• 동의생리병리학회지
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• 제33권2호
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• pp.131-140
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• 2019

The objective of present study is to evaluate concentration-dependent in vitro anti-osteoarthritic (OA) effects of synergic mixed formula consisted of dried pomegranate juice concentrate powder, Eucommiae Cortex aqueous extract and Achyranthis Radix aqueous extract 5:4:1 (g/g) mixture on the primary cultured rat articular chondrocytes. First, any cytotoxic effect of mixture was observed using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium Bromide) assay. Next, cyto-protective effect of test substances was evaluated by using the recombinant human interleukin $(rhIL)-1{\alpha}$ induced chondrocytes. In addition, anti-inflammatory effects were also observed on the lipopolysaccaride (LPS) treated chondrocytes through prostaglandin $E_2(PGE_2)$ productions and 5-lipoxygenase (LPO) activities, and inhibitory effects on matrix metalloproteinase (MMP)-2 and MMP-9 activities were observed on $rhIL-1{\alpha}$ treated chondrocytes with their extracellular matrix (ECM) related mRNA expressions. No obvious cytotoxic effects of mixture were demonstrated. Inflammatory damages of chondrocytes and related ECM degradations induced by treatment of LPS or $rhIL-1{\alpha}$ were significantly and concentration-dependently inhibited by pretreatment of mixture from a concentration level of 0.001 mg/ml to 1 mg/ml. In addition, mixture showed $IC_{50}$ for $rhIL-1{\alpha}-induced$ MMP-2 and MMP-9 activities as 44.01 and $162.47{\mu}g/ml$, and also showed $EC_{50}$ for $rhIL-1{\alpha}-induced$ inhibition of collagen type II, SOX9 and aggrecan mRNA expression as 8.61, 10.79 and $4.47{\mu}g/ml$, respectively. It is observed that mixture showed concentration-dependent anti-inflammatory and cytoprotective ECM preserved effects on the primary cultured rat articular chondrocytes without cytotoxicity.

• Journal of Ginseng Research
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• 제45권3호
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• pp.433-441
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• 2021

Background: Multiple sclerosis (MS) and its animal model, the experimental autoimmune encephalomyelitis (EAE), are primarily characterized as dysfunction of the blood-brain barrier (BBB). Ginsenoside-Rg3-enriched Korean Red Ginseng extract (Rg3-KRGE) is known to exert neuroprotective, anti-inflammatory, and anti-oxidative effects on neurological disorders. However, effects of Rg3-KRGE in EAE remain unclear. Methods: Here, we investigated whether Rg3-KRGE may improve the symptoms and pathological features of myelin oligodendroglial glycoprotein (MOG)_35-55 peptide - induced chronic EAE mice through improving the integrity of the BBB. Results: Rg3-KRGE decreased EAE score and spinal demyelination. Rg3-KRGE inhibited Evan's blue dye leakage in spinal cord, suppressed increases of adhesion molecule platelet endothelial cell adhesion molecule-1, extracellular matrix proteins fibronection, and matrix metallopeptidase-9, and prevented decreases of tight junction proteins zonula occludens-1, claudin-3, and claudin-5 in spinal cord following EAE induction. Rg3-KRGE repressed increases of proinflammatory transcripts cyclooxygenase-2, inducible nitric oxide synthase, interleukin (IL)-1 beta, IL-6, and tumor necrosis factor-alpha, but enhanced expression levels of anti-inflammatory transcripts arginase-1 and IL-10 in the spinal cord following EAE induction. Rg3-KRGE inhibited the expression of oxidative stress markers (MitoSOX and 4-hydroxynonenal), the enhancement of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2) and NOX4, and NADPH activity in the spinal cord of chronic EAE mice. Furthermore, apocynin, a NOX inhibitor, mimicked beneficial effects of Rg3-KRGE in chronic EAE mice. Conclusion: Our findings suggest that Rg3-KRGE might alleviate behavioral symptoms and pathological features of MS by improving BBB integrity through modulation of NOX2/4 expression.

