First Inventory of Fungi in Symptomless and Symptomatic Chinese Mesona Indicates Phytopathological Threat.

Chinese mesona (Platostoma palustre) plays an important role as special crop in Southeast Asia and Taiwan for the production of herbal tea, grass jelly, and further processed food. In order to assess the potential threat of fungi to Chinese mesona, we surveyed isolates from symptomless plants in the area of mesona production, as well as from leaf spots of potted plants in a garden shop and a plantation in a botanical garden in Taiwan. From leaves, stems, and roots of 15 symptomless plants sampled at five collection events over two years, 154 isolates from 810 surface-sterilized plant fragments were obtained and identified based on DNA sequence data of the internal transcribed spacer region, and partially of the ß-tubulin and histone H3 genes. The most common species belonged to the genera Cercospora, Colletotrichum, and Fusarium and were considered to be potential plant pathogens. Latent pathogenicity was confirmed by an infection experiment with an endophytic strain of Corynespora cassiicola. Observation of leaf spot disease associated with Cercospora kikuchii suggested pathogenicity of this fungus, which was also isolated as an endophyte from symptomless leaves. We hypothesize that the most common endophytic fungi are latent pathogens in the host and may cause plant disease when the host becomes weakened by senescence or changed cultivation condition. Leaf spots of plants in the botanical garden were associated with a species of Pseudocercospora, which was not found among the endophytic isolates and is newly described based on morphology and analysis of translation elongation factor 1 alpha gene sequences.

Ouabain promotes immune responses in WEHI-3 cells to generate leukemia mice through enhancing phagocytosis and natural killer cell activities in vivo.

Ouabain, a cardiotonic steroid, was used for the treatment of heart failure and atrial fibrillation and induces cancer cell apoptosis in many human cancer cells including human leukemia cells. However, there are no reports to show the effects on immune responses in a leukemia mouse model. In this study, WEHI-3 cell generated leukemia mice were developed and treated by oral ouabain at 0, 0.75, 1.5, and 3 mg/kg for 15 days. Results indicated that ouabain did not affect body appearance, but decreased liver and spleen weights, B- and T-cell proliferation at all three doses treatment and increased CD19 cells at 3.0 mg/kg treatment, decreased CD3, CD11b, and Mac-3 cells levels compared with positive control. Furthermore, ouabain increased the macrophage phagocytosis from peripheral blood mononuclear cell and peritoneal cavity at all three doses treatment and increased NK cell activities. Ouabain restored GOT, GPT and LDH levels in WEHI-3 leukemia mice in vivo.

Laminarin Promotes Immune Responses and Reduces Lactate Dehydrogenase But Increases Glutamic Pyruvic Transaminase in Normal Mice In Vivo.

BACKGROUND/AIM: Laminarin, a typical component of fungal cell walls, has been shown to induce immune responses in both adult and larval locusts. We investigated the effects of laminarin on immune response and glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT) and lactate dehydrogenase (LDH) levels in normal mice. MATERIALS AND METHODS: Thirty-six normal BALB/c mice were randomly divided into four groups and treatments were provided by gavage. Group I mice acted as normal control; mice of groups II-IV received laminarin at different doses (100 µl at 1, 2.5 and 5.0 mg/mouse in double-distilled water, respectively). All animals were treated for 14 days and were weighed, blood was collected for determination of cell markers, liver and spleen samples were weighed. Spleens were used for phagocytosis and determination of natural killer (NK) cell activity and cell proliferation by flow cytometric assay. RESULTS: Laminarin reduced the body weights and weights of liver and spleen. Laminarin increased CD3, CD19 and Mac-3 cell populations at 2.5 and 5 mg/mouse, however, these did not affect CD11b marker levels. Laminarin (1 and 5 mg/mouse) reduced macrophage phagocytosis from peripheral blood mononuclear cells, but did not affect phagocytosis by macrophages from the peritoneal cavity. At an effector:target ratio of 50:1, laminarin reduced NK cell cytotoxic activity at all levels, but at a ratio of 25:1, only at 1 mg treatment. Laminarin did not affect T-cell and B-cell proliferation. Laminarin increased the level of GPT and reduced that of LDH at all doses, indicating laminarin can protect against liver injury. Laminarin is worthy of investigation in future experiments on improving immune responses.

Ouabain Induces Apoptotic Cell Death Through Caspase- and Mitochondria-dependent Pathways in Human Osteosarcoma U-2 OS Cells.

