Exosome loaded immunomodulatory biomaterials alleviate local immune response in immunocompetent diabetic mice post islet xenotransplantation

Foreign body response (FBR) to biomaterials compromises the function of implants and leads to medical complications. Here, we report a hybrid alginate microcapsule (AlgXO) that attenuated the immune response after implantation, through releasing exosomes derived from human Umbilical Cord Mesenchymal Stem Cells (XOs). Upon release, XOs suppress the local immune microenvironment, where xenotransplantation of rat islets encapsulated in AlgXO led to >170 days euglycemia in immunocompetent mouse model of Type 1 Diabetes. In vitro analyses revealed that XOs suppressed the proliferation of CD3/CD28 activated splenocytes and CD3+ T cells. Comparing suppressive potency of XOs in purified CD3+ T cells versus splenocytes, we found XOs more profoundly suppressed T cells in the splenocytes co-culture, where a heterogenous cell population is present. XOs also suppressed CD3/CD28 activated human peripheral blood mononuclear cells (PBMCs) and reduced their cytokine secretion including IL-2, IL-6, IL-12p70, IL-22, and TNFα. We further demonstrate that XOs mechanism of action is likely mediated via myeloid cells and XOs suppress both murine and human macrophages partly by interfering with NFκB pathway. We propose that through controlled release of XOs, AlgXO provide a promising new platform that could alleviate the local immune response to implantable biomaterials

Defining Immunosuppressive Myeloid Cells in Breast Cancer in Single Cell Resolution

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of innate immunecells that have the capacity to suppress adaptive immune responses. In cancer patients, increased numbers of MDSCs in circulation correlate with advanced clinical stages, increased metastatic progression and immune suppression. MDSCs can be further classified into granulocytic MDSCs (G-MDSCs) and monocytic MDSC (M-MDSCs). However, since MDSCs, normal neutrophil granulocytes, and monocytes are defined by the same flow cytometry markers (CD11b+Gr1+), finding MDSC-specific markers is essential. We use a breast cancer mouse model (MMTV-PyMT) to define the cellular and molecular properties of MDSCs in single cell resolution. To test the capacity of CD11b+Gr1+ cells to inhibit immune responses, CD11b+Gr1+ cells from PyMT and wild type (WT) mice were sorted by fluorescence-activated cell sorting (FACS) from bone marrow, lung, and spleen, and subjected to a T cell activation assay in co-culture with T cells. We found that spleen-derived CD11b+Gr1+ cells from PyMT significantly suppressed CD4 and CD8 T cell proliferation, while CD11b+Gr1+ cells from bone marrow and lung showed no effect on T cell proliferation. Therefore single-cell RNA sequencing (scRNAseq) was used to compare MDSC-containing splenic myeloid cells from breast tumor bearing mice to wildtype controls. Our computational analysis of 14,646 single cell transcriptomes revealed a MDSCs gene signature and identify CD84 as a surface marker for improved detection and enrichment of MDSCs in breast cancers model and human. Additionally, our data provides a MDSC-specific gene signature and identified specific surface markers for MDSCs detection and enrichment in this breast cancer model and human. Our studies provide crucial insight into the biology of MDSCs, which may ultimately reveal novel markers and therapeutic avenues to improve cancer immunotherapy

Defining Immunosuppressive Myeloid Cells in Breast Cancer in Single Cell Resolution

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of innate immunecells that have the capacity to suppress adaptive immune responses. In cancer patients, increased numbers of MDSCs in circulation correlate with advanced clinical stages, increased metastatic progression and immune suppression. MDSCs can be further classified into granulocytic MDSCs (G-MDSCs) and monocytic MDSC (M-MDSCs). However, since MDSCs, normal neutrophil granulocytes, and monocytes are defined by the same flow cytometry markers (CD11b+Gr1+), finding MDSC-specific markers is essential. We use a breast cancer mouse model (MMTV-PyMT) to define the cellular and molecular properties of MDSCs in single cell resolution. To test the capacity of CD11b+Gr1+ cells to inhibit immune responses, CD11b+Gr1+ cells from PyMT and wild type (WT) mice were sorted by fluorescence-activated cell sorting (FACS) from bone marrow, lung, and spleen, and subjected to a T cell activation assay in co-culture with T cells. We found that spleen-derived CD11b+Gr1+ cells from PyMT significantly suppressed CD4 and CD8 T cell proliferation, while CD11b+Gr1+ cells from bone marrow and lung showed no effect on T cell proliferation. Therefore single-cell RNA sequencing (scRNAseq) was used to compare MDSC-containing splenic myeloid cells from breast tumor bearing mice to wildtype controls. Our computational analysis of 14,646 single cell transcriptomes revealed a MDSCs gene signature and identify CD84 as a surface marker for improved detection and enrichment of MDSCs in breast cancers model and human. Additionally, our data provides a MDSC-specific gene signature and identified specific surface markers for MDSCs detection and enrichment in this breast cancer model and human. Our studies provide crucial insight into the biology of MDSCs, which may ultimately reveal novel markers and therapeutic avenues to improve cancer immunotherapy

