Embryonic Stem Cell Microenvironment Research Articles

Modulating DNA damage response in uveal melanoma through embryonic stem cell microenvironment

BackgroundUveal melanoma (UVM) is the most common primary intraocular tumor in adults, with a median survival of 4–5 months following metastasis. DNA damage response (DDR) upregulation in UVM, which could be linked to its frequent activation of the PI3K/AKT pathway, contributes to its treatment resistance. We have reported that embryonic stem cell microenvironments (ESCMe) can revert cancer cells to less aggressive states through downregulation of the PI3K signaling, showing promise in modulating the DDR of UVM.MethodsSince nonhomologous end joining (NHEJ) is the main DNA repair mechanism in UVM, this study utilized gene expression analysis and survival prognosis analysis to investigate the role of NHEJ-related genes in UVM based on public databases. Xenograft mouse models were established to assess the therapeutic potential of ESC transplantation and exposure to ESC-conditioned medium (ESC-CM) on key DNA repair pathways in UVM. Quantitative PCR and immunohistochemistry were used to analyze NHEJ pathway-related gene expression in UVM and surrounding normal tissues. Apoptosis in UVM tissues was evaluated using the TUNEL assay.ResultsPRKDC, KU70, XRCC5, LIG4 and PARP1 showed significant correlations with UM progression. High expression of PRKDC and XRCC5 predicted poorer overall survival, while low PARP1 and XRCC6 expression predicted better disease-free survival in UVM patients. ESCMe treatment significantly inhibited the NHEJ pathway transcriptionally and translationally and promoted apoptosis in tumor tissues in mice bearing UVM. Furthermore, ESC transplantation enhanced DDR activities in surrounding normal cells, potentially mitigating the side effects of cancer therapy. Notably, direct cell-to-cell contact with ESCs was more effective than their secreted factors in regulating the NHEJ pathway.ConclusionsOur results suggest that NHEJ-related genes might serve as prognostic markers and therapeutic targets in UVM. These findings support the therapeutic potential of ESC-based therapy in enhancing UVM sensitivity to radiochemotherapy and improving treatment outcomes while minimizing damage to healthy cells.

Optimized Method for Using Embryonic Microenvironment to Reprogram Cancer Stem Cells

Purpose: The embryonic microenvironment contains many properties that have not yet been fully explored. Our aim in this study is to report an optimized and efficient method that enables investigating the effects of the secretome of pluripotent embryonic stem cells on cancer stem cells.
 Methods: The study is performed with a chimeric model consisted of mouse blastocysts, human prostate cancer stem cells and non-cancer stem cells. Ovulation induced mice were used for blastocyst collection. DU145 prostate cancer cell line was separated into cancer stem cells and non-cancer stem cells according to cancer stem cells biomarker expressions by fluorescent activated cell sorting method. Human prostate cancer stem cells and non-cancer stem cells were microinjected into 4-day blastocyst culture in vitro by intracytoplasmic sperm injection method.
 Results: Chimeric models provide us great convenience in basic oncological studies. In this study, using a chimeric model, we were able to study the secretome of mouse embryonic stem cells and their effect on cancer stem cells. The method is efficient and yield promising result; and could be used to study the effects on other cells as well.
 Conclusion: The embryonic stem cell microenvironment is suggested to have a great regenerative capacity which is, nowadays, the center of attraction for cancer research studies. Ethical issues restrict the human embryo studies, however, mimicking the in vivo human microenvironment with 3D cell cultures or bioprinting are now possible. Finally, optimization of new methods including 3D cell cultures with human cell lines will be a great opportunity for better understanding the reprogramming notion.

The embryonic stem cell microenvironment inhibits mouse glioma cell proliferation by regulating the PI3K/AKT pathway.

