Chia-Chen Lu

Ganoderma lucidum reduces obesity in mice by modulating the composition of the gut microbiota

Obesity is associated with low-grade chronic inflammation and intestinal dysbiosis. Ganoderma lucidum is a medicinal mushroom used in traditional Chinese medicine with putative anti-diabetic effects. Here, we show that a water extract of Ganoderma lucidum mycelium (WEGL) reduces body weight, inflammation and insulin resistance in mice fed a high-fat diet (HFD). Our data indicate that WEGL not only reverses HFD-induced gut dysbiosis—as indicated by the decreased Firmicutes-to-Bacteroidetes ratios and endotoxin-bearing Proteobacteria levels—but also maintains intestinal barrier integrity and reduces metabolic endotoxemia. The anti-obesity and microbiota-modulating effects are transmissible via horizontal faeces transfer from WEGL-treated mice to HFD-fed mice. We further show that high molecular weight polysaccharides (>300 kDa) isolated from the WEGL extract produce similar anti-obesity and microbiota-modulating effects. Our results indicate that G. lucidum and its high molecular weight polysaccharides may be used as prebiotic agents to prevent gut dysbiosis and obesity-related metabolic disorders in obese individuals.

Activation of multiple apoptotic pathways in human nasopharyngeal carcinoma cells by the prenylated isoflavone, osajin.

Osajin is a prenylated isoflavone showing antitumor activity in different tumor cell lines. The underlying mechanism of osajin-induced cancer cell death is not clearly understood. In the present study, the mechanisms of osajin-induced cell death of human nasopharyngeal carcinoma (NPC) cells were explored. Osajin was found to significantly induce apoptosis of NPC cells in a dose- and time-dependent manner. Multiple molecular effects were observed during osajin treatment including a significant loss of mitochondrial transmembrane potential, release of cytochrome c into the cytosol, enhanced expression of Fas ligand (FasL), suppression of glucose-regulated protein 78 kDa (GRP78), and activation of caspases-9, -8, -4 and -3. In addition, up-regulation of proapoptotic Bax protein and down-regulation of antiapoptotic Bcl-2 protein were also observed. Taken together, osajin induces apoptosis in human NPC cells through multiple apoptotic pathways, including the extrinsic death receptor pathway, and intrinsic pathways relying on mitochondria and endoplasmic reticulum stress. Thus, osajin could be developed as a new effective and chemopreventive compound for human NPC.

Activation of an NLRP3 Inflammasome Restricts Mycobacterium kansasii Infection

Mycobacterium kansasii has emerged as an important nontuberculous mycobacterium pathogen, whose incidence and prevalence have been increasing in the last decade. M. kansasii can cause pulmonary tuberculosis clinically and radiographically indistinguishable from that caused by Mycobacterium tuberculosis infection. Unlike the widely-studied M. tuberculosis, little is known about the innate immune response against M. kansasii infection. Although inflammasome activation plays an important role in host defense against bacterial infection, its role against atypical mycobacteria remains poorly understood. In this report, the role of inflammasome activity in THP-1 macrophages against M. kansasii infection was studied. Results indicated that viable, but not heat-killed, M. kansasii induced caspase-1-dependent IL-1β secretion in macrophages. The underlying mechanism was found to be through activation of an inflammasome containing the NLR (Nod-like receptor) family member NLRP3 and the adaptor protein ASC (apoptosis-associated speck-like protein containing a CARD). Further, potassium efflux, lysosomal acidification, ROS production and cathepsin B release played a role in M. kansasii-induced inflammasome activation. Finally, the secreted IL-1β derived from caspase-1 activation was shown to restrict intracellular M. kansasii. These findings demonstrate a biological role for the NLRP3 inflammasome in host defense against M. kansasii.

Identification of CD24 as a Cancer Stem Cell Marker in Human Nasopharyngeal Carcinoma

Cancer stem cells (CSCs) represent a unique sub-population of tumor cells with the ability to initiate tumor growth and sustain self-renewal. Although CSC biomarkers have been described for various tumors, only a few markers have been identified for nasopharyngeal carcinoma (NPC). In this study, we show that CD24+ cells isolated from human NPC cell lines express stem cell genes (Sox2, Oct4, Nanog, Bmi-1, and Rex-1), and show activation of the Wnt/β-catenin signaling pathway. CD24+ cells possess typical CSC characteristics that include enhanced cell proliferation, increased colony and sphere formation, maintenance of cell differentiation potential in prolonged culture, and enhanced resistance to chemotherapeutic drugs. Notably, CD24+ cells produce tumors following inoculation of as few as 500 cells in immunodeficient NOD/SCID mice. CD24+ cells further show increased invasion ability in vitro, which correlates with enhanced expression of matrix metalloproteinase 2 and 9. In summary, our results suggest that CD24 represents a novel CSC biomarker in NPC.

