title: Intracellular pH imaging in cancer cells in vitro and tumors in vivo using the new genetically encoded sensor SypHer2.
abstract: BACKGROUND
  Measuring intracellular pH (pHi) in tumors is essential for the monitoring of cancer progression and the response of cancer cells to various treatments. The purpose of the study was to develop a method for pHi mapping in living cancer cells in vitro and in tumors in vivo, using the novel genetically encoded indicator, SypHer2.
  
  
  METHODS
  A HeLa Kyoto cell line stably expressing SypHer2 in the cytoplasm was used, to perform ratiometric (dual excitation) imaging of the probe in cell culture, in 3D tumor spheroids and in tumor xenografts in living mice.
  
  
  RESULTS
  Using SypHer2, pHi was demonstrated to be 7.34±0.11 in monolayer HeLa cells in vitro under standard cultivation conditions. An increasing pHi gradient from the center to the periphery of the spheroids was displayed. We obtained fluorescence ratio maps for HeLa tumors in vivo and ex vivo. Comparison of the map with the pathomorphology and with hypoxia staining of the tumors revealed a correspondence of the zones with higher pHi to the necrotic and hypoxic areas.
  
  
  CONCLUSIONS
  Our results demonstrate that pHi imaging with the genetically encoded pHi indicator, SypHer2, can be a valuable tool for evaluating tumor progression in xenograft models.
  
  
  CONCLUSIONS
  We have demonstrated, for the first time, the possibility of using the genetically encoded sensor SypHer2 for ratiometric pH imaging in cancer cells in vitro and in tumors in vivo. SypHer2 shows great promise as an instrument for pHi monitoring able to provide high accuracy and spatiotemporal resolution.
authors.affiliation: Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia; Lobachevsky State University of Nizhny Novgorod, 23 Gagarin Ave., 603950 Nizhny Novgorod, Russia. Electronic address: Shirmanovam@mail.ru.
authors.forename: Marina V
authors.initials: MV
authors.lastname: Shirmanova
authors.affiliation: Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia.
authors.forename: Irina N
authors.initials: IN
authors.lastname: Druzhkova
authors.affiliation: Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia; Lobachevsky State University of Nizhny Novgorod, 23 Gagarin Ave., 603950 Nizhny Novgorod, Russia.
authors.forename: Maria M
authors.initials: MM
authors.lastname: Lukina
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, 16/10 Miklukho-Maklaya St., 117997 Moscow, Russia.
authors.forename: Mikhail E
authors.initials: ME
authors.lastname: Matlashov
authors.affiliation: Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia; Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, 16/10 Miklukho-Maklaya St., 117997 Moscow, Russia.
authors.forename: Vsevolod V
authors.initials: VV
authors.lastname: Belousov
authors.affiliation: Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia.
authors.forename: Ludmila B
authors.initials: LB
authors.lastname: Snopova
authors.affiliation: Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia.
authors.forename: Natalia N
authors.initials: NN
authors.lastname: Prodanetz
authors.affiliation: Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia; Lobachevsky State University of Nizhny Novgorod, 23 Gagarin Ave., 603950 Nizhny Novgorod, Russia.
authors.forename: Varvara V
authors.initials: VV
authors.lastname: Dudenkova
authors.affiliation: Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia; Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry RAS, 16/10 Miklukho-Maklaya St., 117997 Moscow, Russia.
authors.forename: Sergey A
authors.initials: SA
authors.lastname: Lukyanov
authors.affiliation: Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia; Lobachevsky State University of Nizhny Novgorod, 23 Gagarin Ave., 603950 Nizhny Novgorod, Russia.
authors.forename: Elena V
authors.initials: EV
authors.lastname: Zagaynova
mesh_terms: D000818:Animals; D015374:Biosensing Techniques; D015687:Cell Hypoxia; D005818:Genetic Engineering; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D051379:Mice; D009369:Neoplasms / Q000378:metabolism* / Q000473:pathology; D018874:Spheroids, Cellular
keywords: Cancer cell; Genetically encoded sensor; Intracellular pH; Ratiometric imaging; SypHer2; Tumor