Exosomal circ_0084043 derived from colorectal cancer-associated fibroblasts promotes in vitro endothelial cell angiogenesis by regulating the miR-140-3p/HIF-1α/VEGF signaling axis.

Background: Circular RNAs (circRNAs) hold potential as diagnostic markers for colorectal cancer (CRC); however, their functional mechanisms remain incompletely elucidated. This work investigates the clinical implications of a unique set comprising six circRNAs derived from serum in CRC. Furthermore, we delve into the role of exosomal circ_0084043, originating from colorectal cancer-associated fibroblasts (CAFs), with a specific focus on its contribution to endothelial cell angiogenesis. Methods: The study analyzed circRNA levels in serum samples obtained from both CRC and control groups using qRT-PCR. Additionally, exosomes originating from colorectal CAFs and normal fibroblasts (NFs) were purified and confirmed by electron microscopy and Western blotting techniques. The proangiogenic effects of CAF-derived exosomal circ_0084043 were assessed in endothelial cells through proliferation, migration, and in vitro capillary tube formation assays. Gain- and loss-of-function experiments were employed to clarify the role of the circ_0084043/miR-140-3p/HIF-1α axis in endothelial cell angiogenesis, utilizing luciferase reporter assay, Western blotting, and ELISA for mechanism elucidation. Results: The candidate circRNAs (circ_0060745, circ_001569, circ_007142, circ_0084043, Circ_BANP, and CiRS-7) exhibited notably elevated expression in CRC patient sera compared to the levels observed in healthy individuals. Except for CiRS-7, all circRNAs showed elevated expression in CRC patients with positive lymph node metastasis and advanced tumor stages. Exosomes released by colorectal CAFs augmented endothelial cell proliferation, migration, and angiogenesis by upregulating VEGF expression and secretion. Circ_0084043 was highly detected in endothelial cells treated with CAF-derived exosomes. Silencing circ_0084043 reduced VEGFA expression and diminished CAF exosome-induced endothelial cell processes, indicating its pivotal role in angiogenesis. Circ_0084043 sponges miR-140-3p, regulating HIF-1α, and a reverse relationship was also identified between miR-140-3p and VEGFA in endothelial cells. Inhibiting miR-140-3p mitigated circ_0084043 knockdown effects in CAF exosome-treated endothelial cells. Co-transfection of si-circ_0084043 and a miR-140-3p inhibitor reversed the inhibited migration and angiogenesis caused by circ_0084043 knockdown in CAF exosome-treated endothelial cells. Inhibiting miR-140-3p rescued reduced VEGFA expression due to circ_0084043 knockdown in endothelial cells exposed to CAF-derived exosomes, indicating modulation of the circ_0084043/miR-140-3p/VEGF signaling in CAF-derived exosome-induced angiogenesis. Conclusions: This study unveiled a distinctive signature of six serum-derived circular RNAs, indicating their potential as promising diagnostic biomarkers for CRC. Importantly, exosomal circ_0084043 originating from colorectal CAFs was identified as playing a crucial role in endothelial cell angiogenesis, exerting its influence through the modulation of the miR-140-3p/HIF-1α/VEGF signaling axis.

Cancer-associated fibroblasts drive colorectal cancer cell progression through exosomal miR-20a-5p-mediated targeting of PTEN and stimulating interleukin-6 production.

