OBJECTIVES: To observe the effect of moxibustion at "Shenshu" (BL23) and "Zusanli" (ST36) on the PTEN-induced putative kinase 1 (PINK1)/E3 ubiquitin ligase (Parkin) pathway and mitophagy in rats with rheumatoid arthritis (RA), so as to explore its possible mechanism in improving RA.
METHODS: Twenty-four SD rats were randomly divided into normal, model, moxibustion, and medication groups, with 6 rats in each group. The RA model was established by Freund's complete adjuvant solution injection combined with freezing and wind-cold dampness method. Suspended moxibustion was applied to BL23 and ST36 for 20 min, once daily for 15 consecutive days. Methotrexate (0.35 mg/kg) was administered via oral gavage twice a week for 15 consecutive days. The morphological changes of mitochondria in synovial tissue were observed by transmission electron microscopy. JC-1 staining was used to detect the level of mitochondrial membrane potential in synovial tissue. The ROS level in serum of rats was detected by fluorescence probe method. The content of ATP in synovial tissue was detected by luciferase assay. The co-localization of mitochondrial outer membrane translocation enzyme 20 (TOMM20) and microtubule-associated protein light chain 3 (LC3) B in synovial tissue was observed by immunofluorescence. The expression levels of Beclin1, selective autophagy adaptor protein (p62), PINK1 and Parkin and the ratio of LC3 Ⅱ/LC3 Ⅰ in synovial tissue were detected by Western blot.
RESULTS: Compared with the normal group, the model group showed swollen mitochondria, disordered structure, disrupted cristae, and increased autophagosomes in synovial tissue;the mitochondrial membrane potential level and ATP content, and the protein expression level of p62 were decreased (<0.01), while the serum ROS level, the co-localization expression of TOMM20 and LC3B in synovial tissue, and the protein expression levels of Beclin1, PINK1, and Parkin and the ratio of LC3 Ⅱ/LC3 Ⅰ were increased (<0.01). In contrast to the model group, the increased and decreased indexes mentioned above were reversed (<0.05,<0.01) in both moxibustion and medication groups;and the mitochondrial membrane potential was obviously higher (<0.05) in the medication group than that in the moxibustion group. P P P P P
CONCLUSIONS: Moxibustion at BL23 and ST36 can ameliorate mitochondrial structural damage and reduce the level of mitochondrial autophagy in RA model rats, which may be related to its function in inhibiting the abnormal activation of PINK1/Parkin pathway.
