인간 재조합 인터루긴-32 면역조절작용에 대한 유세포 분석

이광수,김영관,채정일,심정현,김은미,강형식,김수현,윤도영,명평근 충남대학교 생물공학연구소 2006 생물공학연구지 Vol.12 No.-

Xenotransplantation of porcine organs has the potential to overcome the severe shortage of human tissues and organ available for human transplantation. however, it remains various hurdles for clinical xenotransplantation. In pig and mouse xenotransplantation, porcine xenograft evoke a strong cellular rejection response in immunocompetent host and grafts are destroyed within a week. This cellular immune response could involved both T cells and NK cells. A number of groups have shown that human NK cells can recognize and damage porcine endothelial cells. In addition, human T cells can respond to porcine endothelial cells through both direct and indirect mechanisms. Cellular rejection of porcine tissues requires T cells, particularly CD4^(+) cells. A new cytokine recombinant human interleukin-32α,β(IL-32α,β) has a role innate and acquired immune system. In order to investigate the role of recombinant mouse IL-18 and recombinant human IL-32α,β in xenograft rejection, we transplanted the PK(15) cells to C57BL/6 mice with or without intraperitoneal injection of recombinant mouse IL-18 or recombinant human IL-32 α,β. It was analyzed the population of NK cell, T cell and B cell in the C57BL/6 mice transplanted with PK(15) cells and recombinant mouse IL-18 or recombinant human IL-32α,β by flow cytometry analysis. As a result, lymph node and thymus of PK15/IL18, PK15/IL32α and PK15/IL32β injected group were increased to T cell activation population than normal injected groups. CD8^(+) T cells were decreased in lymph node of PK15/IL18, PK15/IL32α and PK15/IL32β injected groups. CD4^(+) T cells were increased in lymph node cell of PK15/IL32α and PK15/IL32β injected group and also, B cell population were increased in lymph node cell and spleen of PK15/IL18, PK15/IL32α and PK15/IL32β injected group. Therefore, we suggest that recombinant mouse IL-18 and recombinant human IL-32α,β suppress xenograft rejection in cellular xenotransplantation.

자궁내막증 병변에서의 Cytokine mRNA의 발현 양상

이택후(Taek Hoo Lee),김광수(Gwang Soo Kim),김일규(Il Gyu Kim),전상식(Sang Sik Chun),조영래(Young Lae Cho) 대한산부인과학회 1999 Obstetrics & Gynecology Science Vol.42 No.9

