Preexisting Stem Cells Research Articles

Cloning a profibrotic stem cell variant in idiopathic pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressive, irreversible, and rapidly fatal interstitial lung disease marked by the replacement of lung alveoli with dense fibrotic matrices. Although the mechanisms initiating IPF remain unclear, rare and common alleles of genes expressed in lung epithelia, combined with aging, contribute to the risk for this condition. Consistently, single-cell RNA sequencing (scRNA-seq) studies have identified lung basal cell heterogeneity in IPF that might be pathogenic. We used single-cell cloning technologies to generate "libraries" of basal stem cells from the distal lungs of 16 patients with IPF and 10 controls. We identified a major stem cell variant that was distinguished from normal stem cells by its ability to transform normal lung fibroblasts into pathogenic myofibroblasts in vitro and to activate and recruit myofibroblasts in clonal xenografts. This profibrotic stem cell variant, which was shown to preexist in low quantities in normal and even fetal lungs, expressed a broad network of genes implicated in organ fibrosis and showed overlap in gene expression with abnormal epithelial signatures identified in previously published scRNA-seq studies of IPF. Drug screens highlighted specific vulnerabilities of this profibrotic variant to inhibitors of epidermal growth factor and mammalian target of rapamycin signaling as prospective therapeutic targets. This profibrotic stem cell variant in IPF was distinct from recently identified profibrotic stem cell variants in chronic obstructive pulmonary disease and may extend the notion that inappropriate accrual of minor and preexisting stem cell variants contributes to chronic lung conditions.

More than a simple epithelial layer: multifunctional role of echinoderm coelomic epithelium

In echinoderms, the coelomic epithelium (CE) is reportedly the source of new circulating cells (coelomocytes) as well as the provider of molecular factors such as immunity-related molecules. However, its overall functions have been scarcely studied in detail. In this work, we used an integrated approach based on both microscopy (light and electron) and proteomic analyses to investigate the arm CE in the starfish Marthasterias glacialis during different physiological conditions (i.e., non-regenerating and/or regenerating). Our results show that CE cells share both ultrastructural and proteomic features with circulating coelomocytes (echinoderm immune cells). Additionally, microscopy and proteomic analyses indicate that CE cells are actively involved in protein synthesis and processing, and membrane trafficking processes such as phagocytosis (particularly of myocytes) and massive secretion phenomena. The latter might provide molecules (e.g., immune factors) and fluids for proper arm growth/regrowth. No stem cell marker was identified and no pre-existing stem cell was observed within the CE. Rather, during regeneration, CE cells undergo dedifferentiation and epithelial-mesenchymal transition to deliver progenitor cells for tissue replacement. Overall, our work underlines that echinoderm CE is not a “simple epithelial lining” and that instead it plays multiple functions which span from immunity-related roles as well as being a source of regeneration-competent cells for arm growth/regrowth.

Abstract 372: Macrophage contact-dependent stemness induction and progressive CSC enrichment during metastatic dissemination in breast cancer

Abstract Background: Cancer stem cells (CSCs) play an important role during metastatic progression of breast cancer. However, little is known, at the single cell level, about the process of stemness induction in non-stem cells or the dynamic behavior of CSCs during hematogenous dissemination. Methods: Here, we employed high-resolution intravital multiphoton microscopy with a SOX2/OCT4 responsive fluorescent biosensor for stemness to directly observe the induction of stemness in single non-stem cells and their evolution through the metastatic cascade in living animals using orthotopic breast cancer xenograft model. We confirmed our findings in vitro using tumor cell-macrophage co-culture assays. Results: We report that, both in vitro and in vivo, direct physical contact with macrophages induces stemness in non-stem cancer cells via juxtacrine Notch-Jagged1 signaling. In vivo, macrophage depletion with clodronate treatment showed a significant decrease in stem cells. In vitro, using either the fate mapping of non-stem cells with or without macrophage contact, or the origin-mapping of stem cells to find whether they originated from non-stem cells or pre-existing stem cells, we found that there was four-fold increase in new CSC induction after direct macrophage contact. In contrast, we did not see any role of macrophages in the expansion of pre-existing CSCs, both in vivo and in vitro, indicating that macrophage contact-dependent stem induction is the primary mechanism of CSC generation. Using immunohistochemical staining in fixed tissue and live imaging of primary tumors and lungs in mice using optical windows, we found that during the course of dissemination of tumor cells from the primary site, CSCs become progressively enriched in the tumor cell population as they approach dissemination doorways (known as TMEM, Tumor MicroEnvironment of Metastasis), intravasate, circulate and arrive at the lung. Association with and passage through TMEM doorways is the step that generates the greatest enrichment in CSCs (~ 60-fold). On arrival in the lung, CSCs represent more than 75% of the disseminated tumor cell population, greatly enriched compared with their representation in the bulk primary tumor of ~ 1%. Conclusion: Overall, these data indicate, for the first time, that macrophages associated with TMEM induce CSCs and promote TMEM-mediated CSC intravasation and early metastatic seeding. Our results are consistent with the dramatic enrichment of cancer stem cell markers in association with TMEM in breast cancer patients (Kim et al 2020 AACR abstract) and support a strategy for anti-metastatic therapy. Citation Format: Ved P. Sharma, Yarong Wang, Binwu Tang, George S. Karagiannis, Emily A. Xue, David Entenberg, Lucia Borriello, Anouchka Coste, Joan G. Jones, Chinmay R. Surve, Dominic Esposito, Maja H. Oktay, Lalage M. Wakefield, John S. Condeelis. Macrophage contact-dependent stemness induction and progressive CSC enrichment during metastatic dissemination in breast cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 372.

