Antiallergic effects of peiminine through the regulation of inflammatory mediators in HMC-1 cells.

Peiminine is the main biologically active component derived from Fritillaria ussuriensis. Peiminine was investigated in various pulmonary diseases, but its antiallergic effect and the related mechanism have not been reported yet. The present study aimed to evaluate the effect of peiminine on mast cell-mediated allergic inflammation in HMC-1 cells. The pro-inflammatory cytokine production was measured using ELISA, reverse transcription-polymerase chain reaction and nuclear factor-kappaB (NF-κB), mitogen-activated protein kinases (MAPKs) pathway activation, as determined by Western blot analysis. Peiminine inhibits the production of the pro-inflammatory cytokine, such as interleukin (IL)-6, IL-8, tumor necrosis factor-alpha (TNF-α) and IL-1beta (IL-1ß). It was shown to have inhibitory effects on MAPKs phosphorylation and NF-B expression in human mast cells (HMC)-1 using Western blot. HMC-1 cells were observed for confirmation of histamine release. Passive cutaneous anaphylaxis (PCA) reactions were evaluated using an animal model and peiminine demonstrated inhibitory effects on IgE-dependent anaphylaxis. These results suggest that peiminine has regulatory potential for allergic inflammatory reactions mediated by HMC-1 cells.

Seasonal change and subniche dynamics of three Alexandrium species in the Korea Strait.

Some members of the dinoflagellate genus Alexandrium produce toxins responsible for paralytic shellfish poisoning, which causes environmental impacts and large economic losses worldwide. The Outlying Mean Index (OMI) and the Within Outlying Mean Index (WitOMI) were used to examine the ecological niches of three Alexandrium species identifying factors affecting their population dynamics in the Korea Strait (KS). Species niches were divided into seasonal subniches based on species' temporal and spatial patterns, with A. catenella being highest in the spring, A. pacificum in the summer, and A. affine in the autumn. These shifts in abundance are likely due to changes in their habitat preferences and resource availability, as well as the effects of biological constraints. A subniche-based approach, which considers the interactions between the environment and the biological characteristics of a species, was useful in understanding the factors shaping the population dynamics of the individual species. Additionally, a species distribution model was used to predict the phenology and biogeography of the three Alexandrium species in the KS and their thermal niches on a larger scale. The model predicted that, in the KS, A. catenella exists on the warm side of the thermal niche, while A. pacificum and A. affine exist on the cold side, indicating that these species may respond differently to increases in water temperature. However, the predicted phenology was incongruent with the abundance of the species as measured by droplet digital PCR. Overall, the WitOMI analysis and species distribution model can provide valuable insights into how population dynamics are influenced by the integrated interplay of biotic and abiotic processes.

Quantification of the paralytic shellfish poisoning dinoflagellate Alexandrium species using a digital PCR.

A ubiquitous dinoflagellate, Alexandrium, produces paralytic shellfish toxin (PST), and its outbreaks have negative impacts on aquaculture, fisheries, human health, and the marine ecosystem. To minimize such damages, a routine monitoring program of toxic species must be implemented with a suitable analytical technique for their identification and quantification. However, the taxonomic identification and cell quantification of Alexandrium species based on their external morphology under a light microscope, or by using conventional molecular approaches have limited sensitivity and reproducibility. To address these challenges, we have developed an advanced protocol using droplet-digital PCR (ddPCR) for the discrimination and enumeration of three co-occurring Alexandrium species (A. affine, A. catenella, and A. pacificum) in environmental samples. Copies of species-specific internal transcribed spacer (ITS) per cell, which were calculated from environmental samples spiked with various numbers of culture cells, were used to estimate the abundance of species in the field samples. There were no significant differences in ITS copies estimated by the digital PCR assay between environmental samples from different localities, spiked artificially with a consistent number of cells from Alexandrium cultures. This sensitive assay was applied to determine the abundance and vertical distribution of those populations in the southern coastal waters of Korea. In spring, A. catenella was the dominant species, followed by the non-toxic A. affine in summers. A novel digital PCR assay can also be used to monitor other harmful marine protists that require high sample throughput and low detection limit with high accuracy and precision.

Optimized Production of Poly(γ-Glutamic acid) By Bacillus sp. FBL-2 through Response Surface Methodology Using Central Composite Design.

