• Allergy, Asthma & Immunology Research
• /
• v.10 no.6
• /
• pp.698-715
• /
• 2018

Purpose: Hrd1 has recently emerged as a critical regulator of B-cells in autoimmune diseases. However, its role in the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNP) remains largely unexplored. This study aimed to examine Hrd1 expression and B-cell accumulation and their possible roles in CRSwNP. Methods: Quantitative real-time polymerase chain reaction, immunohistochemistry, enzyme-linked immunosorbent assay and Western blotting were used to assess gene and protein expression in nasal tissue extracts. Cells isolated from nasal tissues and peripheral blood mononuclear cells were characterized by flow cytometry. Local antibody production was measured in tissue extracts with a Bio-Plex assay. Additionally, changes in Hrd1 expression in response to specific inflammatory stimuli were measured in cultured dispersed polyp cells. Results: Nasal polyps (NPs) from patients with eosinophilic CRSwNP (ECRS) had increased levels of Hrd1, B-cells and plasma cells compared with NPs from patients with non-eosinophilic CRSwNP (non-ECRS) or other control subjects (P < 0.05). The average Hrd1 levels in B-cells in NPs from ECRS patients were significantly higher than those from non-ECRS patients and control subjects (P < 0.05). NPs also contained significantly increased levels of several antibody isotypes compared with normal controls (P < 0.05). Interestingly, Hrd1 expression in cultured polyp cells from ECRS patients, but not non-ECRS patients, was significantly increased by interleukin-$1{\beta}$, lipopolysaccharide and Poly(I:C) stimulation, and inhibited by dexamethasone treatment (P < 0.05). Conclusions: Differential Hrd1 expression and B-cell accumulation between the ECRS and non-ECRS subsets suggests that they can exhibit distinct pathogenic mechanisms and play important roles in NP.

• Tuberculosis and Respiratory Diseases
• /
• v.45 no.4
• /
• pp.846-860
• /
• 1998

Background: Endotoxin (LPS : lipopolysaccharide), a potent activator of immune system, can induce acute and chronic inflammation through the production of cytokines by a variety of cells, such as monocytes, endothelial cells, lymphocytes, eosinophils, neutrophils and fibroblasts. LPS stimulate the mononucelar cells by two different pathway, the CD14 dependent and independent way, of which the former has been well documented, but not the latter. LPS binds to the LPS-binding protein (LBP), in serum, to make the LPS-LBP complex which interacts with CD14 molecules on the mononuclear cell surface in peripheral blood or is transported to the tissues. In case of high concentration of LPS, LPS can stimulate directly the macrophages without LBP. We investigated to detect the generation of proinflammatory cytokines such as interleukin 1 (IL-1), IL-6 and TNF-$\alpha$ and fibrogenic cytokine, TGF-$\beta$, by peripheral blood mononuclear cells (PBMC) after LPS stimulation under serum-free conditions, which lacks LBPs. Methods : PBMC were obtained by centrifugation on Ficoll Hypaque solution of peripheral venous bloods from healthy normal subjects, then stimulated in the presence of LPS (0.1 ${\mu}g/mL$ to 100 ${\mu}g/mL$ ). The activities of IL-1, IL-6, TNF, and TGF-$\beta$ were measured by bioassaies using cytokines - dependent proliferating or inhibiting cell lines. The cellular sources producing the cytokines was investigated by immunohistochemical stains and in situ hybridization. Results : PBMC started to produce IL-6, TNF-$\alpha$ and TGF-$\beta$ in 1 hr, 4 hrs and 8hrs, respectively, after LPS stimulation. The production of IL-6, TNF-$\alpha$ and TGF-$\beta$ continuously increased 96 hrs after stimulation of LPS. The amount of production was 19.8 ng/ml of IL-6 by $10^5$ PBMC, 4.1 ng/mL of TNF by $10^6$ PBMC and 34.4 pg/mL of TGF-$\beta$ by $2{\times}10^6$ PBMC. The immunoreactivity to IL-6, TNF-$\alpha$ and TGF-$\beta$ were detected on monocytes in LPS-stimulated PBMC. Some of lymphocytes showed positive immunoreactivity to TGF-$\beta$. Double immunohistochemical stain showed that IL-1$\beta$, IL-6, TNF-$\alpha$ expression was not associated with CD14 postivity on monocytes. IL-1$\beta$, IL-6, TNF-$\alpha$ and TGF-$\beta$mRNA expression were same as observed in immunoreactivity for each cytokines. Conclusion: When monocytes are stimulated with LPS under serum-free conditions, IL-6 and TNF-$\alpha$ are secreted in early stage of inflammation. In contrast, the secretion of TGF-$\beta$ arise in the late stages and that is maintained after 96 hrs. The main cells releasing IL-1$\beta$, IL-6, TNF-$\alpha$ and TGF-$\beta$ are monocytes, but also lymphocytes can secret TGF-$\beta$.