BACKGROUND/AIM: Ouabain, a plant-derived product/substance with Na+/K+-ATPase inhibiting properties, has been shown to exert anti-cancer activity on human cancer cells. This is the first study to investigate the effect of ouabain on apoptotic cell death of human osteosarcoma-derived U-2 OS cells. MATERIALS AND METHODS: Flow cytometry was used to examine cell viability, cell cycle, and reactive oxygen species (ROS), Ca2+, mitochondrial membrane potential (MMP) and caspase activity. Morphological changes were examined by contrast-phase microscopy, while apoptosis-associated protein levels were analyzed by western blot. RESULTS: Ouabain, at concentrations of 5-60 µM, significantly decreased the total viable cells and induced cell morphological changes in a time-dependent manner. It also time-dependently decreased G0/G1 phase and increased S and G2/M phase in U-2 OS cells. The production of ROS and the levels of MMPs (ΔΨm) were inhibited, while Ca2+ production in U-2 OS cells was increased. Regarding cell apoptosis, flow cytometry assay revealed increased caspase-3, -8, and -9 activities in U-2 OS cells. Moreover, western blot results showed that ouabain increased the expression of pro-apoptotic protein Bax and decreased the expression of anti-apoptotic protein Bcl-2 in U-2 OS cells. Furthermore, results also showed that ouabain increased cytochrome c release, apoptosis-inducing factor (AIF) and endonuclease (Endo) G that is associated with apoptosis through caspase-dependent and -independent pathway in U-2 OS cells. CONCLUSION: Our findings provide important insight into the cytotoxic effects of ouabain on U-2 OS cells, in vitro, which are mediated at least partly via cell apoptosis induction.

Butylated hydroxyanisole affects immunomodulation and promotes macrophage phagocytosis in normal BALB/c mice.

Butylated hydroxyanisole (BHA), a synthetic antioxidant, has been used in fat and fatty foods to prevent oxidative deterioration. However, the functions of BHA on immune responses in normal mice remain elusive. The aim of the present study was to investigate the effects of oral treatment of BHA on immune responses in normal mice in vivo. BALB/c mice received various treatments. Blood samples were collected and analyzed. Flow cytometry was used to determine the levels of the cell markers. Results showed that BHA did not significantly affect the weight of the animal body and spleen in normal mice. BHA promoted macrophage phagocytosis from peripheral blood mononuclear cells, but did not alter this process in the peritoneal cavity. Furthermore, BHA did not influence natural-killer cell cytotoxicity in normal mice. Notably, BHA promoted the levels of CD3 (T cells) and decreased the level of CD19 (B cells), but did not significantly affect the levels of CD11b (monocytes) and macrophages (Mac-3) in normal mice. Based on these observations it can be concluded that BHA promotes immune responses by increasing T cells and activating phagocytosis by macrophages in normal mice. However, the molecular mechanisms require further investigation.

Danthron induced apoptosis through mitochondria- and caspase-3-dependent pathways in human brain glioblastoma multiforms GBM 8401 cells.

Danthron (1,8-dihydroxyanthraquinone), is one of component from Rheum palmatum L. (Polygonaceae), has been shown several biological activities but did not show to induce apoptosis in human brain tumor cells. The aim of this study is to investigate the mechanisms by danthron for the induction of apoptotic potential on human brain glioblastoma multiforms GBM 8401 cell line. Danthron showed a marked concentration- and time-dependent inhibition of GBM 8401 cell viability and induced apoptosis in a dose-and time-dependent manner. There was an attenuation of mitochondrial membrane potential (DeltaPsi(m)) with the alterations of Bcl-2/Bax protein ratio in GBM 8401 cells, indicating the participation of a mitochondria-related mechanism. Pretreatment of a caspase-8 inhibitor (Z-IETD-FMK), caspase-9 inhibitor (Z-LEHD-FMK) and caspase-3 inhibitor (Z-DEVE-FMK) significantly increased the viable of GBM 8401 cells implied that the participations of caspases. Western blotting analysis also showed the activation of initiator caspase-8 and caspase-9, and executor caspase-3 in GBM 8401 cells. Meanwhile, danthron also promoted the generation of reactive oxygen species (ROS) and cytosolic Ca(2+) in GBM 8401 cells. Taken together, our data showed that danthron induced apoptosis in GBM 8401 cells through mitochondria-related and caspase-related pathways, and it may be further evaluated as a chemotherapeutic agent for human brain cancer.