In the present study, the optimization of poly(γ-glutamic acid) (γ-PGA) production by Bacillus sp. FBL-2 was studied using a statistical approach. One-factor-at-a-time method was used to investigate the effect of carbon sources and nitrogen sources on γ-PGA production and was utilized to select the most significant nutrients affecting the yield of γ-PGA. After identifying effective nutrients, response surface methodology with central composite design (CCD) was used to obtain a mathematical model to identify the optimum concentrations of the key nutrients (sucrose, L-glutamic acid, yeast extract, and citric acid) for improvement of γ-PGA production. The optimum amount of significant medium components appeared to be sucrose 51.73 g/l, L-glutamic acid 105.30 g/l, yeast extract 13.25 g/l, and citric acid 10.04 g/l. The optimized medium was validated experimentally, and γ-PGA production increased significantly from 3.59 g/l (0.33 g/l/h) to 44.04 g/l (3.67 g/l/h) when strain FBL-2 was cultivated under the optimal medium developed by the statistical approach, as compared to non-optimized medium.

Effect of peiminine on DNCB-induced atopic dermatitis by inhibiting inflammatory cytokine expression in vivo and in vitro.

Peiminine (PMN) is the main component derived from Fritillaria ussuriensis and is used in traditional medicine in East Asia. The aim of this study was to evaluate the effects of PMN on atopic dermatitis (AD) induced by a dinitrochlorobenzene (DNCB) in Balb/c mice. Inflammatory cytokine expression of PMN was investigated in vitro. Eosinophil infiltration and the thickness of DNCB-induced AD mouse skin were measured. The levels of IgE, IL-4, IL-6, IL-13, and TNF-α in the serum were measured by ELISA. The effects of PMN on the transcription level of MAPK and nuclear factor (NF)-κB were evaluated in mouse skin. In addition, the inhibitory effect of TNF-α, IL-1ß, COX-2 and PGE2 were measured in RAW264.7 cells; TARC was investigated in HaCaT cells; and ß-hexosaminidase was examined in RBL-2H3 cells. PMN decreased the number of eosinophils in the dermis as well as mast cells and decreased the thickness of the epidermis and dermis. The PMN High group had a significantly reduced serum level of IgE, IL-4, IL-13 and TNF-α. Moreover, P-ERK and P-P38 were inhibited in the PMN High group compared with the DNCB-treated group. PMN additionally attenuated the expression of inflammatory cytokines in cells, including RAW264.7, HaCaT and RBL-2H3 cells. Our results suggest that PMN could be a potential therapeutic candidate for the treatment of AD.

Peimine Inhibits the Production of Proinflammatory Cytokines Through Regulation of the Phosphorylation of NF-κB and MAPKs in HMC-1 Cells.

BACKGROUND: Peimine is a major biologically active component of Fritillaria ussuriensis. Peimine was investigated in chronic inflammation response, but it has not been studied in mast cell-related immediate allergic reaction. The present study aimed to evaluate anti-allergic effect of peimine in human mast cell (HMC-1). MATERIALS AND METHODS: The effect of peimine on cell viability was measured by MTS assay in HMC-1. Histamine release was investigated in rat peritoneal mast cells (RPMCs). Interleukin (IL)-6, IL-8, and tumor necrosis factor-α (TNF-α) expressions were measured by ELISA assay and reverse transcription-polymerase chain reaction. Mitogen-activated protein kinases (MAPKs) and nuclear factor-kappaB (NF-κB) were examined by Western blot. Passive cutaneous anaphylaxis (PCA) reactions were evaluated using Sprague-Dawley (SD) rats. RESULTS: Peimine inhibited the production of pro-inflammatory cytokines, such as IL-6, IL-8, and TNF-α. Moreover, peimine reduced MAPKs phosphorylation and the nuclear NF-κB expression in PMACI-induced HMC-1. Peimine decreased PCA reactions in rats as well. CONCLUSION: Our study proved that peimine might be suitable for the treatment of mast cell-derived allergic inflammatory reactions. SUMMARY: Peimine inhibited the production of pro-inflammatory cytokines, such as IL-6, IL-8, and TNF-αPeimine reduced MAPKs phosphorylation and the nuclear NF-κB expression in PMACI-induced HMC-1Peimine decreased PCA reactions in ratsPeimine has anti-allergic effect through regulation of pro-inflammatory mechanism on mast cell. Abbreviations used: HMC-1: Human mast cell, MTS: 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, RPMCs: Rat peritoneal mast cells. IL-6: Interleukin 6, IL-8: Interleukin 8, TNF-α: Tumor necrosis factor-α, MAPKs: Mitogen-activated protein kinases; NF-κB: Nuclear factor-kappaB, PCA: Passive cutaneous anaphylaxis reactions, SD: Sprague-Dawley.

