黑斑侧褶蛙蝌蚪皮肤抗菌肽最早表达时间的确定

Determination of the Onset Time of Antimicrobial Peptide Expression in the Skin of Black-Spotted Frog Tadpoles

两栖类动物的胚胎和蝌蚪期体表包被着由黏蛋白构成的胶体屏障,使其免受环境微生物的侵袭。这层屏障随着蝌蚪的发育逐渐变薄,保护作用逐渐减弱。与此同时,皮肤开始分泌抗菌肽作为补偿。但是,蝌蚪最早在哪个发育阶段开始分泌抗菌肽尚不清楚。为了回答这一问题,本研究以东北地区黑斑侧褶蛙(Pelophylax nigromaculatus)蝌蚪为对象,研究了各个发育时期皮肤组织结构的变化和抗菌肽的表达情况,结果显示,在蝌蚪孵化之后,皮肤仅由1—2层细胞构成,大量的巨型细胞分泌黏液和微小色素颗粒;随着发育进程,皮肤表皮层逐渐加厚,到T3期(Gosner 26期)之后疏松层和致密层均开始加厚,并出现简单的颗粒腺和黏液腺。也是从这一时期皮肤开始表达抗菌肽。到T4期(Gosner 27—45期)和幼蛙期已经具备了成年蛙近似的皮肤结构和抗菌肽合成功能,形成了完备的先天免疫屏障。这些结果表明,蝌蚪皮肤免疫必须从物理屏障转换为化学屏障才能保证其生存和成功登陆,任何影响这一转换过程的因素都会通过扰动其免疫能力而影响其生存力,这为深入理解气候变化或环境污染等生态背景下两栖类种群波动的机制提供新的视角。

The embryos and tadpoles of amphibians are protected by a mucus barrier from threats of environmental microbes.The protective mucus layer becomes weaker as development progresses.As compensation,the skin synthesizes antimicrobial peptides(AMPs)to inhibit accessing microbes.However,it is still unknown when the synthesis of AMPs starts.In order to answer the question,we investigated histological dynamics and expression of AMPs in the skin of black-spotted frog(Pelophylax nigromaculatus)tadpoles at different developmental stages.Results showed that the skin of the embryos and hatchlings were composed of 1-2 layers of cells.Among these layers,giant mucosal cells and secretion cells produce mucus layers and fine pigment granules on the surface of the skin.Epidermis continued to thicken as development progressed,and up to stage T3(corresponding to Gosner s stage 26),the spongy layer and compact layer were drastically thickened with primary mucus and granular glands being observable.It was at this stage that AMPs were first detected.By stage T4(Gosner s stage 27 to 45)and to the juvenile frog stage,the skin tissue had developed structure similar to that of the adult stage that was available for AMP synthesis.Thus the skin had formed a complete congenital immunologic barrier.These findings suggest that the shift of immunity barrier from physical form to chemical form might be essential for tadpole viability in water and on land.Factors disturbing this process would affect its survival.This provides a new insight into the mechanism of population dynamics in the ecological contexts of climate change and water pollution.