title: Implementing Mass Cytometry at the Bedside to Study the Immunological Basis of Human Diseases: Distinctive Immune Features in Patients with a History of Term or Preterm Birth.
abstract: Single-cell technologies have immense potential to shed light on molecular and biological processes that drive human diseases. Mass cytometry (or Cytometry by Time Of Flight mass spectrometry, CyTOF) has already been employed in clinical studies to comprehensively survey patients' circulating immune system. As interest in the "bedside" application of mass cytometry is growing, the delineation of relevant methodological issues is called for. This report uses a newly generated dataset to discuss important methodological considerations when mass cytometry is implemented in a clinical study. Specifically, the use of whole blood samples versus peripheral blood mononuclear cells (PBMCs), design of mass-tagged antibody panels, technical and analytical implications of sample barcoding, and application of traditional and unsupervised approaches to analyze high-dimensional mass cytometry datasets are discussed. A mass cytometry assay was implemented in a cross-sectional study of 19 women with a history of term or preterm birth to determine whether immune traits in peripheral blood differentiate the two groups in the absence of pregnancy. Twenty-seven phenotypic and 11 intracellular markers were simultaneously analyzed in whole blood samples stimulated with lipopolysaccharide (LPS at 0, 0.1, 1, 10, and 100 ng mL(-1)) to examine dose-dependent signaling responses within the toll-like receptor 4 (TLR4) pathway. Complementary analyses, grounded in traditional or unsupervised gating strategies of immune cell subsets, indicated that the prpS6 and pMAPKAPK2 responses in classical monocytes are accentuated in women with a history of preterm birth (FDR<1%). The results suggest that women predisposed to preterm birth may be prone to mount an exacerbated TLR4 response during the course of pregnancy. This important hypothesis-generating finding points to the power of single-cell mass cytometry to detect biologically important differences in a relatively small patient cohort.
authors.affiliation: Department of Anesthesiology, Perioperative, and Pain Medicine, Stanford University, School of Medicine, Stanford, California, 94305.|Baxter Laboratory in Stem Cell Biology, Department of Microbiology and Immunology, Stanford University, Stanford, California, 94305.
authors.forename: Brice
authors.initials: B
authors.lastname: Gaudillière
authors.affiliation: Department of Anesthesiology, Perioperative, and Pain Medicine, Stanford University, School of Medicine, Stanford, California, 94305.
authors.forename: Edward A
authors.initials: EA
authors.lastname: Ganio
authors.affiliation: Department of Anesthesiology, Perioperative, and Pain Medicine, Stanford University, School of Medicine, Stanford, California, 94305.
authors.forename: Martha
authors.initials: M
authors.lastname: Tingle
authors.affiliation: Department of Anesthesiology, Perioperative, and Pain Medicine, Stanford University, School of Medicine, Stanford, California, 94305.
authors.forename: Hope L
authors.initials: HL
authors.lastname: Lancero
authors.affiliation: Baxter Laboratory in Stem Cell Biology, Department of Microbiology and Immunology, Stanford University, Stanford, California, 94305.|Department of Microbiology and Immunology, Stanford University, Stanford, California, 94305.
authors.forename: Gabriela K
authors.initials: GK
authors.lastname: Fragiadakis
authors.affiliation: Department of Anesthesiology, Perioperative, and Pain Medicine, Stanford University, School of Medicine, Stanford, California, 94305.
authors.forename: Quentin J
authors.initials: QJ
authors.lastname: Baca
authors.affiliation: Baxter Laboratory in Stem Cell Biology, Department of Microbiology and Immunology, Stanford University, Stanford, California, 94305.
authors.forename: Nima
authors.initials: N
authors.lastname: Aghaeepour
authors.affiliation: Department of Pediatrics, Stanford University School of Medicine, Stanford, California, 94305.
authors.forename: Ronald J
authors.initials: RJ
authors.lastname: Wong
authors.affiliation: Department of Pediatrics, Stanford University School of Medicine, Stanford, California, 94305.
authors.forename: Cele
authors.initials: C
authors.lastname: Quaintance
authors.affiliation: Department of Obstetrics and Gynecology, Stanford University School of Medicine, Stanford, California, 94305.
authors.forename: Yasser Y
authors.initials: YY
authors.lastname: El-Sayed
authors.affiliation: Department of Pediatrics, Stanford University School of Medicine, Stanford, California, 94305.
authors.forename: Gary M
authors.initials: GM
authors.lastname: Shaw
authors.affiliation: Department of Pediatrics, Stanford University School of Medicine, Stanford, California, 94305.
authors.forename: David B
authors.initials: DB
authors.lastname: Lewis
authors.affiliation: Department of Pediatrics, Stanford University School of Medicine, Stanford, California, 94305.
authors.forename: David K
authors.initials: DK
authors.lastname: Stevenson
authors.affiliation: Baxter Laboratory in Stem Cell Biology, Department of Microbiology and Immunology, Stanford University, Stanford, California, 94305.|Department of Microbiology and Immunology, Stanford University, Stanford, California, 94305.
authors.forename: Garry P
authors.initials: GP
authors.lastname: Nolan
authors.affiliation: Department of Anesthesiology, Perioperative, and Pain Medicine, Stanford University, School of Medicine, Stanford, California, 94305.
authors.forename: Martin S
authors.initials: MS
authors.lastname: Angst
mesh_terms: D000328:Adult; D003430:Cross-Sectional Studies; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D007231:Infant, Newborn; D007963:Leukocytes, Mononuclear / Q000276:immunology*; D000067716:Point-of-Care Testing; D011247:Pregnancy; D047928:Premature Birth / Q000175:diagnosis* / Q000276:immunology*; D047929:Term Birth / Q000276:immunology*
keywords: CyTOF; TLR4; clinical applications; innate immunity; mass cytometry; monocytes; preterm birth; single cell