Development Of Transgenic Mice Research Articles

The malignant transformation potential of the oncogene STYK1/NOK at early lymphocyte development in transgenic mice

B-cell Chronic Lymphocytic Leukemia (B-CLL) is a malignancy caused by the clonal expansion of mature B lymphocytes bearing a CD5+CD19+ (B1) phenotype. However, the origin of B-CLL remains controversial. We showed previously that STYK1/NOK transgenic mice develop a CLL-like disease. Using this model system in this study, we attempt to define the stage of CLL initiation. Here, we show that the phenotype of STYK1/NOK-induced B-CLL is heterogeneous. The expanded B1 lymphocyte pool was detected within peripheral lymphoid organs and was frequently associated with the expansions of memory B cells. Despite this immunophenotypic heterogeneity, suppression of B cell development at an early stage consistently occurred within the bone marrow (BM) of STYK1/NOK-tg mice. Overall, we suggest that enforced expression of STYK1/NOK in transgenic mice might significantly predispose BM hematopoietic stem cells (HSCs) towards the development of B-CLL.

Potential activity of adiponectin-expressing regulatory T cells against triple-negative breast cancer cells through the cell-in-cell phenomenon.

A population of regulatory T cells (Treg), which reside within thymic nurse cell complexes, express adiponectin and abrogate breast cancer development in transgenic mice. In this study, we examined whether adiponectin-expressing Treg could impair triple-negative breast cancer, which is defined by a lack of estrogen receptors, progesterone receptors, and human epidermal growth factor receptor-2. CD4- and CD25-positive cells were sorted from cultured T lymphocytes of a previously characterized experimental thymic tumor model composed of thymic nurse cells and abundant lymphoid stroma. These sorted cells were examined for FOXP3 and adiponectin immunoreactivity and subsequently exposed to triple-negative breast cancer MDA-MB-157 and -231 cells. Adiponectin-expressing Treg were obtained by CD4- and CD25-positive sorting and cell death was induced in triple-negative breast cancer cells through the cell-in-cell phenomenon. Adiponectin-expressing Treg may be candidates for adoptive cell therapy against triple-negative breast cancer.

The role of SF3B1 mutations in EVI1-rearranged myeloid neoplasms

In acute myeloid leukemia (AML), EVI1 rearrangement represented by inv(3)(q21q26) or t(3;3)(q21;q26) causes EVI1 overexpression via structural rearrangement of an enhancer, and confers poor prognosis. My colleagues and I performed a mutational analysis of EVI1-rearranged myeloid neoplasms and identified SF3B1, a core RNA splicing factor, as the most commonly co-mutated gene. Indeed, latent leukemia development in transgenic mice bearing the humanized inv(3)(q21q26) allele was significantly accelerated by co-occurrence of Sf3b1 mutation. Intriguingly, we found that this SF3B1 mutant induced mis-splicing of EVI1 itself, which generated an aberrant EVI1 isoform with in-frame insertion of 6 amino acids near the DNA-binding domain of EVI1. This aberrant EVI1 isoform exhibited DNA-binding activity different from wild-type EVI1 and significantly enhanced the self-renewal capacity of murine hematopoietic stem cells. We also identified the cryptic branch point and exonic splicing enhancer required for this EVI1 mis-splicing induced by the SF3B1 mutant. These data provide a basis for further elucidation of the molecular mechanism and potential therapeutic candidates for EVI1-rearranged AML.

RelB contributes to the survival, migration and lymphomagenesis of B cells with constitutively active CD40 signaling.

Activation of CD40-signaling contributes to the initiation, progression and drug resistance of B cell lymphomas. We contributed to this knowledge by showing that constitutive CD40-signaling in B cells induces B cell hyperplasia and finally B cell lymphoma development in transgenic mice. CD40 activates, among others, the non-canonical NF-ĸB signaling, which is constitutively activated in several human B cell lymphomas and is therefore presumed to contribute to lymphopathogenesis. This prompted us to study the regulatory role of the non-canonical NF-ĸB transcription factor RelB in lymphomagenesis. To this end, we crossed mice expressing a constitutively active CD40 receptor in B cells with conditional RelB-KO mice. Ablation of RelB attenuated pre-malignant B cell expansion, and resulted in an impaired survival and activation of long-term CD40-stimulated B cells. Furthermore, we found that hyperactivation of non-canonical NF-кB signaling enhances the retention of B cells in the follicles of secondary lymphoid organs. RNA-Seq-analysis revealed that several genes involved in B-cell migration, survival, proliferation and cytokine signaling govern the transcriptional differences modulated by the ablation of RelB in long-term CD40-stimulated B cells. Inactivation of RelB did not abrogate lymphoma development. However, lymphomas occurred with a lower incidence and had a longer latency period. In summary, our data suggest that RelB, although it is not strictly required for malignant transformation, accelerates the lymphomagenesis of long-term CD40-stimulated B cells by regulating genes involved in migration, survival and cytokine signaling.