BackgroundGlioblastoma multiforme (GBM) is the most common malignant tumor of the central nervous system, accounting for 48.6% of malignant tumors. The current standard treatment plan includes the widest range of safe surgical resection, supplemented by local brain radiotherapy and temozolomide concurrent chemotherapy; this can cause serious side effects. Even so, the median survival time of GBM patients is only 8 months, and the 5-year survival rate is only 5.5%. It is imminent to find new treatments. Early studies have shown that chicken and zebrafish embryos can reprogram cancer cells into a non-tumorigenic phenotype through the embryonic microenvironment. However, the effect of embryonic stem cell microenvironment on GBM and its possible mechanism are not clear.MethodsIn this study, the glioblastoma cell line, U118, in the brain was investigated. There were four experimental groups: GB, GE, GA and GT. U118 cells were harvested after culturing for 72 hours. Cell proliferation, apoptosis, reactive oxygen species (ROS) were examined using vasculogenic mimicry assays, quantitative real-time polymerase chain reaction (QRT-PCR), western blotting (WB) and flow cytometry. The differences in the biological function of U118 cells and the PI3K/protein kinase B (AKT) signaling pathway were compared between the groups.ResultsCompared with the GB control group, the GE co-culture group and GT chemotherapy group showed reduced cell proliferation, increased apoptosis, increased ROS, as well as decreased or inhibited vasculogenic mimicry. Expressions of cyclin B1 and cyclin D1 were also notably reduced, while that of Bax, Bcl-2, p53, Caspase-3, GSK-3β, p21, and p27 were significantly increased. Moreover, the expression of PI3K, AKT, and mTOR were markedly decreased, whereas expression of PTEN increased considerably. Also, the expression of positive regulatory factors significantly increased, however negative regulatory factors decreased in the GA group compared to the GE group.ConclusionsThe ESC microenvironment reverses glioma malignancy, partially via inhibition of the PI3K signaling pathway. Our study may have a significant impact and important clinical implications for cell therapy in the treatment of glioma.

Embryonic stem cell microenvironment enhances proliferation of human retinal pigment epithelium cells by activating the PI3K signaling pathway

BackgroundRetinal pigment epithelium (RPE) replacement has been proposed as an efficacious treatment for age-related macular degeneration (AMD), which is the primary cause of vision loss in the elderly worldwide. The embryonic stem cell (ESC) microenvironment has been demonstrated to enable mature cells to gain a powerful proliferative ability and even enhance the stem/progenitor phenotype via activation of the phosphoinositide 3-kinase (PI3K) signaling pathway. As the PI3K signaling pathway plays a pivotal role in proliferation and homeostasis of RPE, we hypothesize that the stemness and proliferative capability of RPE can be enhanced by the ESC microenvironment via activation of the PI3K signaling pathway.MethodsTo investigate whether the ESC microenvironment improves the stem cell phenotype and proliferation properties of human RPE (hRPE) cells by regulating the PI3K signaling pathway, primary hRPE cells were cocultured with either ESCs or human corneal epithelial cells (CECs) for 72 h, after which their proliferation, apoptosis, cell cycle progression, and colony formation were assayed to evaluate changes in their biological characteristics. Gene expression was detected by real-time PCR and protein levels were determined by western blotting or immunofluorescence. LY294002, an antagonist of the PI3K signaling pathway, was used to further confirm the mechanism involved.ResultsIn comparison to hRPE cells cultured alone, hRPE cells cocultured with ESCs had an increased proliferative capacity, reduced apoptotic rate, and higher colony-forming efficiency. The expression of the stem cell-associated marker KLF4 and the differentiation marker CRALBP increased and decreased, respectively, in hRPE cells isolated from the ESC coculture. Furthermore, PI3K pathway-related genes were significantly upregulated in hRPE cells after exposure to ESCs. LY294002 reversed the pro-proliferative effect of ESCs on hRPE cells. In contrast, CECs did not share the ability of ESCs to influence the biological behavior and gene expression of hRPE cells.ConclusionsOur findings indicate that the ESC microenvironment enhances stemness and proliferation of hRPE cells, partially via activation of the PI3K signaling pathway. This study may have a significant impact and clinical implication on cell therapy in regenerative medicine, specifically for age-related macular degeneration.

Embryonic stem cell microenvironment suppresses the malignancy of cutaneous melanoma cells by down-regulating PI3K/AKT pathway.

Malignant cancer cells engage in a dynamic reciprocity with the tumor microenvironment (TME) that promotes tumor growth, development, and resistance to therapy. Early embryonic blastocyst microenvironments can reverse the tumorigenic phenotype of malignant cancer cells via ameliorating of TME. It is potential to apply embryonic stem cell (ESC) microenvironment to suppress the malignant behaviors of cancer cells. This study aimed to investigate a better method and the mechanism of ESC microenvironment supplied by ESCs on suppressing the malignancy of cutaneous melanoma cells. Cutaneous melanoma cell line A2058 were cultured and divided into four groups: (a) A2058‐only (Control); (b) A2058 and ESCs continuously co‐cultured (Group One); (c) A2058 co‐cultured with daily refreshed ESCs (Group two); (d) Group one with VO‐Ohpic, inhibitor of PTEN (VO‐Ohpic Group). The results showed that, compared to control group, A2058 cells in group one exhibited decreased cellular proliferation, migration, invasiveness and vasculogenic mimicry concomitant with an increase in cell apoptosis, accompanied by down‐regulation of PI3K/AKT pathway. Besides, the above mentioned anti‐tumor effects on A2058 cells were significantly enhanced in group two but statistically weakened after administration of VO‐Ohpic compared to group one. We demonstrate that ESC microenvironment reduces the malignancy of A2058 by down‐regulating PI3K/AKT pathway. Notably, such anti‐tumor effects can be enhanced by appropriately increasing the quality and quantity of ESCs in co‐culture system. Our results suggest that ESC microenvironment could be an effective and safe approach to treating cancer.