How the axolotl makes a new limb

Limb Regeneration Unlike most vertebrate limbs, the axolotl limb regenerates the skeleton after amputation. Dermal and interstitial fibroblasts have been thought to provide sources for skeletal regeneration, but it has been unclear whether preexisting stem cells or dedifferentiation of fibroblasts formed the blastema. Gerber et al. developed transgenic reporter animals to compare periskeletal cell and fibroblast contributions to regeneration. Callus-forming periskeletal cells extended existing bone, but fibroblasts built new limb segments. Single-cell transcriptomics and Brainbow-based lineage tracing revealed the lack of a preexisting stem cell. Instead, the heterogeneous population of fibroblasts lost their adult features to form a multipotent skeletal progenitor expressing the embryonic limb program. Science , this issue p. [eaaq0681][1] [1]: /lookup/doi/10.1126/science.aaq0681

Engineering Cardiovascular Regeneration

The human heart has very limited regenerative capacity, making the loss of heart muscle tissue during myocardial infarction essentially irreversible. Regenerative medicine aims at promoting the formation of new functional heart muscle in an injured heart, either by stimulating myocyte proliferation, formation of myocytes from preexisting stem cells, direct reprogramming of fibroblasts into myocytes, or adding new muscle tissue from exogenous stem cell sources. Recent advances in stem cell research make these goals more realistic. This review provides a short overview about different approaches for cardiovascular regeneration, stem cell sources, and modes of application, focusing on current cardiac tissue engineering techniques and their way towards clinical application.

Epithelial-mesenchymal transition and cancer stem cells: a dangerously dynamic duo in breast cancer progression

Aberrant activation of a latent embryonic program - known as the epithelial-mesenchymal transition (EMT) - can endow cancer cells with the migratory and invasive capabilities associated with metastatic competence. The induction of EMT entails the loss of epithelial characteristics and the de novo acquisition of a mesenchymal phenotype. In breast cancer, the EMT state has been associated with cancer stem cell properties including expression of the stem cell-associated CD44+/CD24-/low antigenic profile, self-renewal capabilities and resistance to conventional therapies. Intriguingly, EMT features are also associated with stem cells isolated from the normal mouse mammary gland and human breast reduction tissues as well as the highly aggressive metaplastic and claudin-low breast tumor subtypes. This has implications for the origin of these breast tumors as it remains unclear whether they derive from cells that have undergone EMT or whether they represent an expansion of a pre-existing stem cell population that expresses EMT-associated markers to begin with. In the present review, we consider the current evidence connecting EMT and stem cell attributes and discuss the ramifications of these newly recognized links for our understanding of the emergence of distinct breast cancer subtypes and breast cancer progression.

Stem Cells and Neuroprotection: Understanding the Players

The use of neuroprotective therapies begs the question of how such therapies could affect preexisting stem cell populations within the host, as well as those introduced through cell-replacement therapy. Multiple mechanisms may mediate stem cell responses to neuroprotectants such as host/donor age and gender, cellular lineage/differentiation status, and mitochondrial dynamics. Current therapeutic sources for stem cells are embryonic, somatic, or induced pluripotent, with very little known about the effects of gender, age, cell type, and mitochondrial dynamics. With the advent of therapies to stimulate and recruit endogenous stem cells or transplant donor cells into damage areas in the hopes of recuperative regeneration of lost neurons, it is important to discuss mechanisms that dictate the winning players in the neuroprotection game. This review will focus on our current understanding of the characteristics of renewing stem cells that may affect neuroprotection.