목적: 자궁내막증은 최근에 그 빈도가 급격하게 증가되고 있으나 아직까지 병인과 치료법이 확실하게 정립되지 못하고 있는 부인과 내분비의 대표적인 질환이다. 최근 들어서 자궁내막증 환자의 불임의 원인으로 cytokine이 밀접하게 관여됨이 밝혀지고 있고 자궁내막증의 병인으로 복강내로 역류된 월경혈에 의한 복강내의 국소적 염증에 대한 개체의 반응정도와 면역체계의 변화가 중요하게 인정되고 있으나 아직까지 정확한 기전은 밝혀져 있지 않기 때문에 이에 저자는 자궁내막증 환자의 복수 내에서의 cytokine의 정량측정과 골반내 자궁내막증 병변조직에서의 cytokine mRNA의 발현 양상을 조사하여 자궁내막증 환자의 병인분석을 시도하였다. 연구방법: 자궁내막증 진단을 받고 복강경 혹은 개복 수술을 받은 30명의 환자에서 총 60개의 자궁내막증 병변이라고 의심되는 조직을 채취하여 이를 RT-PCR법을 이용하여 Cytokine 유전자의 발현양상을 조사하였다. 또한 7례의 정상대조군과 23례의 자궁내막증 환자의 복수를 ELISA법을 이용하여 정량분석을 시도하였다. 결과: 자궁내막증 환자의 복수내에서 IL-6와 IL-10의 농도는 자궁내막증의 임상적 중증도에 따라서 의미있게 증가되어 있었으나 IFN-γ, TNF-α, IL-1β, 그리고 IL-5의 농도는 정상 대조군에 비해서 변화가 없었다. 모든 예의 심부 자궁내막증 병변조직과 표재성 자궁내막증 병변조직에서 IL-1β cytokine mRNA의 발현을 볼 수 있었다. IL-5와 IL-6은 표재성 병변 12개 중에서 각각 2개의 black lesion에서만 발현을 볼 수 있었으며 IL-10은 표재성 병변에서는 12개중 2개에서 그리고 심부성 병변에서는 8개중 1개의 조직에서만 발현되었다. IFN-γ는 표재성 병변에서는 전혀 발현이 없었으나 심부성 병변에서는 8개중 4개의 조직에서 발현이 되었으며 TNF-α는 표재성 병변에서는 red 및 black 병변에서 각각 1개의 조직에서만 발현이 되었으나 심부성 병변에서는 역시 8개중 4개의 조직에서 발현이 되었다. 결론: 표재성 병변이 골반강내에 착상하여 염증성 반응이 일어날 수 있는 원인이 제공되면 IL-1β 혹은 TNF-α같은 염증성 cytokine이 분비가 되며 이로 인해 생성되는 단핵세포의 chemotactic factor에 의해 대식세포의 증가와 활성화가 이루어지고 이어서 복강내에 IL-6 등의 cytokine이 증가되며 마지막으로 여러 가지 증가된 cytokine에 대한 반대 반응으로 IL-10이 증가됨을 추정할 수가 있겠으며 이러한 가정은 앞으로 cytokine을 이용한 치료적 응용의 기초적인 연구로서 중요한 의미를 제공할 수 있다고 하겠다. Objective: The pathogenesis of endometriosis is generally accepted that retrograde menstruation and alterations in the local pelvic immune environment. This study was performed to help elucidate what kind of role various cytokines might play in the pathogenesis of endometriosis. Method: Concentrations of peritoneal fluid cytokines were compared in 7 women with normal pelvic finding and 23 women with endometriosis by enzyme-linked immunosorbent assay(ELISA). The patterns of cytokine mRNA expression in 8 ovarian endometrioma and 12 superficial pelvic endometriosis lesions were investigated by reverse transcription-polymerase chain reaction(RT-PCR) amplification method. Result: Both IL-6 and IL-10 levels in peritoneal fluid specimens with endometriosis tended to be higher than normal. However, there were no significant differences between peritoneal fluid concentrations of IFN-γ, TNF-α, IL-1β, and IL-5 of women with and without endometriosis. The levels of IL-6 and IL-10 were significantly higher in peritoneal fluid of women with severe endometriosis compared to women with mild endometriosis. IL-1β mRNA was expressed in all of 8 deep and 12 superficial endometriosis lesions. IL-5 and IL-6 mRNA were expressed in only two black lesions respectively, however, both were not expressed in the all deep lesions. Expressions of IL-10 mRNA occurred in one red and one black lesion while this was expressed in only one of the deep lesions. TNF-α mRNA was expressed in one red and one black lesion of 12 superficial lesions compared with four of the deep lesions. There was the difference between kinds of increased cytokines in the peritoneal fluid and those of expressed cytokines in the endometriotic lesions of patients with endometriosis. Conclusion: This study supports the concept that local immunologic factors may be important in the pathogenesis and pathophysiology of endometriosis. The pattern of cytokine mRNA expression of endometriotic lesions would seem to indicate that proinflammatory cytokines such as IL-1β and TNF-α are responsible for the development or progression of endometriosis.

Interferon-γ Inhibits in vitro Mobilization of Eosinophils by Interleukin-5

Park, Choon-Sik,Choi, Eun Nam,Kim, Jung Sun,Choi, Yun Sung,Rhim, Tai Youn,Chang, Hun Soo,Chung, Il Yup S. Karger AG 2005 International archives of allergy and immunology Vol.136 No.3

<P><I>Background:</I> Th2 cytokines play pivotal roles in allergic inflammation, including eosinophilia, and their actions are antagonized by Th1 cytokines, conferring them therapeutic potential. <I>Methods:</I> In this study, we examined the ability of a number of cytokines to suppress the activation of eosinophils that function as effector cells for allergic airway diseases. <I>Results:</I> Interleukin (IL)-5, IL-6, and tumor necrosis factor (TNF) induced an eosinophil shape change, whereas interferon (IFN)-γ significantly inhibited the shape change. Other cytokines, including IL-1β, IL-4, IL-10 and IL-13, had little or only slightly enhancing or reducing effects on the shape change. We further analyzed the IFN-γ effect, showing that pretreatment with IFN-γ strongly suppressed IL-5-induced eosinophil shape change, and cycloheximide (CHX) abrogated the suppression by IFN-γ, suggesting that new protein synthesis is required for the inhibitory effect by this cytokine. In agreement with these results, IFN-γ blocked the eosinophil migration and ERK phophorylation induced by IL-5, and the addition of CHX restored eosinophil chemotaxis. <I>Conclusions:</I> Collectively, IFN-γ may attenuate eosinophilic inflammation by directly negating eosinophil mobilization.</P><P>Copyright © 2005 S. Karger AG, Basel</P>

