Shirmanova Marina V, Druzhkova Irina N, Lukina Maria M, Matlashov Mikhail E, Belousov Vsevolod V, Snopova Ludmila B, Prodanetz Natalia N, Dudenkova Varvara V, Lukyanov Sergey A, Zagaynova Elena V
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.
Nizhny Novgorod State Medical Academy, 10/1 Minin Pozharsky Sq., 603005 Nizhny Novgorod, Russia.
Biochim Biophys Acta. 2015 Sep;1850(9):1905-11. doi: 10.1016/j.bbagen.2015.05.001. Epub 2015 May 8.
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.
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.
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.
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.
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.
测量肿瘤细胞内pH值（pHi）对于监测癌症进展以及癌细胞对各种治疗的反应至关重要。本研究的目的是开发一种方法，利用新型基因编码指示剂SypHer2，在体外活癌细胞和体内肿瘤中进行pHi测绘。
使用在细胞质中稳定表达SypHer2的HeLa Kyoto细胞系，对细胞培养物、3D肿瘤球体和活体小鼠体内的肿瘤异种移植中的探针进行比率（双激发）成像。
使用SypHer2，在标准培养条件下，体外单层HeLa细胞中的pHi被证明为7.34±0.11。显示出从球体中心到周边pHi梯度增加。我们获得了体内和体外HeLa肿瘤的荧光比率图。将该图与肿瘤的病理形态学和缺氧染色进行比较，发现pHi较高的区域与坏死和缺氧区域相对应。
我们的结果表明，使用基因编码的pHi指示剂SypHer2进行pHi成像可成为评估异种移植模型中肿瘤进展的有价值工具。
我们首次证明了使用基因编码传感器SypHer2在体外癌细胞和体内肿瘤中进行比率pH成像的可能性。SypHer2作为一种能够提供高精度和时空分辨率的pHi监测仪器显示出巨大的前景。
