title: In vivo tracking of Th1 cells by PET reveals quantitative and temporal distribution and specific homing in lymphatic tissue.
abstract: Although T cells can be labeled for noninvasive in vivo imaging, little is known about the impact of such labeling on T-cell function, and most imaging methods do not provide holistic information about trafficking kinetics, homing sites, or quantification.
  
  
  METHODS
  We developed protocols that minimize the inhibitory effects of (64)Cu-pyruvaldehyde-bis(N4-methylthiosemicarbazone) ((64)Cu-PTSM) labeling on T-cell function and permit the homing patterns of T cells to be followed by PET. Thus, we labeled ovalbumin (OVA) T-cell receptor transgenic interferon (IFN)-γ-producing CD4(+) T (Th1) cells with 0.7-2.2 MBq of (64)Cu-PTSM and analyzed cell viability, IFN-γ production, proliferation, apoptosis, and DNA double-strand breaks and identified intracellular (64)Cu accumulation sites by energy dispersive x-ray analysis. To elucidate the fate of Th1 cell homing by PET, 10(7 64)Cu-OVA-Th1 cells were injected intraperitoneally or intravenously into healthy mice. To test the functional capacities of (64)Cu-OVA-Th1 cells during experimental OVA-induced airway hyperreactivity, we injected 10(7 64)Cu-OVA-Th1 cells intraperitoneally into OVA-immunized or nonimmunized healthy mice, which were challenged with OVA peptide or phosphate-buffered saline or remained untreated. In vivo PET investigations were followed by biodistribution, autoradiography, and fluorescence-activated cell sorting analysis.
  
  
  RESULTS
  PET revealed unexpected homing patterns depending on the mode of T-cell administration. Within 20 min after intraperitoneal administration, (64)Cu-OVA-Th1 cells homed to the perithymic lymph nodes (LNs) of naive mice. Interestingly, intravenously administered (64)Cu-OVA-Th1 cells homed predominantly into the lung and spleen but not into the perithymic LNs. The accumulation of (64)Cu-OVA-Th1 cells in the pulmonary LNs (6.8 ± 1.1 percentage injected dose per cubic centimeter [%ID/cm(3)]) 24 h after injection was highest in the OVA-immunized and OVA-challenged OVA airway hyperreactivity-diseased littermates 24 h after intraperitoneal administration and lowest in the untreated littermates (3.7 ± 0.4 %ID/cm(3)). As expected, (64)Cu-OVA-Th1 cells also accumulated significantly in the pulmonary LNs of nonimmunized OVA-challenged animals (6.1 ± 0.5 %ID/cm(3)) when compared with phosphate-buffered saline-challenged animals (4.6 ± 0.5 %ID/cm(3)).
  
  
  CONCLUSIONS
  Our protocol permits the detection of Th1 cells in single LNs and enables temporal in vivo monitoring of T-cell homing over 48 h. This work enables future applications for (64)Cu-PTSM-labeled T cells in clinical trials and novel therapy concepts focusing on T-cell-based immunotherapies of autoimmune diseases or cancer.
authors.affiliation: Department of Preclinical Imaging and Radiopharmacy, Werner Siemens Imaging Center, Eberhard Karls University, Tübingen, Germany.
authors.forename: Christoph M
authors.initials: CM
authors.lastname: Griessinger
authors.forename: Rainer
authors.initials: R
authors.lastname: Kehlbach
authors.forename: Daniel
authors.initials: D
authors.lastname: Bukala
authors.forename: Stefan
authors.initials: S
authors.lastname: Wiehr
authors.forename: Rüdiger
authors.initials: R
authors.lastname: Bantleon
authors.forename: Funda
authors.initials: F
authors.lastname: Cay
authors.forename: Andreas
authors.initials: A
authors.lastname: Schmid
authors.forename: Heidi
authors.initials: H
authors.lastname: Braumüller
authors.forename: Birgit
authors.initials: B
authors.lastname: Fehrenbacher
authors.forename: Martin
authors.initials: M
authors.lastname: Schaller
authors.forename: Martin
authors.initials: M
authors.lastname: Eichner
authors.forename: Julie L
authors.initials: JL
authors.lastname: Sutcliffe
authors.forename: Walter
authors.initials: W
authors.lastname: Ehrlichmann
authors.forename: Oliver
authors.initials: O
authors.lastname: Eibl
authors.forename: Gerald
authors.initials: G
authors.lastname: Reischl
authors.forename: Simon R
authors.initials: SR
authors.lastname: Cherry
authors.forename: Martin
authors.initials: M
authors.lastname: Röcken
authors.forename: Bernd J
authors.initials: BJ
authors.lastname: Pichler
authors.forename: Manfred
authors.initials: M
authors.lastname: Kneilling
mesh_terms: D000818:Animals; D017209:Apoptosis; D015551:Autoimmunity; D002465:Cell Movement; D049109:Cell Proliferation; D002469:Cell Separation; D002470:Cell Survival; D003301:Copper Radioisotopes; D053903:DNA Breaks, Double-Stranded; D005434:Flow Cytometry; D007167:Immunotherapy / Q000379:methods; D007371:Interferon-gamma / Q000378:metabolism; D008198:Lymph Nodes / Q000473:pathology; D008221:Lymphoid Tissue / Q000000981:diagnostic imaging* / Q000187:drug effects*; D051379:Mice; D009942:Organometallic Compounds; D010455:Peptides / Q000737:chemistry; D049268:Positron-Emission Tomography / Q000379:methods*; D018417:Th1 Cells / Q000166:cytology*; D013882:Thiosemicarbazones; D013997:Time Factors; D014018:Tissue Distribution
chemical_list: D003301:Copper Radioisotopes; D009942:Organometallic Compounds; D010455:Peptides; D013882:Thiosemicarbazones; C052629:copper pyruvaldehyde bis(N(4)-methylthiosemicarbazone) complex; D007371:Interferon-gamma
keywords: 64Cu-PTSM; in vivo cell tracking; murine Th1 cells; small animal PET