IPS Cell Origins

Retroviral-driven overexpression of genes encoding four transcription factors ( Oct 3/4 , Sox2 , Klf4 , and c-Myc ) is sufficient to generate pluripotent cells from adult fibroblasts. Whether these induced pluripotent stem (iPS) cells arise from preexisting stem cells or from reprogramming of differentiated cells has not been clear. The group that first developed the iPS cell method reports that iPS cells made from adult mouse epithelial cells from the stomach (gastric epithelia) and liver (primary hepatocytes) were very similar to embryonic stem (ES) cells in several respects, including morphology, proliferation rate, pluripotency, and gene expression profile. Lineage tracing experiments revealed that iPS cells generated from primary hepatocytes were derived from cells that expressed the liver marker albumin , illustrating that differentiated cells can indeed be reprogrammed to give rise to iPS cells. Liver-derived iPS cells expressed ES cell markers more robustly than did those derived from adult fibroblasts, and they exhibited more extensive, although not complete, promoter demethylation, suggesting that the epithelial cells were more efficiently reprogrammed than were the fibroblasts. Chimeric mice containing fibroblast-derived iPS cells had a high risk of developing cancer as adults, presumably because the iPS cells contained a retrovirally encoded copy of the oncogene c-Myc . The incidence of cancer was reduced in chimeras containing liver-derived iPS cells. Furthermore, it was possible to generate epithelium-derived iPS cells without retroviral expression of the oncogene c-Myc , a feature that could be used to further limit the propensity of these cells to cause cancer. T. Aoi, K. Yae, M. Nakagawa, T. Ichisaka, K. Okita, K. Takahashi, T. Chiba, S. Yamanaka, Generation of pluripotent stem cells from adult mouse liver and stomach cells. Science 321 , 699-702 (2008). [Abstract] [Full Text]

Expression of embryonic stem cell marker OCT4 in human thyroid neoplasia

Current investigations suggested that thyroid carcinoma originate from individual mature thyroid follicular epithelial cells. Here we provide the evidence for a stem cell-associated model of thyroid carcinogenesis. We investigated the expression of the octamer-binding transcription factor-4 (OCT4) in human thyroid tissues. OCT4 is a POU domain–containing transcription factor encoded by Pou5f1 and functions as an important regulator in initial cell fate decisions during mammalian development. We assessed the expression of human OCT4 transcripts in 8 normal and 12 adenoma thyroid tissues, and 12 follicular, 20 papillary and 18 anaplastic thyroid carcinoma (Ca). We also investigated OCT4 expression in seven thyroid cancer cell lines. OCT4 transcripts were detected in most anaplastic carcinoma (80%), but weak or absent in normal thyroid (25%), benign thyroid adenoma (50%), follicular (33%) and papillary (30%) thyroid Ca tissues. Furthermore, OCT4 gene activity was absent in a papillary thyroid Ca cell line (B-CPAP), weak in follicular FTC-238, FTC-236, FTC-133, but strong in anaplastic 8305-C, C643, Hth74 Ca cell lines. The stem cell gene OCT4 may play a direct role and serve as a valuable new biomarker in thyroid carcinogenesis. Future research aims to provide further evidence to our hypothesis that thyroid carcinogenesis includes an involvement of pre-existing stem cells within the adult thyroid gland.

Differentiation of islet cells in long-term culture.

Our previous studies in the hamster pancreatic cancer model have shown that exocrine pancreatic cancer arises from ductal/ductular cells, as well as from within the islets, most probably from islet precursor (stem) cells. To identify and characterize these cells, we established a long-term culture from isolated hamster islets and investigated their growth, differentiation, and expression of biomarkers. Islets maintained their original form and structure within the first 14 days in culture. However, beginning at day 7, ductular structures began to form within the islets. At day 21 in culture, acinar cells, intermediary cells, oncocytes, and cells comparable to pancreatic hepatocytes also appeared between ductular and endocrine cells. The number of duct-like cells gradually increased, whereas the number of hormone-producing cells decreased. After 35 days in culture, the exocrine cells disappeared, and undifferentiated cells formed a monolayer. These cells expressed cytokeratins, alpha1-antitrypsin, transforming growth factor-alpha, epidermal growth factor receptor, carbonic anhydrase II, vimentin, laminin, and showed binding to tomato lectin and Phaseolus vulgaris leukoagglutinin. They did not express the regulatory transcriptional factors, insulin-promoting factor 1, NKx6.1, Pax6, and NeuroD. The results thus indicate that islet cells have potential to form exocrine cells. At present, it is not clear whether these cells originate from preexisting stem cells or from transdifferentiated islet cells.

EFFECTS OF ELECTRICALLY GENERATED SILVER IONS ON HUMAN CELLS AND WOUND HEALING

A method of producing local antibiotic effects by means of an iontophoretic technique using free silver ions has been evaluated in vitro and in vivo for more than two decades. The antibiotic properties of the technique have proved useful in both animal and human studies. In the course of determining the optimal clinical methodology for infected open wounds, a significant growth stimulation property resembling local tissue regeneration was noted. This has been traced to either the apparent production of dedifferentiation of normal mature cells or the stimulation of preexisting stem cells in the wound, resulting in the production of large numbers of progenitor cells. This process has now been studied in detail, and the results are presented herewith.