Cyperus Rotundus L. extract suppresses RANKL-induced osteoclastogenesis through NFATc1/c-fos downregulation and prevent bone loss in OVX-induced osteoporosis rat.

ETHNOPHARMACOLOGICAL RELEVANCE: Cyperus Rotundus L. (CyR) has been widely used for the treatment of gynecologic disorder. Recent studies have reported that CyR can prevent the formation of cystic follicles and ovarian malfunction. However, the effects of CyR on osteoclastogenesis and postmenopausal osteoporosis remain unknown. AIM OF THE STUDY: This study was aimed to investigate the preventive effects of CyR on RANKL-induced osteoclast formation and ovariectomy (OVX)-induced bone loss. MATERIALS AND METHODS: In this in vitro study, we investigate the anti-osteoporotic effect of CyR on receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclastogenesis, the formation of tartrate-resistant acid phosphatase (TRAP) multinucleated cells, pit formation, transcription factors such as NFATc1 and c-Fos, and mRNA expression of osteoclast-associated genes were investigated. Forty 12-weeks female Sprague-Dawley rats for in vivo effect of CyR were used and OVX rat model was determined. The rats were randomly assigned into sham group and four OVX groups, i.e. OVX with D.W; OVX with estradiol (E2, 100µg/kg/day), OVX with CyR-L (16mg/kg/day), OVX with CyR-H (160mg/kg/day). The treatment lasted for 8weeks. RESULTS: CyR inhibited osteoclast differentiation and pit formation in the RANKL-induced osteoclastogenesis of RAW 264.7 cells. Reverse transcription polymerase chain reaction analysis also showed that CyR reduced the mRNA expression of osteoclast-associated genes such as carbonic anhydrase II, TRAP, RANK, cathepsin K, matrix metalloproteinase 9, nuclear factor of activated T cells cytoplasmic 1 (NFATc1), and c-Fos. In addition, CyR decreased protein levels of NFATc1 and c-Fos. CyR inhibited trabecular bone loss in the femur caused by OVX. CONCLUSION: The results of this study indicate that CyR inhibits the RANKL-induced osteoclast differentiation in RAW 264.7 cells and trabecular bone loss in OVX rats.

The effects of Lycii Radicis Cortex on RANKL-induced osteoclast differentiation and activation in RAW 264.7 cells.

Post-menopausal osteoporosis is a serious age-related disease. After the menopause, estrogen deficiency is common, and excessive osteoclast activity causes osteoporosis. Osteoclasts are multinucleated cells generated from the differentiation of monocyte/macrophage precursor cells such as RAW 264.7 cells. The water extract of Lycii Radicis Cortex (LRC) is made from the dried root bark of Lycium chinense Mill. and is termed 'Jigolpi' in Korea. Its effects on osteoclastogenesis and post­menopausal osteoporosis had not previously been tested. In the present study, the effect of LRC on receptor activator of nuclear factor-κB (NF-κB) ligand (RANKL)-induced osteoclast differentiation was demonstrated using a tartrate-resistant acid phosphatase (TRAP) assay and pit formation assay. Moreover, in order to analyze molecular mechanisms, we studied osteoclastogenesis-related markers such as nuclear factor of activated T-cells cytoplasmic 1 (NFATc1), c-Fos, receptor activator of NF-κB (RANK), TRAP, cathepsin K (CTK), matrix metallopeptidase-9 (MMP-9), calcitonin receptor (CTR) and carbonic anhydrase Ⅱ (CAII) using RT-qPCR and western blot analysis. Additionally, we also determined the effect of LRC on an ovariectomized (OVX) rat model. We noted that LRC inhibited RANKL-induced osteoclast differentiation via suppressing osteoclastogenesis-related markers. It also inhibited osteoporosis in the OVX rat model by decreasing loss of bone density and trabecular area. These results suggest that LRC exerts a positive effect on menopausal osteoporosis.