Y-018. Placental-derived human sFLT1 accumulates in maternal serum and alters placental and fetal development in transgenic mice

In previous work, we have developed an autoregressive Mixed-Effects model of the evolution of the kinetic gene expression parameters along cell generations, and an identification method simultaneously exploiting single-cell gene expression profiles and known parental relationships among cells (lineage tree data). Here, we extend our modelling and identification approach to explicitly account for stochasticity of promoter activation, and demonstrate via simulation the performance of the method and the improvement relative to the original approach where this source of noise is not accounted for.

TRIP13 modulates protein deubiquitination and accelerates tumor development and progression of B cell malignancies.

Multiple myeloma (MM), a terminally differentiated B cell malignancy, remains difficult to cure. Understanding the molecular mechanisms underlying the progression of MM may identify therapeutic targets and lead to a fundamental shift in treatment of the disease. Deubiquitination, like ubiquitination, is a highly regulated process, implicated in almost every cellular process. Multiple deubiquitinating enzymes (DUBs) have been identified, but their regulation is poorly defined. Here, we determined that TRIP13 increases cellular deubiquitination. Overexpression of TRIP13 in mice and cultured cells resulted in excess cellular deubiquitination by enhancing the association of the DUB USP7 with its substrates. We show that TRIP13 is an oncogenic protein because it accelerates B cell tumor development in transgenic mice. TRIP13-induced resistance to proteasome inhibition can be overcome by a USP7 inhibitor in vitro and in vivo. These findings suggest that TRIP13 expression plays a critical role in B cell lymphoma and MM by regulating deubiquitination of critical oncogenic (NEK2) and tumor suppressor (PTEN, p53) proteins. High TRIP13 identifies a high-risk patient group amenable to adjuvant anti-USP7 therapy.

Pathological Aspects of Neuronal Hyperploidization in Alzheimer's Disease Evidenced by Computer Simulation.

When subjected to stress, terminally differentiated neurons are susceptible to reactivate the cell cycle and become hyperploid. This process is well documented in Alzheimer’s disease (AD), where it may participate in the etiology of the disease. However, despite its potential importance, the effects of neuronal hyperploidy (NH) on brain function and its relationship with AD remains obscure. An important step forward in our understanding of the pathological effect of NH has been the development of transgenic mice with neuronal expression of oncogenes as model systems of AD. The analysis of these mice has demonstrated that forced cell cycle reentry in neurons results in most hallmarks of AD, including neurofibrillary tangles, Aβ peptide deposits, gliosis, cognitive loss, and neuronal death. Nevertheless, in contrast to the pathological situation, where a relatively small proportion of neurons become hyperploid, neuronal cell cycle reentry in these mice is generalized. We have recently developed an in vitro system in which cell cycle is induced in a reduced proportion of differentiated neurons, mimicking the in vivo situation. This manipulation reveals that NH correlates with synaptic dysfunction and morphological changes in the affected neurons, and that membrane depolarization facilitates the survival of hyperploid neurons. This suggests that the integration of synaptically silent, hyperploid neurons in electrically active neural networks allows their survival while perturbing the normal functioning of the network itself, a hypothesis that we have tested in silico. In this perspective, we will discuss on these aspects trying to convince the reader that NH represents a relevant process in AD.