• Clinical and Experimental Reproductive Medicine
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• 제50권1호
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• pp.44-52
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• 2023

Objective: The DNA integrity of spermatozoa that attach to fallopian tube (FT) cells is higher than spermatozoa that do not attach. FT epithelial cells can distinguish normal and abnormal sperm chromatin. This study investigated the effects of sperm with a high-DNA fragmentation index (DFI) from men with unexplained repeated implantation failure (RIF) on the Toll-like receptor (TLR) signaling pathway in human FT cells in vitro. Methods: Ten men with a RIF history and high-DFI and 10 healthy donors with low-DFI comprised the high-DFI (>30%) and control (<30%) groups, respectively. After fresh semen preparation, sperm were co-cultured with a human FT epithelial cell line (OE-E6/E7) for 24 hours. RNA was extracted from the cell line and the human innate and adaptive immune responses were tested using an RT2 profiler polymerase chain reaction (PCR) array. Results: The PCR array data showed significantly higher TLR-1, TLR-2, TLR-3, TLR-6, interleukin 1α (IL-1α), IL-1β, IL-6, IL-12, interferon α (IFN-α), IFN-β, tumor necrosis factor α (TNF-α), CXCL8, GM-CSF, G-CSF, CD14, ELK1, IRAK1, IRAK2, IRAK4, IRF1, IRF3, LY96, MAP2K3, MAP2K4, MAP3K7, MAP4K4, MAPK8, MAPK8IP3, MYD88, NFKB1, NFKB2, REL, TIRAP, and TRAF6 expression in the high-DFI group than in the control group. These factors are all involved in the TLR-MyD88 signaling pathway. Conclusion: The MyD88-dependent pathway through TLR-1, TLR-2, and TLR-6 activation may be one of the main inflammatory pathways activated by high-DFI sperm from men with RIF. Following activation of this pathway, epithelial cells produce inflammatory cytokines, resulting in neutrophil infiltration, activation, phagocytosis, neutrophil extracellular trap formation, and apoptosis.

• Molecules and Cells
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• 제45권3호
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• pp.134-147
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• 2022

The anti-oxidant enzyme heme oxygenase-1 (HO-1) is known to exert anti-inflammatory effects. From a library of pyrazolo[3,4-d]pyrimidines, we identified a novel compound KKC080096 that upregulated HO-1 at the mRNA and protein levels in microglial BV-2 cells. KKC080096 exhibited anti-inflammatory effects via suppressing nitric oxide, interleukin1β (IL-1β), and iNOS production in lipopolysaccharide (LPS)-challenged cells. It inhibited the phosphorylation of IKK and MAP kinases (p38, JNK, ERK), which trigger inflammatory signaling, and whose activities are inhibited by HO-1. Further, KKC080096 upregulated anti-inflammatory marker (Arg1, YM1, CD206, IL-10, transforming growth factor-β [TGF-β]) expression. In 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridinetreated mice, KKC080096 lowered microglial activation, protected the nigral dopaminergic neurons, and nigral damage-associated motor deficits. Next, we elucidated the mechanisms by which KKC080096 upregulated HO-1. KKC080096 induced the phosphorylation of AMPK and its known upstream kinases LKB1 and CaMKKbeta, and pharmacological inhibition of AMPK activity reduced the effects of KKC080096 on HO-1 expression and LPS-induced NO generation, suggesting that KKC080096-induced HO-1 upregulation involves LKB1/AMPK and CaMKKbeta/AMPK pathway activation. Further, KKC080096 caused an increase in cellular Nrf2 level, bound to Keap1 (Nrf2 inhibitor protein) with high affinity, and blocked Keap1-Nrf2 interaction. This Nrf2 activation resulted in concurrent induction of HO-1 and other Nrf2-targeted antioxidant enzymes in BV-2 and in dopaminergic CATH.a cells. These results indicate that KKC080096 is a potential therapeutic for oxidative stress-and inflammation-related neurodegenerative disorders such as Parkinson's disease.

