Wonseok Chung

Capillarisin inhibits constitutive and inducible STAT3 activation through induction of SHP-1 and SHP-2 tyrosine phosphatases.

Signal transducers and activators of transcription (STAT)-3 is a latent cytosolic transcription factor that has been closely associated with survival, proliferation, chemoresistance, and metastasis of tumor cells. Whether the anti-proliferative, pro-apoptotic, and anti-metastatic effects of capillarisin (CPS), derived from Artemisia capillaris (Compositae), are linked to its capability to inhibit STAT3 activation was investigated. We found that CPS specifically inhibited both constitutive and inducible STAT3 activation at tyrosine residue 705 but not at serine residue 727 in human multiple myeloma cells. Besides the inhibition of STAT3 phosphorylation, CPS also abrogated STAT3 constitutive activity and nuclear translocation. The suppression of STAT3 was mediated through the inhibition of activation of upstream JAK1, JAK2, and c-Src kinases. Treatment with the protein tyrosine phosphatase (PTP) inhibitor pervanadate treatment reversed the CPS-induced down-regulation of JAK1/2 and STAT3, thereby suggesting the involvement of a PTP. Indeed, knockdown of the SHP-1 and SHP-2 genes by small interfering RNA suppressed the ability of CPS to inhibit JAK1 and STAT3 activation, suggesting the critical role of both SHP-1 and SHP-2 in its possible mechanism of action. CPS downregulated the expression of STAT3-regulated antiapoptotic and proliferative gene products; and this correlated with suppression of cell viability, the accumulation of cells in sub-G1 phase of cell cycle and induction of apoptosis. Moreover, CPS potentiated bortezomib-induced apoptotic effects in MM cells, and this correlated with down-regulation of various gene products that mediate cell proliferation (Cyclin D1 and COX-2), cell survival (Bcl-2, Bcl-xl, IAP1, IAP2, and Survivin), invasion (MMP-9), and angiogenesis (VEGF). Thus, overall, our results suggest that CPS is a novel blocker of STAT3 activation and thus may have a potential in negative regulation of growth, metastasis, and chemoresistance of tumor cells.

Capillarisin inhibits iNOS, COX-2 expression, and proinflammatory cytokines in LPS-induced RAW 264.7 macrophages via the suppression of ERK, JNK, and NF-κB activation.

The aerial parts of Artemisia capillaris (Compositae) have been used in traditional Korean medicine as a cholagogic, antipyretic, anti-inflammatory, and diuretic purposes. In our previous study, ethanolic extracts of the plant demonstrated a marked anti-inflammatory effect in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells (J. Korean Soc. Appl. Biol. Chem., 2010, 53, 275–282). In the present study, capillarisin (CPS), a flavone, main constituent of A. capillaris, was examined for its anti-inflammatory activity in the cells. We found that CPS highly suppressed LPS-induced nitric oxide (NO) without exerting cytotoxic effects on RAW 264.7 cells. CPS inhibited the expression of LPS-induced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein and their mRNA in a dose-dependent manner. Also, tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-1β, and prostaglandin E2 (PGE2) secretion were decreased by CPS in LPS-stimulated macrophages. As a result, CPS inhibited proinflammatory cytokines, iNOS, and COX-2, which is attributed to the suppression of LPS-induced ERK, JNK, and nuclear factor-κB (NF-κB) activation. Therefore, we demonstrate here that CPS potentially inhibits the biomarkers related to inflammation through the abrogation of ERK, JNK, and NF-κB p65 activation, and it may be a potential therapeutic candidate for the treatment of inflammatory diseases.

Bergamottin, a natural furanocoumarin obtained from grapefruit juice induces chemosensitization and apoptosis through the inhibition of STAT3 signaling pathway in tumor cells

Persistent activation of signal transducers and activator of transcription 3 (STAT3) has been closely related to growth, survival, proliferation, metastasis, and angiogenesis of various cancer cells, and thus its inhibition can be considered a potential therapeutic strategy. In this study, we investigated the role of bergamottin (BGM) obtained from grapefruit juice in abrogating the constitutive STAT3 activation in multiple myeloma (MM) cells. This suppression was mediated through the inhibition of phosphorylation of Janus-activated kinase (JAK) 1/2 and c-Src. Pervanadate reversed the BGM induced down-regulation of STAT3 activation, suggesting the involvement of a protein tyrosine phosphatase (PTP). Furthermore, BGM induced the expression of the tyrosine phosphatase SHP-1, and gene silencing of the SHP-1 by small interfering RNA abolished the ability of BGM to inhibit STAT3 activation, suggesting a critical role for SHP-1 in the action of BGM. BGM also downregulated the expression of STAT3-regulated gene products such as COX-2, VEGF, cyclin D1, survivin, IAP-1, Bcl-2, and Bcl-xl in MM cells. This correlated with induction of substantial apoptosis as indicated by an increase in the sub-G1 cell population and caspase-3 induced PARP cleavage. Also, this agent significantly potentiated the apoptotic effects of bortezomib and thalidomide in MM cells. Overall, these results suggest that BGM is a novel blocker of STAT3 activation pathway thus may have a potential in therapy of MM and other cancers.