BACKGROUND: This study evaluated the clinical relevance of a set of five serum-derived circulating microRNAs (miRNAs) in colorectal cancer (CRC). Additionally, we investigated the role of miR-20a-5p released by exosomes derived from cancer-associated fibroblasts (CAFs) in the context of CRC. METHODS: The expression levels of five circulating serum-derived miRNAs (miR-20a-5p, miR-122-5p, miR-139-3p, miR-143-5p, and miR-193a-5p) were quantified by real-time quantitative PCR (RT-qPCR), and their associations with clinicopathological characteristics in CRC patients were assessed. The diagnostic accuracy of these miRNAs was determined through Receiver Operating Characteristic (ROC) curve analysis. CAFs and normal fibroblasts (NFs) were isolated from tissue samples, and subsequently, exosomes derived from these cells were isolated and meticulously characterized using electron microscopy and Western blotting. The cellular internalization of fluorescent-labeled exosomes was visualized by confocal microscopy. Gain- and loss-of-function experiments were conducted to elucidate the oncogenic role of miR-20a-5p transferred by exosomes derived from CAFs in CRC progression. The underlying mechanisms were uncovered through luciferase reporter assay, Western blotting, enzyme-linked immunosorbent assays, as well as proliferation and migration assays. RESULTS: The expression levels of serum-derived circulating miR-20a-5p and miR-122-5p were significantly higher in CRC and were positively correlated with advanced stages of tumorigenesis and lymph node metastasis (LNM). In contrast, circulating miR-139-3p, miR-143-5p, and miR-193a-5p were down-regulated in CRC and associated with early tumorigenesis. Except for miR-139-3p, they showed a negative correlation with LNM status. Among the candidate miRNAs, significantly elevated levels of miR-20a-5p were observed in both cellular and exosomal fractions of CAFs. Our findings indicated that miR-20a-5p induces the expression of EMT markers, partly by targeting PTEN. Exosomal miR-20a secreted by CAFs emerged as a key factor enhancing the proliferation and migration of CRC cells. The inhibition of miR-20a impaired the proliferative and migratory potential of CAF-derived exosomes in SW480 CRC cells, suggesting that the oncogenic effects of CAF-derived exosomes are mediated through the exosomal transfer of miR-20a. Furthermore, exosomes originating from CAFs induced increased nuclear translocation of the NF-kB p65 transcription factor in SW480 CRC cells, leading to increased interleukin-6 (IL-6) production. CONCLUSIONS: We established a set of five circulating miRNAs as a non-invasive biomarker for CRC diagnosis. Additionally, our findings shed light on the intricate mechanisms underpinning the oncogenic impacts of CAF-derived exosomes and underscore the pivotal role of miR-20a-5p in CRC progression.

Exosomes released by oxidative stress-induced mesenchymal stem cells promote murine mammary tumor progression through activating the STAT3 signaling pathway.

Mesenchymal stem cells (MSCs) may play a pivotal role in shaping the tumor microenvironment (TME), influencing tumor growth. Nonetheless, conflicting evidence exists regarding the distinct impacts of MSCs on tumor progression, with some studies suggesting promotion while others indicate suppression of tumor cell growth. Considering that oxidative stress is implicated in the dynamic interaction between components of the TME and tumor cells, we investigated the contribution of exosomes released by hydrogen peroxide (H2O2)-treated MSCs to murine mammary tumor growth and progression. Additionally, we aimed to identify the underlying mechanism through which MSC-derived exosomes affect breast tumor growth and angiogenesis. Our findings demonstrated that exosomes released by H2O2-treated, stress-induced MSCs (St-MSC Exo) promoted breast cancer cell progression by inducing the expression of vascular endothelial growth factor (VEGF) and markers associated with epithelial-to-mesenchymal transition. Further clarification revealed that the promoting effect of St-MSC Exo on VEGF expression may, in part, depend on activating STAT3 signaling in BC cells. In contrast, exosomes derived from untreated MSCs retarded JAK1/STAT3 phosphorylation and reduced VEGF expression. Additionally, our observations revealed that the activation of the transcription factor NF-κB in BC cells, stimulated with St-MSC Exo, occurs concurrently with an increase in intracellular ROS production. Moreover, we observed that the increase in VEGF secretion into the conditioned media of 4T1 BC, mediated by St-MSC Exo, positively influenced endothelial cell proliferation, migration, and vascular behavior in vitro. In turn, our in vivo studies confirmed that St-MSC Exo, but not exosomes derived from untreated MSCs, exhibited a significant promoting effect on breast tumorigenicity. Collectively, our findings provide new insights into how MSCs may contribute to modulating the TME. We propose a novel mechanism through which exosomes derived from oxidative stress-induced MSCs may contribute to tumor progression and angiogenesis.

TET1 regulates stem cell properties and cell cycle of Cancer stem cells in triple-negative breast cancer via DNA demethylation.

