Aeyung Kim

A Novel Herbal Medicine, KIOM-C, Induces Autophagic and Apoptotic Cell Death Mediated by Activation of JNK and Reactive Oxygen Species in HT1080 Human Fibrosarcoma Cells

KIOM-C was recently demonstrated to have anti-metastatic activity in highly malignant cancer cells via suppression of NF-κB-mediated MMP-9 activity. In addition, it was reported to be effective for clearance of the influenza virus by increasing production of anti-viral cytokines, such as TNF-α and IFN-γ, and efficacious in the treatment of pigs suffering from porcine circovirus-associated disease (PCVAD). In this study, we investigated whether KIOM-C induces cancer cell death and elucidated the underlying anti-cancer mechanisms. In addition, we examined whether KIOM-C oral administration suppresses in vivo tumor growth of HT1080 cells in athymic nude mice. We initially found that KIOM-C at concentrations of 500 and 1000 µg/ml caused dose- and time-dependent cell death in cancer cells, but not normal hepatocytes, to approximately 50% of control levels. At the early stage of KIOM-C treatment (12 h), cells were arrested in G1 phase, which was accompanied by up-regulation of p21 and p27, down-regulation of cyclin D1, and subsequent increases in apoptotic and autophagic cells. Following KIOM-C treatment, the extent of caspase-3 activation, PARP cleavage, Beclin-1 expression, and LC3-II conversion was remarkably up-regulated, but p62 expression was down-regulated. Phosphorylation of AMPK, ULK, JNK, c-jun, and p53 was increased significantly in response to KIOM-C treatment. The levels of intracellular ROS and CHOP expression were also increased. In particular, the JNK-specific inhibitor SP600125 blocked KIOM-C-induced ROS generation and CHOP expression almost completely, which consequently almost completely rescued cell death, indicating that JNK activation plays a critical role in KIOM-C-induced cell death. Furthermore, daily oral administration of 85 and 170 mg/kg KIOM-C efficiently suppressed the tumorigenic growth of HT1080 cells, without systemic toxicity. These results collectively suggest that KIOM-C efficiently induces cancer cell death by both autophagy and apoptosis via activation of JNK signaling pathways, and KIOM-C represents a safe and potent herbal therapy for treating malignancies.

A Novel Herbal Medicine KIOM-MA Exerts an Anti-Inflammatory Effect in LPS-Stimulated RAW 264.7 Macrophage Cells

KIOM-MA was recently reported as a novel herbal medicine effective for atopic dermatitis and asthma. In this study, we have demonstrated the inhibitory effect of KIOM-MA on proinflammatory mediator produced in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. KIOM-MA significantly inhibited the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) as well as nitric oxide (NO) and prostaglandin E2 (PGE2). Consistent with the inhibitory effect on PGE2, KIOM-MA suppresses the LPS-induced migration of macrophages and gelatinase activity and the expression of matrix metalloprotease-9 (MMP-9) in a dose-dependent manner. Additionally, KIOM-MA showed a strong suppressive effect on the inflammatory cytokines production such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). We also found that KIOM-MA inhibits the activation of nuclear factor-κB (NF-κB) and represses the activity of extracellular signal-regulated kinase (ERK), p38, and c-Jun NH2-terminal kinase (JNK) mitogen-activated protein kinases (MAPKs). Taken together, we elucidated the mechanism of anti-inflammatory effect of KIOM-MA using RAW 264.7 cells stimulated by LPS.

Suppression of the invasive potential of highly malignant tumor cells by KIOM-C, a novel herbal medicine, via inhibition of NF-κB activation and MMP-9 expression.

KIOM-C, a novel herbal formula, was recently reported to be effective for treating pigs suffering from porcine circovirus-associated disease (PCVAD). In addition, administration of KIOM-C promoted clearance of influenza virus via production of antiviral cytokines, such as TNF-α and IFN-γ. Since metastasis is the major cause of cancer-related death and the greatest challenge in cancer treatment, we investigated the effect of KIOM-C on the metastatic potential of HT1080 and B16F10 cells. We observed inhibitory properties of KIOM-C in colony-forming activity, migration and invasion. Matrix metalloproteinase-9 (MMP-9) activity in the resting and PMA-stimulated state in HT1080 cells was dose-dependently decreased by KIOM-C treatment via suppression of NF-κB activation. In addition, daily oral administration of KIOM-C at doses of 170 and 510 mg/kg, the corresponding human adult daily doses, efficiently blocked lung metastasis in C57BL/6J mice following injection of B16F10 cells in the tail veins. In particular, none of the mice administered KIOM-C during the experimental period exhibited systemic toxicity, such as body weight loss or liver and kidney dysfunction. Collectively, our results suggest that KIOM-C is a potential therapeutic formula useful as a safe herbal medicine for controlling metastatic cancer.