복막 중피 세포에서 IL-1β 자극에 의한 MCP-1과 RANTES의 생성

송인숙,이상구,박정식,양원석,김순배,윤견일 대한신장학회 2000 Kidney Research and Clinical Practice Vol.19 No.5

Human peritoneal mesothelial cells may have a great potential to secrete chemokines, growth factors, adhesion molecules, and various cytokines stimulated with proinflammatory cytokines during peritoneal infection. In the course of peritonitis, rapid neutrophil cell influx and subsequent monocytic cell influx can be observed. It has been demonstrated that human peritoneal mesothelial cells secrete a C-X-C chemokine, IL-8, which contributes to the recruitment of neutrophil influx during peritoneal infection. However, the production and role of C-C chemokines have not been fully defined in human peritoneal mesothelial cells. This study was performed to evaluate the production of MCP-1 and RANTES and their influence on the chemotaxis of monocytes when human peritoneal mesothelial cells were stimulated with IL-1β. Mesothelial cells obtained by enzymatic digestion of pieces of human omentum and stimulated with a various doses and times of IL-1β. The expression of MCP-1 and RANTES mRNA was measured by Northern bloassay and the expression of their proteins was analyzed by ELISA. To evaluate their function, monocytes chemotaxis assay was performed using a 48-well chemotactic chamber. Cultured human peritoneal mesothelial cells appeared to be polygonal at confluence using phase contrast microscope. Indirect immunofluorescent staining demonstrated that the mesothelial cells reacted positively with anti-cytokeratin antibody and anti-vimentin antibody. The expression of MCP-1 and RANTES mRNA increased in response to IL-1β in time and dose dependent manner. The protein levels of MCP-1 and RANTES with stimulation of 1.0ng/mL of IL-1β for 24 hours were higher than those without(30.0±2.22 vs 3.55±0.74ng/105cells and 1.53±0.41 vs 0.11±0.02ng/105cells respectively, p$lt;0.05, n=6). Chemotaxis assay showed that the supernatants from human peritoneal mesothelial cells with stimulation of IL-1β for 24 hours had significantly higher chemotaxis of monocytes than those without(71±3.4% vs 50±2.9%, p$lt;0.05, n=6). Coincubation of sup with stimulation and antibodies to MCP-1 or RANTES(20μL/mL, lOμL/mL, respectively) resulted in a significant inhibition of chemotaxis of monocytes by 33% and 12%(47±3.1% and 62±3.0% respectively, p$lt;0.05, n=6). Human peritoneal mesothelial cells are capable of the expression of MCP-1 and RANTES mRNA and the production of their proteins in response to IL-1β. Functionally, mesothelial cells derived Mand RANTES may contribute to the recruitment of monocytes and amplify the inflammatory process. Thus, human peritoneal mesothelial cells play an important role during peritoneal infection.

Chronological changes in inflammatory cytokines immunoreactivities in the mouse hippocampus after systemic administration of high dosage of tetanus toxin.

Yan, Bing Chun,Park, Joon Ha,Kim, In Hye,Shin, Bich Na,Ahn, Ji Hyeon,Yoo, Ki-Yeon,Lee, Deuk-Sik,Kim, Myong Jo,Kang, Il-Jun,Won, Moo-Ho Springer-Verlag 2012 Experimental brain research Vol.223 No.2

