Effect of chloroquine on the formation of tight junctions in cultured immature rat Sertoli cells.
Effect of chloroquine on the formation of tight junctions in cultured immature rat Sertoli cells.
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DOI：
10.1002/j.1939-4640.1996.tb01780.x
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发表时间：
1996-05
影响因子：
--
通讯作者：
Abayomi Okanlawon;M. Dym
中科院分区：
文献类型：
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作者：
Abayomi Okanlawon;M. Dym
生精上皮中相邻的未成熟支持细胞形成紧密的连接复合体，导致血睾屏障的发育。蛋白酶和抗蛋白酶活性与紧密连接的形成过程有关。在这里，我们报告了氯喹（一种具有抗蛋白酶活性的抗疟药物）对培养的未成熟大鼠支持细胞中细胞间紧密连接发育的影响。对于阳性对照，使用经典的溶酶体剂氯化铵。将支持细胞以 3 x 10(6) 细胞/0.64-cm2 孔的密度接种在 Matrigel 覆盖的 Millicell-HA 过滤器上的无血清限定培养基中。在培养的第 1 天或第 7 天，将浓度范围为 25 至 100 µM 的氯喹添加到两室系统的外室中。通过使用阻抗计每隔 24 小时测量跨上皮电阻 (TER) 来监测紧密连接的形成。未经处理的对照组的 TER 在第 1 天为 50 欧姆/平方厘米；到第 7 天，它逐渐增加到 80 ohms/cm2，并稳定到第 12 天。从第 1 天开始用氯喹处理的细胞也显示出 TER 的剂量依赖性逐渐增加，直到第 9 天，在用 100 µM 浓度处理的细胞中达到 225 ohms/cm2。与对照组相比，TER 的增加明显更高。在从培养第 7 天开始用氯喹处理的细胞中，与未处理的对照相比，TER 没有观察到差异。这些观察结果表明，氯喹和氯化铵会增加两室室中未成熟支持细胞的 TER。
Adjoining immature Sertoli cells in the seminiferous epithelium form a tight junctional complex leading to the development of the blood-testis barrier. Protease and antiprotease activities have been implicated in the process of formation of tight junctions. Here, we report the effect of chloroquine, an antimalarial drug with antiprotease activity, on the development of intercellular tight junctions in cultured immature rat Sertoli cells. For positive control, the classical lysosomotropic agent ammonium chloride was used. Sertoli cells were seeded in serum-free defined medium at a density of 3 x 10(6) cells/0.64-cm2 well on Matrigel-covered Millicell-HA filters. Chloroquine at concentrations ranging from 25 to 100 microM was added to the outer chamber of the bicameral system on either day 1 or 7 of the culture. The formation of the tight junction was monitored by the measurement of the transepithelial resistance (TER) at 24-hour intervals using an impedance meter. TER in untreated controls was 50 ohms/cm2 on day 1; it increased progressively to 80 ohms/cm2 by day 7 and plateaued until day 12. The cells treated from day 1 with chloroquine also showed a dose-dependent progressive increase in TER until day 9, reaching 225 ohms/cm2 in cells treated with the 100 microM concentration. In comparison to controls, the increase in TER was significantly higher. In cells treated with chloroquine starting from day 7 of culture onwards, there was no observable difference in TER from the untreated control. These observations demonstrate that chloroquine and ammonium chloride increase the TER of immature Sertoli cells in the bicameral chamber.