Monascin ameliorate inflammation in the lipopolysaccharide-induced BV-2 microglial cells via suppressing the NF-κB/p65 pathway.

OBJECTIVES: The pathophysiology of neurodegenerative diseases is complicated, in which inflammatory reactions play a vital role. Microglia cells activation, an essential process of neuroinflammation, can produce neurotoxic molecules and neurotrophic factors, which aggravate inflammation and neuronal injury. Monascin, a major component of red yeast rice, is an azaphilonoid pigment with potential anti-inflammatory effects; however, the effects in central nervous system have not been evaluated. Our goal in this project was to explore the therapeutic effect and the underlying mechanism of Monascin, which may be via anti-inflammatory action. MATERIALS AND METHODS: We used lipopolysaccharide to induce BV-2 microglial cells in order to form an inflammation model in vitro. The anti-inflammatory effects of Monascin were measured by enzyme-linked immunosorbent assay (ELISA), real time-polymerase chain reaction (RT-PCR), Western Blot and Immunofluorescent staining. RESULTS: Our data indicated that inflammatory cytokines including interleukin-1ß (IL-1ß), IL-6, tumor necrosis factor-alpha (TNF-α) and nitric oxide were suppressed by Monascin treatment. Furthermore, the related pro-inflammatory genes were inhibited consistent with the results of ELISA assay. Western blotting results showed that the phosphorylation of nuclear factor kappa B (NF-κB/p65) was reduced by Monascin treatment may be through suppressing the activation of IκB. Furthermore, immunofluorescence staining showed that the translocation of NF-κB/p65 to the cellular nuclear was blockaded after Monascin treatment. CONCLUSION: Taken together, Monascin exerts anti-inflammatory effect and suppressed microglia activation, which suggested its potential therapeutic effect for inflammation-related diseases.

p204 Is Required for Canonical Lipopolysaccharide-induced TLR4 Signaling in Mice.

p204, a murine member of an interferon-inducible p200 family, was reported to recognize intracellular viral and bacterial DNAs, however, its role in the innate immunity in vivo remains unknown due to the lack of p204-deficient animal models. In this study we first generated the p204-/- mice. Unexpectedly, p204 deficiency led to significant defect in extracellular LPS signaling in macrophages, as demonstrated by dramatic reductions of LPS-mediated IFN-ß and pro-inflammatory cytokines. The serum levels of IFN-ß and pro-inflammatory cytokines were also significantly reduced in p204-/- mice following LPS challenge. In addition, p204-/- mice were resistant to LPS-induced shock. LPS-activated NF-ĸB and IRF-3 pathways were all defective in p204-deficient macrophages. p204 binds to TLR4 through its Pyrin domain, and it is required for the dimerization of TLR4 following LPS-challenge. Collectively, p204 is a critical component of canonical LPS-TLR4 signaling pathway, and these studies also suggest that p204 could be a potential target to prevent and treat inflammatory and infectious diseases.

Influence of dissolved organic matter on dissolved vanadium speciation in the Churchill River estuary (Manitoba, Canada).

Diffusive gradients in thin films (DGT) devices were used to investigate the temporal and spatial changes in vanadium (V) speciation in the Churchill estuary system (Manitoba). Thirty-six DGT sets and 95 discrete water samples were collected at 8 river and 3 estuary sites during spring freshet and summer base flow. Dissolved V concentration in the Churchill River at summer base flow was approximately 5 times higher than those during the spring high flow (27.3 ± 18.9 nM vs 4.8 ± 3.5 nM). DGT-labile V showed an opposite trend with greater values found during the spring high flow (2.6 ± 1.8 nM vs 1.4 ± 0.3 nM). Parallel factor analysis (PARAFAC) conducted on 95 excitation-emission matrix spectra validated four humic-like (C1C4) and one protein-like (C5) fluorescent components. Significant positive relationship was found between protein-like DOM and DGT-labile V (r = 0.53, p < 0.05), indicating that protein-like DOM possibly affected the DGT-labile V concentration in Churchill River. Sediment leachates were enriched in DGT-labile V and protein-like DOM, which can be readily released when river sediment began to thaw during spring freshet.

Astaxanthin reduces matrix metalloproteinase expression in human chondrocytes.

Astaxanthin is a red carotenoid pigment which exerts multiple biological activities. However, little is known about the effects of astaxanthin on matrix metalloproteinases (MMPs) in OA. The present study investigated the effects of astaxanthin on MMPs in human chondrocytes. Human chondrocytes were pretreated with astaxanthin at 1, 10 or 50µM, then, cells were stimulated with IL-1ß (10ng/ml) for 24h. MMP-1, MMP-3 and MMP-13 were observed. We found that astaxanthin reduced the expression of MMP-1, MMP-3 and MMP-13 as well as the phosphorylation of two mitogen-activated protein kinases (MAPK) (p38 and ERK1/2) in IL-1ß-stimulated chondrocytes. Astaxanthin also blocked the IκB-α degradation. These results suggest that astaxanthin may be beneficial in the treatment of OA.