Effect of Korean Herbal Medicine Combined with a Probiotic Mixture on Diarrhea-Dominant Irritable Bowel Syndrome: A Double-Blind, Randomized, Placebo-Controlled Trial

Introduction. Although combination therapy with herbal medicine and probiotics is gaining popularity for controlling diarrhea-dominant irritable bowel syndrome (D-IBS) symptoms, few studies have investigated its clinical effects. Materials and Methods. Fifty-three patients with D-IBS were randomly allocated into 1 of the following 4 groups: herbal medicine (Gwakhyangjeonggisan; GJS) plus probiotics (Duolac7S; DUO), GJS plus placebo DUO, placebo GJS plus DUO, and placebo GJS plus placebo DUO. The study period consisted of a 2-week run-in, 8 weeks of administration, and 2 weeks of follow-up. The primary outcomes were weekly adequate relief (AR) of overall IBS symptoms and the proportion of responders (PR) during the administration period. The secondary outcomes included individual IBS symptoms, stool assessment, and quality of life. Changes of intestinal microbiota and intestinal permeability were also analyzed. Results and Discussion. Weekly AR was not different among the 4 groups throughout the treatment period. However, the 3 treatment groups exhibited significant improvements in PR compared to the findings in the placebo group. In the intestinal microbiota assessment, herbal medicine and probiotics synergistically increased beneficial bacteria counts. Conclusion. Combination therapy with herbal medicine and probiotics appears to relieve overall IBS symptoms by synergistically increasing beneficial intestinal microbe counts.

β-Caryophyllene oxide potentiates TNFα-induced apoptosis and inhibits invasion through down-modulation of NF-κB-regulated gene products

We have recently reported that β-caryophyllene oxide (CPO) can induce apoptosis, suppress tumor growth, and inhibit metastasis through the suppression of signal transducer and activator of transcription 3, PI3K/AKT/mTOR/S6K1 signaling cascades and ROS-mediated MAPKs activation. In the present study, we found that CPO potentiated the apoptosis induced by tumor necrosis factor α (TNFα) and chemotherapeutic agents, suppressed TNFα-induced tumor cell invasion, all of which are known to require NF-κB activation. We found that TNFα stimulated the expression of gene products involved in anti-apoptosis (IAP1, IAP2, Bcl-2, Bcl-xL, and survivin), proliferation (COX-2, cyclin D1, and c-Myc), invasion (MMP 9 and ICAM-1), and angiogenesis (VEGF) and that CPO treatment suppressed their expression. Because these gene products are also regulated by proinflammatory transcription factor NF-κB, we postulated that CPO may mediate its effects by modulating the NF-κB pathway. We found that CPO blocked both inducible and constitutive NF-κB activation in a wide variety of tumor cells. CPO was also found to inhibit the TNFα-induced degradation of IκBα through the inhibition of activation of IκBα kinase and p65 nuclear translocation and phosphorylation. Interestingly, CPO failed to potentiate the apoptotic effect induced by TNFα in p65−/− cells as compared to the wild-type. Thus, overall, our results indicate that the inhibition of NF-κB is one of major mechanisms by which CPO enhances TNFα-induced apoptosis and suppresses invasion.

Antiinflammatory effect of Oldenlandia diffusa and its constituent, hentriacontane, through suppression of caspase-1 activation in mouse peritoneal macrophages.

Oldenlandia diffusa (OD) has been used as a natural drug for the treatment of cancer in Asia and specifically in Korea. However, the antiinflammatory mechanisms employed by OD have yet to be completely understood. This study attempted to determine the effects of OD and hentriacontane, one of the constituent compounds of OD, on lipopolysaccharide (LPS)‐induced inflammatory responses in mouse peritoneal macrophages. The findings of this study showed that OD inhibited the production of tumor necrosis factor (TNF)‐α, interleukin (IL)‐6 and prostaglandin E2 (PGE2). The OD inhibited the enhanced levels of cyclooxygenase (COX)‐2 and inducible nitric oxide synthase (iNOS) induced by LPS. It was shown that the antiinflammatory effect of OD occurs via the regulation of the activation of nuclear factor (NF)‐κB and caspase‐1. Moreover, hentriacontane was shown to ameliorate the expression of inflammatory mediators (TNF‐α, IL‐6, PGE2, COX‐2 and iNOS) and the activation of NF‐κB and caspase‐1 in LPS‐stimulated peritoneal macrophages. These results provide novel insights into the pharmacological actions of OD as a potential candidate for the development of new drugs for the treatment of inflammatory diseases. Copyright

6-Shogaol exerts anti-proliferative and pro-apoptotic effects through the modulation of STAT3 and MAPKs signaling pathways

