Effects of Fulvic Acid on Homocysteine-Induced Upregulation of Cyclooxygenase-2 and PGE 2 in Human Monocytes

( Tzong-shean Chin ),( Cheng-nan Chen ),( Jan-jeng Huang ) 한국농업기계학회 2018 한국농업기계학회 학술발표논문집 Vol.23 No.1

Homocysteine and pro-inflammatory mediators such as cyclooxygenase-2 (COX-2) have been linked to vascular dysfunction and risks of cardiovascular diseases. Fulvic acid (FA), a class of compounds of humic substances, possesses various pharmacological properties. However, the effect of FA on inflammatory responses of the monocytes remains unclear. We investigated the regulatory effect of FA on homocysteine-induced COX-2 expression in human monocytes. The effects of FA on mitogen-activated protein kinase (MAPK) and nuclear factor (NF)-κB pathways in homocysteine-induced U937 monocytes were also identified. Pretreating monocytes with FA inhibited the homocysteine-induced COX-2 expression in a dose-dependent manner. Stimulation of U937 monocytes with homocysteine induced rapid increases in the phosphorylation of ERK and JNK; the inhibitor for ERK and JNK attenuated this homocysteine-induced COX-2 expression. Transcription factor ELISA and chromatin immunoprecipitation assays showed that FA blocked the homocysteine-induced increases in the binding activity and in vivo promoter binding of NF-κB in monocytes. Our findings provide a molecular mechanism by which FA inhibits homocysteine-induced COX-2 expression in monocytes, and a basis for using FA in pharmaceutical therapy against inflammation.

CD45+/CD11b+ Monocytes are Required for Mesenchymal Stem Cell Proliferation In Vitro

홍현숙,안우성,손영숙 한국조직공학과 재생의학회 2014 조직공학과 재생의학 Vol.11 No.3

Mesenchymal stem cells (MSCs) have been used to treat patients with incurable diseases. However, thelow yield of MSCs even after long periods of ex vivoculture remains a serious problem. Several attempts were madeto increase cell expansion rate of MSCs in cultures. In this study, we examined the stimulatory effect of monocyteson the proliferation and colony forming efficiency of MSCs by co-culturing CD29+/CD45- MSCs with CD45+/CD11b+ monocytes. In presence of CD45+/CD11b+ monocytes, CD29+/CD45- MSCs proliferated rapidly to formlarge and compact colonies within 10 days. In the absence of CD45+/CD11b+ monocytes, CD29+/CD45- MSC col-ony formation was much delayed, indicating that the presence of CD45+/CD11b+ monocytes is required for CD29+/CD45- MSC proliferation and stemness in vitro. Similar growth-stimulating effect on MSC culture was also foundwhen conditioned medium from CD45+/CD11b+ monocyte cultures was supplied instead of CD45+/CD11b+monocytes. These results suggest that secretory molecules of CD45+/CD11b+ monocytes may mainly mediate theenhanced growth of CD29+/CD45- MSCs. Collectively, this study reveals that co-culture of CD29+/CD45- MSCswith CD45+/CD11b+ monocytes stimulate MSC growth via paracrine factors, which could improve colony formingcapacity of MSC. Furthermore, conditioned medium of CD45+/CD11b+ monocytes may also be developed for cellculture supplement for MSC cell therapeutics.

Apoptosis of Peripheral Blood Cells Stimulated by Lipopolysaccharide, Cytokines, or Escherichia coli

Choi, Jung-Hyun,Kim, Hee-Jung,Huh, Dong-Ho,Shin, Wan-Shik 가톨릭대학교 2000 Bulletin of The Catholic Research Institutes of Me Vol.28 No.-

