inactivation of some LESC clones. In this study we used computer simulations to show that these results could also be explained by stochastic replacement of LESCs by neighbouring LESCs, leading to neutral drift of LESC populations. This was shown to reduce the number of coherent clones of LESCs and hence would coarsen the mosaic pattern in the corneal epithelium without reducing the absolute number of LESCs. Simulations also showed that corrected stripe numbers declined more slowly when LESCs were grouped non-randomly and that mosaicism was rarely lost unless simulated LESC numbers were unrealistically low. Possible reasons why age-related changes differ between mosaic corneal epithelia and other systems, such as adrenal cortices and intestinal crypts, are discussed.
Project description:The limbal epithelial stem cell (LESC) hypothesis proposes that LESCs in the corneal limbus maintain the corneal epithelium both during normal homeostasis and wound repair. The alternative corneal epithelial stem cell (CESC) hypothesis proposes that LESCs are only involved in wound repair and CESCs in the corneal epithelium itself maintain the corneal epithelium during normal homeostasis. We used tamoxifen-inducible, CreER-loxP lineage tracing to distinguish between these hypotheses. Clones of labelled cells were induced in adult CAGG-CreER;R26R-LacZ reporter mice and their distributions analysed after different chase periods. Short-lived clones, derived from labelled transient amplifying cells, were shed during the chase period and long-lived clones, derived from stem cells, expanded. At 6 weeks, labelled clones appeared at the periphery, extended centripetally as radial stripes and a few reached the centre by 14 weeks. Stripe numbers depended on the age of tamoxifen treatment. Stripes varied in length, some were discontinuous, few reached the centre and almost half had one end at the limbus. Similar stripes extended across the cornea in CAGG-CreER;R26R-mT/mG reporter mice. The distributions of labelled clones are inconsistent with the CESC hypothesis and support the LESC hypothesis if LESCs cycle between phases of activity and quiescence, each lasting several weeks.
Project description:Identification of defined cell populations with stem/progenitor properties is key for understanding prostate development and tumorigenesis. Here we show that the polycomb repressor protein Bmi1 marks a population of castration-resistant luminal epithelial cells enriched in the mouse proximal prostate. We employ lineage tracing to show that these castration-resistant Bmi1-expressing cells (or CARBs) are capable of tissue regeneration and self-renewal. Notably, CARBs are distinct from the previously described luminal castration-resistant Nkx3.1-expressing cells (CARNs). CARBs can serve as a prostate cancer cell-of-origin upon Pten deletion, yielding luminal prostate tumours. Clonal analysis using the R26R-confetti allele indicates preferential tumour initiation from CARBs localized to the proximal prostate. These studies identify Bmi1 as a marker for a distinct population of castration-resistant luminal epithelial cells enriched in the proximal prostate that can serve as a cell of origin for prostate cancer.
Project description:Squamous cell carcinoma in the head and neck (HNSCC) is a common yet poorly understood cancer, with adverse clinical outcomes due to treatment resistance, recurrence, and metastasis. Putative cancer stem cells (CSCs) have been identified in HNSCC, and BMI1 expression has been linked to these phenotypes, but optimal treatment strategies to overcome chemotherapeutic resistance and eliminate metastases have not yet been identified. Here we show through lineage tracing and genetic ablation that BMI1+ CSCs mediate invasive growth and cervical lymph node metastasis in a mouse model of HNSCC. This model and primary human HNSCC samples contain highly tumorigenic, invasive, and cisplatin-resistant BMI1+ CSCs, which exhibit increased AP-1 activity that drives invasive growth and metastasis of HNSCC. Inhibiting AP-1 or BMI1 sensitized tumors to cisplatin-based chemotherapy, and it eliminated lymph node metastases by targeting CSCs and the tumor bulk, suggesting potential regimens to overcome resistance to treatments and eradicate HNSCC metastasis.
Project description:Colon cancers are composed of phenotypically heterogeneous tumor cell subpopulations with variable expression of putative stem cell and differentiation antigens. While in normal colonic mucosa, clonal repopulation occurs along differentiation gradients from crypt base toward crypt apex, the clonal architecture of colon cancer and the relevance of tumor cell subpopulations for clonal outgrowth are poorly understood. Using a multicolor lineage tracing approach in colon cancer xenografts that reflect primary colon cancer architecture, we here demonstrate that clonal outgrowth is mainly driven by tumor cells located at the leading tumor edge with clonal axis formation toward the tumor center. While our findings are compatible with lineage outgrowth in a cancer stem cell model, they suggest that in colorectal cancer tumor cell position may be more important for clonal outgrowth than tumor cell phenotype.
Project description:A central question in stem cell biology is whether organ homeostasis is maintained in adult organs through undifferentiated stem cells or self-duplication of specialized cell populations. To address this issue in the exocrine pancreas we analyzed the Bmi1-labeled cell lineage of pancreatic acinar cells. Previously, we had shown that inducible linage tracing with Bmi1-Cre-estrogen receptor (ER) in the small intestine specifically, labels "classical" undifferentiated intestinal stem cells. In this article we demonstrate that the Bmi1-Cre-ER system labels a subpopulation of differentiated acinar cells in the exocrine pancreas whose derivatives are still present, at a steady-state level, 1 year after a single TM pulse. This study suggests that Bmi1 is a marker for a subpopulation of self-renewing acinar cells, indicating that self-renewal is not an exclusive feature of adult undifferentiated stem cells. Further, the extended period that Bmi1-labeled acinar cells retain a pulse of BrdU suggests that some of this subpopulation of cells are not continuously replicating, but rather are set aside until needed. This cellular behavior is again reminiscent of behavior normally associated with more classical adult stem cells. Setting aside cells capable of self-renewal until needed retains the advantage of protecting this subpopulation of cells from DNA damage induced during replication.
Project description:Lineage tracing has become the method of choice to study the fate and dynamics of stem cells (SCs) during development, homeostasis, and regeneration. However, transgenic and knock-in Cre drivers used to perform lineage tracing experiments are often dynamically, temporally, and heterogeneously expressed, leading to the initial labeling of different cell types and thereby complicating their interpretation. Here, we developed two methods: the first one based on statistical analysis of multicolor lineage tracing, allowing the definition of multipotency potential to be achieved with high confidence, and the second one based on lineage tracing at saturation to assess the fate of all SCs within a given lineage and the "flux" of cells between different lineages. Our analysis clearly shows that, whereas the prostate develops from multipotent SCs, only unipotent SCs mediate mammary gland (MG) development and adult tissue remodeling. These methods offer a rigorous framework to assess the lineage relationship and SC fate in different organs and tissues.