Jennifer Pasquier

Preferential transfer of mitochondria from endothelial to cancer cells through tunneling nanotubes modulates chemoresistance

Our vision of cancer has changed during the past decades. Indeed tumors are now perceived as complex entities where tumoral and stromal components interact closely. Among the different elements of tumor stroma the cellular component play a primordial role. Bone Marrow derived mesenchymal cells (MSCs) are attracted to tumor sites and support tumor growth. Endothelial cells (ECs) play a major role in angiogenesis. While the literature documents many aspects of the cross talk between stromal and cancer cells, the role of direct hetero-cellular contact is not clearly established. Recently, Tunneling nanotubes (TnTs) have been shown to support cell-to-cell transfers of plasma membrane components, cytosolic molecules and organelles within cell lines. Herein, we have investigated the formation of heterocellular TnTs between stromal (MSCs and ECs) and cancer cells. We demonstrate that TnTs occur between different cancer cells, stromal cells and cancer-stromal cell lines. We showed that TnTs-like structure occurred in 3D anchorage independent spheroids and also in tumor explant cultures. In our culture condition, TnTs formation occurred after large membrane adhesion. We showed that intercellular transfers of cytoplasmic content occurred similarly between cancer cells and MSCs or ECs, but we highlighted that the exchange of mitochondria occurred preferentially between endothelial cells and cancer cells. We illustrated that the cancer cells acquiring mitochondria displayed chemoresistance. Our results illustrate the perfusion-independent role of the endothelium by showing a direct endothelial to cancer cell mitochondrial exchange associated to phenotypic modulation. This supports another role of the endothelium in the constitution of the metastatic niche.

Epithelial to Mesenchymal Transition in a Clinical Perspective

Tumor growth and metastatic dissemination rely on cellular plasticity. Among the different phenotypes acquired by cancer cells, epithelial to mesenchymal transition (EMT) has been extensively illustrated. Indeed, this transition allows an epithelial polarized cell to acquire a more mesenchymal phenotype with increased mobility and invasiveness. The role of EMT is quite clear during developmental stage. In the neoplastic context in many tumors EMT has been associated with a more aggressive tumor phenotype including local invasion and distant metastasis. EMT allows the cell to invade surrounding tissues and survive in the general circulation and through a stem cell phenotype grown in the host organ. The molecular pathways underlying EMT have also been clearly defined and their description is beyond the scope of this review. Here we will summarize and analyze the attempts made to block EMT in the therapeutic context. Indeed, till today, most of the studies are made in animal models. Few clinical trials are ongoing with no obvious benefits of EMT inhibitors yet. We point out the limitations of EMT targeting such tumor heterogeneity or the dynamics of EMT during disease progression.

Microparticles mediated cross-talk between tumoral and endothelial cells promote the constitution of a pro-metastatic vascular niche through Arf6 up regulation

The tumor stroma plays an essential role in tumor growth, resistance to therapy and occurrence of metastatic phenotype. Tumor vessels have been considered as passive conducts for nutrients but several studies have demonstrated secretion of pro-tumoral factors by endothelial cells. The failure of anti-angiogenic therapies to meet expectations raised by pre-clinical studies prompt us to better study the cross-talk between endothelial and cancer cells. Here, we hypothesized that tumor cells and the endothelium secrete bio-active microparticles (MPs) participating to a functional cross-talk. We characterized the cancer cells MPs, using breast and ovarian cancer cell lines (MCF7, MDA-MB231, SKOV3, OVCAR3 and a primary cell lines, APOCC). Our data show that MPs from mesenchymal-like cell lines (MDA-MB231, SKOV3 and APOCC) were able to promote an activation of endothelial cells through Akt phosphorylation, compared to MPs from epithelial-like cell lines (OVCAR3 and MCF7). The MPs from mesenchymal-like cells contained increased angiogenic molecules including PDGF, IL8 and angiogenin. The endothelial activation was associated to increased Arf6 expression and MPs secretion. Endothelial activation functionalized an MP dependent pro-tumoral vascular niche promoting cancer cells proliferation, invasiveness, stem cell phenotype and chemoresistance. MPs from cancer and endothelial cells displayed phenotypic heterogeneity, and participated to a functional cross-talk where endothelial activation by cancer MPs resulted in increased secretion of EC-MPs sustaining tumor cells. Such cross-talk may play a role in perfusion independent role of the endothelium.