The role of cancer stem cells in metastasis, recurrence, and resistance to conventional therapies is significant. Addressing these cells could potentially decrease cancer reoccurrences and mortality rates. TET1, a crucial gene involved in stem cell self-renewal and potency, may also play a part in cancer stem cells, which warrants further research. To explore the role of TET1 in cancer stem cells, we conducted experiments involving loss and gain. We then analyzed factors such as migration, invasion, cell cycle, cell viability, mammosphere formation, and the CD44+/CD24- subpopulation of cancer cells. We also investigate the influence of TET1 on CCNB1, CDK1, and OCT4. Our study reveals that TET1 can regulate the phenotype of cancer stem cells via OCT4. Additionally, it can control the cell cycle by increasing CDK1 and CCNB1 levels. These findings suggest that targeting DNA methylation and TET1 could be an effective strategy to overcome obstacles posed by Cancer stem cells. Our research also indicates that TET1 can influence the phenotype of cancer stem cells and the cell cycle of breast cancer cells potentially through OCT4, CCNB1, and CDK1. This highlights the importance of TET1 in breast cancer cases and suggests a potential therapeutic approach through DNA methylation and modulation of TET1.

Colorectal cancer-secreted exosomal circ_001422 plays a role in regulating KDR expression and activating mTOR signaling in endothelial cells by targeting miR-195-5p.

BACKGROUND: As non-coding RNAs, exosomal circular RNAs (circRNAs) regulate colorectal cancer (CRC) progression, although the functional mechanisms by which such molecules affect the tumor microenvironment are still elusive. Herein, we aimed to explore the potential clinical significance of a signature of five serum-derived circRNAs in CRC and investigated the mechanisms underlying endothelial cell angiogenesis mediated by CRC-secreted exosomal circ_001422. METHODS: The expression of a signature of five serum-derived circRNAs (circ_0004771, circ_0101802, circ_0082333, circ_0072309, and circ_001422) were measured by RT-qPCR, and their associations with tumor staging and lymph node metastasis were further evaluated in CRC patients. In silico analysis was used to show the relationship between circ_001422, miR-195-5p, and KDR, validated by dual-luciferase reporter and Western blotting assays. CRC cell-derived exosomes were isolated and characterized by scanning electron microscopy and Western blotting. Endothelial cell uptake of PKH26-labeled exosomes was demonstrated using a spectral confocal microscope. In vitro genetic strategies were used to exogenously alter the expression level of circ_001422 and miR-195-5p expression. Cell proliferation assay, transwell migration assay, and capillary tube formation assay were conducted to explore the role of CRC-secreted exosomal circ_001422 in endothelial cell function in vitro. RESULTS: The expression levels of serum-derived circ_0004771, circ_0101802, circ_0082333, and circ_001422 were significantly higher in CRC and were positively correlated with the lymph node metastasis status. However, circ_0072309 showed a significant down-regulation in CRC than in healthy individuals. Furthermore, a higher expression level of circ_001422 in both cellular and exosomal fractions was found in HCT-116 CRC cells. We found that HCT-116 exosomes considerably enhanced proliferation and migration of endothelial cells through shuttling of circ_001422. We also observed that exosomes derived from HCT-116 cell, but not non-aggressive Caco-2 CRC cells, increased in vitro tubulogenesis of endothelial cells. Importantly, knockdown of circ_001422 impaired the capability of endothelial cells to form the capillary-like tube structures. CRC-secreted circ_001422 acted as an endogenous miR-195-5p sponge to inhibit miR-195-5p activity, which led to increased KDR expression and mTOR signaling activation in endothelial cells. Importantly, ectopic expression of miR-195-5p mimicked the effect of circ_001422 silencing on KDR/mTOR signaling in endothelial cells. CONCLUSION: This study attributed a biomarker role for circ_001422 in CRC diagnosis and proposed a novel mechanism whereby circ_001422 up-regulates KDR through sponging miR-195-5p. These interactions may give rise to the activation of mTOR signaling and may be a possible clarification for the pro-angiogenesis effects of CRC-secreted exosomal circ_001422 on endothelial cells.

Monocytes educated by cancer-associated fibroblasts secrete exosomal miR-181a to activate AKT signaling in breast cancer cells.