<P>Tetanus toxin (TeT) is an exotoxin and has a capacity for neuronal binding and internalization. In the present study, we compared changes in the immunoreactivities and protein levels of interleukin (IL-) 2 as a pro-inflammatory cytokine and IL-4 as an anti-inflammatory cytokine in the hippocampus proper (HP) and dentate gyrus (DG) after systemic treatment of 10 or 100?ng/kg TeT into mice. In this study, we could not find any neuronal damage or loss in any subregions of the hippocampus after TeT treatment. In the control groups, strong IL-2 immunoreactivity was shown in the stratum pyramidal (SP) of the HP and in the granule cell layer (GCL) of the DG. At 6?h post-treatment, IL-2 immunoreactivity was hardly detected in the SP and GCL; however, strong IL-2 immunoreactivity was shown in the stratum oriens of the HP in both the groups. Thereafter, intermediate IL-2 immunoreactivity was shown in the SP and GCL. On the other hand, intermediate IL-4 immunoreactivity was detected in the SP and GCL of the control groups. At 6?h post-treatment, IL-4 immunoreactivity in the SP and GCL was apparently increased. Thereafter, IL-4 immunoreactivity was lower than that at 6?h post-treatment. In brief, IL-2 and 4 immunoreactivities were easily detected in SP and GCL in the controls and dramatically decreased and increased at 6?h post-treatment, respectively.</P>

Activation of Transient Receptor Potential Melastatin Family Member 8 (TRPM8) Receptors Induces Proinflammatory Cytokine Expressions in Bronchial Epithelial Cells

김주희,Young Sook Jang,Hwan Il Kim,박지영,박성훈,Yong Il Hwang,Seung Hun Jang,Ki-Suck Jung,Hae Sim Park,Choon-Sik Park 대한천식알레르기학회 2020 Allergy, Asthma & Immunology Research Vol.12 No.4

Purpose: Cold air is a major environmental factor that exacerbates asthma. Transient receptor potential melastatin family member 8 (TRPM8) is a cold-sensing channel expressed in the airway epithelium. However, its role in airway inflammation remains unknown. We investigated the role of TRPM8 in innate immune responses in bronchial epithelial cells and asthmatic subjects. Methods: The TRPM8 mRNA and protein expression on BEAS2B human bronchial epithelial cells was examined by real-time polymerase chain reaction (PCR), immunofluorescence staining and western blotting. Additionally, interleukin (IL)-4, IL-6, IL-8, IL-13, IL-25 and thymic stromal lymphopoietin (TSLP) levels before and after menthol, dexamethasone and N-(4-tert-butylphenyl)-4-(3-chloropyridin-2-yl) piperazine-1-carboxamide (BCTC) treatments were measured via real-time PCR. TRPM8 protein levels in the supernatants of induced sputum from asthmatic subjects and normal control subjects were measured using enzyme-linked immunosorbent assay, and mRNA levels in sputum cell lysates were measured using real-time PCR. Results: Treatment with up to 2 mM menthol dose-dependently increased TRPM8 mRNA and protein in BEAS2B cells compared to untreated cells (P < 0.001) and concomitantly increased IL-25 and TSLP mRNA (P < 0.05), but not IL-33 mRNA. BCTC (10 μM) significantly abolished menthol-induced up-regulation of TRPM8 mRNA and protein and IL-25 and TSLP mRNA (P < 0.01). TRPM8 protein levels were higher in the supernatants of induced sputum from asthmatic subjects (n = 107) than in those from healthy controls (n = 19) (P < 0.001), and IL-25, TSLP and IL-33 mRNA levels were concomitantly increased (P < 0.001). Additionally, TRPM8 mRNA levels correlated strongly with those of IL-25 and TSLP (P < 0.001), and TRPM8 protein levels were significantly higher in bronchodilator-responsive asthmatic subjects than in nonresponders. Conclusions: TRPM8 may be involved in the airway epithelial cell innate immune response and a molecular target for the treatment of asthma.

Genetic effect of <i>CCR3</i> and <i>IL5RA</i> gene polymorphisms on eosinophilia in asthmatic patients

Lee, June-Hyuk,Chang, Hun Soo,Kim, Ji Hyun,Park, Se-Min,Lee, Yong Mok,Uh, Soo Taek,Rhim, Taiyoun,Chung, Il Yup,Kim, Yong-Hoon,Park, Byung Lae,Park, Choon-Sik,Shin, Hyoung Doo Elsevier 2007 The journal of allergy and clinical immunology Vol.120 No.5

