Epon 812

Gastrocnemius were immediately fixed in 2.5% glutaraldehyde and post-fixed in 1% osmium tetroxide. After ethanol gradient dehydration, the samples were embedded in Epon 812 (SPI-90529-77-4, Servicebio, Inc.). Ultrathin sections (50–70 nm) were cut and stained with toluidine blue dye. Then, the sections were observed under a transmission electron microscope (hitachi-HT7700; Hitachi Ltd., Tokyo, Japan). The number of mitochondria was counted. A total of six different fields of view (magnification, x5000) were randomly selected from each section.

Intestinal mucosal sections were fixed in 2.5% glutaraldehyde (Sinopharm Chemical Reagent Co., Ltd., Shanghai, China) for 24 h and 1% osmic acid (Sinopharm Chemical Reagent Co., Ltd) for 2 h, dehydrated through a graded series of ethanol (Sinopharm Chemical Reagent Co., Ltd) and acetone solution (Sinopharm Chemical Reagent Co., Ltd), embedded in Epon812 (Servicebio), sectioned with ultramicrotome (Leica, Weztlar, Germany), and stained with uranyl acetate (Sinopharm Chemical Reagent Co., Ltd) followed by lead citrate (Sinopharm Chemical Reagent Co., Ltd), then observed with transmission electron microscopy (Hitachi, Tokyo, Japan). A total of 20 tissue sections were analyzed for each animal.