Time dependent degeneration of the nigrostriatal tract in mice with 6-OHDA lesioned medial forebrain bundle and the effect of activin A on l-Dopa induced dyskinesia

BackgroundAccurately assessing promising therapeutic interventions for human diseases depends, in part, on the reproducibility of preclinical disease models. With the development of transgenic mice, the rapid adaptation of a 6-OHDA mouse model of Parkinson’s disease that was originally described for the use in rats has come with a lack of a comprehensive characterization of lesion progression. In this study we therefore first characterised the time course of neurodegeneration in the substantia nigra pars compacta and striatum over a 4 week period following 6-OHDA injection into the medial forebrain bundle of mice. We then utilised the model to assess the anti-dyskinetic efficacy of recombinant activin A, a putative neuroprotectant and anti-inflammatory that is endogenously upregulated during the course of Parkinson’s disease.ResultsWe found that degeneration of fibers in the striatum was fully established within 1 week following 6-OHDA administration, but that the loss of neurons continued to progress over time, becoming fully established 3 weeks after the 6-OHDA injection. In assessing the anti-dyskinetic efficacy of activin A using this model we found that treatment with activin A did not significantly reduce the severity, or delay the time-of-onset, of dyskinesia.ConclusionFirst, the current study concludes that a 3 week duration is required to establish a complete lesion of the nigrostriatal tract following 6-OHDA injection into the medial forebrain bundle of mice. Second, we found that activin A was not anti-dyskinetic in this model.

Premature aging and cancer development in transgenic mice lacking functional CYLD.

CYLD is a deubiquitinating enzyme known for its role as a tumor suppressor whose mutation leads to skin appendages tumors and other cancers. In this manuscript we report that the tumor suppressor CYLD, similarly to other renowned tumor suppressor genes, protects from premature aging and cancer. We have generated transgenic mice expressing the mutant CYLDC/S protein, lacking its deubiquitinase function, under the control of the keratin 5 promoter, the K5-CYLDC/S mice. These mice express the transgene in different organs, including those considered to be more susceptible to aging, such as skin and thymus. Our results show that K5-CYLDC/S mice exhibit epidermal, hair follicle, and sebaceous gland alterations; and, importantly, they show signs of premature aging from an early age. Typically, 3-month-old K5-CYLDC/S mice exhibit a phenotype characterized by alopecia and kyphosis, and, the histological examination reveals that transgenic mice show signs of accelerated aging in numerous organs such as skin, thymus, pancreas, liver and lung. Additionally, they spontaneously develop tumors of diverse origin. Over-activation of the NF-κB pathway, along with hyperactivation of Akt, JNK and c-Myc, and chronic inflammation, appear as the mechanisms responsible for the premature aging of the K5-CYLDC/S mice.

Female Mouse Model of Neisseria gonorrhoeae Infection.

Mouse models of infection are important tools in the study of infectious disease or host the development of products to prevent or treat infections. The estradiol-treated mouse model of Neisseria gonorrhoeae genital tract infection has proved to be a valuable system for determining the importance of gonococcal factors that mediate evasion of host innate effectors in vivo or host gonococcal adaptation to hormonally driven host factors in females. Examination of mechanisms that Neisseria gonorrhoeae uses to subvert the host immune response also has been greatly aided by this whole model system, as have studies on the consequence of antibiotic resistance mutations on gonococcal fitness in vivo and the search for new antibiotics to treat antibiotic-resistant infections. The strict human specificity of N. gonorrhoeae limits the ability of experimental murine infection to mimic human infection. However, in recent years, the development of transgenic mice and protocols for supplementing mice with human factors has improved animal modeling of gonorrhea. To date, however, because the mouse estrous cycle is much shorter than the human reproductive cycle, all reported gonorrhea mouse models require treatment with estradiol and antibiotics to maintain an estrus-like state and suppress the overgrowth of inhibitory commensal flora that occurs under the influence of estrogen to allow sustained N. gonorrhoeae infection. In this chapter, we detail the methods used to (1) prepare the mice for experimental infection with N. gonorrhoeae, (2) inoculate mice and quantitatively culture vaginal swabs for noncompetitive and competitive infection experiments, and (3) monitor the host innate immune response to infection.

Immunomodulatory Roles of PACAP and VIP: Lessons from Knockout Mice.

A bidirectional cross-talk is established between the nervous and immune systems through common mediators including neuropeptides, neurotransmitters, and cytokines. Among these, PACAP and VIP are two highly related neuropeptides widely distributed in the organism with purported immunomodulatory actions. Due to their well-known anti-inflammatory properties, administration of these peptides has proven to be beneficial in models of acute and chronic inflammatory diseases. Nevertheless, the relevance of the endogenous source of these peptides in the modulation of immune responses remains to be elucidated. The development of transgenic mice with specific deletions in the genes coding for these neuropeptides (Vip and Adcyap1) or for their G-protein-coupled receptors VPAC1, VPAC2, and PAC1 (Vipr1, Vipr2, Adcyap1r1) has allowed to address this question, underscoring the complexity of the immunoregulatory properties of PACAP and VIP. The goal of this review is to integrate the existing information on the immune phenotypes of mice deficient for PACAP, VIP, or their receptors, to provide a global view on the roles of these endogenous neuropeptides during immunological health and disease.