Human Embryonic Stem Cell Derived Mesenchymal Progenitors Express Cardiac Markers but Do Not Form Contractile Cardiomyocytes

Mesenchymal progenitors or stromal cells have shown promise as a therapeutic strategy for a range of diseases including heart failure. In this context, we explored the growth and differentiation potential of mesenchymal progenitors (MPs) derived in vitro from human embryonic stem cells (hESCs). Similar to MPs isolated from bone marrow, hESC derived MPs (hESC-MPs) efficiently differentiated into archetypical mesenchymal derivatives such as chondrocytes and adipocytes. Upon treatment with 5-Azacytidine or TGF-β1, hESC-MPs modified their morphology and up-regulated expression of key cardiac transcription factors such as NKX2-5, MEF2C, HAND2 and MYOCD. Nevertheless, NKX2-5+ hESC-MP derivatives did not form contractile cardiomyocytes, raising questions concerning the suitability of these cells as a platform for cardiomyocyte replacement therapy. Gene profiling experiments revealed that, although hESC-MP derived cells expressed a suite of cardiac related genes, they lacked the complete repertoire of genes associated with bona fide cardiomyocytes. Our results suggest that whilst agents such as TGF-β1 and 5-Azacytidine can induce expression of cardiac related genes, but treated cells retain a mesenchymal like phenotype.

Role of the Microenvironment in Ovarian Cancer Stem Cell Maintenance

Despite recent progresses in cancer therapy and increased knowledge in cancer biology, ovarian cancer remains a challenging condition. Among the latest concepts developed in cancer biology, cancer stem cells and the role of microenvironment in tumor progression seem to be related. Indeed, cancer stem cells have been described in several solid tumors including ovarian cancers. These particular cells have the ability to self-renew and reconstitute a heterogeneous tumor. They are characterized by specific surface markers and display resistance to therapeutic regimens. During development, specific molecular cues from the tumor microenvironment can play a role in maintaining and expanding stemness of cancer cells. The tumor stroma contains several compartments: cellular component, cytokine network, and extracellular matrix. These different compartments interact to form a permissive niche for the cancer stem cells. Understanding the molecular cues underlying this crosstalk will allow the design of new therapeutic regimens targeting the niche. In this paper, we will discuss the mechanisms implicated in the interaction between ovarian cancer stem cells and their microenvironment.

Endothelial Cells Provide a Notch-Dependent Pro-Tumoral Niche for Enhancing Breast Cancer Survival, Stemness and Pro-Metastatic Properties

Treating metastasis has been challenging due to tumors complexity and heterogeneity. This complexity is partly related to the crosstalk between tumor and its microenvironment. Endothelial cells -the building blocks of tumor vasculature- have been shown to have additional roles in cancer progression than angiogenesis and supplying oxygen and nutrients. Here, we show an alternative role for endothelial cells in supporting breast cancer growth and spreading independent of their vascular functions. Using endothelial cells and breast cancer cell lines MDA-MB231 and MCF-7, we developed co-culture systems to study the influence of tumor endothelium on breast tumor development by both in vitro and in vivo approaches. Our results demonstrated that endothelial cells conferred survival advantage to tumor cells under complete starvation and enriched the CD44HighCD24Low/- stem cell population in tumor cells. Moreover, endothelial cells enhanced the pro-metastatic potential of breast cancer cells. The in vitro and in vivo results concordantly confirmed a role for endothelial Jagged1 to promote breast tumor through notch activation. Here, we propose a role for endothelial cells in enhancing breast cancer progression, stemness, and pro-metastatic traits through a perfusion-independent manner. Our findings may be beneficial in developing novel therapeutic approaches.

