Androgen receptor inhibition alleviated inflammation in experimental autoimmune myocarditis by increasing autophagy in macrophages.

OBJECTIVE: Experimental autoimmune myocarditis (EAM) is characterized by pronounced macrophage infiltration, cardiac necrosis, and cardiac fibrosis. Our previous studies have demonstrated that suppressed androgen receptor (AR) enables anti-inflammation to promote tissue repair by decreasing M1 macrophages and increasing M2 macrophages in an EAM model. Given that autophagy mediates inflammatory response in macrophages, we investigated whether AR inhibition executes its protective role in inflammation through the autophagy pathway in EAM. MATERIALS AND METHODS: To determine whether AR inhibition can perform its anti-inflammatory effects by upregulating autophagy, we pre-treated mice with 3-methyl adenine (3-MA), a pharmacological inhibitor of autophagy. Immunofluorescence assay and Western blot were used to detect autophagy levels and autophagy activity in five different groups. Immunofluorescence marked F4/80 and LC3 to illustrate the autophagy level in macrophages. TUNEL assays were used to detect the apoptosis level in heart tissue of five different groups. RESULTS: We demonstrated that AR inhibition resolves injury with sustained inhibition of inflammatory cytokines associated with enhanced autophagy, especially in macrophages. Increased LC3II/I expression corroborated complete autolysosome formation detected by electron microscopy and correlated with degradation of SQSTM1/p62 in the AR inhibition group by Western blot. These effects could be reversed within 3-MA, a pharmacological inhibitor of autophagy. Specifically, pharmacological inhibition of autophagy increased apoptosis and inflammation, which could be attenuated by AR inhibition. CONCLUSIONS: AR inhibition alleviates the inflammatory response and tissue apoptosis by enhancing autophagy, especially in macrophages.

[Isolation of high-quality genomic DNA from plants].

In order to isolate high-quality genomic DNA from medicinal plant tissues enriching polyphenols and polysaccharides, a simple and rapid method based on CTAB extraction for isolating high-quality intact DNA was established by modifying several existing methods. With this technique, the absorbance ratio (A260/A280) of DNAs obtained from fresh and/or dried roots of Panax ginseng, P. Quinquefolius and P. notoginseng was 1.8 approximately. The restriction fragments of DNAs were directly digested with restriction enzyme (EcoR I/Mse I), linked up by T4 DNA ligase and amplified by nested PCR. Reproducible amplified fragment length polymorphism (AFLP) genomic DNA fingerprinting profiles were established with the isolated DNAs. The results demonstrate that the modified technique may be efficient and reliable in isolating high-quality and high-molecular-weight DNAs from fresh and/or dried medicinal plants containing a high content of polyphenols and polysaccharides. We expect that this method can also be applied to other plants.