Antiallergic effect of gami-hyunggyeyeongyotang on ovalbumin-induced allergic rhinitis in mouse and human mast cells.

BACKGROUND: Gami-hyunggyeyeongyotang (GMHGYGT) is a polyherbal medicine derived from an oriental prescription traditionally used in the treatment of allergic diseases such as allergic rhinitis (AR). This study aimed to evaluate the effects of GMHGYGT on ovalbumin (OVA) sensitization/challenge-induced AR in BALB/C mice, through examination of allergic inflammatory response regulation, as well as examination of human mast cells (HMC-1). METHODS: Nasal symptoms were evaluated in the OVA-induced allergic rhinitis mouse model, and total immunoglobulin (Ig)E and OVA-specific IgE levels in serum were investigated. Eosinophil infiltration and thickness of the nasal mucosa, and levels of interleukin (IL)-1ß and caspase-1 were also measured by immunohistochemistry. Additionally, the effect of GMHGYGT on the phorbol-12-myristate-13-acetate plus calcium ionophore A23187-induced phosphorylation of extracellular signal-regulated kinase, C-Jun N-terminal kinase and p38 in HMC-1 cells was investigated. RESULTS: GMHGYGT was demonstrated to have antiallergic effects on the nasal symptoms of the OVA-induced mouse model, decreasing serum levels of OVA-specific IgE and levels of the cytokines IL-5, IL-6, IL-1ß, monocyte chemotactic protein-1, and macrophage inflammatory protein-2. GMHGYGT reduced the number of eosinophils in the nasal mucosa and thickness of the nasal septum, and inhibited the expression of IL-1ß and caspase-1. Moreover, it inhibited the phosphorylation of extracellular signal-regulated kinase and C-Jun N-terminal kinase, as well as the activation of nuclear factor-κB on protein level in HMC-1 cells. CONCLUSION: These results suggest that GMHGYGT has therapeutic potential for the treatment of allergic rhinitis.

Xanthii fructus inhibits inflammatory responses in LPS-stimulated RAW 264.7 macrophages through suppressing NF-κB and JNK/p38 MAPK.

ETHNOPHARMACOLOGICAL RELEVANCE: Xanthii fructus (XF) has long been used to treat a variety of inflammatory conditions in Korean traditional medicine, but the underlying mechanisms that could explain the anti-inflammatory actions of XF remain largely unknown. AIM OF THE STUDY: This study aimed to elucidate the anti-inflammatory effects of X. fructus (XF) and to examine its underlying molecular mechanisms in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. MATERIALS AND METHODS: The effect of XF on LPS-induced mRNA and protein expressions of inflammatory mediators and cytokines were determined. Moreover, the activation of the nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways and the expression of heme oxygenase-1 (HO-1) were explored to elucidate the anti-inflammatory mechanisms. RESULTS: XF significantly inhibited LPS-induced production of inflammatory mediators, interleukin-6 (IL-6), nitric oxide (NO), and prostaglandin E2 (PGE2), without any cytotoxicity. However, it did not affect tissue necrosis factor (TNF)-α or IL-1ß production in LPS-stimulated RAW 264.7 cells. Expression levels of inducible nitric oxide synthase (iNOS) mRNA and protein were inhibited dose-dependently by XF in LPS-stimulated RAW 264.7 cells, but there were no changes in cyclooxygenase-2 (COX-2) mRNA and protein. XF significantly attenuated LPS-induced phosphorylation and degradation of inhibitory kappa Bα (IκBα) and consequently reduced the nuclear translocation of p65 NF-κB. Pretreatment with XF also strongly inhibited the LPS-induced phosphorylation of p38 kinase and JNK, whereas the phosphorylation of ERK1/2 was not affected. In addition, XF led to an increase in HO-1 expression. CONCLUSION: Taken together, our findings support that XF inhibits LPS-induced inflammatory responses by blocking NF-κB activation, inhibiting JNK/p38 MAPK phosphorylation, and enhancing HO-1 expression in macrophages, suggesting that it could be an attractive therapeutic candidate for various inflammatory diseases.