Notch pathway is involved in the suppression of colorectal cancer by embryonic stem cell microenvironment.

Objectives: Recently, embryonic microenvironment is being known for its non-permissive property for tumor growth. However, the regulatory mechanism to maintain the balance between differentiation and tumorigenicity of cancer cell in microenvironment is not well understood.Materials and Methods: qRT-PCR was performed to detect the levels of gene expression in HT29, LoVo and Caco-2 colorectal cancer cells, and Western blot was used to measure the protein levels. Cell migration and apoptosis were measured by Transwell and flow cytometry assays. Cancer cell markers were detected using immunohistochemical staining. In vivo tumor formation assay was conducted by subcutaneous injection of embryonic microenvironment-treated cancer cells.Results: Colorectal cancer cell lines were treated with human embryonic stem cell conditioned culture and then collected for in vivo tumor formation assay and in vitro assays assessing the aggressive properties. We found exposure of cancer cells in human ES cultures resulted in inhibition of growth, migration of tumor cells. Moreover, we found that manipulation of Notch pathway in the ES cells microenvironment could influence the stemness of tumor. We specifically discovered that some factor in the embryonic microenvironment could suppress Notch1 pathway in the cancer cells, leading to a reduction in tumorigenesis and invasiveness.Conclusions: This study may provide another evidence to understand the crosstalk between tumor cells and embryonic environment and may offer new therapeutic strategies to inhibit colorectal cancer progression.

Reprogramming Malignant Cancer Cells toward a Benign Phenotype following Exposure to Human Embryonic Stem Cell Microenvironment

The embryonic microenvironment is well known to be non-permissive for tumor development because early developmental signals naturally suppress the expression of proto-oncogenes. In an analogous manner, mimicking an early embryonic environment during embryonic stem cell culture has been shown to suppress oncogenic phenotypes of cancer cells. Exosomes derived from human embryonic stem cells harbor substances that mirror the content of the cells of origin and have been reported to reprogram hematopoietic stem/progenitor cells via horizontal transfer of mRNA and proteins. However, the possibility that these embryonic stem cells-derived exosomes might be the main effectors of the anti-tumor effect mediated by the embryonic stem cells has not been explored yet. The present study aims to investigate whether exosomes derived from human embryonic stem cells can reprogram malignant cancer cells to a benign stage and reduce their tumorigenicity. We show that the embryonic stem cell-conditioned medium contains factors that inhibit cancer cell growth and tumorigenicity in vitro and in vivo. Moreover, we demonstrate that exosomes derived from human embryonic stem cells display anti-proliferation and pro-apoptotic effects, and decrease tumor size in a xenograft model. These exosomes are also able to transfer their cargo into target cancer cells, inducing a dose-dependent increase in SOX2, OCT4 and Nanog proteins, leading to a dose-dependent decrease of cancer cell growth and tumorigenicity. This study shows for the first time that human embryonic stem cell-derived exosomes play an important role in the tumor suppressive activity displayed by human embryonic stem cells.

Engineering Embryonic Stem Cell Microenvironments for Tailored Cellular Differentiation

The rapid progress of embryonic stem cell (ESCs) research offers great promise for drug discovery, tissue engineering, and regenerative medicine. However, a major limitation in translation of ESCs technology to pharmaceutical and clinical applications is how to induce their differentiation into tailored lineage commitment with satisfactory efficiency. Many studies indicate that this lineage commitment is precisely controlled by the ESC microenvironment in vivo. Engineering and biomaterial-based approaches to recreate a biomimetic cellular microenvironment provide valuable strategies for directing ESCs differentiation to specific lineages in vitro. In this review, we summarize and examine the recent advances in application of engineering and biomaterial-based approaches to control ESC differentiation. We focus on physical strategies (e.g., geometrical constraint, mechanical stimulation, extracellular matrix (ECM) stiffness, and topography) and biochemical approaches (e.g., genetic engineering, soluble bioactive factors, coculture, and synthetic small molecules), and highlight the three-dimensional (3D) hydrogel-based microenvironment for directed ESC differentiation. Finally, future perspectives in ESCs engineering are provided for the subsequent advancement of this promising research direction.