Resveratrol Enhances Perforin Expression and NK Cell Cytotoxicity Through NKG2D-Dependent Pathways

In a previous report, we showed that the in vivo cytotoxic activity of the natural killer (NK) cells isolated from resveratrol‐pretreated rats is significantly enhanced compared with that of the non‐pretreated rats; however, the underlying mechanism remains unclear. In the present study, we use cultured NK92 cell line to examine the possible signaling pathways underlying the resveratrol‐induced activation. Using cultured K562, HepG2, and A549 cells as targets, we show that resveratrol pretreatment increases NK cell cytotoxicity in a dose‐dependent manner. The enhanced cytotoxic effect is accompanied by increases in JNK and ERK‐1/2 MAP kinase activity and perforin expression. Moreover, the expression of NKG2D, an upstream signaling molecule of the MAP kinases pathway, is also enhanced. Resveratrol‐enhanced perforin expression and cytotoxic activity are effectively inhibited by pretreatment with the inhibitors of JNK (SP600125), ERK‐1/2 (PD98059), or by siRNAs against JNK‐1 and ERK‐2. However, the inhibitors or siRNA to p38 exhibits no effect. Since IL‐2 has been shown to induce NKG2D expression and perforin release, we therefore, examined whether IL‐2 and resveratrol act in parallel. We show that IL‐2 also stimulates perforin expression, however, when treated together with resveratrol, they exhibit no additive effect. The results suggest that in NK92 cells, resveratrol may act via a similar or overlapping pathway as that of IL‐2, to enhance perforin expression and cytotoxic activity. Data presented strongly indicate that resveratrol act via NKG2D‐dependent JNK and ERK‐1/2 pathways. J. Cell. Physiol. 223: 343–351, 2010.

RssAB-FlhDC-ShlBA as a major pathogenesis pathway in Serratia marcescens.

ABSTRACT Serratia marcescens has long been recognized as an important opportunistic pathogen, but the underlying pathogenesis mechanism is not completely clear. Here, we report a key pathogenesis pathway in S. marcescens comprising the RssAB two-component system and its downstream elements, FlhDC and the dominant virulence factor hemolysin ShlBA. Expression of shlBA is under the positive control of FlhDC, which is repressed by RssAB signaling. At 37°C, functional RssAB inhibits swarming, represses hemolysin production, and promotes S. marcescens biofilm formation. In comparison, when rssBA is deleted, S. marcescens displays aberrant multicellularity favoring motile swarming with unbridled hemolysin production. Cellular and animal infection models further demonstrate that loss of rssBA transforms this opportunistic pathogen into hypervirulent phenotypes, leading to extensive inflammatory responses coupled with destructive and systemic infection. Hemolysin production is essential in this context. Collectively, a major virulence regulatory pathway is identified in S. marcescens.

Anti-obesogenic and antidiabetic effects of plants and mushrooms

Obesity is reaching global epidemic proportions as a result of factors such as high-calorie diets and lack of physical exercise. Obesity is now considered to be a medical condition, which not only contributes to the risk of developing type 2 diabetes mellitus, cardiovascular disease and cancer, but also negatively affects longevity and quality of life. To combat this epidemic, anti-obesogenic approaches are required that are safe, widely available and inexpensive. Several plants and mushrooms that are consumed in traditional Chinese medicine or as nutraceuticals contain antioxidants, fibre and other phytochemicals, and have anti-obesogenic and antidiabetic effects through the modulation of diverse cellular and physiological pathways. These effects include appetite reduction, modulation of lipid absorption and metabolism, enhancement of insulin sensitivity, thermogenesis and changes in the gut microbiota. In this Review, we describe the molecular mechanisms that underlie the anti-obesogenic and antidiabetic effects of these plants and mushrooms, and propose that combining these food items with existing anti-obesogenic approaches might help to reduce obesity and its complications.