BACKGROUND: Cancer-associated fibroblasts (CAFs), one of the major components of the tumor stroma, contribute to an immunosuppressive tumor microenvironment (TME) through the induction and functional polarization of protumoral macrophages. We have herein investigated the contribution of CAFs to monocyte recruitment and macrophage polarization. We also sought to identify a possible paracrine mechanism by which CAF-educated monocytes affect breast cancer (BC) cell progression. METHODS: Monocytes were educated by primary CAFs and normal fibroblast (NF); the phenotypic alterations of CAF- or NF-educated monocytes were measured by flow cytometry. Exosomes isolated from the cultured conditioned media of the educated monocytes were characterized. An in vivo experiment using a subcutaneous transplantation tumor model in athymic nude mice was conducted to uncover the effect of exosomes derived from CAF- or NF-educated monocytes on breast tumor growth. Gain- and loss-of-function experiments were performed to explore the role of miR-181a in BC progression with the involvement of the AKT signaling pathway. Western blotting, enzyme-linked immunosorbent assay, RT-qPCR, flow cytometry staining, migration assay, immunohistochemical staining, and bioinformatics analysis were performed to reveal the underlying mechanisms. RESULTS: We illustrated that primary CAFs recruited monocytes and established pro-tumoral M2 macrophages. CAF may also differentiate human monocyte THP-1 cells into anti-inflammatory M2 macrophages. Besides, we revealed that CAFs increased reactive oxygen species (ROS) generation in THP-1 monocytes, as differentiating into M2 macrophages requires a level of ROS for proper polarization. Importantly, T-cell proliferation was suppressed by CAF-educated monocytes and their exosomes, resulting in an immunosuppressive TME. Interestingly, CAF-activated, polarized monocytes lost their tumoricidal abilities, and their derived exosomes promoted BC cell proliferation and migration. In turn, CAF-educated monocyte exosomes exhibited a significant promoting effect on BC tumorigenicity in vivo. Of clinical significance, we observed that up-regulation of circulating miR-181a in BC was positively correlated with tumor aggressiveness and found a high level of this miRNA in CAF-educated monocytes and their exosomes. We further clarified that the pro-oncogenic effect of CAF-educated monocytes may depend in part on the exosomal transfer of miR-181a through modulating the PTEN/Akt signaling axis in BC cells. CONCLUSIONS: Our findings established a connection between tumor stromal communication and tumor progression and demonstrated an inductive function for CAF-educated monocytes in BC cell progression. We also proposed a supporting model in which exosomal transfer of miR-181a from CAF-educated monocytes activates AKT signaling by regulating PTEN in BC cells.

SMAD4 contributes to chondrocyte and osteocyte development.

Different cellular and molecular mechanisms contribute to chondrocyte and osteocyte development. Although vital roles of the mothers against decapentaplegic homolog 4 (also called 'SMAD4') have been discussed in different cancers and stem cell-related studies, there are a few reviews summarizing the roles of this protein in the skeletal development and bone homeostasis. In order to fill this gap, we discuss the critical roles of SMAD4 in the skeletal development. To this end, we review the different signalling pathways and also how SMAD4 defines stem cell features. We also elaborate how the epigenetic factors-ie DNA methylation, histone modifications and noncoding RNAs-make a contribution to the chondrocyte and osteocyte development. To better grasp the important roles of SMAD4 in the cartilage and bone development, we also review the genotype-phenotype correlation in animal models. This review helps us to understand the importance of the SMAD4 in the chondrocyte and bone development and the potential applications for therapeutic goals.

Colorectal cancer cell-derived extracellular vesicles transfer miR-221-3p to promote endothelial cell angiogenesis via targeting suppressor of cytokine signaling 3.

BACKGROUND: Secreted microRNAs (miRNAs) can serve as promising diagnostic markers for colorectal cancer (CRC). Herein, we evaluated the potential clinical significance of a signature of four circulating serum-derived miRNAs in CRC. We also demonstrated that extracellular vesicles (EVs) containing miR-221-3p could facilitate endothelial cell angiogenesis. METHODS: The expressions of four circulating serum-derived miRNAs (miR-19a-3p, miR-203-3p, miR-221-3p, and let-7f-5p) were measured by real-time quantitative PCR, and their associations with lymph node metastasis were determined in CRC patients. Receiver operating characteristic curve analysis was used to determine their diagnostic accuracy. EVs were isolated and characterized from the conditioned media of human CRC cells (HCT116 and Caco2). Cell proliferation, transwell migration, and tube formation assays were performed to investigate the pro-angiogenic effect of miR-221-3p transferred by CRC-EVs into the endothelial cells. In silico analysis was used to show the regulatory functions of miR-221-3p on SOCS3, validated by luciferase and Western blotting assays. RESULTS: The expression levels of serum-derived miR-19a-3p, miR-203-3p, miR-221-3p, and let-7f-5p were significantly higher in CRC than in healthy individuals. The expression of miR-19a-3p, miR-203-3p, and miR-221-3p were positively correlated with the lymph node metastasis status. Moreover, SOCS3 was identified as a direct target of miR-221-3p and the secreted miR-221-3p shuttled by CRC-EVs regulated STAT3/VEGFR-2 signaling axis by targeting SOCS3 in endothelial cells. CRC-EVs promoted endothelial cell proliferation, migration, and the formation of vessel-like structures. The proangiogenic effect of CRC-EVs on the cells was recapitulated by miR-221-3p overexpression, showing the importance of EVs-derived miR-221-3p in promoting endothelial cell angiogenesis. CONCLUSION: We introduced a signature of four-circulating miRNAs (miR-19a-3p, miR-203-3p, miR-221-3p, and let-7f-5p) as a novel diagnostic biomarker for CRC. Besides, we revealed that miR-221-3p induces endothelial cell angiogenesis in vitro by targeting SOCS3.

