A modified fluorescent sensor for reporting glucose concentration in the airway lumen.
A modified fluorescent sensor for reporting glucose concentration in the airway lumen.
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DOI：
10.1371/journal.pone.0254248
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发表时间：
2021
期刊：
影响因子：
3.7
通讯作者：
Baines DL
中科院分区：
文献类型：
--
作者：
Bearham J;Krutrök N;Lindberg B;Woodall M;Astrand A;Taylor JD;Biggart M;Vasiljevs S;Tarran R;Baines DL
我们已经修改了大肠杆菌的周质葡萄糖/半乳糖结合蛋白（GBP）和环境敏感的荧光标记，以进一步探索其作为传感器的潜力，用于评估气道表面液体（ASL）中的葡萄糖浓度。我们鉴定了用N，N '-二甲基-N-（碘乙酰基）-N'-（7-硝基苯-2-氧杂-1，3-二唑-4-基）乙二胺（IANBD，最大发射波长536 nm）标记的E149 C/A213 R GBP，其对D-葡萄糖的Kd为1.02 mM，荧光动态范围为5.8。该传感器对D-葡萄糖具有特异性，并在实验中表现出数小时的荧光稳定性。E149 C/A213 R GBP-IANBD在气-液界面（ALI）处生长的气道细胞的ASL中的使用检测到在将基底侧葡萄糖从5 mM升高至15 mM后10分钟葡萄糖浓度的增加。与人支气管上皮细胞（HBEC）相比，该传感器还报告了响应于H441气道细胞中基底外侧葡萄糖增加的ASL葡萄糖浓度的更大变化，并且HBEC数据的变异性小于H441，表明HBEC更有效地调节葡萄糖进入ASL。传感器检测到糖尿病db/db小鼠的支气管肺泡灌洗液（BALf）中的葡萄糖，但未检测到正常血糖野生型小鼠，表明传感器在葡萄糖浓度<50μM时的灵敏度有限。使用鼻吸入传感器和光谱解混来生成图像，在小鼠远端肺的冷冻切片的管腔区域中检测到E149 C/A213 R GBP-IANBD荧光，其以db/db计大于野生型小鼠。总之，该传感器为进一步开发提供了有用的工具，以测量肺/气道模型中的管腔葡萄糖浓度，以探索其在疾病中如何变化。
We have modified the periplasmic Escherichia coli glucose/galactose binding protein (GBP) and labelled with environmentally sensitive fluorophores to further explore its potential as a sensor for the evaluation of glucose concentration in airway surface liquid (ASL). We identified E149C/A213R GBP labelled with N,N’-Dimethyl-N-(iodoacetyl)-N’-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)ethylenediamine (IANBD, emission wavelength maximum 536nm) with a Kd for D-glucose of 1.02mM and a fluorescence dynamic range of 5.8. This sensor was specific for D-glucose and exhibited fluorescence stability in experiments for several hours. The use of E149C/A213R GBP-IANBD in the ASL of airway cells grown at air-liquid-interface (ALI) detected an increase in glucose concentration 10 minutes after raising basolateral glucose from 5 to 15mM. This sensor also reported a greater change in ASL glucose concentration in response to increased basolateral glucose in H441 airway cells compared to human bronchial epithelial cells (HBEC) and there was less variability with HBEC data than that of H441 indicating that HBEC more effectively regulate glucose movement into the ASL. The sensor detected glucose in bronchoalveolar lavage fluid (BALf) from diabetic db/db mice but not normoglycaemic wildtype mice, indicating limited sensitivity of the sensor at glucose concentrations <50μM. Using nasal inhalation of the sensor and spectral unmixing to generate images, E149C/A213R GBP-IANBD fluorescence was detected in luminal regions of cryosections of the murine distal lung that was greater in db/db than wildtype mice. In conclusion, this sensor provides a useful tool for further development to measure luminal glucose concentration in models of lung/airway to explore how this may change in disease.
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跨囊性纤维化气道上皮单层的细胞旁葡萄糖通量升高是铜绿假单胞菌生长的重要因素。
影响因子：
3.7
作者：
Garnett JP;Gray MA;Tarran R;Brodlie M;Ward C;Baker EH;Baines DL
通讯作者：
Baines DL
影响因子：
4.6
作者：
Garnett JP;Kalsi KK;Sobotta M;Bearham J;Carr G;Powell J;Brodlie M;Ward C;Tarran R;Baines DL
通讯作者：
Baines DL
影响因子：
3.7
作者：
Pezzulo AA;Gutiérrez J;Duschner KS;McConnell KS;Taft PJ;Ernst SE;Yahr TL;Rahmouni K;Klesney-Tait J;Stoltz DA;Zabner J
通讯作者：
Zabner J
DOI：
10.1016/j.bbrc.2013.07.111
发表时间：
2013-08-30
影响因子：
3.1
作者：
Khan, Faaizah;Pickup, John C.
通讯作者：
Pickup, John C.
影响因子：
5
作者：
Hermanns, MI;Unger, RE;Kirkpatrick, CJ
通讯作者：
Kirkpatrick, CJ