Introduction and Methods : Apoptosis is a natural suicidal mechanism of eukaryotic cells to maintain the homeostasis of hosts. It comprises important pathogenesis of autoimmune diseases, cancers, and some infectious diseases. To access the role of apoptosis in the pathogenesis of infectious diseases, we studied the difference of apoptotic patterns between polymorphonuclear leukocytes and of monocytes to various stimuli such as lipopolysaccharide(LPS), tumor necrosis factor (TNF)-α, interleukin (IL)-6, granulocyte, macrophage-colony stimulating factor (GM-CSF), or Escherichia coli. The cell survival and expression of Fas and Fas ligand (FasL), bcl-2, and IL-1β converting enzyme (ICE) were evaluated by flow cytometry, northern blotting or western blotting analysis. Results : The survival of polymorphonuclear leukocytes and monocytes prolonged after stimulation with LPS and cytokines. Stimulation of Fas with Fas monoclonal antibody induced the apoptosis of polymorphonuclear leukocytes and monocytes. Expressions of Fas and FasL were dominant in polymorphonuclear leukocytes than monocytes. bcl-2 was expressed only in monocytes and it was associated with the delay of apoptosis. ICE is a common pathway to apoptosis and thus pretreatment of ICE inhibitor could partially inhibit the apoptosis of both cells. Interestingly, E. coli had prolonged the life-span of polymorphonuclear leukocytes, but it accelerated apoptosis of monocytes in spite of the over-expression of bcl-2. Conclusion : Polymorphonuclear leukocytes and monocytes might have different apoptotic mechanisms to maintain their homeostasis. Polymorphonuclear leukocytes have only one way mechanism to apoptosis namely via Fas-FasL, whereas, monocytes have both Fas-FasL and bcl-2 mechanisms. These differences may be associated with their primary functions and natural life-span. Direct stimulation by live E. coli accelerated apoptosis in monocytes in spite of the over-expression of inhibitor of apoptosis, i.e. bcl-2. It could be due to activation of other apoptotic mechanisms by E. coli which may detour anti-apoptotic action of bcl-2, or direct inhibition of bcl-2 function. (Korean Journal of Infectious Disease 31:279-290, 1999)

Enhanced Neovascularization by Simultaneous Transplantation of Peripheral Blood CD34+ Hematopoietic Stem Cells and CD14+ Monocytes

( Ji Eun Lee ),( Hyun Ok Kim ),( Si Young Song ),( Han Soo Kim ) 한국조직공학과 재생의학회 2010 조직공학과 재생의학 Vol.7 No.2

Formation of new blood vessels is required for normal embryonic development and healing of damaged tissues, but also is essential for tumor growth. Although CD34+VEGF-R2(KDR)+ endothelial progenitor cells and CD14+ monocytes in human peripheral blood are known to actively participate in angiogenesis and vasculogenesis, the clear role of monocytes in the process of neovascularization is matter of debate. Here, we investigated whether a combination of two types of cells shows synergism in tumor-induced neovascularization. Fluorescently labeled purified CD34+ HSCs, CD14+ monocytes or combination of CD34+ HSCs and CD14+ monocytes were intratumorally injected into nude mice bearing human tumor of pancreatic adenocarcinoma. CD14+ monocytes or combination of CD34+ HSCs and CD14+ monocytes. Injection of a mixture of the 2 subsets resulted in improved neovascularization in vivo to any single-cell-type transplantation. These data demonstrate that human CD14+ monocytes as well as CD34+ HSCs can differentiate along the endothelial lineage in a specific permissive environment and thus this combination represent an autologous transplantable cell source for therapeutic neovasculogenesis.

Decreased thrombomodulin mRNA expression on peripheral monocytes in disseminated intravascular coagulation patients relates to poor outcomes: The ex vivo effects of lipopolysaccharide and thrombin on monocyte thrombomodulin and CD14 mRNA

Hong, S.K.,Kim, J.E.,Han, K.S.,Kim, H.K. Pergamon Press ; Elsevier Science Ltd 2013 Thrombosis research Vol.132 No.3

Background: Monocytes express substantial amounts of thrombomodulin, which is consumed throughout ongoing thrombin generation. The modulation of thrombomodulin may aggravate intravascular fibrin deposition and the clinical course of disseminated intravascular coagulation (DIC). Although thrombomodulin restoration has received considerable attention, no reports have been published on the in vivo expression status of thrombomodulin. CD14 expression on monocytes is important for regulation of the inflammatory response. We used an ex vivo stimulation study to evaluate the association of the levels of monocyte-expressed thrombomodulin and CD14 messenger RNA (mRNA) with the severity and prognosis of disseminated intravascular coagulation. Methods: A total of 78 patients with suspected DIC were enrolled. Thrombomodulin and CD14 mRNA levels were measured in peripheral blood by real-time quantitative reverse-transcription polymerase chain reaction. Thrombomodulin and CD14 mRNA were also assessed in ex vivo cultures of peripheral whole blood that were stimulated by lipopolysaccharide or thrombin. Results: The levels of monocyte-expressed thrombomodulin mRNA were significantly lower in the non-survivors than in the survivors. A low level of monocyte-expressed thrombomodulin mRNA was a significant prognostic marker, but CD14 did not possess prognostic power. Monocyte-expressed CD14 mRNA correlated significantly with the severity of DIC in survivors. In addition, stimulation of ex vivo cultures of whole blood demonstrated that thrombin upregulates both thrombomodulin and CD14 mRNA, and lipopolysaccharide downregulates thrombomodulin mRNA. Conclusions: The downregulation of thrombomodulin on monocytes reflects the decompensated status of physiological defenses against hypercoagulopathy and represents the poor prognosis in DIC. The expression levels of thrombomodulin on monocytes may be a useful marker to screen for candidates eligible for recombinant thrombomodulin therapy in future.