Upregulation of the long noncoding RNAs DSCAM-AS1 and MANCR is a potential diagnostic marker for breast carcinoma.

Breast cancer (BC) is one of the most common malignancies among women in the world. There is a global attempt to diagnose BC as early as possible. Long noncoding RNAs (lncRNAs) are emerging as novel targets and biomarkers for BC diagnosis and prognosis. Aberrant expression of lncRNAs is associated with BC development, making them a potential tumor marker for BC. To investigate this possibility, we determined the expression levels of Down syndrome cell adhesion molecule-antisense RNA-1 (DSCAM-AS1) and mitotically-associated long non-coding RNA (MANCR) lncRNAs in BC tissues. This case-control study included 50 paired tumor and adjacent nontumor tissues from female BC patients. The total RNA was isolated and the expression levels of MANCR and DSCAM-AS1 lncRNAs were assessed using quantitative real-time reverse transcription-PCR. Potential correlations between lncRNA levels and clinicopathological characteristics were also analyzed. DSCAM-AS1 and MANCR lncRNAs were significantly upregulated in BC tumor tissues compared with the adjacent nontumor tissues. We also found the significant upregulation of DSCAM-AS1 in advanced tumor-node-metastasis stage (TNM III) of BC tumor tissues. Furthermore, the expression of DSCAM-AS1 and MANCR in HER-2 positive patients was significantly higher than HER-2 negative affected individuals. Receiver operating characteristic curve analysis showed a satisfactory diagnostic efficacy (P value < 0.0001), which means that DSCAM-AS1 and MANCR lncRNAs can potentially serve as a biomarker. The present study might provide further approval for the clinical diagnostic significance of DSCAM-AS1 and MANCR lncRNAs that their high expressions were associated with aggressive clinical parameters of BC.

Docosahexaenoic acid reverses the promoting effects of breast tumor cell-derived exosomes on endothelial cell migration and angiogenesis.

AIM: As a natural compound, docosahexaenoic acid (DHA) exerts anti-cancer and anti-angiogenesis functions through exosomes; however, little is known about the molecular mechanisms. MAIN METHODS: Breast cancer (BC) cells were treated with DHA (50 µM) and then tumor cell-derived exosomes (TDEs) were collected and characterized by electron microscopy, dynamic light scattering, and western blot analyses. By the time the cells were treated with DHA, RT-qPCR was used to investigate the expression of vascular endothelial growth factor (VEGF) and the selected pro- and anti-angiogenic microRNAs (miRNAs). The quantification of secreted VEGF protein was measured by enzyme-linked immunosorbent assay (ELISA). The effects of TDEs on endothelial cell angiogenesis were explored by transwell cell migration and in vitro vascular tube formation assays. KEY FINDINGS: DHA treatment caused a significant and time-dependent decrease in the expression and secretion of VEGF in/from BC cells. This also increased expression of anti-angiogenic miRNAs (i.e. miR-34a, miR-125b, miR-221, and miR-222) while decreased levels of pro-angiogenic miRNAs (i.e. miR-9, miR-17-5p, miR-19a, miR-126, miR-130a, miR-132, miR-296, and miR-378) in exosomes derived from DHA-treated BC cells, TDE (DHA+). While treatment with exosomes (100 µg/ml) obtained from untreated BC cells, TDE (DHA-), enhanced the expression of VEGF-A in human umbilical vein endothelial cells (HUVECs), incubation with DHA or TDE (DHA+) led to the significant decrease of VEGF-A transcript level in these cells. We indicated that the incubation with TDE (DHA+) could significantly decrease endothelial cell proliferation and migration and also the length and number of tubes made by HUVECs in comparison with endothelial cells incubated with exosomes obtained from untreated BC cells. SIGNIFICANCE: DHA alters angiogenesis by shifting the up-regulation of exosomal miRNA contents from pro-angiogenic to anti-angiogenic, resulting in the inhibition of endothelial cell angiogenesis. These data can help to figure out DHA's anti-cancer function, maybe its use in cancer therapy.

