Shinnick K.M.,Feinberg Cardiovascular Research Institute |
Shinnick K.M.,Robert rie Cancer Center |
Eklund E.A.,Robert rie Cancer Center |
Eklund E.A.,Jesse Brown Medical Center |
And 2 more authors.
Journal of Clinical Investigation | Year: 2010
HSCs maintain the circulating blood cell population. Defects in the orderly pattern of hematopoietic cell division and differentiation can lead to leukemia, myeloproliferative disorders, or marrow failure; however, the factors that control this pattern are incompletely understood. Geminin is an unstable regulatory protein that regulates the extent of DNA replication and is thought to coordinate cell division with cell differentiation. Here, we set out to determine the function of Geminin in hematopoiesis by deleting the Geminin gene (Gmnn) from mouse bone marrow cells. This severely perturbed the pattern of blood cell production in all 3 hematopoietic lineages (erythrocyte, megakaryocyte, and leukocyte). Red cell production was virtually abolished, while megakaryocyte production was greatly enhanced. Leukocyte production transiently decreased and then recovered. Stem and progenitor cell numbers were preserved, and Gmnn-/- HSCs successfully reconstituted hematopoiesis in irradiated mice. CD34+ Gmnn-/- leukocyte precursors displayed DNA overreplication and formed extremely small granulocyte and monocyte colonies in methylcellulose. While cultured Gmnn-/- megakaryocyte-erythrocyte precursors did not form erythroid colonies, they did form greater than normal numbers of megakaryocyte colonies. Gmnn-/- megakaryocytes and erythroblasts had normal DNA content. These data led us to postulate that Geminin regulates the relative production of erythrocytes and megakaryocytes from megakaryocyte-erythrocyte precursors by a replication-independent mechanism.
McMahon K.M.,Northwestern University |
McMahon K.M.,Institute for BioNanotechnology and Medicine IBNAM |
Mutharasan R.K.,Feinberg Cardiovascular Research Institute |
Tripathy S.,Northwestern University |
And 15 more authors.
Nano Letters | Year: 2011
We report a gold nanoparticle-templated high density lipoprotein (HDL AuNP) platform for gene therapy that combines lipid-based nucleic acid transfection strategies with HDL biomimicry. For proof-of-concept, HDL AuNPs are shown to adsorb antisense cholesterylated DNA. The conjugates are internalized by human cells, can be tracked within cells using transmission electron microscopy, and regulate target gene expression. Overall, the ability to directly image the AuNP core within cells, the chemical tailorability of the HDL AuNP platform, and the potential for cell-specific targeting afforded by HDL biomimicry make this platform appealing for nucleic acid delivery. © 2011 American Chemical Society.
Sahoo S.,Feinberg Cardiovascular Research Institute |
Losordo D.W.,Feinberg Cardiovascular Research Institute |
Losordo D.W.,Northwestern Memorial Hospital |
Losordo D.W.,Neostem, Inc.
Circulation Research | Year: 2014
Myocardial infarction is a leading cause of death among all cardiovascular diseases. The analysis of molecular mechanisms by which the ischemic myocardium initiates repair and remodeling indicates that secreted soluble factors are key players in communication to local and distant tissues, such as bone marrow. Recently, actively secreted membrane vesicles, including exosomes, are being recognized as new candidates with important roles in intercellular and tissue-level communication. In this review, we critically examine the emerging role of exosomes in local and distant microcommunication mechanisms after myocardial infarction. A comprehensive understanding of the role of exosomes in cardiac repair after myocardial infarction could bridge a major gap in knowledge of the repair mechanism after myocardial injury. © 2014 American Heart Association, Inc.
Shah A.,University of Toronto |
Shah A.,Toronto General Research Institute |
Xia L.,University of Toronto |
Xia L.,Toronto General Research Institute |
And 12 more authors.
