Effects of dietary phytoncides extracted from Korean pine (Pinus koraiensis) cone on performance, egg quality, gut microflora, and immune response in laying hens.

This study was conducted to evaluate the effects of dietary phytoncides extracted from discarded Korean pine cones (Pinus koraiensis) on the performance, egg quality, immune response and gut microflora in laying hens. A total of 400 Hy-Line brown laying hens (50-week old) were allotted into four dietary treatments including a control diet or a diet supplemented with phytoncides at 0.002%, 0.004% and 0.008%. During the 6 weeks of experimental feeding, 0.008% of dietary phytoncides improved egg production, feed conversion ratio (p < 0.05), but not feed intake, egg weight or feed efficiency. Although dietary phytoncides had no effect on egg quality, decreases in Haugh units depending on storage periods were improved by 0.008% of dietary phytoncides (p < 0.05). To investigate the roles of dietary phytoncides on the alteration of the immune response during inflammation, lipopolysaccharide (LPS) or saline was intraperitoneally injected into 10 hens per diet group on the end date of the experimental feeding period. Serum immunoglobulins and splenic cytokine expression at mRNA levels were then measured at 4 hr postinjection. Although the levels of IgA were decreased by LPS injection in all dietary groups, dietary phytoncides at 0.008% showed a higher level of IgA by LPS (p < 0.05). Interestingly, although LPS injection resulted in an enhanced expression of proinflammatory cytokines such as IL-1ß and IL-6, dietary phytoncides at 0.008% showed less increased levels of them (p < 0.05). Gut microflora was examined from 10 hens per diet group at the end of the experimental period. While the number of Lactobacillus spp. was increased (p < 0.05), Escherichia coli counts in the cecal contents were decreased by 0.008% of dietary phytoncides. Taken together, these results demonstrate that dietary supplementation of 0.008% phytoncides improved the egg production, immune responses during inflammation and gut microflora in laying hens.

Induction of intercellular adhesion molecule-1 by water-soluble components of Hericium erinaceum in human monocytes.

AIM OF THE STUDY: Hericium erinaceum is a medicinal mushroom that has been traditionally used in Asian countries for the treatment of cancers and infectious diseases. Although the immunomodulating activity of H. erinaceum is considered to be responsible for its medicinal activity, its action mechanisms are poorly understood. In the present study, we investigated the capability of water-extracted H. erinaceum (WEHE) to induce the expression of intercellular adhesion molecule-1 (ICAM-1), which regulates the migration of immune cells. MATERIALS AND METHODS: THP-1, a human monocytic cell-line, or human peripheral blood mononuclear cells (PBMC) were stimulated with WEHE (0-30 µg/mL) and subsequently analyzed using flow cytometry to examine the surface expression of ICAM-1 protein. Steady-state levels of ICAM-1 mRNA were estimated using real-time reverse transcription-polymerase chain reaction analysis. Electrophoretic mobility shift assay was conducted to examine transcription factors involved in ICAM-1 transcription. RESULTS: WEHE induced ICAM-1 expression at both protein and mRNA levels in THP-1 cells in a dose- and time-dependent fashion. A similar pattern of ICAM-1 induction was also observed in CD14(+) monocytes in human PBMC that were stimulated with WEHE. The ICAM-1 expression on THP-1 cells stimulated with WEHE was suppressed by specific inhibitors for extracellular signal-regulated kinases (ERK) and reactive oxygen species (ROS). Additionally, exposure of THP-1 cells to WEHE increased the DNA binding activities of NF-κB, AP-1, SP-1 and STAT-1 transcription factors, all of which are known to be required for ICAM-1 gene expression. CONCLUSIONS: These results suggest that WEHE induces ICAM-1 expression in human monocytes through ERK- and ROS-dependent signaling pathways, resulting in the subsequent activations of NF-κB, AP-1, SP-1, and STAT-1 transcription factors.

Induction of ICAM-1 by Armillariella mellea is mediated through generation of reactive oxygen species and JNK activation.

ETHNOPHARMACOLOGICAL RELEVANCE: Armillariella mellea is an edible mushroom that has been traditionally used as an alternative medicine in many countries because of its anti-microbial and anti-cancer effects. AIM OF THE STUDY: In this study, we examined the ability of Armillariella mellea to induce the expression of intercellular adhesion molecule (ICAM)-1, an important cellular adhesion molecule for the recruitment of immune cells to regional inflammatory sites. MATERIALS AND METHODS: A human monocytic cell line, THP-1 or human peripheral blood mononuclear cells (PBMC) were stimulated with Armillariella mellea extract (AME) and subjected to flow cytometry to examine the expression of ICAM-1 protein on the cell surface. Steady-state mRNA level of ICAM-1 was determined by real-time reverse transcription-polymerase chain reaction. The phosphorylation of JNK protein was examined by Western blot analysis using antibodies specific for non-phosphorylated and phosphorylated forms of JNK. For the analysis of transcription factors regulating ICAM-1 transcription, the nuclear fraction was extracted from AME-treated THP-1 cells and subjected to electrophoretic mobility shift assay. RESULTS: AME induced expression of ICAM-1 and its mRNA in THP-1 cells in dose- and time-dependent manners. AME-induced ICAM-1 expression was also observed on CD14-positive monocytes in human PBMC. Interestingly, AME-induced ICAM-1 production was inhibited by the specific inhibitors of reactive oxygen species (ROS) and JNK, whereas no inhibitory effect was observed when inhibitors of ERK, p38 kinase, phosphatidylinositol 3-kinase, or protein kinase C were used. Concomitantly, AME increased phosphorylation of JNK in a time-dependent fashion. DNA binding activities of NF-kappaB, AP-1, SP-1, and STAT-1 were increased by AME treatment. CONCLUSION: These results suggest that AME induces ICAM-1 expression in human monocytic cells through ROS/JNK-dependent signaling pathways leading to the activation of NF-kappaB, AP-1, SP-1, and STAT-1 transcription factors.

