Transmission electron microscope

T24 and EJ cells were seeded in 6-well plates, and treated with or without 10 μM PPM-18 for 18 h, then washed with PBS, trypsinized, and harvested. Cell pellets were fixed with 2.5% glutaraldehyde, and ultrathin sections were prepared. The formation of autophagosomes in bladder cancer cells was observed under the transmission electron microscope (Olympus, Japan).

After eight weeks of treatment, rats in each group were anesthetized with an intraperitoneal injection of 1% pentobarbital sodium. Then, blood samples were drawn from the abdominal aorta, transferred into dried tubes containing EDTA as an anticoagulant, and centrifuged at 3000 rpm for 10 min at 4 °C. The supernatant liquid of blood was separated and stored at − 80 °C, until being assayed for the plasma SSAO activity, and levels of MA, FA, plasma nitrate/nitrite NO(x)-, and endothelin-1 (ET-1) in the plasma. Meanwhile, the aorta was quickly removed through a thoracotomy, select the 1.5 cm artery segment at the origin of aortic arch, cut the artery longitudinally with the ophthalmic scissors and attach the endothelium to the filter paper, rinse the normal asline and place in formalin solution. Fixation, routine dehydration, paraffin embedding, HE staining, analyzed by optical microscopy (Olympus, Tokyo, Japan). Afterward, cuted approximately 1 cm long aorta into 4% glutaraldehyde phosphate buffer (pH 7.4), were examined under a transmission electron microscope (Olympus, Tokyo, Japan). The other aortic segments were used for determining the SSAO activity. Additionally, the right kidney was quickly removed, fixed with 10% formaldehyde, and stained with H&E, while the cortex of left kidney was dissected out for subsequent examination by electron microscopy.

Treated HT22 cells were then fixed in 2.5% (v/v) glutaraldehyde and collected in a centrifuge tube, following which they were fixed in osmic acid, dehydrated via graded ethanol solutions, and embedded in epoxy resin. Embedded cells were sectioned into ultraslices, stained with uranyl acetate and lead citrate, and observed using a transmission electron microscope (Olympus, Tokyo, Japan).