Journal of the American Society of Nephrology | Year: 2015
Expression of thioredoxin-interacting protein (TxNIP), an endogenous inhibitor of the thiol oxidoreductase thioredoxin, is augmented by high glucose (HG) and promotes oxidative stress.We previously reported that TxNIP-deficient mesangial cells showed protection from HG-induced reactive oxygen species, mitogenactivated protein kinase phosphorylation, and collagen expression.Here,we investigated the potential roleof TxNIP in the pathogenesis of diabetic nephropathy (DN) in vivo. Wild-type (WT) control, TxNIP-/-, and TxNIP+/- mice were rendered equally diabetic with low-dose streptozotocin. In contrast to effects in WT mice, diabetes did not increase albuminuria, proteinuria, serum cystatin C, or serum creatinine levels in TxNIP-/- mice.Whereas morphometric studies of kidneys revealed a thickened glomerular basement membrane and effaced podocytes in the diabetic WT mice, these changes were absent in the diabetic TxNIP-/-mice. Immunohistochemical analysis revealed significant increases in the levels of glomerular TGF-β1, collagen IV, and fibrosis only inWTdiabeticmice. Additionally, onlyWTdiabeticmice showed significant increases in oxidative stress (nitrotyrosine, urinary 8-hydroxy-2-deoxy-guanosine) and inflammation (IL-1bmRNA, F4/80 immunohistochemistry). Expression levels ofNox4-encodedmRNA and protein increased only in the diabetic WT animals. A significant loss of podocytes, assessed by Wilms' tumor 1 and nephrin staining and urinary nephrin concentration, was found in diabetic WT but not TxNIP-/- mice. Furthermore, in cultured human podocytes exposed to HG, TxNIP knockdown with siRNA abolished the increased mitochondrial O2 2 generation and apoptosis. These data indicate that TxNIP has a critical role in the progression ofDNandmay be a promising therapeutic target. Copyright © 2015 by the American Society of Nephrology.
PubMed | University of Oulu, Feinberg Cardiovascular Research Institute, University of Edinburgh, Institute for Molecular Medicine and Soochow University of China
Type: Journal Article | Journal: Experimental cell research | Year: 2015
The indifferent mammalian embryonic gonad generates an ovary or testis, but the factors involved are still poorly known. The Wnt-4 signal represents one critical female determinant, since its absence leads to partial female-to-male sex reversal in mouse, but its signalling is as well implicated in the testis development. We used the Wnt-4 deficient mouse as a model to identify candidate gonadogenesis genes, and found that the Notum, Phlda2, Runx-1 and Msx1 genes are typical of the wild-type ovary and the Osr2, Dach2, Pitx2 and Tacr3 genes of the testis. Strikingly, the expression of these latter genes becomes reversed in the Wnt-4 knock-out ovary, suggesting a role in ovarian development. We identified the transcription factor Runx-1 as a Wnt-4 signalling target gene, since it is expressed in the ovary and is reduced upon Wnt-4 knock-out. Consistent with this, introduction of the Wnt-4 signal into early ovary cells ex vivo induces Runx-1 expression, while conversely Wnt-4 expression is down-regulated in the absence of Runx-1. We conclude that the Runx-1 gene can be a Wnt-4 signalling target, and that Runx-1 and Wnt-4 are mutually interdependent in their expression. The changes in gene expression due to the absence of Wnt-4 in gonads reflect the sexually dimorphic role of this signal and its complex gene network in mammalian gonad development.
Biyashev D.,Northwestern University |
Biyashev D.,Feinberg Cardiovascular Research Institute |
Veliceasa D.,Northwestern University |
Topczewski J.,Northwestern University |
And 8 more authors.
Blood | Year: 2012
We discovered that miR-27b controls 2 critical vascular functions: it turns the angiogenic switch on by promoting endothelial tip cell fate and sprouting and it promotes venous differentiation.We have identified its targets, a Notch ligand Deltalike ligand 4 (Dll4) and Sprouty homologue 2 (Spry2). miR-27b knockdown in zebrafish and mouse tissues severely impaired vessel sprouting and filopodia formation. Moreover, miR-27b was necessary for the formation of the first embryonic vein in fish and controlled the expression of arterial and venous markers in human endothelium, including Ephrin B2 (EphB2), EphB4, FMS-related tyrosine kinase 1 (Flt1), and Flt4. In zebrafish, Dll4 inhibition caused increased sprouting and longer intersegmental vessels and exacerbated tip cell migration. Blocking Spry2 caused premature vessel branching. In contrast, Spry2 overexpression eliminated the tip cell branching in the intersegmental vessels. Blockade of Dll4 and Spry2 disrupted arterial specification and augmented the expression of venous markers. Blocking either Spry2 or Dll4 rescued the miR-27b knockdown phenotype in zebrafish and in mouse vascular explants, pointing to essential roles of these targets downstream of miR-27b. Our study identifies critical role of miR-27b in the control of endothelial tip cell fate, branching, and venous specification and determines Spry2 and Dll4 as its essential targets. © 2012 by The American Society of Hematology.