ATDC/TRIM29 Drives Invasive Bladder Cancer Formation through miRNA-Mediated and Epigenetic Mechanisms

Bladder cancer is a common and deadly malignancy but its treatment has advanced little due to poor understanding of the factors and pathways that promote disease. ATDC/TRIM29 is a highly expressed gene in several lethal tumor types, including bladder tumors, but its role as a pathogenic driver has not been established. Here we show that overexpression of ATDC in vivo is sufficient to drive both noninvasive and invasive bladder carcinoma development in transgenic mice. ATDC-driven bladder tumors were indistinguishable from human bladder cancers, which displayed similar gene expression signatures. Clinically, ATDC was highly expressed in bladder tumors in a manner associated with invasive growth behaviors. Mechanistically, ATDC exerted its oncogenic effects by suppressing miR-29 and subsequent upregulation of DNMT3A, leading to DNA methylation and silencing of the tumor suppressor PTEN. Taken together, our findings established a role for ATDC as a robust pathogenic driver of bladder cancer development, identified downstream effector pathways, and implicated ATDC as a candidate biomarker and therapeutic target.

Disruption of neurogenesis and cortical development in transgenic mice misexpressing Olig2, a gene in the Down syndrome critical region

The basic helix-loop-helix (bHLH) transcription factor Olig2 is crucial for mammalian central nervous system development. Human ortholog OLIG2 is located in the Down syndrome critical region in trisomy 21. To investigate the effect of Olig2 misexpression on brain development, we generated a developmentally regulated Olig2-overexpressing transgenic line with a Cre/loxP system. The transgenic mice with Olig2 misexpression in cortical neural stem/progenitor cells exhibited microcephaly, cortical dyslamination, hippocampus malformation, and profound motor deficits. Ectopic misexpression of Olig2 impaired cortical progenitor proliferation and caused precocious cell cycle exit. Massive neuronal cell death was detected in the developing cortex of Olig2-misexpressing mice. In addition, Olig2 misexpression led to a significant downregulation of neuronal specification factors including Ngn1, Ngn2 and Pax6, and a defect in cortical neurogenesis. Chromatin-immunoprecipitation and sequencing (ChIP-Seq) analysis indicates that Olig2 directly targets the promoter and/or enhancer regions of Nfatc4, Dscr1/Rcan1 and Dyrk1a, the critical neurogenic genes that contribute to Down syndrome phenotypes, and inhibits their expression. Together, our study suggests that Olig2 misexpression in neural stem cells elicits neurogenesis defects and neuronal cell death, which may contribute to developmental disorders including Down syndrome, where OLIG2 is triplicated on chromosomal 21.

B-cell expansion and lymphomagenesis induced by chronic CD40 signaling is strictly dependent on CD19.

CD40, a member of the TNF receptor family, is expressed on all mature B cells and on most B-cell lymphomas. Recently, we have shown that constitutive activation of CD40 signaling in B cells induced by a fusion protein consisting of the transmembrane part of the Epstein-Barr viral latent membrane protein 1 (LMP1) and the cytoplasmic part of CD40 (LMP1/CD40) drives B-cell lymphoma development in transgenic mice. Because LMP1/CD40-expressing B cells showed an upregulation of CD19, we investigated CD19's function in CD40-driven B-cell expansion and lymphomagenesis. Here, we demonstrate that ablation of CD19 in LMP1/CD40 transgenic mice resulted in a severe loss and reduced lifespan of mature B cells and completely abrogated development of B-cell lymphoma. CD19 is localized to lipid rafts and constitutively activated by the LMP1/CD40 fusion protein in B cells. We provide evidence that the improved survival and malignant transformation of LMP1/CD40-expressing B cells are dependent on activation of the MAPK Erk that is mediated through CD19 in a PI3K-dependent manner. Our data suggest that constitutively active CD40 is dependent on CD19 to transmit survival and proliferation signals. Moreover, we detected a similarly functioning prosurvival pathway involving phosphorylated CD19 and PI3K-dependent Erk phosphorylation in human diffuse large B-cell lymphoma cell lines. Our data provide evidence that CD19 plays an important role in transmitting survival and proliferation signals downstream of CD40 and therefore might be an interesting therapeutic target for the treatment of lymphoma undergoing chronic CD40 signaling.