Prominent Inflammatory Features of Monocytes/Macrophages in Acute Calcium Pyrophosphate Crystal Arthritis: a Comparison with Acute Gouty Arthritis

정지혜,정재형,이정선,오지선,김용길,이창근,유빈,홍석찬 대한면역학회 2019 Immune Network Vol.19 No.3

Calcium pyrophosphate (CPP) crystals can present as acute inflammatory arthritis which is known as an acute CPP crystal arthritis. Although monocytes/macrophages have been shown to play a role in the initiation of crystal-mediated inflammatory responses, differences in their phenotypes between acute CPP crystal arthritis and acute gouty arthritis have not yet been investigated. We examined the immunological characteristics of synovial monocytes/macrophages in patients with acute CPP crystal and acute gouty arthritis. CD14+CD3−CD19−CD56− cell frequencies in synovial fluid mononuclear cells (SFMCs) were measured. Expression of pro- and anti-inflammatory cytokines and markers was determined. The SFMCs were dominated by a population of monocytes/macrophages in acute CPP crystal arthritis similar to that in acute gout. Synovial monocytes/macrophages showed the phenotypes of infiltrated monocytes as shown by expression of CD88, C-C chemokine receptor type 2, myeloid-related protein (MRP)8 and MRP14 but not proto-oncogene tyrosine-protein kinase MER. Comparatively, the CD14+ cells from patients with acute CPP crystal arthritis had similar high levels of IL-1β and TNF-α production but significantly lower expression of IL-10 and M2 marker (CD163). The monocytes/macrophages had the capacity to produce IL-8 in response to CPP crystals. Proinflammatory features were more dominant in monocytes/macrophages during acute CPP crystal arthritis than those during acute gouty arthritis.

내독소, Cytokines, 대장균 자극에 의한 말초 혈구의 Apoptosis

최정현,신완식,김희정,허동호 대한감염학회 1999 감염 Vol.31 No.4

Exacerbation of Japanese Encephalitis by CD11chi Dendritic Cell Ablation Is Associated with an Imbalance in Regulatory Foxp3+ and IL-17+CD4+ Th17 Cells and in Ly-6Chi and Ly-6Clo Monocytes

최진영,김진형,Ajit Mahadev Patil,김성범,Erdenebileg Uyangaa,Ferdaus Mohd Altaf Hossain,어성국 대한면역학회 2017 Immune Network Vol.17 No.3

Japanese encephalitis (JE) is neuroinflammation characterized by uncontrolled infiltration of peripheral leukocytes into the central nervous system (CNS). We previously demonstrated exacerbation of JE following CD11chi dendritic cell (DC) ablation in CD11c-DTR transgenic mice. Moreover, CD11chi DC ablation led to abnormal differentiation of CD11b+Ly-6Chi monocytes and enhanced permeability of the blood-brain barrier (BBB), resulting in promoting the progression of JE. Here, we examined changes in lymphoid and myeloid-derived leukocyte subpopulations associated with pro- and anti-inflammation during JE progression. The analyses of this study focused on regulatory CD4+Foxp3+ regulatory T cells (Tregs), IL-17+CD4+ Th17 cells, and CD11b+Ly-6Chi and Ly-6Clo monocytes. CD11chi DC ablation resulted in the accumulation of IL-17+CD4+ Th17 cells in the CNS, thereby leading to lower ratio of Tregs to Th17 cells. This result was corroborated by the higher expression levels of IL-17 and RORgT in CD4+ T cells from the brains of CD11chi DC-ablated mice. In addition, CD11chi DC-ablated mice showed higher frequency and total number of inflammatory CD11b+Ly-6Chi monocytes, whereas CD11b+Ly-6Clo monocytes were detected with lower frequency and total number in CD11chi DC-ablated mice. Furthermore, CD11chi DC ablation altered the phenotype and function of CD11b+Ly-6Clo monocytes, resulting in lower levels of activation marker and anti-inflammatory cytokine (IL-10 and TGF-b) expression. Collectively, these results indicate that CD11chi DC ablation caused an imbalance in CD4+ Th17/Treg cells and CD11b+Ly-6Chi/Ly-6Clo monocytes in the lymphoid tissue and CNS during JE progression. This imbalanced orchestration of pro- and anti-inflammatory leukocytes following CD11chi DC ablation may contribute to the exacerbation of JE.