• 생명과학회지
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• 제26권4호
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• pp.431-438
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• 2016

TRAIL은 정상세포에서는 세포독성을 나타내지 않는 반면, 암세포에서는 사멸을 유도하므로 항암제로 각광받고 있지만 많은 암세포에서 TRAIL에 저항성을 가지고 있는 것으로 알려져 있으므로 이를 극복해야하는 큰 어려움이 남아있다. 본 연구에서는 TRAIL에 저항성을 가지는 인간 방광암 세포주인 5637 세포를 이용하여 histone deacetylase 억제제인 sodium butyrate (SB)와 TRAIL을 혼합처리하였을 경우 유발되는 세포사멸 효과와 이와 관련된 분자생물학적 메카니즘을 연구하였다. 세포독성이 없는 조건의 TRAIL과 SB를 혼합처리 하였을 경우 SB 단독처리군 보다 세포사멸이 현저하게 증가하는 것으로 확인되었다. TRAIL과 SB의 혼합처리는 caspases (caspase-3, -8 and -9)의 활성화 및 PARP의 단편화를 유발하였다. 하지만 caspase 억제제에 의하여 TRAIL과 SB의 혼합처리에 의하여 유발되는 apoptosis가 현저하게 억제되는 것으로 나타났다. 또한 TRAIL과 SB의 혼합처리는 세포표면에 존재하는 DR5의 발현 증가 및 c-FLIP의 발현 감소를 유발하였으며, pro-apoptotic protein인 Bax와 세포질 cytochrome c의 발현 증가 및 anti- apoptotic protein인 Bcl-xL의 발현감소와 함께 tBid의 형성을 유발하였다. 이는 SB와 TRAIL의 혼합처리가 안전하고 선택적으로 TRAIL에 저항성을 가지는 방광암 세포에서 치료하는데 효과적인 전략임을 제시하는 결과이다.

• Journal of Periodontal and Implant Science
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• 제23권1호
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• pp.109-126
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• 1993

Refractory periodontitis manifest progressive attachment loss in a rapid and unrelenting manner regardless of the type or frequency of therapy applied. The purpose of this study was ta evaluate the relation between the level of cytokines in GCF and periodontopathic microflora with disease activity of refractory periodontitis. Selection of patients with refractory periodontitis (7 males, 3 females) were made by long term clinical observation including conventional clinical history and parameters. Teeth that showed pocket depth greater than 6mm were selected as sample teeth. Subjects were examined at baseline and after 3 months. Prior to baseline test, individual acrylic stent was fabricated. Reference grooves were made on each sample tooth site. Pocket depth and attachment loss were measured by Florida Probe. Gingival index was measured at 4 sites each sample teeth. Disease activity was defined as attachment loss of ${\ge}$ 2.1mm, as determined by sequential probing and tolerance method. The pattern and amount of alveolar bone resorption was observed with quantitative digital subtraction image processing radiography. Morphological analysis of subgingival bacteria was taken by phase contrast microscopy. Predominant cultivable bacterial distribution and frequency were compared between disease-active and disease-inactive site using immunofluorescence microscopy and selective microbial culturing. Levels of $interleukin-l{\beta}$, 2, 4, 6 and $TNF-{\alpha}$ in GCF and blood serum sample were quantified by ELISA. In active sites, P. intermedia was significantly increased to compare with inactive site. $IL-1{\beta}$, IL-2, IL-6 and $TNF-{\alpha}$ in GCF were increased in active sites and IL-2 in serum was increased in active patients significantly. Alveolar bone loss in active site was correlated with $IL-1{\beta}$, IL-2 in GCF. And loss of attachment in active site was correlated with IL-2 in GCF. These results demonstrate that IL-2 in serum, $IL-1{\beta}$, IL-2, IL-6 and $TNF-{\alpha}$ in GCF, P, intermedia might be used as possible predictors of disease activity in refractory periodontitis before it is clinically expressed as attachment loss and quantitative alveolar bone change.