A panel of six-circulating miRNA signature in serum and its potential diagnostic value in colorectal cancer.

AIM: Colorectal carcinoma (CRC) is one of the most prevalent cancers throughout the world. Circulating serum-derived microRNAs (miRNAs, miRs) can be used as non-invasive biomarkers for CRC diagnosis. This study aimed to identify a panel of six serum exosomal miRNAs as novel diagnostic biomarkers for CRC. MAIN METHODS: Exosomes were isolated and characterized from the conditioned media of the human colon adenocarcinoma cells (HCT-116 and Caco2). Sera were isolated from peripheral blood of 45 CRC and also 45 healthy individuals. The expression levels and diagnostic value of candidate circulating miRNAs (miR-19a, miR-20a, miR-150, miR-143, miR-145, and let-7a) were measured through quantitative real-time PCR. Receiver operating characteristic (ROC) curves were applied to evaluate the diagnostic accuracy of selected miRNAs. The association of candidate miRNAs and clinicopathological characteristics e.g. tumor node metastasis (TNM) staging and lymph node metastasis (LNM) were further evaluated. KEY FINDINGS: Circulating serum miR-19a, miR-20a, miR-150, and let-7a were significantly up-regulated in CRC patients, while miR-143 and miR-145 showed a significant down-regulation. The higher levels of miR-143 and miR-145 in patients with TNM stage I-II were detected, whereas miR-19a, miR-20a, miR-150, and let-7a were highly expressed in TNM stage III. The expression levels of miR-19a, miR-20a, and miR-150 were positively correlated with LNM status, while the expression levels of miR-143 and miR-145 were lower in patients with LNM. Area under the ROC curves of miR-19a, miR-20a, miR-150, miR-143, miR-145, and let-7a were 0.87, 0.83, 0.75, 0.76, 0.78 and 0.71, respectively. SIGNIFICANCE: We established a panel of six-circulating miRNA signature (i.e. miR-19a, miR-20a, miR-143, miR-145, miR-150, and let-7a) in serum as a non-invasive biomarker for CRC diagnosis. These findings confirm that serum-derived miRNAs have a strong potential to be a diagnostic biomarker for patients with CRC.

Regulation of MicroRNAs by Phytochemicals: A Promising Strategy for Cancer Chemoprevention.

A growing body of evidence suggests that phytochemicals are potentially able to affect a variety of cellular processes, including proliferation, apoptosis, cell-cycle control, angiogenesis, inflammation, and DNA repair. Phytochemicals may typically play pleiotropic regulatory roles in cancer cells. Chemoprevention, which can be achieved by using these natural agents, has emerged as a helpful strategy to manage a variety of malignancies. With regard to cancer-associated chemopreventive mechanisms, phytochemicals can act by modulating microRNAs (miRNAs) and their target genes. This review aims to present an overview of recent findings on the effects of some wellcharacterized bioactive phytochemicals on miRNA regulation in different cancer types. The potential use of these phytochemicals for the chemoprevention and treatment of cancer is also discussed.

MicroRNA-100 shuttled by mesenchymal stem cell-derived exosomes suppresses in vitro angiogenesis through modulating the mTOR/HIF-1α/VEGF signaling axis in breast cancer cells.