<P><B>Background</B></P><P>Eosinophilic infiltration and peripheral blood eosinophilia in asthma require the cooperation of eosinophil-specific cytokines and chemokines and their receptors.</P><P><B>Objective</B></P><P>We investigated the association of polymorphisms in <I>CCR3</I> and <I>IL5RA</I> with asthma susceptibility or peripheral blood eosinophilia and the effects of the polymorphisms on receptor expression.</P><P><B>Methods</B></P><P>Polymorphisms in <I>CCR3</I> and <I>IL5RA</I> were identified and genotyped in 576 asthmatic patients and 180 healthy control subjects. CCR3 and IL-5 receptor α (IL-5Rα) protein expression on eosinophils was measured by means of flow cytometry.</P><P><B>Results</B></P><P>Although polymorphisms in <I>CCR3</I> were not associated with asthma susceptibility, the <I>CCR3</I> haplotype <I>ht2</I> showed a negative gene dose effect on the eosinophil count (<I>P</I> = .003–.009). <I>IL5RA c.−5091G>A</I> was weakly associated with eosinophil count. The effects of <I>ht2</I> were greater when paired with <I>IL5RA c.−5091A</I> (<I>P</I> = .001–.002). CCR3 protein expression was higher on eosinophils of asthmatic patients without <I>ht2</I> than in those with <I>ht2</I>. Asthmatic patients with the <I>IL5RA c.−5091A</I> allele showed higher IL-5Rα expression than those who were homozygous for the G allele.</P><P><B>Conclusion</B></P><P>The genetic association between <I>CCR3</I> polymorphisms and the number of circulating eosinophils was revealed as a novel finding. These associations were more pronounced when the <I>CCR3</I> polymorphisms were paired with polymorphisms in <I>IL5RA</I>. The protein expression levels of CCR3 and IL-5Rα on peripheral blood eosinophils are associated with the polymorphisms on their own genes.</P><P><B>Clinical implications</B></P><P>The identification of single nucleotide polymorphisms and haplotypes of <I>CCR3</I> and <I>IL5RA</I> might be useful in developing markers for intermediate phenotypes of eosinophil number and in designing strategies to control diseases related to hypereosinophilia.</P>

OK-432(Picibanilⓡ)와 외과적 절제술을 이용한 선천성 림프관종의 치험례

김일규,이성호,오성섭,최진호,오남식,김왕식,임영일,양동환 대한악안면성형재건외과학회 2001 Maxillofacial Plastic Reconstructive Surgery Vol.23 No.3

Lymphangiomas are relatively rare benign tumors of the lymphatic system, characterized by congenital lymphatic malformation such as anastomosis or obstuction of the lymphatic channels. There are most frequently diagnosed in children younger than 3years. In contrast to that of the hemangioma, the sex distribution of the lymphangioma is nearly evenly divided. The head and neck lymphangioma represents about 70∼75% of all lymphangiomas, and they are difficult to manage. Spontaneous regression is rare, and rapid intermittent enlargement occurs secondary to infection or trauma. Enlargement may cause serious sequelae such as airway obstruction, feeding difficulties, and cosmetic problems. Treatments previously used for lymphangiomas include surgical excision and intralesional injection of sclerosants. Problems associated with surgical excision include the risk of cosmetically unacceptable scarring and the risk of damage to surrounding vital stricture and the high risk of incomplete excision. The sclerosants previously used have numerous other local and systemic side effects. This report describes a case that was successfully treated using OK-432 as a new sclerosant drug and secondary surgical excision for congential cavenous lymphangioma extensively enlarged to tongue, mouth floor and submandibular region.

Comparison of Cytokine and Nitric Oxide Induction in Murine Macrophages between Whole Cell and Enzymatically Digested Bifidobacterium sp. Obtained from Monogastric Animals

Kim, Dong-Woon,Cho, Sung-Back,Lee, Hyun-Jeong,Chung, Wan-Tae,Kim, Kyoung-Hoon,HwangBo, Jong,Nam, In-Sik,Cho, Yong-Il,Yang, Mhan-Pyo,Chung, Il-Byung The Microbiological Society of Korea 2007 The journal of microbiology Vol.45 No.4