Luminol luminescence-based theranostics for pre-clinical breast adenocarcinoma

Master of ScienceDepartment of Anatomy & PhysiologyDeryl L. TroyerBreast cancer ranks second as a cause of cancer death in women in the USA. Detection of early tumors and tumor-targeted treatments could decrease the problems associated with breast cancer management. Photodynamic therapy (PDT) is a cancer treatment that uses a photosensitizer and a specific wavelength of light and is currently in clinical trials for breast cancer. When tumor cells which have absorbed photosensitizer are exposed to the correct wavelength of light, reactive oxygen species are generated, resulting in tumor cell death. Poor tissue penetration of light is a major limitation in PDT, restricting its use to treatment of localized tumors. Light generation at the tumor area might increase the effectiveness of PDT. Polymorphonuclear neutrophils (PMNs) are known to often infiltrate breast adenocarcinoma, and their activatation in tumor stroma produces luminescence in the presence of luminol. Here, we hypothesized that luminol can be used as a theranostic agent for luminescence-based early tumor detection (diagnosis) and in situ PDT (treatment). BALB/c mice were transplanted with 4T1 mammary adenocarcinoma cells to establish a breast adenocarcinoma model. The early tumor detection objective was tested by daily intraperitoneal injection of luminol and in vivo luminescence imaging. To test the PDT treatment objective,the photosensitizer 5-aminolevulinic acid (ALA) and luminol were administered to mice through intraperitoneal and intravenous routes, respectively. This treatment regimen was repeated six times and ALA alone/luminol alone/saline treated tumor-bearing mice were used as controls. Results demonstrated that luminol allowed detection of activated PMNs only two days after 4T1 cell transplantation, even though tumors were not yet palpable. Relative differences in the increase of tumor volume and final tumor weights were analyzed to test the in situ PDT. Analysis of the data showed luminol treatments resulted in breast adenocarcinoma tumor growth attenuation. In conclusion this study provides evidence that luminol can be a theranostic agent for breast adenocarcinoma

Luminol-based bioluminescence imaging of mouse mammary tumors

Polymorphonuclear neutrophils (PMNs) are the most abundant circulating blood leukocytes. They are part of the innate immune system and provide a first line of defense by migrating toward areas of inflammation in response to chemical signals released from the site. Some solid tumors, such as breast cancer, also cause recruitment and activation of PMNs and release of myeloperoxidase. In this study, we demonstrate that administration of luminol to mice that have been transplanted with 4T1 mammary tumor cells permits the detection of myeloperoxidase activity, and consequently, the location of the tumor. Luminol allowed detection of activated PMNs only two days after cancer cell transplantation, even though tumors were not yet palpable. In conclusion, luminol-bioluminescence imaging (BLI) can provide a pathway towards detection of solid tumors at an early stage in preclinical tumor models

Defining the emergence of myeloid-derived suppressor cells in breast cancer using single-cell transcriptomics

Myeloid-derived suppressor cells (MDSCs) are innate immune cells that acquire the capacity to suppress adaptive immune responses during cancer. It remains elusive how MDSCs differ from their normal myeloid counterparts, which limits our ability to specifically detect and therapeutically target MDSCs during cancer. Here, we sought to determine the molecular features of breast cancer-associated MDSCs using the widely studied mouse model based on the mouse mammary tumor virus (MMTV) promoter-driven expression of the polyomavirus middle T oncoprotein (MMTV-PyMT). To identify MDSCs in an unbiased manner, we used single-cell RNA sequencing to compare MDSC-containing splenic myeloid cells from breast tumor-bearing mice with wild-type controls. Our computational analysis of 14,646 single-cell transcriptomes revealed that MDSCs emerge through an aberrant neutrophil maturation trajectory in the spleen that confers them an immunosuppressive cell state. We establish the MDSC-specific gene signature and identify CD84 as a surface marker for improved detection and enrichment of MDSCs in breast cancers

A nanobiosensor for the detection of arginase activity

A nanobiosensor for arginase detection was designed and synthesized. It features a central dopamine-coated iron/iron oxide nanoparticle to which sulfonated cyanine 7.0 is tethered via a stable amide bond. Cyanine 5.5 is linked to the N-terminal of the peptide sequence GRRRRRRRG. Arginine (R) reacts to ornithine (O) in the presence of arginase. Based on calibration with commercially obtained arginase II, the limit of detection (LOD) is picomolar. It is noteworthy that the nanobiosensor for arginase detection does not show a fluorescence increase when incubated with the enzyme NO-reductase, which also uses arginase as substrate, but is indicative of an inflammatory response by the host to cancer and infections. Arginase activity was determined in a syngeneic mouse model for aggressive breast cancer (4 T1 tumors in BALB/c mice). It was found that the arginase activity is systemically enhanced, but especially pronounced in the active tumor regions. (C) 2016 Elsevier Inc. All rights reserved