BACKGROUND: Human mesenchymal stem cells (MSCs) have been shown to be involved in the formation and modulation of tumor stroma and in interacting with tumor cells, partly through their secretome. Exosomes are nano-sized intraluminal multi-vesicular bodies secreted by most types of cells and have been found to mediate intercellular communication through the transfer of genetic information via coding and non-coding RNAs to recipient cells. Since exosomes are considered as protective and enriched sources of shuttle microRNAs (miRNAs), we hypothesized that exosomal transfer of miRNAs from MSCs may affect tumor cell behavior, particularly angiogenesis. METHODS: Exosomes derived from MSCs were isolated and characterized by scanning electron microscopy analyses, dynamic light scattering measurements, and Western blotting. Fold changes in miR-100 expression levels were calculated in exosomes and their corresponding donor cells by qRT-PCR. The effects of exosomal transfer of miR-100 from MSCs were assessed by qRT-PCR and Western blotting of the mTOR/HIF-1α/VEGF signaling axis in breast cancer cells. The quantification of secreted VEGF protein was determined by enzyme-linked immunosorbent assay. The putative paracrine effects of MSC-derived exosomes on tumor angiogenesis were explored by in vitro angiogenesis assays including endothelial cell proliferation, migration and tube formation assays. RESULTS: We found that MSC-derived exosomes induce a significant and dose-dependent decrease in the expression and secretion of vascular endothelial growth factor (VEGF) through modulating the mTOR/HIF-1α signaling axis in breast cancer-derived cells. We also found that miR-100 is enriched in MSC-derived exosomes and that its transfer to breast cancer-derived cells is associated with the down-regulation of VEGF in a time-dependent manner. The putative role of exosomal miR-100 transfer in regulating VEGF expression was substantiated by the ability of anti-miR-100 to rescue the inhibitory effects of MSC-derived exosomes on the expression of VEGF in breast cancer-derived cells. In addition, we found that down-regulation of VEGF mediated by MSC-derived exosomes can affect the vascular behavior of endothelial cells in vitro. CONCLUSIONS: Overall, our findings suggest that exosomal transfer of miR-100 may be a novel mechanism underlying the paracrine effects of MSC-derived exosomes and may provide a means by which these vesicles can modulate vascular responses within the microenvironment of breast cancer cells.

Prostate cancer stem cells: from theory to practice.

None of the generally accepted theories on prostate cancer development can fully explain many distinguishing features of the disease, such as intratumoral heterogeneity, metastatic growth, drug resistance and tumor relapse. Prostate stem cells are a heterogeneous and small subpopulation of self-renewing cells which can actively proliferate in response to changes in the androgen level and give rise to all the cell lineages that build the prostate epithelium. According to the cancer stem cell hypothesis, prostate cancer could be a stem cell disease. Prostate cancer stem cells, which represent only a minimal percentage of the tumor mass, are characterized by a markedly increased clonogenicity and therapeutic resistance. These tumor-initiating cells reside in dynamic niches distributed within the prostate but at a higher concentration in proximal regions of the prostatic ducts. Several markers have been used to identify prostate cancer stem cells. Nevertheless, a definitive profile has not yet been established owing to specificity issues. As cancer stem cells play determining roles in the birth and burst of prostate malignancy, strategies that selectively target them have gained huge clinical attention. Unraveling the mechanisms underlying the physiological functions of cancer stem cells and gaining fundamental insights into their putative involvement in the pathogenesis of prostate tumors provide novel opportunities for the development of efficient and sophisticated therapeutic strategies in the future.

Novel strategies for targeting leukemia stem cells: sounding the death knell for blood cancer.

BACKGROUND: Cancer stem cells (CSCs), also known as tumor-initiating cells (TICs), are characterized by high self-renewal and multi-lineage differentiation capacities. CSCs are thought to play indispensable roles in the initiation, progression and metastasis of many types of cancer. Leukemias are thought to be initiated and maintained by a specific sub-type of CSC, the leukemia stem cell (LSC). An important feature of LSCs is their resistance to standard therapy, which may lead to relapse. Increasing efforts are aimed at developing novel therapeutic strategies that selectively target LSCs, while sparing their normal counterparts and, thus, minimizing adverse treatment-associated side-effects. These LSC targeting therapies aim to eradicate LSCs through affecting mechanisms that control their survival, self-renewal, differentiation, proliferation and cell cycle progression. Some LSC targeting therapies have already been proven successful in pre-clinical studies and they are now being tested in clinical studies, mainly in combination with conventional treatment regimens. CONCLUSIONS: A growing body of evidence indicates that the selective targeting of LSCs represents a promising approach to improve disease outcome. Beyond doubt, the CSC hypothesis has added a new dimension to the area of anticancer research, thereby paving the way for shaping a new trend in cancer therapy.