Recent research on tendinopathy has focused on its relationship to programmed cell death. Increased autophagy has been observed in ruptured rotator cuff tendon tissues, suggesting a causal relationship. We investigated whether autophagy occurs in human rotator cuff tenofibroblast death induced by oxidative stress and whether antioxidants protect against autophagic cell death. We used H 2 O 2 (0.75 mM) as oxidative stressor, cyanidin (100 mg/ml) as antioxidant, zVAD (20 mM) as apoptosis inhibitor, and 3-MA (10 mM) as autophagy inhibitor. We evaluated cell viability and known autophagic markers: LC3-II expression, GFP-LC3 puncta formation, autolysosomes, and Atg5-12 and Beclin 1 expression. H 2 O 2 exposure increased the rates of cell death, LC3-II expression, GFP-LC3 puncta formation, and autolysosomes. After we induced apoptosis arrest using zVAD, H 2 O 2 exposure still induced cell death, LC3-II expression, and GFP-LC3 puncta formation. H 2 O 2 exposure also increased Atg5-12 and Beclin 1 expressions, indicating autophagic cell death. However, cyanidin treatment reduced H 2 O 2 -induced cell death, LC3-II expression, GFP-LC3 puncta formation, and autolysosomes. Cyanidin and 3-MA similarly reduced the cell-death rate, and Atg5-12 and Beclin 1 expression. This study demonstrated that H 2 O 2 , an oxidative stressor, induces autophagic cell death in rotator cuff tenofibroblasts, and that cyanidin, a natural antioxidant, inhibits autophagic cell death. ß

Local anesthetics (LA) are among the drugs most frequently used for musculoskeletal problems, in procedures ranging from diagnosis to postoperative pain control. The cytotoxicity of LA is an emerging area of concern. The purpose of this study was to determine whether cyanidin, an antioxidant, exerts cytoprotective effects against tenofibroblast death induced by LA. Primary cultured human rotator cuff tenofibroblasts were used to evaluate the cytotoxicity of these LA: Ropivacaine (0.075%), Bupivacaine (0.05%), and Lidocaine (0.2%). The effects of cyanidin (100 mg/ml) on the cytotoxicity induced by these LA were investigated. Cell viability, ROS production, caspase-3/7 activity, and expressions of phospho-extracellular signal-regulated kinases (ERK), phospho-p38, phospho-c-Jun N-terminal kinase (JNK), and cleaved PARP-1 were evaluated. Exposure to LA significantly induced cell death (p < 0.001), ROS production (p 0.04), the activation of caspase-3/7 (p < 0.001), and the increased expressions of phospho-ERK, phospho-p38, phospho-JNK, and cleaved PARP-1. These LA-induced cytotoxic effects were reduced by cyanidin. These data indicate that cyanidin, an antioxidant, has cytoprotective effects against LA-induced cytotoxicity to rotator cuff tenofibroblasts. ß

This study was carried out to determine the biological activities of Wa-song (Orostachys japonicus) hot water extracts. Four types of extract samples were prepared, including Wa-song, traditional plants mixture [PM; mixture of Baekbokryung (Poria cocos), Changchul (Atractylodis rhizoma), and Sa-in (Amomum xanthoides)], and two different ratio composites of these (mixture of PM and Wa-song extract, 1:1 (v/v); PMO-1 and 1:3 (v/v); PMO-3). Their biological activities were measured using various in vitro assays. Total phenolic and flavonoid contents of PM were higher compared to those of Wa-song, and those of PMO-1 were higher than those of PMO-3. Further, PMO-1 contained higher ABTS and DPPH radical scavenging, reducing power, and nitrite scavenging activities than PMO-3. On the contrary, PMO-3 contained higher tyrosinase and inhibitory activities of MCF-7 and HT-29 cancer cells than PMO-1. According to the results, biological activities of PMOs were significantly higher than those of Wa-song extract and PM in in vitro assays. Therefore, we expect that PMOs could show higher biological activities than Wa-song extract alone in vivo.

Local anesthetics (LA) are among the drugs most frequently used for musculoskeletal problems, in procedures ranging from diagnosis to postoperative pain control. Chondrocyte toxicity induced by LA is an emerging area of concern. The purpose of this study was to determine whether N-acetyl cysteine (NAC), an antioxidant, will exert cytoprotective effects against chondrocyte death induced by LA. Primary cultured human chondrocytes were used for this study. This study used control, NAC, LA, and NAC-LA groups. Cytotoxicity was induced in the LA subgroups and their paired NAC-LA subgroups through exposure to ropivacaine (0.075%), bupivacaine (0.05%), or lidocaine (0.2%) for 24 h. The NAC-LA subgroups were exposed to 10 mM NAC for 1 h, before LA exposure. These study groups were evaluated for rates of cell viability, apoptosis, necrosis, intracellular ROS production, and caspase-3/7 activity. Cell viability in all LA subgroups was significantly lower than in the control group (p < 0.001). Cell viability in the NAC-LA subgroups was significantly higher than in their paired LA subgroups (p < 0.001). In the LA subgroups, rates of apoptosis and necrosis, intracellular ROS production, and caspase-3/7 activity were significantly higher than in the control group (p ≤ 0.029). In the NAC-LA subgroups, rates of apoptosis and necrosis, intracellular ROS production, and caspase-3/7 activity were significantly lower than in their paired LA subgroups (p ≤ 0.023). These results indicate that N-acetyl cysteine, an antioxidant, has cytoprotective effects against LA-induced toxicity to chondrocytes in vitro. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:297-303, 2017.

Garlic, a once-a-year crop, is mass-produced in a single event. Most of the garlic harvested during the year, unless consumed or processed immediately, should be stored. Stored raw garlic (SRG) can be used to make black garlic (BG) via aging, and storage may affect the properties and quality of the BG compared with the use of raw garlic that has not been stored. This study was performed to identify the effect of long-term storage of raw garlic on the quality of BG products. SRG was aged for 21 days at 40–86 °C for BG production. Moisture content and pH gradually decreased with the aging period. Total phenolic, total flavonoid, and fructose contents were significantly increased during the aging period. Compared with non-stored raw garlic (NSRG), alliin and S-allylcysteine (SAC) contents were 1.7-fold and 5.9-fold higher in SRG, respectively, and γ-glutamyl-S-allylcysteine (γ-GSAC) content was 2.8-fold lower in SRG. The contents of alliin and γ-GSAC reduced as the aging period of SRG and NSRG progressed. However, the SAC content of NSRG increased with aging, but the SAC content of SRG decreased or increased slightly with extended aging. The antioxidant activity was also higher in BG made from NSRG rather than SRG. These results show that the SAC content is relatively low in BG manufactured from SRG compared with NSRG. Our findings suggest that it is necessary to establish an aging method suitable for SRG in BG production with high SAC content, a representative indicator of BG.