Probe for the measurement of cell surface pH in vivo and ex vivo
Probe for the measurement of cell surface pH in vivo and ex vivo
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DOI：
10.1073/pnas.1608247113
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发表时间：
2016-07-19
影响因子：
11.1
通讯作者：
Andreev, Oleg A.
中科院分区：
文献类型：
--
作者：
Anderson, Michael;Moshnikova, Anna;Andreev, Oleg A.
我们已经开发了一种方法来测量细胞表面的pH值，通过定位的pH敏感的荧光染料，sNARF或（SNARF），共轭的pH值低插入肽（pHLIP）。已经观察到许多病变组织是酸性的，肿瘤尤其如此。这些作用的组合使肿瘤细胞内部酸化，细胞泵出乳酸和质子以维持细胞内pH值，使细胞外空间酸化。碳酸酐酶在细胞表面的过表达进一步促进酸化。因此，预期肿瘤细胞表面附近的pH值较低，并且随着与膜的距离而增加，因此本体pH值测量将不会报告表面酸度。我们的新表面pH测量工具在球状体中生长的癌细胞、体内小鼠肿瘤模型和切除的肿瘤中得到了验证。我们发现，表面pH值是敏感的细胞糖酵解活性：pH值降低，在高葡萄糖和增加，如果葡萄糖被替换为非代谢脱氧葡萄糖。对于高转移性癌细胞，当周围外部pH为7.4时，在表面处测量的pH为6.7-6.8。该方法足够灵敏，可以在体内检测由腹膜内注射葡萄糖诱导的肿瘤中的0.2-0.3 pH单位变化。发现肿瘤内高转移性细胞表面的pH值约为6.1-6.4，而在非转移性肿瘤中，pH值为6.7-6.9，这可能创造了一种区分更具侵袭性和侵袭性较低的肿瘤的方法。既然有针对性的测量是可能的，那么表面酸度的其他生物作用也可能被发现。
We have developed a way to measure cell surface pH by positioning a pH-sensitive fluorescent dye, seminaphtharhodafluor (SNARF), conjugated to the pH low insertion peptide (pHLIP). It has been observed that many diseased tissues are acidic and that tumors are especially so. A combination of effects acidifies tumor cell interiors, and cells pump out lactic acid and protons to maintain intracellular pH, acidifying the extracellular space. Overexpression of carbonic anhydrases on cell surfaces further contributes to acidification. Thus, the pH near tumor cell surfaces is expected to be low and to increase with distance from the membrane, so bulk pH measurements will not report surface acidity. Our new surface pH-measurement tool was validated in cancer cells grown in spheroids, in mouse tumor models in vivo, and in excised tumors. We found that the surface pH is sensitive to cell glycolytic activity: the pH decreases in high glucose and increases if glucose is replaced with nonmetabolized deoxyglucose. For highly metastatic cancer cells, the pH measured at the surface was 6.7-6.8, when the surrounding external pH was 7.4. The approach is sensitive enough to detect 0.2-0.3 pH unit changes in vivo in tumors induced by i.p. injection of glucose. The pH at the surfaces of highly metastatic cells within tumors was found to be about 6.1-6.4, whereas in nonmetastatic tumors, it was 6.7-6.9, possibly creating a way to distinguish more aggressive from less aggressive tumors. Other biological roles of surface acidity may be found, now that targeted measurements are possible.