Oyaksungisan, a Traditional Herbal Formula, Inhibits Cell Proliferation by Induction of Autophagy via JNK Activation in Human Colon Cancer Cells

Oyaksungisan (OY) is a traditional herbal formula broadly used to treat beriberi, vomiting, diarrhea, and circulatory disturbance in Asian countries from ancient times. The effect of OY on cancer, however, was not reported until now. In this study, we have demonstrated that OY inhibits cell proliferation and induces cell death via modulating the autophagy on human colon cancer cells. In HCT116 cells, OY increased the ratio of LC3-II/LC3-I, a marker of autophagy, and treatment with 3-MA, an inhibitor of autophagy, and considerably reduced the formation of autophagosomes. In addition, OY regulated mitogen-activated protein kinase (MAPK) cascades; especially, JNK activation was closely related with autophagy effect by OY in HCT116 cells. Our results indicate that autophagy induction is responsible for the antiproliferative effect by OY, despite the weak apoptosis induction in HCT116 cells. In conclusion, OY might have a potential to be developed as an herbal anticancer remedy.

Induction of apoptotic cell death by betulin in multidrug-resistant human renal carcinoma cells.

Betulin, a triterpene from the bark of various species of birch tree, has various biological effects, including antiviral, antifungal and anticancer activities. The aim of the present study was to elucidate the mechanisms underlying the apoptotic effect of betulin in RCC4 multidrug-resistant human renal carcinoma cells. To evaluate anticancer activity, we performed cell viability and caspase activity assays, a proteome profiler array and western blot analysis in RCC4 cells. Betulin significantly decreased RCC4 cell viability in a time- and concentration-dependent manner. Betulin activated caspase family proteins, including caspase-3, -7, -8 and -9, and increased the expression of apoptosis-related proteins, including PARP and Bcl-2 family members. In an apoptosis array, betulin activated the tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors TRAIL R1/DR4 and R2/DR5, and tumour necrosis factor receptor 1 (TNFR1), suggesting that betulin treatment leads to induction of apoptosis through both intrinsic and extrinsic apoptosis pathways in RCC4 cells. Notably, betulin significantly enhanced cytotoxicity and PARP cleavage in etoposide-treated RCC4 cells, and downregulated the expression of multidrug resistance protein 1 (MDR1). Taken together, our findings suggest that the anticancer effects of betulin involve induction of apoptosis and sensitisation of RCC4 cells, providing potentially useful information applicable to the use of betulin in renal cancer treatment.

Fermented So-Cheong-Ryong-Tang (FCY) induces apoptosis via the activation of caspases and the regulation of MAPK signaling pathways in cancer cells

AbstractBackgroundSo-Cheong-Ryong-Tang (CY), a traditional herbal formula, mainly has been shown to possess allergic rhinitis and asthma for hundreds of years in Asian countries. Although this medicine has been attracted Asian scientists with investigating mechanisms of action against inflammatory-related diseases, there is a little available information on the anti-cancer effect of CY, especially on the fermented form (FCY). In this study, we explored the chemopreventive/chemotherapeutic efficacy of FCY against cancer cells and proved the efficacy of FCY through performing in vivo xenograft assay.MethodsCY was fermented with bacteria and lyophilized. For analysis of the constituents of CY and FCY, high performance liquid chromatography (HPLC)-DAD system was performed. To detect the anti-cancer effect of FCY, cell viability assay, caspase activity assay, cell cycle analysis, and Western blot analysis were performed in AGS human gastric cancer cells. The inhibitory effects of tumor growth by CY and FCY were evaluated in athymic nude mice inoculated with HCT116 human colon cancer cells.ResultsAs a result of analyzing the 11components present in CY and FCY, the contents of ephedrine HCl, glycyrrhizin, gingerol, schisandrin, and gomisin A were respectively increased by fermentation in FCY. The treatment of CY or FCY inhibited the viability of AGS cells, interestingly, the inhibition of cancer cell growth was enhanced by fermentation of CY. FCY induced the apoptosis through activating the caspase-3, −8, and −9. Additionally, FCY regulated the activation of mitogen-activated protein kinases (MAPKs) including extracellular signal-regulated kinase (ERK), p38, and c-Jun NH2-terminal kinase (JNK). In vivo xenografts, administration of FCY significantly inhibited the tumor formation, and improved the anti-tumor effect compared to that of CY in athymic nude mice.ConclusionsFCY indicated significant anti-cancer effects, and its efficacy against tumor formation was improved than that of CY, therefore, FCY might be used for applications of traditional medicine against cancer in modern complementary and alternative therapeutics. Graphical Abstractᅟ