AN IN VITRO CELL POPULATION DYNAMICS MODEL INCORPORATING CELL SIZE, QUIESCENCE, AND CONTACT INHIBITION

In this paper, we construct a model to describe the spatial motion of a monolayer of cells occupying a two-dimensional dish. By taking care of nonlocal contact inhibition, quiescence phenomenon, and the cell cycle, we derive porous media-like equation with nonlocal reaction terms. The first part of this paper is devoted to the construction of the model. In the second part we study the well-posedness of the model. We conclude the paper by presenting some numerical simulations of the model and we observe the formation of colonies.

Epigenetics and Cardiovascular Disease in Diabetes

Type 2 diabetes has become a major health issue worldwide. Chronic hyperglycemia induces a low-grade inflammation that, on top of other mechanisms, leads to endothelial dysfunction. Mounting evidence suggests that DNA methylation, post-translational modifications of histones, and long non-coding RNAs play an important role in the initiation, maintenance, and progression of both macro- and micro-vascular complications of diabetes. Long-term exposure to hyperglycemia induces epigenetic changes that could become irreversible, a phenomenon known as the ‘metabolic memory.’ Whether epigenetic-based therapies could be used to slow or limit the progression of cardiovascular disease remains unclear. While non-coding RNAs are currently investigated as potential biomarkers that predict diabetic cardiovascular disease incidence and progression, their therapeutic role is only hypothetical. In this review, we highlight the latest findings in experimental and clinical studies relevant to epigenetics and cardiovascular disease in diabetes.

Structural and functional analysis of tunneling nanotubes (TnTs) using gCW STED and gconfocal approaches

Tunneling nanotubes (TnTs) are thin plasma membrane bridges mediating transfers of materials and signals between cells. Heterogeneity of heterocellular and homocellular TnTs is largely described but ultrafine imaging of these light‐sensitive floating nanometric structures represents a real challenge in microscopy. We propose here imaging strategies designed to dissect structural and dynamic aspects of TnT formation and function in fixed or living PC12 cells.

SIRT1 Limits Adipocyte Hyperplasia through c-Myc Inhibition

The expansion of fat mass in the obese state is due to increased adipocyte hypertrophy and hyperplasia. The molecular mechanism that drives adipocyte hyperplasia remains unknown. The NAD+-dependent protein deacetylase sirtuin 1 (SIRT1), a key regulator of mammalian metabolism, maintains proper metabolic functions in many tissues, counteracting obesity. Here we report that differentiated adipocytes are hyperplastic when SIRT1 is knocked down stably in mouse 3T3-L1 preadipocytes. This phenotype is associated with dysregulated adipocyte metabolism and enhanced inflammation. We also demonstrate that SIRT1 is a key regulator of proliferation in preadipocytes. Quantitative proteomics reveal that the c-Myc pathway is altered to drive enhanced proliferation in SIRT1-silenced 3T3-L1 cells. Moreover, c-Myc is hyperacetylated, levels of p27 are reduced, and cyclin-dependent kinase 2 (CDK2) is activated upon SIRT1 reduction. Remarkably, differentiating SIRT1-silenced preadipocytes exhibit enhanced mitotic clonal expansion accompanied by reduced levels of p27 as well as elevated levels of CCAAT/enhancer-binding protein β (C/EBPβ) and c-Myc, which is also hyperacetylated. c-Myc activation and enhanced proliferation phenotype are also found to be SIRT1-dependent in proliferating mouse embryonic fibroblasts and differentiating human SW872 preadipocytes. Reducing both SIRT1 and c-Myc expression in 3T3-L1 cells simultaneously does not induce the adipocyte hyperplasia phenotype, confirming that SIRT1 controls adipocyte hyperplasia through c-Myc regulation. A better understanding of the molecular mechanisms of adipocyte hyperplasia will open new avenues toward understanding obesity.