• IMMUNE NETWORK
• /
• v.3 no.1
• /
• pp.1-7
• /
• 2003

Type II collagen (CII), major component of hyaline cartilage, has been considered as an auto-antigen in rheumatoid arthritis (RA). However, the clinical and biological significances with regard to the CII autoimmunity need to be clarified in human RA. The presence of antibodies to CII has been identified in sera, synovial fluid, and cartilage of patients with RA. In our study, the increased titer of IgG anti-CII in sera was well correlated with C-reactive protein, suggesting that this antibody may reflect the inflammatory status of RA. The titer of anti-CII antibodies (anti-CII Abs) tended to be higher in early stages of diseases. In our extending study, among 997 patients with RA, 269 (27.0%) were positive for circulatory IgG antibody to CII, those levels were fluctuated over time. It is hard to assess the significant amount of T cell responses to CII and CII (255~274) in RA. By using a sensitive method of antigen specific mixed lymphocyte culture, we can detect the presence of CII-reactive T cells in peripheral blood mononuclear cells of RA patients. Sixty seven (46.9%) of 143 patients showed positive CII reactive T cell responses to CII or CII (255~274). The frequencies of CII reactive T cells were more prominent in inflamed synovial fluid (SF) than in peripheral blood. These T cells could be clonally expanded after consecutive stimulation of CII with feeding of autologous irradiated antigen presenting cells (APC). Moreover, the production of Th1-related cytokine, such as IFN-${\gamma}$, was strongly up-regulated by CII reactive T cells. These data suggest that T cells responding to CII, which are probably presenting the IFN-${\gamma}$ producing cells, may play an important role in the perpetuation of inflammatory process in RA. To evaluate the effector function of CII reactive T cells, we investigated the effect of CII reactive T cells and fibroblasts-like synoviocytes (FLS) interaction on the production of pro-inflammatory cytokines. When the CII reactive T cells were co-cultured with FLS, the production of IL-15 and TNF-${\alpha}$ from FLS were significantly increased (2 to 3 fold increase) and this increase was clearly presented in accord to the expansion of CII reactive T cells. In addition, the production of IFN-${\gamma}$ and IL-17, T cell derived cytokines, were also increased by the co-incubation of CII reactive T cells with FLS. We also examined the impact of CII reactive T cells on chemokines production. When FLS were co-cultured with CII stimulated T cells, the production of IL-8, MCP-1, and MIP-1${\alpha}$ were significantly enhanced. The increased production of these chemokines was strongly correlated with increase the frequency of CII reactive T cells. Conclusively, immune response to CII was frequently found in RA. Activated T cells in response to CII contributed to increase the production of proinflammatory cytokines and chemokines, which were critical for inflammatory responses in RA. The interaction of CII-reactive T cells with FLS further augmented this phenomenon. Taken together, our recent studies have suggested that autoimmunity to CII could play a crucial role not only in the initiation but amplification/perpetuation of inflammatory process in human RA.

• Korean Journal of Poultry Science
• /
• v.47 no.1
• /
• pp.49-59
• /
• 2020

The present study determined the effect of dietary cultivated microalgae (Parachlorella sp.) on the growth and immune responses of pre-starter broilers. A total of 320 one-day-old birds (Ross 308) were allocated to 4 treatments with 8 blocks in a randomized complete block design. The four experimental diets consisted of a corn-soybean meal-based control diet, and three diets contained 0.5%, 1.0%, and 1.5% microalgae powder at the expense of cornstarch in the control diet. After feeding the experimental diets for 7 days, the body weight and feed intake of all birds were measured, and 8 birds were randomly selected from each treatment. Peripheral blood mononuclear cells (PBMCs) and serum were harvested for immune profile assessment, including cytokines and cell migration receptors. No differences in growth performance were observed among the treatments. The birds that were fed diets containing graded levels of microalga showed a linear increase in the mRNA expression of cytokine genes in PBMCs, including that of IL2, IL1β, and IL18 (P<0.05). With respect to the chemokine receptor genes in PBMCs, mRNA expression of CCR2, CCR9, and ITGA4 changed quadratically (P<0.05), but that of CCR7 increased linearly (P<0.01). Cytokine protein secretion in blood, including that of IL-1β and IL-6, increased linearly (P<0.01) with an increase in the microalgal content. Overall, the present results show that the indigenous microalgae powder used in this study could stimulate immunity with no detrimental effects on the growth performance of pre-starter broiler chickens.