• 대한약침학회지
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• 제20권2호
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• pp.93-99
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• 2017

Objectives: Selaginella bryopteris L. (family: Selaginaceae), is often used in traditional Indian systems of medicine for the prevention and cure of several disorders and for the treatment of patient with spermatorrhoea, venereal disease, constipation, colitis, urinary tract infections, fever, epilepsy, leucorrhoea, beri-beri and cancer. It is also used as a strength tonic. This study aimed to evaluate the mechanisms underlying the anti-inflammatory effects of topically administered aqueous, polar and non-polar methanolic fractions ($10mg/20{\mu}L$) of Selaginella bryopteris. Methods: An acute oral toxicity study of Selaginella bryopteris at doses from 250 to 2,000 mg/kg body weight (bw) was performed. Aqueous, polar and non-polar methanolic extracts ($10mg/20{\mu}L$) applied topically for 5 days were evaluated for their anti-inflammatory effects against 12-tetra-O-decanoyl phorbol acetate (TPA)- induced inflammation by using the redness in the ear, the ear's weight (edema), oxidative stress parameters, such as lipid-peroxide (LPO) and nitric oxide (NO), and the pro-inflammatory cytokines involved in inflammation, such as tumour necrosis factor $(TNF)-{\alpha}$, interleukin $(IL)-1{\beta}$ and IL-6. Indomethacine ($0.5mg/20{\mu}L$) was used for the positive control. Results: Selaginella bryopteris produced no mortalities when administered orally at doses from 250 to 2,000 mg/kg bw. Topical treatment with the non-polar methanolic fraction ($10mg/20{\mu}L$) significantly suppressed redness ($2.4{\pm}0.5$) and edema ($30.4{\pm}1$) and effectively reduced the LPO level ($32.3{\pm}3.3$). The NO level was ($8.07{\pm}0.55$), and the $TNF-{\alpha}$, $IL-1{\beta}$, and IL-6 levels were decreased to $69.6{\pm}15.5$, $7.7{\pm}4.8$ and $82.6{\pm}5.9$, respectively. Conclusion: This study demonstrated for the first time the mechanisms underlying the anti-inflammatory effect of medicinal plants like Selaginella bryopteris and quantified the pharmacological interactions between them. The present study showed this herbal product to be a promising anti-inflammatory phytomedicine for the treatment of patients with inflammatory skin diseases.

• Toxicological Research
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• 제34권1호
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• pp.65-73
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• 2018

Pretreatment of low-dose lipopolysaccharide (LPS) induces a hyporesponsive state to subsequent secondary challenge with high-dose LPS in innate immune cells, whereas super-low-dose LPS results in augmented expression of pro-inflammatory cytokines. However, little is known about the difference between super-low-dose and low-dose LPS pretreatments on immune cell-mediated inflammatory and hepatic acute-phase responses to secondary LPS. In the present study, RAW 264.7 cells, EL4 cells, and Hepa-1c1c7 cells were pretreated with super-low-dose LPS (SL-LPS: 50 pg/mL) or low-dose LPS (L-LPS: 50 ng/mL) in fresh complete medium once a day for 2~3 days and then cultured in fresh complete medium for 24 hr or 48 hr in the presence or absence of LPS ($1{\sim}10{\mu}g/mL$) or concanavalin A (Con A). SL-LPS pretreatment strongly enhanced the LPS-induced production of tumor necrosis factor (TNF)-${\alpha}$, interleukin (IL)-6, TNF-${\alpha}$/IL-10, prostaglandin E2 ($PGE_2$), and nitric oxide (NO) by RAW 264.7 cells compared to the control, whereas L-LPS increased IL-6 and NO production only. SL-LPS strongly augmented the Con A-induced ratios of interferon (IFN)-${\gamma}$/IL-10 in EL4 cells but decreased the LPS-induced ratios of IFN-${\gamma}$/IL-10 compared to the control, while L-LPS decreased the Con A- and LPS-induced ratios of IFN-${\gamma}$/IL-10. SL-LPS enhanced the LPS-induced production of IL-6 by Hepa1c1c-7 cells compared to the control, while L-LPS increased IL-6 but decreased IL-$1{\beta}$ and C reactive protein (CRP) levels. SL-LPS pretreatment strongly enhanced the LPS-induced production of TNF-${\alpha}$, IL-6, IL-10, $PGE_2$, and NO in RAW 264.7 cells, and the IL-6, IL-$1{\beta}$, and CRP levels in Hepa1c1c-7 cells, as well as the ratios of IFN-${\gamma}$/IL-10 in LPS- and Con A-stimulated EL4 cells compared to L-LPS. These findings suggest that pre-conditioning of SL-LPS may contribute to the mortality to secondary infection in sepsis rather than pre-conditioning of L-LPS.

