title: Single-Cell Detection of Secreted Aβ and sAPPα from Human IPSC-Derived Neurons and Astrocytes.
abstract: Secreted factors play a central role in normal and pathological processes in every tissue in the body. The brain is composed of a highly complex milieu of different cell types and few methods exist that can identify which individual cells in a complex mixture are secreting specific analytes. By identifying which cells are responsible, we can better understand neural physiology and pathophysiology, more readily identify the underlying pathways responsible for analyte production, and ultimately use this information to guide the development of novel therapeutic strategies that target the cell types of relevance. We present here a method for detecting analytes secreted from single human induced pluripotent stem cell (iPSC)-derived neural cells and have applied the method to measure amyloid β (Aβ) and soluble amyloid precursor protein-alpha (sAPPα), analytes central to Alzheimer's disease pathogenesis. Through these studies, we have uncovered the dynamic range of secretion profiles of these analytes from single iPSC-derived neuronal and glial cells and have molecularly characterized subpopulations of these cells through immunostaining and gene expression analyses. In examining Aβ and sAPPα secretion from single cells, we were able to identify previously unappreciated complexities in the biology of APP cleavage that could not otherwise have been found by studying averaged responses over pools of cells. This technique can be readily adapted to the detection of other analytes secreted by neural cells, which would have the potential to open new perspectives into human CNS development and dysfunction.
  
  
  CONCLUSIONS
  We have established a technology that, for the first time, detects secreted analytes from single human neurons and astrocytes. We examine secretion of the Alzheimer's disease-relevant factors amyloid β (Aβ) and soluble amyloid precursor protein-alpha (sAPPα) and present novel findings that could not have been observed without a single-cell analytical platform. First, we identify a previously unappreciated subpopulation that secretes high levels of Aβ in the absence of detectable sAPPα. Further, we show that multiple cell types secrete high levels of Aβ and sAPPα, but cells expressing GABAergic neuronal markers are overrepresented. Finally, we show that astrocytes are competent to secrete high levels of Aβ and therefore may be a significant contributor to Aβ accumulation in the brain.
authors: Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, and.
authors: Mei-Chen
authors: MC
authors: Liao
authors: Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, and.
authors: Christina R
authors: CR
authors: Muratore
authors: Koch Institute for Integrative Cancer Research and.
authors: Todd M
authors: TM
authors: Gierahn
authors: Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, and.
authors: Sarah E
authors: SE
authors: Sullivan
authors: Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, and.
authors: Priya
authors: P
authors: Srikanth
authors: Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, and.
authors: Philip L
authors: PL
authors: De Jager
authors: Koch Institute for Integrative Cancer Research and Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139.
authors: J Christopher
authors: JC
authors: Love
authors: Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, and tyoung@rics.bwh.harvard.edu.
authors: Tracy L
authors: http://orcid.org/0000-0001-7846-1542
authors: TL
authors: Young-Pearse
mesh_terms: D016229:Amyloid beta-Peptides / Q000032:analysis / Q000378:metabolism; D016564:Amyloid beta-Protein Precursor / Q000032:analysis / Q000378:metabolism*; D000818:Animals; D001253:Astrocytes / Q000737:chemistry / Q000378:metabolism*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000737:chemistry / Q000378:metabolism*; D008297:Male; D009474:Neurons / Q000737:chemistry / Q000378:metabolism*; D059010:Single-Cell Analysis / Q000379:methods*
chemical_list: C545487:APP protein, human; D016229:Amyloid beta-Peptides; D016564:Amyloid beta-Protein Precursor
keywords: APP; Alzheimer's disease; Aβ; iPSC; microengraving; single cell