Safety and efficacy of embryonic stem cell microenvironment in a leukemia mouse model.

The embryonic stem cell (ESC) microenvironment can promote the proliferation of terminal cells and reduce the invasiveness of tumor cells. However, implanting ESCs directly in vivo can result in tumorigenicity, immune rejection after differentiation, and graft-versus-host reaction. Therefore, safety is very important in the clinical application of ESCs. We injected ESCs modified with a suicide gene into a leukemia mouse model through peripheral blood to observe the treatment effectiveness. In addition, according to the pre-test, we set the time point of differentiation after transplantation and then activated the suicide gene to kill the ESCs that we had initially implanted, hoping to avoid the risks mentioned earlier. Our results indicated that the body weight and survival rates of mice treated with an ESC microenvironment increased, and leukemic cells in peripheral blood and bone marrow decreased compared with untreated mice. There was no obvious teratoma in mice that received ESC therapy and induced the suicide gene at the proper time during the observation period, while an apparent teratoma was observed in the lungs of mice which received ESC therapy and never induced the suicide gene. Therefore, the ESC microenvironment can promote self-healing of the in vivo microenvironment. Inducing the suicide gene at the appropriate time can reduce or even avoid tumorigenicity and immune rejection after transplantation of ESCs in vivo and improve the safety of their clinical application.

Abstract 3241: Analysis of transcription factor activation by the embryonic microenvironment in metastatic breast cancer cells

Abstract Introduction: The embryonic microenvironment is an important source of signals that program multipotent cells to adopt a particular fate. Numerous studies have shown that components of the embryonic stem cell microenvironment can induce a reversion of cancer cell phenotype from a more aggressive to less aggressive state through mechanisms that are largely unknown. We have examined the phenotypic transformation associated with the culture of metastatic breast cancer cells in mouse embryonic stem cell conditioned media and have implemented a novel cell array technology to characterize the signaling pathways associated with this transformation. Methods: We used phase contrast microscopy to determine the time course of structure formation in MDA-MB-231 cells cultured in 3D with control or conditioned mouse embryonic stem cell media. Cell proliferation of study cells stably expressing Gaussia Luciferase was quantified by measuring Gluc production over time. Cell migration was measured for treated and untreated cells using a transwell migration assay. A live cell array was implemented to quantify changes in activity of 19 cancer-related transcription factors over a period of 8 days. Results: MDA-MB-231 cells cultured in stem cell conditioned media showed decreased area covered by spindle-like projections over a period of 10 days. Study cells cultured in stem cell conditioned media, but not in control media or mouse embryonic fibroblast conditioned media, had decreased proliferation over time. Cells cultured in stem cell conditioned media had decreased cell migration relative to cells cultured with regular media. Using a cell array to report on TF activity, cells cultured in conditioned stem cell media, when compared to control media, had varied activity in 7 of the 19 constructs investigated, including metastasis-related factors SP1 and GATA1. Conclusion: Our findings support the normalizing influence provided by the stem cell microenvironment through decreased MDA-MB-231 spindle-like projections, cell proliferation, and cell migration. Of the 19 transcription factor reporter constructs tested using the novel live cell array, GATA1 and SP1 emerged as 2 of the transcription factors impacted by the embryonic microenvironment. These factors may reflect potential paracrine signaling pathways stimulated by the microenvironment that can influence transformation and may be targets for therapeutic intervention. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3241. doi:1538-7445.AM2012-3241

Embryonic stem cell microenvironment's effects on tumors

It's considered now that the neoplasm is originated from the immortalized stem cell with un-regulated self-renewal capability. Microenvironment of stem cell determine the fate of stem cell, abnormal microenvironment can induce the neoplastic stem cell, while ameliorated microenvironment can inhibit carcinoma-tous changes. Recent researches have found that embryonic microenvironment capable of keeping the balance between differentiation and self- renewal of stem cell lineage should reprogram the neoplasm. Results show that the embryonic stem cell microenvironment can suppress neoplasm cells' aggressive behaviors, reverse their malig-nant phenotypes, which is of good guiding significance and applicational prospect in the field of tumor's treatment. Key words: Neoplasms; Neoplastic stem cells; The embryonic stem cell microenvironment