Overexpression of p204 leads to abnormal embryos and osteogenesis in zebrafish.

p204, an inteferon-inducible protein, is known to play an important role in modulating cell proliferation, cell cycling, and the differentiation of various tissues, including osteoblasts. In order to determine the role of p204 during development in vivo, the teleost zebrafish (Danio rerio), an established vertebrate model for developmental studies, was employed. p204 cDNA was introduced into zebrafish by microinjection, and p204 was ectopically expressed throughout the whole embryo during the early stages of zebrafish embryogenesis, then its expression gradually decreased, mainly in ventrally located cells and retina capsules. Importantly, overexpression of p204 in zebrafish resulted in striking malformations such as bent spine and expanded belly. Furthermore, the expressions of some genes (vent, runx2b, osn) involved in dorsoventral patterning and osteogenesis were significantly upregulated after p204 injection. This study provides not only the in vivo evidences demonstrating the role of p204 during embryonic development, but also new insights into the molecular mechanism by which p204 mediate osteogenesis.

[Reconstruction of embryo using an improved nuclear transfer method].

Previous methods used for nuclear transplantation were further investigated to develop a method that was both easy to carryout and did not require any special apparatus, such as Piezoimpact or Spindle-View. Following the puncture of zona pellucida with two holes by injection pipette that contained donor nuclei or cells, the injection pipette was pulled back to the perivitelline space while the negative pressure was increased in the holding pipette until the polar body and karyoplasm were wiped off completely. Then a reconstructed embryo was completed by the direct injection of the donor nucleus or cell without pulling out the injection pipette. 200 oocytes were manipulated using this method and it cost about 40 seconds with nucleus injection and about 30 seconds with cell injection to complete a reconstructed embryo. The success rates were 62.6% and 86. 0%, respectively, and enucleation rate was about 73.3% validated by Hoechst 33342. Using this method, the nucleus was completely eliminated and another was injected using the microscope and micromanipulator. Moreover, the efficiency of nuclear transplantation and survival rate of reconstructed embryos were greatly improved. Furthermore, it is very easy to manipulate and popularize in practice.

Factors derived from mouse embryonic stem cells promote self-renewal of goat embryonic stem-like cells.

Goat embryonic stem (ES)-like cells could be isolated from primary materials-inner cell masses (ICMs) and remain undifferentiated for eight passages in a new culture system containing mouse ES cell conditioned medium (ESCCM) and on a feeder layer of mouse embryo fibroblasts (MEFs). However, when cultured in medium without mouse ESCCM, goat ES-like cells could not survive for more than three passages. In addition, no ES-like cells could be obtained when ICMs were cultured on goat embryo fibroblasts or the primary materials-whole goat blastocysts were cultured on MEFs. Goat ES-like cells isolated from ICMs had a normal karyotype and highly expressed alkaline phosphatase. Multiple differentiation potency of the ES-like cells was confirmed by differentiation into neural cells and fibroblast-like cells in vitro. These results suggest that mouse ES cells might secrete factors playing important roles in promoting goat ES-like cells' self-renewal, moreover, the feeder layers and primary materials could also influence the successful isolation of goat ES-like cells.

Macrophage-associated erythropoiesis and lymphocytopoiesis in mouse fetal liver: ultrastructural and ISH analysis.

To elucidate the process of fetal liver hematopoiesis, the relationships between stroma and hematopoietic cells involved in maturation were investigated. Cultured mouse fetal liver explants were established for morphological analysis of the interactions between fetal liver stroma and hematopoietic cells ex vivo. Fetal liver stroma comprised epithelial cells and macrophages, which occupied most of the culture surface. Macrophages proliferated extensively in primary culture, but almost disappeared after 3 passages. Close morphological and functional relationships were established between macrophages and hemopoietic cells, whereas epithelial cells did not interact with blood cells. Scanning electron microscopy revealed that macrophages were in close contact with erythroblasts and formed a three-dimensional network. In each erythroblastic island, 2-3 lymphocytes were also in contact with the macrophages; erythroblasts, lymphocytes and macrophages formed close, firm associations through their cytoplasmic membranes. This cell orientation was confirmed by transmission electron microscopy of fetal liver in vivo. In situ hybridization revealed that the macrophages expressed jagged-1, an important ligand of the Notch signaling system in hematopoiesis.