Barry K.A.,Feinberg Cardiovascular Research Institute |
Schultz K.M.,Feinberg Cardiovascular Research Institute |
Payne C.J.,Childrens Memorial Research Center |
McGarry T.J.,Feinberg Cardiovascular Research Institute
Developmental Biology | Year: 2012
Spermatogonial stem cells divide throughout life, maintaining their own population and giving rise to differentiated gametes. The unstable regulatory protein Geminin is thought to be one of the factors that determine whether stem cells continue to divide or terminally differentiate. Geminin regulates the extent of DNA replication and is thought to maintain cells in an undifferentiated state by inhibiting various transcription factors and chromatin remodeling proteins. To examine how Geminin might regulate spermatogenesis, we developed two conditional mouse models in which the Geminin gene (Gmnn) is deleted from either spermatogonia or meiotic spermatocytes. Deleting Geminin from spermatogonia causes complete sterility in male mice. Gmnn(-/-) spermatogonia disappear during the initial wave of mitotic proliferation that occurs during the first week of life. Gmnn(-/-) spermatogonia exhibit more double-stranded DNA breaks than control cells, consistent with a defect in DNA replication. They maintain expression of genes associated with the undifferentiated state and do not prematurely express genes characteristic of more differentiated spermatogonia. In contrast, deleting Geminin from spermatocytes does not disrupt meiosis or the differentiation of spermatids into mature sperm. In females, Geminin is not required for meiosis, oocyte differentiation, or fertility after the embryonic period of mitotic proliferation has ceased. We conclude that Geminin is absolutely required for mitotic proliferation of spermatogonia but does not regulate their differentiation. Our results suggest that Geminin maintains replication fidelity during the mitotic phase of spermatogenesis, ensuring the precise duplication of genetic information for transmission to the next generation. © 2012 Elsevier Inc.
Kannan Mutharasan R.,Feinberg Cardiovascular Research Institute |
Foit L.,Northwestern University |
Shad Thaxton C.,Northwestern University
Journal of Materials Chemistry B | Year: 2015
High-density lipoproteins (HDL) are a class of natural nanostructures found in the blood and are composed of lipids, proteins, and nucleic acids (e.g. microRNA). Their size, which appears to be well-suited for both tissue penetration/retention as well as payload delivery, long circulation half-life, avoidance of endosomal sequestration, and potential low toxicity are all excellent properties to model in a drug delivery vehicle. In this review, we consider high-density lipoproteins for therapeutic delivery systems. First we discuss the structure and function of natural HDL, describing in detail its biogenesis and transformation from immature, discoidal forms, to more mature, spherical forms. Next we consider features of HDL making them suitable vehicles for drug delivery. We then describe the use of natural HDL, discoidal HDL analogs, and spherical HDL analogs to deliver various classes of drugs, including small molecules, lipids, and oligonucleotides. We briefly consider the notion that the drug delivery vehicles themselves are therapeutic, constituting entities that exhibit "theralivery." Finally, we discuss challenges and future directions in the field. © The Royal Society of Chemistry 2016.
PubMed | Northwestern University, University of Wyoming and Feinberg Cardiovascular Research Institute
Type: | Journal: Scientific reports | Year: 2015
Exosomes are nanoscale vesicles that mediate intercellular communication. Cellular exosome uptake mechanisms are not well defined partly due to the lack of specific inhibitors of this complex cellular process. Exosome uptake depends on cholesterol-rich membrane microdomains called lipid rafts, and can be blocked by non-specific depletion of plasma membrane cholesterol. Scavenger receptor type B-1 (SR-B1), found in lipid rafts, is a receptor for cholesterol-rich high-density lipoproteins (HDL). We hypothesized that a synthetic nanoparticle mimic of HDL (HDL NP) that binds SR-B1 and removes cholesterol through this receptor would inhibit cellular exosome uptake. In cell models, our data show that HDL NPs bind SR-B1, activate cholesterol efflux, and attenuate the influx of esterified cholesterol. As a result, HDL NP treatment results in decreased dynamics and clustering of SR-B1 contained in lipid rafts and potently inhibits cellular exosome uptake. Thus, SR-B1 and targeted HDL NPs provide a fundamental advance in studying cholesterol-dependent cellular uptake mechanisms.
PubMed | Northwestern University and Feinberg Cardiovascular Research Institute
Type: Journal Article | Journal: Journal of materials chemistry. B, Materials for biology and medicine | Year: 2016
High-density lipoproteins (HDL) are a class of natural nanostructures found in the blood and are composed of lipids, proteins, and nucleic acids (e.g. microRNA). Their size, which appears to be well-suited for both tissue penetration/retention as well as payload delivery, long circulation half-life, avoidance of endosomal sequestration, and potential low toxicity are all excellent properties to model in a drug delivery vehicle. In this review, we consider high-density lipoproteins for therapeutic delivery systems. First we discuss the structure and function of natural HDL, describing in detail its biogenesis and transformation from immature, discoidal forms, to more mature, spherical forms. Next we consider features of HDL making them suitable vehicles for drug delivery. We then describe the use of natural HDL, discoidal HDL analogs, and spherical HDL analogs to deliver various classes of drugs, including small molecules, lipids, and oligonucleotides. We briefly consider the notion that the drug delivery vehicles themselves are therapeutic, constituting entities that exhibit theralivery. Finally, we discuss challenges and future directions in the field.