6‐shogaol (6SG), one of active ingredients in ginger (Zingiber officinale), is known to exhibit anti‐proliferative, anti‐metastatic, and pro‐apoptotic activities through a mechanism that is not fully elucidated. Because the aberrant activation of STAT3 and MAPKs have been associated with regulation of proliferation, invasion, and metastasis of tumors, we hypothesized that 6SG modulates the activation of STAT3 and MAPKs activation in tumor cells. We found that 6SG strongly inhibited constitutive phosphorylation of STAT3 through inhibition of the activation of upstream JAK2 and c‐Src kinases and nuclear translocation of STAT3 on both MDA‐MB231 and DU145 cells. Also, 6SG caused the activation of JNK, p38 MAPK, and ERK. Inhibition of ROS generation by N‐acetylcysteine (NAC) significantly prevented 6SG‐induced apoptosis. 6SG induced apoptosis as characterized by cleavage of PARP, accumulation of cells in subG1 phase, positive Annexin V binding, down‐regulation of STAT3‐regulated proteins, and activation of caspase‐8, ‐9, ‐3 in both MDA‐MB231 cells. Compared with other analogues of 6SG, such as 6‐gingerol (6G), 8‐gingerol (8G), and 10‐gingerol (10G), 6SG was found to be the most potent blocker of STAT3 activation. We observed that the administration of 6SG alone significantly suppressed the growth of the tumor. As compared to the vehicle control, 6SG also suppressed the expression of STAT3‐regulated gene products such as Bcl‐2, Bcl‐xL, and Survivin in tumor tissues. Overall, these findings suggest that 6SG can interfere with multiple signaling cascades involved in tumorigenesis and can be used as a potential therapeutic candidate for both the prevention and treatment of cancer.

The Protective Effect of Cassia obtusifolia on DSS-Induced Colitis

Cassia obtusifolia (CO) has been traditionally used in Korea to treat eye inflammation, photophobia, and lacrimation. However, the regulatory effect and molecular mechanism of CO in intestinal inflammation has not been understood. In this study, we investigate the protective effect of CO in dextran sulfate sodium (DSS)-induced colitis. CO reduced clinical signs of DSS-induced colitis, including body weight loss, shortened colon length, and increased disease activity index. The results show that CO significantly suppressed the levels of interleukin (IL)-6 and expression of cyclooxygenase-2 in DSS-treated colon tissues. Additionally, we observed that CO reduced the activation of transcription nuclear factor-κB p65 in DSS-treated colon tissues. Taken together, these findings suggest that CO has improving effects on DSS-induced ulcerative colitis, which may explain its beneficial effect in the regulation of chronic intestinal inflammation.

Transcriptomic analysis of the bitter taste receptor-mediated glucagon-like peptide-1 stimulation effect of quinine

Quinine is a bitter taste receptor agonist that has been studied its anti-pyretic, anti-malarial, anti-pain, and anti-inflammatory activity. In this study, glucagon-like peptide-1 (GLP-1) stimulation effect of quinine was investigated. Bitter taste receptors are G protein-coupled receptor (GPCR), which transfer the molecular signal through its downstream cascade. The activation of bitter taste receptor, which expressed in the enteroendocrine L cells, stimulates the GLP-1 secretion and therefore can be a therapeutic target of the type-2 diabetes mellitus (T2DM). Here, we studied GLP-1 stimulation effect of quinine on the endocrine differentiated NCI-H716 cells. To investigate the molecular mode-of-action of the GLP-1 stimulation effect of quinine in the enteroendocrine L cells, transcriptomic analysis was performed. Our data suggest that quinine stimulates the GLP-1 secretion through the bitter taste receptor-signaling pathway, and thus has the possibility of therapeutic agent of T2DM.

Immune-enhancing effect of Danggwibohyeoltang, an extract from Astragali Radix and Angelicae gigantis Radix, in vitro and in vivo.

Danggwibohyeoltang (DGBHT) is an oriental herbal prescription consisting of two herbs: Astragali Radix and Angelicae gigantis Radix. We examined the immune-enhancing effect of DGBHT in mice using the forced swimming test (FST) and in vitro tests in peritoneal macrophages. After daily oral administration of DGBHT, blood biochemical parameters related to fatigue were measured after the FST. The immobility time in the FST was significantly decreased in the DGBHT-treated group (200 mg/kg) on the 10th and 14th days. DGBHT (100~~200 mg/kg) treatment significantly increased glucose levels, acting as an energy source. Lactic dehydrogenase levels, which are accurate indicators of muscle damage, tended to decline after DGBHT administration (100~200 mg/kg). When DGBHT (200 mg/kg) was orally administered to mice, creatine kinase levels tended to decrease; however, this decrease was not significant. DGBHT did not have any effects on the variation of total protein and blood urea nitrogen levels. Further, we examined how DGBHT regulates cytokine production, nitric oxide (NO) production, and nuclear factor-kappa B (NF-κB) activation in mouse peritoneal macrophages. When DGBHT was used in combination with recombinant interferon-gamma (rIFN-γ), there was a noticeable cooperative induction of NO production and NF-κB activation. Moreover, rIFN-γ plus DGBHT treatment of peritoneal macrophages significantly increased the production of tumor necrosis factor-alpha (TNF-α) and interleukin-12 (IL-12). These results suggest that DGBHT improves immune function through the changes in indicators related to fatigue and the regulatory effects on immunological parameters, such as TNF-α, IL-12, NO production, and NF-κB activation.