Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Rd., Qingdao, Shandong 266071, China.
Fish Shellfish Immunol. 2013 Mar;34(3):855-64. doi: 10.1016/j.fsi.2012.12.023. Epub 2013 Jan 11.
In the present study, the expression of some immune-related genes was examined as indicator to understand the development of immune defense system during the ontogenesis of scallop Chlamys farreri. The mRNA transcripts of pattern-recognition receptors (PRRs) including CfPGRP-S1, CfLGBP, CfLec-1 and CfLec-3 were observed at a low level or even undetected at early developmental stages from eggs to blastula, and then began to increase overwhelmingly in trochophore. For the genes of immune effector including CfLYZ, CfLBP/BPI, CfSOD and CfCAT, their mRNA transcripts were higher expressed in embryos, and increased significantly in D-hinged or early veliger larvae. The whole-mount immunofluorescence assay revealed two immunoreactive spots of CfPGRP-S1 were first observed in the mid-ventral region of prototroch in trochophore, and the immunopositive fluorescence of CfLGBP, CfLec-1 and CfLec-3 appeared at the same spots in early D-hinged larvae. Most of the PRRs were located in velum, mouth, esophagus and stomach region in early and mid-veliger larvae, and especially the strong immunopositive fluorescence of CfLec-3 was observed in velum. The immunoreactive areas of CfLYZ, CfLBP/BPI, CfSOD and CfCAT were observed in trochophore and early D-hinged larvae. After D-hinged larvae, they distributed in different tissues from the edge of velum, mouth, esophagus to the region around digestive gland. After bacterial challenge, the mRNA expression of CfLGBP, CfLec-1 and CfLec-3 did not change significantly in trochophore, while a down-regulation of CfPGRP-S1 was observed at 6 h (P < 0.05). The expression of CfPGRP-S1 and CfLGBP decreased or increased inversely in D-hinged and late veliger larvae respectively, whereas CfLec-1 and CfLec-3 increased significantly during 6-24 h after bacterial challenge in the two stages (P < 0.05). In contrast, the expressions of immune effectors in trochophore and late veliger larvae were significant up-regulated at 6 h, 12 h or 24 h after bacterial challenge (P < 0.05). However, in late D-hinged larvae, CfLYZ and CfSOD expressions were significantly down-regulated at 6 h, while CfLBP/BPI expression was up-regulated at 6 h and 24 h post challenge (P < 0.05). These results indicated that the immune defense system of scallop might appear firstly in the mid-ventral region of prototroch in trochophore, and developed maturely after late D-hinged larvae. The developing immune system in the D-hinged and late veliger larvae could respond to the immune stimulation in different manner.
在本研究中，我们检测了一些免疫相关基因的表达情况，以了解海湾扇贝（Chlamys farreri）个体发育过程中免疫系统的发育情况。在从卵到囊胚的早期发育阶段，模式识别受体（PRRs）的 mRNA 转录物（包括 CfPGRP-S1、CfLGBP、CfLec-1 和 CfLec-3）的表达水平较低，甚至无法检测到，然后在担轮幼虫期急剧增加。对于免疫效应因子（包括 CfLYZ、CfLBP/BPI、CfSOD 和 CfCAT）的基因，它们的 mRNA 转录物在胚胎中表达较高，并在 D 型铰合或早期面盘幼虫中显著增加。整体免疫荧光分析显示，CfPGRP-S1 的两个免疫反应性斑点首先在担轮幼虫的中腹区观察到，而 CfLGBP、CfLec-1 和 CfLec-3 的免疫阳性荧光在早期 D 型铰合幼虫的同一部位出现。大多数 PRRs 位于早期和中期面盘幼虫的翼瓣、口、食管和胃区，尤其是 CfLec-3 的强烈免疫阳性荧光出现在翼瓣中。CfLYZ、CfLBP/BPI、CfSOD 和 CfCAT 的免疫反应性区域在担轮幼虫和早期 D 型铰合幼虫中均有观察到。D 型铰合幼虫后，它们在翼瓣边缘、口、食管到消化腺周围的不同组织中分布。在细菌攻毒后，CfLGBP、CfLec-1 和 CfLec-3 在担轮幼虫中的表达没有显著变化，而 CfPGRP-S1 的表达在 6 h 时下调（P<0.05）。CfPGRP-S1 和 CfLGBP 的表达在 D 型铰合和晚期面盘幼虫中分别呈相反的下调或上调，而 CfLec-1 和 CfLec-3 在这两个阶段的 6-24 h 内显著增加（P<0.05）。相比之下，在担轮幼虫和晚期面盘幼虫中，免疫效应因子在细菌攻毒后 6 h、12 h 或 24 h 显著上调（P<0.05）。然而，在晚期 D 型铰合幼虫中，CfLYZ 和 CfSOD 的表达在 6 h 时显著下调，而 CfLBP/BPI 的表达在攻毒后 6 h 和 24 h 时上调（P<0.05）。这些结果表明，扇贝的免疫防御系统可能首先出现在担轮幼虫的中腹区，然后在晚期 D 型铰合幼虫后成熟。D 型铰合和晚期面盘幼虫中的发育中的免疫系统可能以不同的方式对免疫刺激做出反应。
