Hong Jeong Hee, Lee Syng-Ill, Kim Kyu-Earn, Yong Tai-Soon, Seo Jeong Taeg, Sohn Myung Hyun, Shin Dong Min
Department of Oral Biology, Oral Science Research Center, Brain Korea 21 Project for Medical Sciences, Yonsei University College of Dentistry, Seoul, Korea.
J Allergy Clin Immunol. 2004 Feb;113(2):315-9. doi: 10.1016/j.jaci.2003.11.026.
The German cockroach has been reported to act as an allergen that might be associated with a protease reaction in asthma. However, the molecular identities of the antigens in German cockroach extract (GCE) with protease activity and the protease-activated receptors (PARs) that are activated by GCE in human airway epithelial cells have not been characterized.
We investigated the direct effect of GCE on Ca(2+) signaling in human airway epithelial cells and the type of PARs activated by GCE.
The Ca(2+)-sensitive dye Fura2 was used to determine intracellular Ca(2+) concentration (Ca(2+)) by means of spectrofluorometry.
GCE induced a baseline type of Ca(2+) oscillations in a dose-dependent manner. The oscillations persisted for long periods of time in the absence of Ca(2+) entry across the plasma membrane, suggesting that the observed Ca(2+) increases were due to Ca(2+) release from intracellular stores. Accordingly, after depleting endoplasmic reticulum Ca(2+) with thapsigargin, an endoplasmic reticulum Ca(2+) ATPase inhibitor, the GCE-mediated Ca(2+) signals were abolished. Whereas desensitization of PAR-1, PAR-3, and PAR-4 had no effect on GCE-mediated Ca(2+) mobilization, no GCE-mediated Ca(2+) increase was observed after desensitization of PAR-2.
These results indicate that GCE has a direct effect on human airway epithelial cells, in particular generating Ca(2+) oscillations through Ca(2+) release from thapsigargin-sensitive Ca(2+) stores through activation of PAR-2.
据报道，德国小蠊可作为一种变应原，可能与哮喘中的蛋白酶反应有关。然而，德国小蠊提取物（GCE）中具有蛋白酶活性的抗原以及在人气道上皮细胞中被GCE激活的蛋白酶激活受体（PARs）的分子特性尚未得到表征。
我们研究了GCE对人气道上皮细胞中Ca(2+)信号的直接影响以及被GCE激活的PARs类型。
使用Ca(2+)敏感染料Fura2通过荧光分光光度法测定细胞内Ca(2+)浓度（[Ca(2+)]i）。
GCE以剂量依赖性方式诱导[Ca(2+)]i出现基线型振荡。在没有Ca(2+)跨质膜进入的情况下，振荡持续很长时间，这表明观察到的[Ca(2+)]i增加是由于细胞内储存的Ca(2+)释放所致。因此，在用内质网Ca(2+)ATP酶抑制剂毒胡萝卜素耗尽内质网Ca(2+)后，GCE介导的[Ca(2+)]i信号被消除。虽然PAR-1、PAR-3和PAR-4的脱敏对GCE介导的Ca(2+)动员没有影响，但在PAR-2脱敏后未观察到GCE介导的[Ca(2+)]i增加。
这些结果表明，GCE对人气道上皮细胞有直接影响，特别是通过激活PAR-2从毒胡萝卜素敏感的Ca(2+)储存中释放Ca(2+)来产生[Ca(2+)]i振荡。
