非损伤微测技术测量——在上皮细胞和内皮细胞研究上的应用

随着“非损伤微测技术”的发展，以其操作简单、实用已经在很多领域发挥作用，在创面周围组织、单层上皮细胞、内皮细胞的研究上同样有很好的应用。

1.研究老鼠雄性生殖系统输精管上皮细胞。

输精管直径约为100 μ m ，用离子选择性电极扫描输精管表面，在人为刺激、化学刺激处理下检测氢离子变化，用以研究上皮组织结构物质合成的生化途径及运输。

2．应用“非损伤微测技术”可以研究上皮细胞表面单个细胞的活动。应用这种试验方法已经对肺上皮细胞参与水分代谢的氯离子转运进行了研究[2]。

3．应用“非损伤微测技术”检测内皮组织,如检测出小毛细血管的温度依赖性氯离子和肌浆蛋白收缩的相关性[3]。

举例：用氢离子电极检测不同处理对老鼠输精管上皮细胞泌氢的影响（Journal of Biological Chemistry）

FIG. 1. Gelsolin localization in the epididymis. A, double immunofluorescence staining showing gelsolin (red) and V-ATPase (yellow) in rat cauda epididymidis. Strong gelsolin staining was observed in clear cells, identified by their positive immunoreactivity for V-ATPase. Adjacent principal cells were negative for gelsolin. B, higher magnification showing gelsolin (green) and actin (red). Nuclei were stained blue by 4_,6-diamidino-2-phenylindole. Although very strong actin staining was detected in the stereocilia of principal cells, no detectable actin was observed in the apical microvilli of clear cells. In these cells, gelsolin was detected throughout the cytoplasm as well as in the apical microvilli. C, detection of gelsolin by Western blotting in total homogenates from rat epididymis. 20 _g of protein were loaded onto the gel. A strong band at _90  kDa was detected, indicating high levels of gelsolin in the epididymis (arrow). Bars _ 25 _m (A) and 10 _m (B).

FIG. 2. Effect of jasplakinolide on proton secretion. A, representative traces showing net proton secretion in cut-open proximal vas deferens under control conditions (upper trace) and upon jasplakinolide addition (lower trace). Apical proton flux was detected using an extracellular self-referencing proton-selective electrode. Upper trace, at time 0, addition of the vehicle (methanol; arrow) induced a rapid increase in the signal due to transient disturbance of the proton gradient. The signal then rapidly recovered to the control value as the proton gradient was re-established and remained stable for periods of up to 1 h. A marked inhibition of net proton secretion was observed when bafilomycin A1 or concanamycin A was added at the end of the experimental period, indicating the participation of V-ATPase in this process. Concanamycin A had no further effect after inhibition by bafilomycin. Lower trace, jasplakinolide induced a progressive inhibition of proton secretion to a level that was not further inhibited by bafilomycin A1 or concanamycin A. B, histogram showing the mean effect of jasplakinolide (Jas) on bafilomycin-sensitive proton secretion (mean _ S.E., n _ 7) measured 40 min after addition of jasplakinolide. The control value was measured in the same vas deferens prior to addition of jasplakinolide. Both control and jasplakinolide values were corrected for the residual value measured after bafilomycin A1 addition. *, p _ 0.001.

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