• Pediatric Gastroenterology, Hepatology & Nutrition
• /
• v.27 no.1
• /
• pp.26-36
• /
• 2024

Purpose: We investigated the role of CD8⁺T cells as host immune factors in pediatric patients with Helicobacter pylori gastritis. Methods: Gastric mucosal tissue and blood samples were collected from 39 children, including 11 children with H. pylori infection and 28 children as controls. Anti-CD8 and anti-T-bet antibodies were used for immunohistochemistry of the gastric mucosa. For the cell surface and intracellular staining, peripheral blood mononuclear cells were stained with anti-IL7Rα, anti-CX3CR1, anti-CD8, anti-T-bet, and anti-IFN-γ antibodies. Cytokines of sera such as tumor necrosis factor alpha (TNF-α) and CX3CL1 were analyzed using enzyme- linked immunosorbent assay (ELISA). Results: In the immunohistochemistry of gastric mucosa, the frequency of CD8⁺ and T-bet⁺ T cells cells was higher in the H. pylori-positive group than in the control group (26.9± 7.8% vs. 16.9±3.3%, p<0.001; 5.0±2.5% vs. 2.2±0.7%, p=0.001). Between the control and H. pylori-positive groups, the frequency of IL-7Rα^lowCX3CR1⁺ CD8⁺ and T-bet⁺ INF-γ⁺ CD8⁺ T cells were not significantly different between surface and intracellular staining, respectively (40.4±24.0% vs. 38.2±17.8%, p=0.914; 40.4±24.0% vs. 38.2±17.8%, p=0.914). In the ELISA, no significant differences in TNF-α and CX3CL1 concentrations were observed between the control and H. pylori-positive groups (34.3±12.1 pg/mL vs. 47.0±22.6 pg/mL, p=0.114/0.5± 0.1 pg/mL vs. 0.5±0.1 pg/mL, p=0.188). Conclusion: CD8⁺ T and Th1 cells, which secrete IFN-γ, might play important roles in the mucosal immunity of the stomach in children with H. pylori infection.

• Microbiology and Biotechnology Letters
• /
• v.38 no.1
• /
• pp.93-104
• /
• 2010

In this study, we investigated the effects of frankincense essential oil (BSEO) on the immune cell change in the lung, BALF and PBMC using a mouse model of asthma. BALB/c mice after intraperitoneal OVA sensitization (day 1) were challenged intratracheally with OVA on day 14. Then, the asthma was induced by repeated OVA inhalation challenged. The asthma induced mice group inhaled 0.3% BSEO for 30 minutes per trial, three times a week, for 8 weeks using the nebulizer. After 12 weeks from the experiment, the mice was killed and the lung, bronchoalveolar lavage fluid (BALF) and peripheral blood mononuclear cell (PBMC) were obtained. Next, the change of immune cells inside the separated tissues was observed to identity the effects of BSEO on the allergic asthma mice. In conclusion, the hypersensitive reaction of airway to the bronchoconstrictor in the allergic asthma induced mice was effectively suppressed in Frankincense group, in Bermagot, Eucalyptus, Chamomile, Marjoram and Frankincense groups, the natural aromatic essential oil groups. Furthermore, it was also confirmed that the weight of lung, total number of alveolus cells and the number of BALF, MNL and DLN increased after inducing allergic asthma were reduced. BSEO suppressed the percentage of $CD3e^+/CD19^-$, $B220^+/CD23^+$ and $CD11b^+/Gr-1^+$ cells in the lung tissue of allergic asthma mice. Moreover, BSEO also reduced the percentage of $CD4^+/CD8^-$, $B220^+/CD23^+$ and $CD3^+/CCR3^+$ cells in BALF. In addition, the percentage of $CD3e^+/CD19^-$, $CD3^+/CD69^+$ and $B220^+/CD23^+$ cells in PBMC was reduced. The results of this study indicate that BSEO would be effective to treat allergic asthma by the immune control suppressing the activity of immune cells in each tissue.

