家蚕蛹变态期的中肠发育变化观察及细胞凋亡与自噬相关基因的表达检测

Exploration of the Developmental Changes of the Midgut of Silkworm Larvae during Pupal Metamorphosis

为探究家蚕中肠在蛹变态过程中的形态发育变化,于家蚕5龄幼虫期、蛹期及蛾期以24h为间隔,对其中肠进行解剖观察,结果显示:家蚕5龄幼虫的中肠为长圆筒形,在食桑期呈绿色、膨大的管状,进入熟蚕期停止食桑后,中肠开始变黄、缩短;在熟蚕营茧化蛹的过程中,中肠持续缩短、变小,并随着发育时期的延长,最终呈椭圆形颗粒状,表面颜色由浅黄色变为棕黄色。进一步观察家蚕不同发育时期中肠组织的石蜡切片,发现幼虫中肠细胞在进入蛹变态期发生了多次凋亡与再生的过程,在上蔟3~4d、上蔟7~8d、上蔟12d~蛾期都形成了新的中肠上皮细胞,这些新的中肠上皮细胞可能是由再生细胞分化更新而来的。对家蚕中肠组织中细胞凋亡基因(Caspase-1、BmDredd)与自噬基因(Atg1、Atg13)表达模式的检测结果显示,Caspase-1、BmDredd在上蔟2d、7d、12d的表达量均降低,Atg1、Atg13在上蔟7d、9d、12d的表达量均降低,表明在家蚕的这些变态发育时期,其中肠细胞的凋亡受到了抑制,与切片观察新的中肠上皮细胞形成时期基本一致。研究结果呈现了家蚕中肠在蛹变态时期的发育变化过程,说明家蚕幼虫的中肠在蛹发育变态过程中并未彻底消亡,成虫的中肠是幼虫中肠在蛹变态时期经过多次消亡与再生过程发育而来的。

In order to explore the developmental changes of silkworm larvae in the process of pupal metamorphosis,the midgut of the fifth instar larvae,pupal and moth stages of silkworm was anatomically observed at 24 h intervals.The results showed that the midgut of the fifth instar larvae of silkworms was long cylindrical,green and enlarged in the mulberry feeding stage,and the midgut began to turn yellow and shorten after the silkworm larvae stopped eating mulberry at the mature silkworm stage.In the process of molting and metamorphosing into a pupa of mature silkworms,the midgut continues to shorten and become smaller,and with the prolongation of the development period,it finally becomes oval granular,and the surface color changes from light yellow to brownish yellow.Further observation of paraffin sections of midgut tissues at different developmental stages of silkworms showed that larval midgut cells underwent multiple processes of apoptosis and regeneration during pupal metamorphosis.New midgut epithelial cells are formed at 3-4 days,7-8 days,and 12 days-moth periods in cocooning,and these new midgut epithelial cells may be differentiated and renewed from regenerative cells.The expression patterns of apoptosis genes(Caspase-1,BmDredd)and autophagy genes(Atg1,Atg13)showed that the expression levels of Caspase-1 and BmDredd were decreased at 2 days,7 days and 12 days in cocooning,and the expression levels of Atg1 and Atg13 were decreased at 7 days,9 days,and 12 days in cocooning,suggesting that apoptosis of intestinal cells was inhibited during these periods,which were largely consistent with the formation of new midgut epithelial cells in the section results.The results of this study showed the developmental changes of the larval midgut during the pupal metamorphosis period,indicating that the midgut of the silkworm larval did not completely die out during the pupal metamorphosis,and the adult midgut was developed by the larval midgut after multiple deaths and regeneration during the pupal metamorphosis period.