NK cells kill mycobacteria directly by releasing perforin and granulysin

Although the mechanisms underlying the cytotoxic effect of NK cells on tumor cells and intracellular bacteria have been studied extensively, it remains unclear how these cells kill extracellular bacterial pathogens. In this study, we examine how human NK cells kill Mycobacterium kansasii and M.tb. The underlying mechanism is contact dependent and requires two cytolytic proteins: perforin and granulysin. Mycobacteria induce enhanced expression of the cytolytic proteins via activation of the NKG2D/NCR cell‐surface receptors and intracellular signaling pathways involving ERK, JNK, and p38 MAPKs. These results suggest that NK cells use similar cellular mechanisms to kill both bacterial pathogens and target host cells. This report reveals a novel role for NK cells, perforin, and granulysin in killing mycobacteria and highlights a potential alternative defense mechanism that the immune system can use against mycobacterial infection.

The Anti-Tumorigenic Mushroom Agaricus blazei Murill Enhances IL-1β Production and Activates the NLRP3 Inflammasome in Human Macrophages

Agaricus blazei Murill (AbM) has been reported to possess immune activity against tumors and infections through stimulation of mononuclear phagocytes. Recently, AbM extract was shown to induce the production of the pro-inflammatory cytokine, interleukin-1β (IL-1β), in human monocytes. IL-1β is a key pro-inflammatory cytokine produced by activated macrophages and monocytes and its secretion is strictly controlled by the inflammasome. The purpose of this study is to investigate the effect of AbM water extracts on the regulation of IL-1β production and activation of the NLRP3 inflammasome in human THP-1 macrophages. The NLRP3 inflammasome consists of an NLRP3 receptor, an adaptor protein called ASC, and the inflammatory protease, caspase-1. Typically, stimulation of immune cells with microbial products results in production of pro-IL-1β, but a second stress-related signal activates the inflammasome and caspase-1, leading to processing and secretion of IL-1β. Our results show that AbM enhances transcription of IL-1β and triggers NLRP3 inflammasome-mediated IL-1β secretion in human THP-1 macrophages. AbM-mediated IL-1β secretion was markedly reduced in macrophages deficient in NLRP3 and ASC, demonstrating that the NLRP3 inflammasome is essential for AbM-induced IL-1β secretion. In addition, caspase-1 was activated and involved in proteolytic cleavage and secretion of IL-1β in AbM-treated macrophages. AbM-mediated IL-1β secretion also decreased in cells treated with cathepsin B inhibitor, suggesting that AbM can induce the release of cathepsin B. Furthermore, our data show that AbM-induced inflammasome activation requires the release of ATP, binding of extracellular ATP to the purinergic receptor P2X7, the generation of reactive oxygen species, and efflux of potassium. Taken together, these findings reveal that AbM activates the NLRP3 inflammasome via multiple mechanisms, resulting in the secretion of IL-1β.

Resveratrol induces apoptosis of human nasopharyngeal carcinoma cells via activation of multiple apoptotic pathways

Resveratrol, a naturally occurring dietary compound with chemopreventive properties has been reported to trigger a variety of cancer cell types to apoptosis. Whether resveratrol shows any activity on human nasopharyngeal carcinoma (NPC) cells remained to be determined. The aim of this study was to investigate the effect and mechanism of resveratrol on human NPC cells. Treatment of resveratrol resulted in significant decrease in cell viability of NPC cell lines in a dose‐ and time‐dependent manner. A dose‐dependent apoptotic cell death was also measured by flow cytometery analysis. Molecular mechanistic studies of apoptosis unraveled resveratrol treatment resulted in a significant loss of mitochondrial transmembrane potential, release of cytochrome c, enhanced expression of Fas ligand (FasL), and suppression of glucose‐regulated protein 78 kDa (GRP78). These were followed by activation of caspases‐9, ‐8, ‐4, and ‐3, subsequently leading to DNA fragmentation and cell apoptosis. Furthermore, up‐regulation of proapoptotic Bax and down‐regulation of antiapoptotic Bcl‐2 protein were also observed. Taken together, resveratrol induces apoptosis in human NPC cells through regulation of multiple apoptotic pathways, including death receptor, mitochondria, and endoplasmic reticulum (ER) stress. Resveratrol can be developed as an effective compound for human NPC treatment. J. Cell. Physiol. 226: 720–728, 2011.