Effect of Fractions from Lycopus lucidus Turcz. Leaves on Genomic DNA Oxidation and Matrix Metalloproteinase Activity

Aim and Objective: We investigated the inhibitory effects of fractions from Lycopus lucidus Turcz. leaves on genomic DNA oxidation, nitric oxide (NO) production and matrix metalloproteinase (MMP) activity. Material and Methods: Oxidative damage of genomic DNA was detected after Fenton reaction with H2O2 using DNA electrophoresis. Western blotting was performed to compare the expression levels of MMP-2 in phorbol 12-myristate 13-acetate (PMA)-induced HT-1080 cells. Lipopolysacchride (LPS)-induced NO production in RAW 264.7 cells was measured using Griess reagent. Results: ll fractions (n-Hexane, 85% aq. MeOH, n-BuOH, and water fractions) from the leaves of L. lucidus Turcz. significantly inhibited intracellular production of reactive oxygen species (ROS) (p<0.05). Particularly, 85% aq. MeOH and n-BuOH fractions showed higher ROS inhibitory activity than the other fractions. n-Hexane, 85% aq. MeOH, n-BuOH and water (0.05 mg/mL) fractions significantly inhibited oxidative DNA damage by 57.97%, 68.48%, 58.97%, and 68.39%, respectively (p <0.05). Treatment of RAW 264.7 cells with each fraction reduced LPS-induced NO production in a dose-dependent manner (p<0.05). n-Hexane and 85% aq. MeOH fractions notably reduced MMP-2 secretion levels of in the culture supernatants from HT-1080 cells. Conclusion: Overall, these results indicated that L. lucidus Turcz. leaves can be exploited as plant based sources of antioxidants in the pharmaceutical, cosmetic, nutraceutical and food industries.

In Vitro and In Silico Studies of Soluble Epoxide Hydrolase Inhibitors from the Roots of Lycopus lucidus

Soluble epoxide hydrolase (sEH) is an enzyme that is considered a potential therapeutic target in human cardiovascular disease. Triterpenes (1–4) and phenylpropanoids (5–10) were isolated from Lycopus lucidus to obtain sEH inhibitors through various chromatographic purificationtechniques. The isolated compounds were evaluated for their inhibitory activity against sEH, and methyl rosmarinate (7), martynoside (8), dimethyl lithospermate (9) and 9″ methyl lithospermate (10) showed remarkable inhibitory activity, with the IC50 values ranging from 10.6 ± 3.2 to 35.7 ± 2.1 µM. Kinetic analysis of these compounds revealed that 7, 9 and 10 were competitive inhibitors bound to the active site, and 8 was the preferred mixed type inhibitor for allosteric sites. Additionally, molecular modeling has identified interacting catalytic residues and bindings between sEH and inhibitors. The results suggest that these compounds are potential candidates that can be used for further development in the prevention and treatment for cardiovascular risk.

Lycopus lucidus Turcz Inhibits the Osteoclastogenesis in RAW 264.7 Cells and Bone Loss in Ovariectomized Rat Model

Lycopus lucidus (LL) is a perennial herb that is traditionally used in Asia to treat edema, wound healing, and gynecological diseases such as irregular menstruation and menstrual pain. We hypothesized that LL would decrease the risk of developing osteoporosis, which is a condition related to gynecological diseases. In this study, we aimed to investigate the effect of a water extract of LL on osteoclastogenesis in vitro and osteoporosis in vivo. In vitro study, we used RAW 264.7 cells as osteoclast precursor cell. Osteoclast differentiation was induced by receptor activator nuclear factor-kappa B ligand (RANKL). We investigated the effect of LL on RANKL-induced osteoclastogenesis, tartrate-resistant acid phosphatase (TRAP) activity, and osteoclast-related genes. In vivo study, we used ovariectomized- (OVX-) induced osteoporosis rat model. OVX-induced Sprague-Dawley rats were randomly separated into sham, OVX, 17β-estradiol (100 μg/kg), wLL-L (15.2 mg/kg), and wLL-H (152 mg/kg) groups. Drugs were administered orally once daily for 9 weeks. wLL inhibited the formation of TRAP-positive osteoclasts; TRAP activity; pit formation; transcription factors (the nuclear factor of activated T-cell cytoplasmic 1 and c-fos); and osteoclast-related genes such as TRAP, carbonic anhydrase II, cathepsin K, osteoclast-associated receptor, and the d2 isoform of the vacuolar ATPase Vo domain. Also, wLL prevented loss of the trabecular area in the OVX femur without change of estrogen level. These results indicate that wLL is able to inhibit osteoclastogenesis and protect bone loss in the OVX-induced osteoporosis model without the influence of hormones like estrogen.

Bioassay-directed fractionation of a blood coagulation factor Xa inhibitor, betulinic acid from Lycopus lucidus

Thrombosis is a major cause of morbidity and mortality worldwide and plays a pivotal role in the pathogenesis of several cardiovascular disorders, including acute coronary syndrome, unstable angina, myocardial infarction, sudden cardiac death, peripheral arterial occlusion, ischemic stroke, deep-vein thrombosis, and pulmonary embolism. Anticoagulants, antiplatelet agents, and fibrinolytics can reduce the risks of these clinical events. Especially, the blood coagulation factor Xa (FXa) inhibitor is a proven anticoagulant. Promoting blood circulation, using traditional Chinese medicine (TCM), for the treatment of these diseases has been safely used for thousands of years in clinical practice. Therefore, highly safe and effective anticoagulant ingredients, including FXa inhibitors, could be found in TCM for activating the blood circulation. One FXa inhibitor, a pentacyclic triterpene (compound 1, betulinic acid) characterized by IR, MS and NMR analyses, was isolated from the ethyl acetate fraction of Lycopus lucidus by bioassay-directed fractionation. Compound 1 exhibited an inhibitory effect on FXa with IC50 25.05 μmol/L and reduced the thrombus weight in an animal model at 25-100 mg/kg. These results indicate that betulinic acid could be the potential for anticoagulant therapy.