• Journal of Neurogastroenterology and Motility
• /
• 제24권4호
• /
• pp.628-642
• /
• 2018

Background/Aims A Subset of patients with irritable bowel syndrome (IBS) may have mild inflammation due to immune activation. Toll-like receptors (TLRs) and cytokines may cause intestinal inflammation. We studied their expression in relation to gut microbiota. Methods Expression of TLRs and cytokines was assessed in 47 IBS patients (Rome III) and 25 controls using quantitative real-time polymerase chain reaction. Immunohistochemistry was further performed to confirm the expression of TLR-4 and TLR-5. Results Of 47 patients with IBS, 20 had constipation (IBS-C), 20 diarrhea (IBS-D), and 7 unclassified (IBS-U). The mRNA levels of TLR-4 and TLR-5 were up-regulated in IBS patients than controls (P = 0.013 and P < 0.001, respectively). Expression of TLR-4 and TLR-5 at protein level was 4.2-folds and 6.6-folds higher in IBS-D than controls. The mRNA levels of IL-6 (P = 0.003), C-X-C motif chemokine ligand 11 (CXCL-11) (P < 0.001) and C-X-C motif chemokine receptor 3 (CXCR-3) (P < 0.001) were higher among IBS patients than controls. Expression of IL-6 (P = 0.002), CXCL-11 (P < 0.001), and CXCR-3 (P < 0.001) were up-regulated and IL-10 (P = 0.012) was down-regulated in IBS-D patients than controls. Positive correlation was seen between TLR-4 and IL-6 (P = 0.043), CXCR-3, and CXCL-11 (P = 0.047), and IL-6 and CXCR-3 (P = 0.003). Stool frequency per week showed positive correlation with mRNA levels of TLR-4 (P = 0.016) and CXCR-3 (P = 0.005), but inversely correlated with IL-10 (P = 0.002). Copy number of Lactobacillus (P = 0.045) and Bifidobacterium (P = 0.011) showed correlation with IL-10 in IBS-C, while Gram-positive (P = 0.031) and Gram-negative bacteria (P = 0.010) showed correlation with CXCL-11 in IBS-D patients. Conclusions Altered immune activation in response to dysbiotic microbiota may promote intestinal inflammation in a subset of patients with IBS.

• The Korean Journal of Physiology and Pharmacology
• /
• 제13권6호
• /
• pp.475-482
• /
• 2009

Rifampicin is a macrocyclic antibiotic which is used extensively for treatment against Mycobacterium tuberculosis and other mycobacterial infections. Recently, a number of studies have focused on the immune-regulatory effects of rifampicin. Therefore, we hypothesized that rifampicin may influence the TLR2 expression in LPS-activated RAW 264.7 cells. In this study, we determined that rifampicin suppresses LPS-induced TLR2 mRNA expression. The down-regulation of TLR2 expression coincided with decreased production of TNF-$\alpha$ Since NF-${\kappa}B$ is a major transcription factor that regulates genes for TLR2 and TNF-$\alpha$, we examined the effect of rifampicin on the LPS-induced NF-${\kappa}B$ activation. Rifampicin inhibited NF-${\kappa}B$ DNA-binding activity in LPS-activated RAW 264.7 cells, while it did not affect IKK$\alpha/\beta$ activity. However, rifampicin slightly inhibited the nuclear translocation of NF-${\kappa}B$ p65. In addition, rifampicin increased physical interaction between pregnane X receptor, a receptor for rifampicin, and NF-${\kappa}B$ p65, suggesting pregnane X receptor interferes with NF-${\kappa}B$ binding to DNA. Taken together, our results demonstrate that rifampicin inhibits LPS-induced TLR2 expression, at least in part, via the suppression of NF-${\kappa}B$ DNA-binding activity in RAW 264.7 cells. Thus, the present results suggest that the rifampicin-mediated inhibition of TLR2 via the suppression of NF-${\kappa}B$ DNA-binding activity may be a novel mechanism of the immune-suppressive effects of rifampicin.

• Journal of Microbiology and Biotechnology
• /
• 제29권6호
• /
• pp.863-876
• /
• 2019