Jaeumganghwa-Tang Induces Apoptosis via the Mitochondrial Pathway and Lactobacillus Fermentation Enhances Its Anti-Cancer Activity in HT1080 Human Fibrosarcoma Cells.

Jaeumganghwa-tang (JGT, Zi-yin-jiang-huo-tang in Chinese and Jiin-koka-to in Japanese) is an oriental herbal formula that has long been used as a traditional medicine to treat respiratory and kidney diseases. Recent studies revealed that JGT exhibited potent inhibitory effects on allergies, inflammation, pain, convulsions, and prostate hyperplasia. Several constituent herbs in JGT induce apoptotic cancer cell death. However, the anti-cancer activity of JGT has not been examined. In this study, we investigated the anti-cancer effects of JGT using highly tumorigenic HT1080 human fibrosarcoma cells and elucidated the underlying mechanisms. In addition, we examined whether the Lactobacillus fermentation of JGT enhanced its anti-cancer activity using an in vivo xenograft model because fermentation of herbal extracts is thought to strengthen their therapeutic effects. Data revealed that JGT suppressed the growth of cancer cells efficiently by stimulating G1 cell cycle arrest and then inducing apoptotic cell death by causing mitochondrial damage and activating caspases. The phosphorylation of p38 and ERK also played a role in JGT-induced cell death. In vitro experiments demonstrated that JGT fermented with Lactobacillus acidophilus, designated fJGT162, elicited similar patterns of cell death as did non-fermented JGT. Meanwhile, the daily oral administration of 120 mg/kg fJGT162 to HT1080-bearing BALB/c nude mice suppressed tumor growth dramatically (up to 90%) compared with saline treatment, whereas the administration of non-fermented JGT suppressed tumor growth by ~70%. Collectively, these results suggest that JGT and fJGT162 are safe and useful complementary and alternative anti-cancer herbal therapies, and that Lactobacillus fermentation improves the in vivo anti-cancer efficacy of JGT significantly.

The novel herbal cocktail MA128 suppresses tumor growth and the metastatic potential of highly malignant tumor cells

MA128, a novel herbal medicine, was previously identified and its effectiveness in the treatment of asthma and atopic dermatitis (AD) was demonstrated. In particular, post-inflammatory hyperpigmentation (PIH) in AD mice was improved by treatment with MA128. In addition, MA128 exhibited anti-melanogenic activity by inhibiting tyrosinase activity via the p38 MAPK and protein kinase A signaling pathways in B16F10 cells. In the present study, we examined whether oral administration of MA128 suppressed the in vivo tumor growth of HT1080 cells in athymic nude mice. The results showed that the daily oral administration of 75 and 150 mg/kg MA128 suppressed the tumorigenic growth of HT1080 cells efficiently. Since metastasis is a major cause of cancer-associated mortality and the greatest challenge during cancer treatment, we investigated the effect of non-toxic concentrations of MA128 on the metastatic potential of HT1080 cells. MA128 inhibited anchorage-independent colony formation, migration and invasion. Matrix metalloproteinase-9 (MMP-9) activity under resting and PMA-stimulated conditions was decreased in a dose-dependent manner by MA128 in HT1080 cells. In addition, the daily oral administration of MA128 at doses of 75 and 150 mg/kg efficiently blocked the lung metastasis of B16F10 cells that had been injected into the tail veins of C57BL/6 mice. In particular, none of the mice treated with MA128 exhibited systemic toxicity, such as body weight loss or liver and kidney dysfunction. MA128 also inhibited tumor‑induced angiogenesis. Taken together, the results suggest that MA128 is a potential therapeutic agent and a safe herbal medicine for controlling malignant and metastatic cancer.