Zebrafish as a Model System to Study Skin Biology and Pathology

INTRODUCTION Several animal models have been developed to recapitulate the features of specific skin diseases, and such model systems have provided valuable insight into the pathomechanistic details of these diseases. They have also provided useful systems for testing of treatment modalities for a number of conditions. The preferred platform for such model development has been the mouse, often through the development of “knockout” mice or by development of transgenic mice with expression of the mutant genes. Although the mouse models have often demonstrated remarkable similarity to human diseases, mice as a model system have considerable limitations, including relatively long lifespan and high cost of development. Also, the genetic background of the mouse strains can have a major influence on the development of the disease phenotype. In some cases, development of the mouse model is not feasible because of the absence of the corresponding gene in the mouse genome. These considerations, particularly in conjunction with cost-containment issues, have prompted the search for an alternative model system to study skin diseases. In fact, Vanchieri (2001) suggested, “Move over mouse: make way for the woodchucks, ferrets, and zebrafish,” and Lieschke and Currie (2007) said, “Animal models of human disease: zebrafish swim into view.”

Longitudinal in vivo two‐photon fluorescence imaging

Fluorescence microscopy is an essential technique for the basic sciences, especially biomedical research. Since the invention of laser scanning confocal microscopy in the 1980s, which enabled imaging both fixed and living biological tissue with 3D precision, high-resolution fluorescence imaging has revolutionized biological research. Confocal microscopy, by its very nature, has one fundamental limitation. Due to the confocal pinhole, deep tissue fluorescence imaging is not practical. In contrast (no pun intended), two-photon fluorescence microscopy allows, in principle, the collection of all emitted photons from fluorophores in the imaged voxel, dramatically extending our ability to see deep into living tissue. Since the development of transgenic mice with genetically encoded fluorescent protein in neocortical cells in 2000, two-photon imaging has enabled the dynamics of individual synapses to be followed for up to 2 years. Since the initial landmark contributions to this field in 2002, the technique has been used to understand how neuronal structure are changed by experience, learning, and memory and various diseases. Here we provide a basic summary of the crucial elements that are required for such studies, and discuss many applications of longitudinal two-photon fluorescence microscopy that have appeared since 2002.

Postnatal cardiac development in transgenic mice with altered expression of the DNA glycosylase Ogg1 (573.4)

A switch from cardiomyocyte hyperplasia to hypertrophy underlies postnatal cardiac development in rodents. Interestingly, this transition is associated with robust mitochondrial biogenesis, which, paradoxically, is accompanied by oxidative mitochondrial (mt) DNA damage and repair (Pohjoismaki et al, NAR, 2012). Whether mtDNA damage and repair are functionally linked to mitochondrial biogenesis is unknown, but previous experiments showed that controlled DNA damage and repair are important for transcription of nuclear genes (Al‐Mehdi et al, Sci Signaling, 2012). To determine if a similar mechanism could be operative in the mitochondrial genome during postnatal cardiac development, we measured oxidative damage in the mtDNA D‐loop transcriptional regulatory sequence and the coding region as a function of time in hearts derived from wild type (WT) mice, knock‐out mice deficient in the DNA glycosylase Ogg1 (KO), and in KO mice transgenically over‐expressing human OGG1 targeted to mitochondria. In comparison to WT animals, KO mice displayed higher mtDNA damage which preceded an exaggerated postnatal increase in heart‐to‐body mass ratio. Conversely, KO mice over‐expressing mitochondria‐targeted OGG1 displayed decreased mtDNA oxidation and a blunted elevation in cardiac mass relative to KO mice. These results suggest mtDNA damage and repair may be functionally linked to postnatal cardiac development.Grant Funding Source: Supported by the NIH R21HL102789, R01HL113614

An Enteroendocrine Cell – Enteric Glia Connection Revealed by 3D Electron Microscopy