Farm animals such as piglets are often affected by environmental stress, which can disturb the gut ecosystem. Antibiotics were commonly used to prevent diarrhea in weaned piglets, but this was banned by the European Union due to the development of antibiotic resistance. However, the use of probiotics instead of antibiotics may reduce the risk posed by pathogenic microorganisms and reduce the incidence of gastrointestinal diseases. Therefore, this study was conducted to investigate the effects of Lactobacillus casei Zhang on the mechanical barrier and immune function of early-weaned piglets infected using Escherichia coli K88 based on histomorphology and immunology. Fourteen-day-old weaned piglets were divided into a control group and experimental groups that were fed L. casei Zhang and infected with E. coli K88 with or without prefeeding and/or postfeeding of L. casei Zhang. The L. casei Zhang dose used was $10^7CFU/g$ diet. Jejunum segments were obtained before histological, immunohistochemical, and western blot analyses were performed. In addition, the relative mRNA expression of toll receptors and cytokines was measured. Piglets fed L. casei Zhang showed significantly increased jejunum villus height, villus height-crypt depth ratio, muscle thickness, and expression of proliferating cell nuclear antigen and tight junction proteins ZO-1 and occludin. The use of L. casei Zhang effectively reduced intestinal inflammation after infection. We found that L. casei Zhang feeding prevented the jejunum damage induced by E. coli K88, suggesting that it may be a potential alternative to antibiotics for preventing diarrhea in early-weaned piglets.

• Asian Pacific Journal of Cancer Prevention
• /
• 제14권4호
• /
• pp.2295-2299
• /
• 2013

Background: To explore the role of caveolin-1(CAV-1) gene silencing on MAPK activation in lipopolysaccharide (LPS)-challenged human mammary epithelial cells. Methods: We established a MCF-10ACE of CAV-1 gene silencing from human mammary epithelial cell line MCF-10A by RNAi technology. DNA Microarray were used to detect the expression of inflammation-associated genes in MCF10ACE. Western blotting was used to examine the activation of MAPK in lipopolysaccharide(LPS)-challenged MCF-10A and MCF-10ACE. Moreover, immunofluorescence and Western bloting were performed to detect the co-localization of CAV-1 and toll-like receptor 4 (TLR4) in human mammary epithelial cells. Results: MCF-10ACE exhibited significant increases in inflammation-associated gene expression, especially IL-6 (~7-fold) and IL6R (~17-fold). In addition, LPS-induced p38 MAPK and JNK MAPK activation was significantly increased in MCF-10ACE. Furthermore, CAV-1 co-localized with TLR4 and appeared a negative correlation trend. Conclusion: CAV-1 gene silencing promotes MAPK activation via TLR4 signaling in human mammary epithelial cells response to LPS.

• 대한의생명과학회지
• /
• 제19권2호
• /
• pp.168-172
• /
• 2013

Toll-like receptors (TLRs) play an important role for host defense against invading pathogens. TLR4 has been identified as the receptor for lipopolysaccharide (LPS), which is a cell wall component of gram-negative bacteria. The activation of TLR4 signaling by LPS leads to the activation of NF-${\kappa}B$ and the expression of pro-inflammatory gene products such as cytokines, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2). To evaluate the therapeutic potential of (E)-1-(2-(2-nitrovinyl)phenyl)pyrrolidine (NVPP), previously synthesized in our laboratory, NF-${\kappa}B$ activation and iNOS and COX-2 expression induced by LPS were examined. NVPP inhibited the activation of NF-${\kappa}B$ induced by LPS. NVPP also suppressed the iNOS expression induced by LPS but it did not suppress COX-2 expression induced by LPS. These results suggest that NVPP has the specific mechanism for anti-inflammatory responses.

• 동의생리병리학회지
• /
• 제36권1호
• /
• pp.28-34
• /
• 2022

Echinacea purpurea (Asteraceae family) is widely used in the European countries and the United States due to its proven immune enhancement and anti-inflammatory effects. Echinacea purpurea has been reported prevent and treat upper respiratory tract infections and common cold, but the underlying molecular mechanisms are not well understood. In the present study, we examined the anti-inflammatory effects and molecular mechanisms of Echinacea purpurea (EP) extract using lipopolysaccharide (LPS)-stimulated signal pathways in RAW264.7 cells. Our results suggest that EP extract exerts anti-inflammatory effects by down-regulating the expression of LPS-induced toll-like receptor 4 (TLR4), subsequently inhibiting the activation of nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK) signaling pathways and suppression of the release of pro-inflammatory cytokines. These results suggest that EP extract is a potential therapeutic agent for inflammatory diseases.

• Biomolecules & Therapeutics
• /
• 제17권3호
• /
• pp.276-281
• /
• 2009

Lactic acid bacteria (LAB) are considered as probiotics with immunostimulatory property. In this study, we investigated the molecular mechanism of its immunostimulating potency on macrophages using combined preparation of LAB (cpLAB). cpLAB is able to strongly stimulate nitric oxide (NO) production as well as inducible NO synthase (iNOS) expression from macrophage-like RAW264.7 cells. The cpLAB-induced NO release seemed to be mediated by extracellular signal-regulated kinase (ERK) but not p38 and C-Jun N-terminal kinase (JNK), since U0126, an ERK inhibitor, clearly suppressed NO production. cpLAB significantly diminished the binding of toll like receptor (TLR)-2 antibody up to 25%, implying that cpLAB-mediated activation of macrophages may be required for the functional activation of TLR-2, but not TLR-4. Therefore, our data suggest that cpLAB may directly allow macrophages to immunostimulating potency via activation of TLR-2 and ERK.