The principal objective of this study was to compare the effects of whole and hydrolyzed cells (bifidobacteria) treated with gastrointestinal digestive enzymes on the activation of cloned macrophages. Seven different strains of Bifidobacterium obtained from swine, chickens, and rats, were digested with pepsin followed by pancreatin and the precipitate (insoluble fraction) and supernatant (soluble fraction) obtained via centrifugation. The RAW 264.7 murine macrophages were incubated with either whole cells, the precipitate, or supernatant at various concentrations. Pronounced increases in the levels of nitric oxide (NO), interleukin $(IL)-1{\beta}$, IL-6, IL-12, and tumor necrosis factor $(TNF)-{\alpha}$ were observed in the whole cells and precipitates, but these effects were less profound in the supernatants. The precipitates also evidenced a slight, but significant, inductive activity for NO and all tested cytokines, with the exception of $(TNF)-{\alpha}$ in the macrophage model as compared with the whole cells. By way of contrast, $(TNF)-{\alpha}$ production when cultured with whole cells (100 ng/ml) resulted in marked increases as compared with what was observed with the precipitates. The results of this study indicated, for the first time, that digested Bifidobacterium sp. can induce the production of NO and several cytokines in RAW 264.7 murine macrophage cells. In the current study, it was demonstrated that Bifidobacterium strains treated with digestive enzymes, as compared with whole cells, are capable of stimulating the induction of macrophage mediators, which reflects that they may be able to modulate the gastrointestinal immune functions of the host.

<i>In vivo</i> genotoxicity evaluation of lung cells from Fischer 344 rats following 28 days of inhalation exposure to MWCNTs, plus 28 days and 90 days post-exposure

Kim, Jin Sik,Sung, Jae Hyuck,Choi, Byung Gil,Ryu, Hyeon Yeol,Song, Kyung Seuk,Shin, Jae Hoon,Lee, Jong Seong,Hwang, Joo Hwan,Lee, Ji Hyun,Lee, Gun Ho,Jeon, Kisoo,Ahn, Kang Ho,Yu, Il Je Informa Healthcare 2014 INHALATION TOXICOLOGY Vol. No.

<P>Despite their useful physico-chemical properties, carbon nanotubes (CNTs) continue to cause concern over occupational and human health due to their structural similarity to asbestos. Thus, to evaluate the toxic and genotoxic effect of multi-wall carbon nanotubes (MWCNTs) on lung cells <I>in vivo</I>, eight-week-old rats were divided into four groups (each group = 25 animals), a fresh air control (0 mg/m<SUP>3</SUP>), low (0.17 mg/m<SUP>3</SUP>), middle (0.49 mg/m<SUP>3</SUP>), and high (0.96 mg/m<SUP>3</SUP>) dose group, and exposed to MWCNTs <I>via</I> nose-only inhalation 6 h per day, 5 days per week for 28 days. The count median length and geometric standard deviation for the MWCNTs determined by TEM were 330.18 and 1.72 nm, respectively, and the MWCNT diameters ranged from 10 to 15 nm. Lung cells were isolated from five male and five female rats in each group on day 0, day 28 (only from males) and day 90 following the 28-day exposure. The total number of animals used was 15 male and 10 female rats for each concentration group. To determine the genotoxicity of the MWCNTs, a single cell gel electrophoresis assay (Comet assay) was conducted on the rat lung cells. As a result of the exposure, the olive tail moments were found to be significantly higher (<I>p</I> < 0.05) in the male and female rats from all the exposed groups when compared with the fresh air control. In addition, the high-dose exposed male and middle and high-dose exposed female rats retained DNA damage, even 90 days post-exposure (<I>p</I> < 0.05). To investigate the mode of genotoxicity, the intracellular reactive oxygen species (ROS) levels and inflammatory cytokine levels (TNF-α, TGF- β, IL-1, IL-2, IL-4, IL-5, IL-10, IL-12 and IFN-γ) were also measured. For the male rats, the H<SUB>2</SUB>O<SUB>2</SUB> levels were significantly higher in the middle (0 days post-exposure) and high- (0 days and 28 days post-exposure) dose groups (<I>p</I> < 0.05). Conversely, the female rats showed no changes in the H<SUB>2</SUB>O<SUB>2</SUB> levels. The inflammatory cytokine levels in the bronchoalveolar lavage (BAL) fluid did not show any statistically significant difference. Interestingly, the short-length MWCNTs deposited in the lung cells were persistent at 90 days post-exposure. Thus, exposing lung cells to MWCNTs with a short tube length may induce genotoxicity.</P>