• 한국약용작물학회지
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• 제26권1호
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• pp.42-54
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• 2018

Background: Allergic diseases like such as atopic dermatitis, asthma, and rhinitis have recently increased both domestically and globally. The present study was undertaken to select candidates with anti-allergic activity from plant resources. Methods and Results: Fifty-six plant extracts at $20{\mu}g/m{\ell}$ were screened against ${\beta}$-hexosaminidase production and interleukin (IL)-4 release in degranulated rat basophilic leukemia (RBL)-2H3 cells. The anti-allergy activity of three plant extracts selected from the preliminary screening experiment, Polygonatum sibiricum F. Delaroche (root), Pyrus pytifolia var. culta (Makino) Nakai (leaf), and Rehmannia glutinosa (Gaertn.) Libosch. ex Steud (root) were measured at concentrations of $2-250{\mu}g/m{\ell}$ in three cell lines as RBL-2H3, HaCaT and Jurkcat T cells. The assay showed the root extract of R. glutinosa to have an inhibitory activity of 4.2% - 28.6% on ${\beta}$-hexosaminidase production from IgE-sensitized RBL-2H3 cells. Each extract of P. sibiricum and R. glutinosa reduced IL-4 release in IgE-sensitized RBL-2H3 cells, respectively. The leaf extract of P. pyrifolia var. culta showed a significantly potent suppressive effect of 10.2% - 74.7% on the mRNA expression of tumor necrosis factor (TNF)-${\alpha}$ in HaCaT cells sensitized with TNF-a and INF-g, and showed inhibitory effect of -8.6% - 90.9% on the mRNA expression of IL-2 in Jurkat T cells sensitized with PMA and A23187. Conclusions: The results showed that the root of R. glutinosa and leaf of P. pyrifolia var. culta could be useful candidates as antiallergy materials.

• BMB Reports
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• 제45권2호
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• pp.79-84
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• 2012

In asthma, T helper 2 (TH2)-type cytokines such as interleukin (IL)-4, IL-5, and IL-13 are produced by activated $CD^{4+}$ T cells. Dendritic cells played an important role in determining the fate of naive T cells into either $T_H1$ or $T_H2$ cells. We determined whether RG-II regulates the $T_H1/T_H2$ immune response by using an ovalbumin-induced murine model of asthma. RG-II reduced IL-4 production but increased interferon-gamma production, and inhibited GATA-3 gene expression. RG-II also inhibited asthmatic reactions including an increase in the number of eosinophils in bronchoalveolar lavage fluid, an increase in inflammatory cell infiltration in lung tissues, airway luminal narrowing, and airway hyperresponsiveness. This study provides evidence that RG-II plays a critical role in ameliorating the pathogenic process of asthmatic inflammation in mice. These findings provide new insights into the immunotherapeutic role of RG-II in terms of its effects in a murine model of asthma.