• International Journal of Oral Biology
• /
• 제33권3호
• /
• pp.77-82
• /
• 2008

Innate immune response is initiated by the recognition of unique microbial molecular patterns through pattern recognition receptors (PRRs). The purpose of this study is to dissect the expression of various PRRs in gingival epithelial cells of differentiated versus undifferentiated states. Differentiation of immortalized human gingival epithelial HOK-16B cells was induced by culture in the presence of high $Ca^{2+}$ at increased cell density. The expression levels of various PRRs in HOK-16B cells were examined by realtime reverse transcription polymerase chain reaction (RTPCR) and flow cytometry. In addition, the expression of human beta defensins (HBDs) was examined by real time RT-PCR and the amounts of secreted cytokines were measured by enzyme linked immunosorbent assay. In undifferentiated HOK-16B cells, NACHT-LRR-PYDcontaining protein (NALP) 2 was expressed most abundantly, and toll like receptor (TLR) 2, TLR4, nucleotide-binding oligomerization domain (NOD) 1, and NOD2 were expressed in substantial levels. However, TLR3, TLR7, TLR8, TLR9, ICE protease-activating factor (IPAF), and NALP6 were hardly expressed. In differentiated cells, the levels of NOD2, NALP2, and TLR4 were different from those in undifferentiated cells at RNA but not at protein levels. Interestingly, differentiated cells expressed the increased levels of HBD-1 and -3 but secreted reduced amount of IL-8. In conclusion, the repertoire of PRRs expressed by gingival epithelial cells is limited, and undifferentiated and differentiated cells express similar levels of PRRs.

• 한국식품영양학회지
• /
• 제24권2호
• /
• pp.268-272
• /
• 2011

전곡류의 섭취와 만성질환의 유병율은 음의 상관관계가 있는 것으로 알려져 있다. 본 연구에서는 선행 연구 결과, 지방축적억제능이 우수한 소재로 선정된 기장의 항염증능 여부를 검증하고자 하였다. 이를 위해 RAW264.7 세포에 기장열수분획($1\;{\mu}g/m\ell$ 및 $10\;{\mu}g/m\ell$)과 lipopolysaccharide(LPS)를 함께 처리한 후, 24시간 배양시켜 염증매개인자들의 분비량 및 mRNA 발현 정도를 측정하였다. 또한 LPS 자극에 대한 첫 번째 신호전달인자로 알려져 있는 interleukin-1 receptor associated kinase-4(IRAK-4)의 단백질 발현 정도를 측정하였다. 본 연구결과, 기장의 열수분획($10\;{\mu}g/m\ell$)은 LPS로 유도된 NO, $PGE_2$, TNF-${\alpha}$, IL-6 및 MCP-1의 생성량 및 mRNA 발현량을 유의적으로 억제하였다(p<0.05). 특히 이들 지표 중 pro-inflammatory cytokine인 TNF-${\alpha}$와 IL-6의 mRNA 발현량이 효과적으로 감소하였다(p<0.01). IRAK-4의 단백질 발현량 또한 유의적으로 감소하여 LPS 자극에 대한 기장열수분획의 항염증능은 toll-like receptor(TLR)를 통한 IRAK-4를 매개로 하는 신호전달체계 조절에 기인하는 것으로 사료된다.

• Journal of Microbiology and Biotechnology
• /
• 제23권7호
• /
• pp.1023-1030
• /
• 2013

Lipoteichoic acid (LTA), uniquely expressed on gram-positive bacteria, is recognized by Toll-like receptor 2 (TLR2) on not only antigen-presenting cells but also activated T cells. Therefore, it is reasonable to assume that LTA is acting on T cells. However, little is known about the effect of LTA on T-cell regulation. In the present study, we investigated the immunomodulatory effects of LTA on $CD4^+$ T cells. Effector $CD4^+$ T cells, induced after co-culture with S. aureus-pulsed dendritic cells, produced high levels of interferon-${\gamma}$, CD25, CD69, and TLRs 2 and 4. When effector $CD4^+$ T cells were treated with LTA, the expressions of the membrane-bound form of transforming growth factor (TGF)-${\beta}$ and forkhead box P3 increased. Coincidently, the proliferation of effector $CD4^+$ T cells was declined after LTA treatment. When TGF-${\beta}$ signaling was blocked by the TGF-${\beta}$ receptor 1 kinase inhibitor, LTA failed to suppress the proliferation of effector $CD4^+$ T cells. Therefore, the present results suggest that LTA suppresses the activity of effector $CD4^+$ T cells by enhancing TGF-${\beta}$ production.