The enteroendocrine cell is the cornerstone of gastrointestinal chemosensation. In the intestine and colon, this cell is stimulated by nutrients, tastants that elicit the perception of flavor, and bacterial by-products; and in response, the cell secretes hormones like cholecystokinin and peptide YY – both potent regulators of appetite. The development of transgenic mice with enteroendocrine cells expressing green fluorescent protein has allowed for the elucidation of the apical nutrient sensing mechanisms of the cell. However, the basal secretory aspects of the enteroendocrine cell remain largely unexplored, particularly because a complete account of the enteroendocrine cell ultrastructure does not exist. Today, the fine ultrastructure of a specific cell can be revealed in the third dimension thanks to the invention of serial block face scanning electron microscopy (SBEM). Here, we bridged confocal microscopy with SBEM to identify the enteroendocrine cell of the mouse and study its ultrastructure in the third dimension. The results demonstrated that 73.5% of the peptide-secreting vesicles in the enteroendocrine cell are contained within an axon-like basal process. We called this process a neuropod. This neuropod contains neurofilaments, which are typical structural proteins of axons. Surprisingly, the SBEM data also demonstrated that the enteroendocrine cell neuropod is escorted by enteric glia – the cells that nurture enteric neurons. We extended these structural findings into an in vitro intestinal organoid system, in which the addition of glial derived neurotrophic factors enhanced the development of neuropods in enteroendocrine cells. These findings open a new avenue of exploration in gastrointestinal chemosensation by unveiling an unforeseen physical relationship between enteric glia and enteroendocrine cells.

The development of transgenic mice for the expression of large amounts of human lysozyme in milk

Human lysozyme (hLYZ) has important potential applications as antimicrobial medicine and food additive. To develop a robust expression vector that ensures expression of large amounts of hLYZ in milk, here a 26,267bp chimeric mouse whey acidic protein (mWAP)::hLYZ cassette was constructed and used as a mammary gland-specific expression vector, in which a 3,010bp genomic sequence in the 24,466bp mWAP gene locus was substituted by a 4,811bp genomic sequence of hLYZ, exactly from the start codon to the stop codon. Corresponding transgenic mice were generated, and enzymatically-active hLYZ was expressed at 18.4-35gl(-1) in the milk of most transgenic mouse lines. Our transgenic mice carrying chimeric mWAP::hLYZ represent a model system for cost-effective production of hLYZ.

Abstract S2-06: Extracellular matrix stiffness modulates tissue inflammation to promote breast tumor aggression

Abstract Tumor progression is influenced by the dynamic interplay between the genetically-modified epithelium and the associated microenvironment. We previously showed that the extra-cellular matrix (ECM) progressively stiffens as mammary tumors evolve, and that stiffening the ECM promotes malignant transformation while inhibiting ECM stiffening reduces tumor progression (Levental et al., 2009; reviewed in Butcher et al., 2009). Yet breast cancers also exhibit elevated influx of inflammatory cells and tissue inflammation promotes tumor progression by fostering angiogenesis and enhancing tumor cell growth and motility (Ruffell et al., 2011). Tumor-associated macrophages additionally stimulate stromal fibroblasts, implying that tissue inflammation could promote malignancy by stiffening the ECM. To address this possibility, we explored the relationship between ECM tension and macrophages in human breast cancer and during mammary tumor development in transgenic mice. Using fresh and archived tissue samples, we quantified a progressive increase in ECM remodeling and stiffening as human breast tumors and mouse mammary cancers developed. We found that the ECM associated with triple negative breast cancers was almost twice as stiff as the ECM adjacent to ER+/PR+ tumors and that ECM stiffness correlated significantly with increased numbers of infiltrating activated macrophages. Consistently, we established a positive correlation between tumor progression, elevated mechanosignaling, ECM stiffness and tissue inflammation in both human and mouse tissue. However surprisingly, when macrophages were depleted from the mammary glands of MMTV-PyMT tumors, ECM tension did not change. Instead we found that inhibiting lysyl oxidase activity to decrease collagen cross-linking and ECM stiffness reduced the activation state of the macrophages within the mammary glands of 14-week old mice. Consistently, in vitro studies showed that ECM tension potentiates the expression of pro-inflammatory chemokines and increases levels of phosphoStat3 and that ECM tension directly modifies macrophage polarity. These findings suggest that ECM tension may promote malignancy either by directly regulating macrophage activity or indirectly through enhancing expression of pro-inflammatory chemokines. Further studies are now underway to explore these findings and to clarify their impact on tumor progression and response to therapy. Acknowledgements: supported by W81XWH-05-1-0330 and R01 CA138818-01A1 to VMW, 1U01 ES019458-01 to VMW and ZW, and P50 CA 58207 to JG, VW, SH and LC, U54CA143836-01 to JL and VW, and Susan G. Komen for the Cure PSF12230246 to IA. Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr S2-06.