Sosiho‑tang ameliorates cachexia‑related symptoms in mice bearing colon 26 adenocarcinoma by reducing systemic inflammation and muscle loss

Cachexia accompanied by muscle wasting is a key determinant of poor prognosis in cancer patients and cancer‑related death. Previous studies have demonstrated that inflammatory cytokines such as interleukin‑6 (IL‑6), tumor necrosis factor‑α (TNF‑α), IL‑1 and interferon‑γ (IFN‑γ) secreted from host cells and tumor cells participate in skeletal muscle wasting followed by severe loss of body weight. Therefore, blockade of the inflammatory response is thought to be a logical target for pharmacological and nutritional interventions to preserve skeletal muscle mass under cachectic conditions. Sosiho‑tang (SO; Xiaocharihu‑tang in Chinese and Sho‑saiko‑to in Japanese) is an Oriental herbal medicine that has been used to treat chronic hepatic diseases and to control fever. In recent studies, SO inhibited the production of inflammatory cytokines in lipopolysaccharide (LPS)‑stimulated macrophages, prevented thrombus formation and suppressed cancer progression. However, the anti‑cachectic activity of SO in tumor‑bearing mice has not yet been examined. In the present study, we characterized the effect of SO administration on cancer‑induced cachexia in CT‑26‑bearing mice, and elucidated the anti‑cachectic mechanisms. Daily oral administration of SO at doses of 50 and 100 mg/kg to CT‑26‑bearing mice significantly retarded tumor growth and prevented the loss of final body weight, carcass weight, heart weight, gastrocnemius muscle, and epididymal fat, compared with saline‑treated control mice. In addition, serum IL‑6 levels elevated by cancer were decreased by SO administration. In the J774A.1 macrophage cell line, SO efficiently suppressed LPS‑mediated increases in inducible nitric oxide synthase (iNOS) expression, nitric oxide (NO), and procachectic inflammatory cytokine production through inhibition of nuclear factor‑κB (NF‑κB) and p38 activation. In addition, SO attenuated muscle atrophy caused by cancer cells by affecting myoblast proliferation and differentiation, and C2C12 myotube wasting. Taken together, these results suggest that SO is a safe and useful anti‑cachectic therapy for cancer patients with severe weight loss.

Screening of aqueous extracts of medicinal herbs for antimicrobial activity against oral bacteria

Background Dental caries is considered to be a preventable disease, and various antimicrobial agents have been developed for the prevention of dental diseases; however, many bacteria show resistance to existing agents. In this study, 14 medicinal herbs were evaluated for antimicrobial activity against five common oral bacteria as a screen for potential candidates for the development of natural antibiotics. Methods Aqueous extracts of medicinal herbs were tested for activity against Enterococcus faecalis, Actinomyces viscosus, Streptococcus salivarius, Streptococcus mutans, and Streptococcus sanguis grown in brain heart infusion (BHI) broth. A broth microdilution assay was used to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). A disk diffusion assay was performed by inoculating bacterial cultures on BHI agar plates with paper disks soaked in each of the medicinal herb extracts. Inhibition of the synthesis of water-insoluble glucans by S. mutans was also investigated. Results The aqueous extracts of many of the 14 medicinal herbs demonstrated antimicrobial activity against the five types of pathogenic oral bacteria. The extracts of Sappan Lignum, Coptidis Rhizoma, and Psoraleae Semen effectively inhibited the growth of oral bacteria and showed distinct bactericidal activity. The extracts of Notoginseng Radix, Perillae Herba, and Psoraleae Semen decreased the synthesis of water-insoluble glucans by the S. mutans enzyme glucosyltransferase (GTase). The present study is the first to confirm the antimicrobial activity of the extract of Sappan Lignum against all five species of oral bacteria strains. Conclusion These results suggest that certain herbal medicines with proven antimicrobial effects, such as Sappan Lignum and Psoraleae Semen, may be useful for the treatment of dental diseases.