Induction of IL-8 expression by Cordyceps militaris grown on germinated soybeans through lipid rafts formation and signaling pathways via ERK and JNK in A549 cells.

AIM OF THE STUDY: In order to elucidate immunoregulatory mechanisms of Cordyceps militaris, a methanol extract of Cordyceps militaris grown on germinated soybeans was prepared and its immunoregulatory effect in the human lung epithelial cells was investigated by examining its ability to induce IL-8 expression. MATERIALS AND METHODS: Cordyceps militaris grown on germinated soybeans was extracted with 80% methanol (GSC4M) and used for stimulation of a human lung epithelial cell-line, A549. An enzyme-linked immunosorbent assay and reverse transcription-polymerase chain reaction were performed to examine the production of IL-8 protein and its mRNA, respectively. For the analysis of transcription factors regulating IL-8 transcriptional activation, the nuclear fraction was extracted from GSC4M-treated A549 cells and subjected to electrophoretic mobility shift assay. RESULTS: GSC4M induced IL-8 protein secretion and its mRNA expression from A549 cells in a dose- and time-dependent manner. GSC4M-induced IL-8 expression was inhibited by an inhibitor for lipid rafts formation but not by that for clathrin-coated pits. In addition, signaling pathways for GSC4M-induced IL-8 expression were mediated through ERK and JNK but hardly through p38 kinase. Furthermore, GSC4M augmented the DNA-binding activity of the transcription factors AP-1, NF-IL6, and NF-kappaB, all of which are involved in the transcriptional activation of the IL-8 gene. CONCLUSION: Cordyceps militaris grown on germinated soybeans stimulates lung epithelial cells to produce IL-8 through lipid rafts formation and signaling pathways via ERK and JNK.

Armillariella mellea induces maturation of human dendritic cells without induction of cytokine expression.

ETHNOPHARMACOLOGICAL RELEVANCE: Armillariella mellea, an edible and medicinal mushroom possessing immuno-modulating potential, has been frequently used for the treatment of infectious diseases or cancers. AIM OF THE STUDY: In order to elucidate immune-regulatory mechanisms of Armillariella mellea, we investigated the effect of water-soluble components from Armillariella mellea (AME) on the regulation of human dendritic cell (DC) maturation and activation. MATERIALS AND METHODS: Immature DCs (iDCs) were prepared by differentiating human peripheral blood CD14-positive cells with GM-CSF and IL-4. Then, iDCs were treated with AME at 2-20 microg/ml for 48 h and subjected to flow cytometry to analyze the expression of DC markers. Dextran-FITC uptake assay and enzyme-linked immunosorbent assay were performed to examine the endocytic capacity of AME-stimulated DC and their production of cytokines, respectively. RESULTS: iDCs stimulated with AME showed representative features during DC maturation such as up-regulated expression of CD80, CD83, CD86, both MHC class I and II molecules, and CD205, with a simultaneous decrease in the expression of CD206 and the endocytic capacity. Interestingly, AME was not able to induce the production of TNF-alpha, IL-12p40, or IL-10, whereas lipopolysaccharides induced a substantial increase of all of the cytokines. CONCLUSION: Armillariella mellea induces maturation of human DCs through a unique mechanism without inducing cytokine expression.

Peptidoglycan-mediated IL-8 expression in human alveolar type II epithelial cells requires lipid raft formation and MAPK activation.

Staphylococcus aureus, a major sepsis-causing Gram-positive bacterium, invades pulmonary epithelial cells and causes lung diseases. In the lung, alveolar type II epithelial cells play an important role in innate immunity by secreting chemokines and antimicrobial peptides upon bacterial infection whereas type I cells mainly function in gas-exchange. In this study, we investigated the ability of S. aureus peptidoglycan (PGN) to induce expression of a chemokine, IL-8, in a human alveolar type II epithelial cell line, A549. PGN induces IL-8 mRNA and protein expression in a dose- and time-dependent manner. Supplementation of soluble CD14 further enhanced the PGN-induced IL-8 expression. Interestingly, PGN-induced IL-8 expression was inhibited by nystatin, a specific inhibitor for lipid rafts, but not by chlorpromazine, a specific inhibitor for clathrin-coated pits. Furthermore, PGN-induced IL-8 expression was attenuated by inhibitors for MAP kinases such as ERK, p38 kinase, and JNK/SAPK, whereas no inhibitory effect was observed by inhibitors for reactive oxygen species or protein kinase C. Electrophoretic mobility shift assay demonstrates that PGN increased the DNA binding of the transcription factors, AP-1 and NF-kappaB while minimally, NF-IL6, all of which are involved in the transcription of IL-8. Taken together, these results suggest that PGN induces IL-8 expression in a CD14-enhanced manner in human alveolar type II epithelial cells, through the formation of lipid rafts and the activation of MAP kinases, which ultimately leads to activation of AP-1, NF-kappaB, and NF-IL6.