• Clinical and Experimental Pediatrics
• /
• v.54 no.4
• /
• pp.157-162
• /
• 2011

Purpose: Exaggerated pro-inflammatory reactions during the acute phase of Kawasaki disease (KD) suggest the role of immune dysregulation in the pathogenesis of KD. We investigated the profiles of T regulatory cells and their correlation with the clinical course of KD. Methods: Peripheral blood mononuclear cells were collected from 17 KD patients during acute febrile and subacute afebrile phases. T cells expressing CD4, CD25, and Foxp3 were analyzed using flow cytometry, and the results were correlated with the clinical course of KD. Results: The percentage of circulating $CD4^+CD25^{high}Foxp3^+$ T cells among $CD4^+$ T cells was Significantly higher during the subacute afebrile phase than during the acute febrile phase ($1.10%{\pm}1.22%$ vs. $0.55%{\pm}0.53%$, P=0.049). Although levels of $CD4^+CD25^{low}Foxp3^+$ T cells and $CD4^+CD25^-Foxp3^+$ T cells were only slightly altered, the percentage of $CD4^+CD25^+Foxp3^-$ T cells among $CD4^+$ T cells was significantly lower during the subacute afebrile phase than during the acute febrile phase ($2.96%{\pm}1.95%$ vs. $5.64%{\pm}5.69%$, P=0.036). Consequently, the ratio of $CD25^{high}Foxp3^+$ T cells to $CD25^+Foxp3^-$ T cells was higher during the subacute afebrile phase than during the acute febrile phase ($0.45%{\pm}0.57%$ vs. $0.13%{\pm}0.13%$, P=0.038). Conclusion: Decreased $CD4^+CD25^{high}Foxp3^+$ T cells and/or an imbalanced ratio of $CD4^+CD25^{high}Foxp3^+$ T cells to $CD4^+CD25^+Foxp3^-$ T cells might playa role in KD development. Considering that all KD patients were treated with intravenous immunoglobulin (IVIG), recovery of $CD4^+CD25^{high}Foxp3^+$ T cells during the subacute afebrile phase could be a mechanism of IVIG.

• Asian-Australasian Journal of Animal Sciences
• /
• v.33 no.7
• /
• pp.1167-1179
• /
• 2020

Objective: This study was conducted to fathom the underlying mechanisms of nutrition intervention and redox sensitive transcription factors regulated by Antrodia cinnamomea fermented product (FAC) dietary supplementation in broiler chickens. Methods: Four hundreds d-old broilers (41±0.5 g/bird) assigned to 5 groups were examined after consuming control diet, or control diet replaced with 5% wheat bran (WB), 10% WB, 5% FAC, and 10% FAC. Liver mRNA expression of antioxidant, inflammatory and lipid metabolism pathways were analyzed. Prostaglandin E2 (PGE₂) concentration in each group were tested in the chicken peripheral blood mononuclear cells (cPBMCs) of 35-d old broilers to represent the stress level of the chickens. Furthermore, these cells were stimulated with 2,2'-Azobis(2-amidinopropane) dihydrochloride (AAPH) and lipopolysaccharide (LPS) to evaluate the cell stress tolerance by measuring cell viability and oxidative species. Results: Heme oxygenase-1, glutathione S-transferase, glutamate-cysteine ligase, catalytic subunit, and superoxide dismutase, and nuclear factor (erythroid-derived 2)-like 2 (Nrf2) that regulates the above antioxidant genes were all up-regulated significantly in FAC groups. Reactive oxygen species modulator protein 1 and NADPH oxygenase 1 were both rather down-regulated in 10% FAC group as comparison with two WB groups. Despite expressing higher level than control group, birds receiving diet containing FAC had significantly lower expression level in nuclear factor-kappa B (NF-κB) and other genes (inducible nitric oxide synthase, tumor necrosis factor-α, interleukin-1β, nucleotide-binding domain, leucine-richcontaining family, pyrin domain-containing-3, and cyclooxygenase 2) involving in inflammatory pathways. Additionally, except for 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase that showed relatively higher in both groups, the WB, lipoprotein lipase, Acetyl-CoA carboxylase, fatty acid synthase, fatty acid binding protein, fatty acid desaturase 2 and peroxisome proliferator-activated receptor alpha genes were expressed at higher levels in 10% FAC group. In support of above results, promoted Nrf2 and inhibited NF-κB nuclear translocation in chicken liver were found in FAC containing groups. H₂O₂ and NO levels induced by LPS and AAPH in cPBMCs were compromised in FAC containing diet. In 35-d-old birds, PGE₂ production in cPBMCs was also suppressed by the FAC diet. Conclusion: FAC may promote Nrf2 antioxidant pathway and positively regulate lipid metabolism, both are potential inhibitor of NF-κB inflammatory pathway.