Expression of Toll-like receptors 3, 7, 9 and cytokines in feline infectious peritonitis virus-infected CRFK cells and feline peripheral monocytes

Megat Hamzah Megat Mazhar Khair,Gayathri Thevi Selvarajah,Abdul Rahman Omar,Farina Mustaffa-Kamal 대한수의학회 2022 Journal of Veterinary Science Vol.23 No.2

Background: The role of Toll-like receptors (TLRs) in a feline infectious peritonitis virus(FIPV) infection is not completely understood. Objectives: This study examined the expression of TLR3, TLR7, TLR9, tumor necrosis factoralpha(TNF-α), interferon (IFN)-β, and interleukin (IL)-10 upon an FIPV infection in Crandell-Reese feline kidney (CRFK) cells and feline monocytes. Methods: CRFK cells and monocytes from feline coronavirus (FCoV)-seronegative cats andFCoV-seropositive cats were infected with type II FIPV-79-1146. At four, 12, and 24 hours postinfection(hpi), the expression of TLR3, TLR7, TLR9, TNF-α, IFN-β, and IL-10, and the viralload were measured using reverse transcription quantitative polymerase chain reaction. Viralprotein production was confirmed using immunofluorescence. Results: FIPV-infected CRFK showed the upregulation of TLR9, TNF-α, and IFN-βexpression between 4 and 24 hpi. Uninfected monocytes from FCoV-seropositive cats showedlower TLR3 and TLR9 expression but higher TLR7 expression compared to uninfectedmonocytes from FCoV-seronegative cats. FIPV-infected monocytes from FCoV-seropositivecats downregulated TLR7 and TNF-α expression between 4 and 24 hpi, and 4 and 12 hpi,respectively. IFN-β was upregulated early in FIPV-infected monocytes from FCoV-seropositivecats, with a significant difference observed at 12 hpi compared to FCoV-seronegative cats. The viral load in the CRFK and FIPV-infected monocytes in both cohorts of cats was similarover time.ConclusionTLR7 may be the key TLR involved in evading the innate responseagainst inhibiting TNF-α production. Distinct TLR expression profiles between FCoVseronegativeand FCoV-seropositive cats were observed. The associated TLR that plays a rolein the induction of IFN-β needs to be explored further.

개 말초혈액(末稍血液)에서 monocytes 분리(分離)

김정배,이방환,Kim, Jeoung-bae,Lee, Bang-whan 대한수의학회 1989 大韓獸醫學會誌 Vol.29 No.2

Pure separation of various leukocytes is required for the assessment of their roles in immunological and phisiological function. In this study, pure separation of monocytes from canine peripheral blood was attempted. At first, mononuclear cells (PBMC) were separated by ficoll-hypaque gradient method and then monocytes were recovered from PBMC suspensions in sucrose gradient Sol. (PBMC-Sucrose), autologous plasma (PBMC-Plasma) and autologous serum (PBMC-Serum) incubated at $37^{\circ}C$ for 2 hours. 1. In the separation of PBMC by ficoll-hypaque gradient method in canine blood, higher relative centrifugal force (RCF) was required, as high as more than 1,300xg RCF for 40 minutes, for clear formation of PBMC layer than that in human blood as usually used 400xg RCF for 40 minutes. 2. In monocytes-separation from three PBMC suspensions following PBMC separation, recovery-, purity- and viability-rate of monocytes showed better results in PBMC-Plasma and PBMC-Serum than in PBMC-Sucrose suspension, particulary showing better results from PBMC suspensions performed by centrifugation at 1,500xg RCF for 40 minutes.