• The Journal of Pediatrics of Korean Medicine
• /
• v.22 no.2
• /
• pp.81-114
• /
• 2008

Objectives : The purpose of this study is to investigate the effect of Jeseupwiryeongtang-gagam(JWTG) on atopic dermatitis by in vivo experiment using NC/Nga atopic dermatitis mouse, which has histological and clinical similarities to the atopic dermatitis of human. Methods : To investigate the effect of JWTG on AD, we evaluated atopic dermatitis-like skin lesions by clinical skin index and analyzed immunological parameters in peripheral blood mononuclear cells(PBMCs), splenocytes, draining lymph node(DLN) and performed skin histology in ears and dorsal skin of atopic dermatitis-like skin NC/Nga mouse in vivo. Results and Conclusions : In vivo, clinical skin severity score were significantly lower in JWTG group than control group. IgE, IL-6, $TNF-{\alpha}$, IgM, IgG2a and IgG2b levels in serum were decreased remarkably in JWTG group than control group and $IFN-{\gamma}$ production, secreted in Th1 cell were increased by JWTG. After experiment ended, we analyzed immunological cells ($CD3^+$, $CD19^+$, $CD4^+$, $CD8^+$, $CD3^+$$CD69^+$, $CD4^+$$CD25^+$ and $CD49b^+$) by flow cytometry. It resulted that total absolute number of $CD3^+$, $CD19^+$, $CD4^+$ and $CD8^+$ cells were recovered as normal and $CD3^+$$CD69^+$ were decreased significantly compared with control group in isolated DLN and PBMCs from NC/Nga mouse and total absolute number of $Gr-1^+$, $CD11b^+$ and $CD3^+$ in dorsal skin of NC/Nga mouse were decreased by JWTG. We analyzed ear, DLN, and neck-back skin after biopsy and dyeing by hematoxyline/eosin(H&E) and toluidine staining (mast cells marker) and obtained results that JWTG were effective to histological symptoms (dermal and epidermal thickening, hyperkeratosis and inflammatory cell infiltration). Ear thickness was decreased significantly than the control group and the size of inflammatory lymphocytes cells(ILC) and plasma cells(PC) in DLN were also decreased.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.23 no.5
• /
• pp.1106-1115
• /
• 2009

Mori Ramulus has multiple applications in Korean traditional medicine prescription because it has antioxidant and anti-inflammatory effects by reducing macrophage activities. Yet, no studies on the anti-arthritic activity of EMR (extract of Mori Ramulus) have been reported in vitro and in vivo. Rheumatoid arthritis (RA) is a systemic autoimmune disease with chronic inflammation characterized by hyperplasia of synovial cells in affected joints, which ultimately leads to the destruction of cartilage and bone. Because collagen-induced arthritis (CIA) is similar to RA in pathological symptoms and immune reactions, there have been several reports concerning RA using CIA mouse model. Here, we investigated the effects of Mori Ramulus on RA using CIA mice. The importance of CD4+ Th1 cells in RA progress was previously indicated and studies further showed that Th17 cells play a prime role in severity of disease. Accordingly, the present study was focused on CIA associated with CD4+ Th1 cells and Th1 7 cells. DBA/1OlaHsd mice were immunized with bovine type II collagen (CII). After a second collagen immunization, mice were treated with EMR once a day for 4 weeks. The severity of arthritis within the paw joints was evaluated by histological assessment of cartilage destruction and pannus formation. Immune cells in peripheral blood mononuclear cells (PBMC), draining lymph node (DLN) and paw joints, cytokine production and gene expression were assessed from CIA mouse using ELISA, FACS and real-time PCR analysis. Administration of EMR significantly suppressed the progression of CIA and inhibited the production of TNF-$\alpha$, IL-6 and IL-17 in the serum. The erosion of cartilage was dramatically reduced in mouse knees after treatment with EMR. In conclusion, our results demonstrate that EMR significantly suppressed the progression of CIA and that this action was mediated by the decreased production of TNF-$\alpha$, IL-6, IL-17 and collagen II-specific antibody in the serum. EMR suppressed Th17 cells and reduced level of IL-6 via B cell suppression, and thus, the levels of autoantibodies produced from B cells were decreased. Furthermore, EMR suppressed NKT cells which directly stimulate B cells and develop imbalance of Th1/Th2 cell. Oral administration of EMR (100 mg/kg or 200 mg/kg) significantly suppressed the progression of CIA, which is comparable to that of methotrexate (MTX, 0.3 mg/kg) used as a positive control. We are currently studying the mechanism underlying the therapeutic role for EMR in CIA mice.