Chemokines and chemokine receptors play a role in migration of circulating leukocytes to the region of inflammation. Human LZIP is an uncharacterized transcription factor and is known to participate in leukotactin (Lkn)-1/CCL15-induced cell migration. We investigated the role of human LZIP in expression of CC chemokine receptors (CCRs) and its involvement in monocyte migration. RNase protection analysis showed that LZIP increased mRNA expression of CCR2 and CCR1 in THP-1 cells. Surface expressions of both CCR2 and CCR1 were also increased by LZIP. Results from an electrophoretic mobility shift assay showed that LZIP binds to the C/EBP element in the CCR2 promoter. LZIP also enhanced the chemotactic activities of monocyte chemoattractant protein-1/CCL2 and Lkn-1. These results suggest that LZIP regulates expression of chemokine receptors that are involved in monocyte migration.
Atherosclerosis/drug therapy/etiology ; CCAAT-Enhancer-Binding Proteins/genetics/immunology/*metabolism ; Cell Line, Tumor ; Cell Movement/drug effects/*physiology ; Chemokine CCL2/*pharmacology ; Chemokines, CC/pharmacology ; Cyclic AMP Response Element-Binding ; Humans ; Macrophage Inflammatory Proteins/pharmacology ; Monocytes/drug effects/metabolism ; Promoter Regions, Genetic ; Protein Binding ; RNA, Messenger/analysis ; Receptors, CCR1/biosynthesis/genetics ; Receptors, CCR2/*biosynthesis/genetics ; Transcriptional Activation/drug effects ; Transfection ; Transgenes

This study was aimed to explore the expression of MIP-1alpha, MCP-1 and their receptors CCR-1, CCR-2 in bcr/abl fusion gene positive CML cells, and to study the effects of P210(bcr/abl) fusion protein tyrosine kinase on expression of MIP-1alpha, MCP-1 and their receptors CCR-1, CCR-2 mRNAs in chronic myeloid leukemia cells. The expression levels of MIP-1alpha, MCP-1 and their receptors CCR-1, CCR-2 mRNA were detected by semi-quantitative RT-PCR in bcr/abl negative cells, bcr/abl positive cells, and P210(bcr/abl)-Rb-C-Box positive cells. The results showed that MIP-1alpha and CCR-1 mRNAs were expressed in bcr/abl negative cells, but not in positive cells. Both MCP-1 and CCR-2 mRNA cannot be detected in both bcr/abl positive and negative cells. After inhibiting P210(bcr/abl) tyrosine kinase activity by Rb-C-Box, expressions of MIP-1alpha and CCR-1 mRNAs were restored to normal (similar to P210(bcr/abl) negative cells). It is concluded that P210(bcr/abl) fusion protein inhibits the expression of MIP-1alpha and CCR-1 in chronic myeloid leukemia cells, but does not inhibit MCP-1 and CCR-2 mRNA expressions in these leukemia cells.
Chemokine CCL2 ; biosynthesis ; genetics ; Chemokine CCL3 ; Chemokine CCL4 ; Humans ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; metabolism ; Macrophage Inflammatory Proteins ; biosynthesis ; genetics ; Receptors, CCR1 ; Receptors, CCR2 ; Receptors, Chemokine ; biosynthesis ; genetics ; Tumor Cells, Cultured

Cells, Cultured ; Chemokine CCL4 ; Chemotaxis, Leukocyte ; drug effects ; Endothelial Cells ; cytology ; metabolism ; Homocysteine ; pharmacology ; Humans ; Macrophage Inflammatory Proteins ; biosynthesis ; genetics ; Monocytes ; physiology ; RNA, Messenger ; biosynthesis ; genetics ; Umbilical Veins ; cytology

OBJECTIVETo investigate the effect of homocysteine (HCY) on activation of nuclear factor (NF-kappaB) and inhibitory factor IkappaB-alpha in human monocyte cell line THP-1, as well as its association with macrophage inflammatory protein (MIP-1alpha) upregulation.

METHODSTHP-1 monocytes were incubated with HCY, with and without NF-kappaB inhibitor pyrolidine dithiocarbamate (PDTC) pretreatment. Northern blot analysis and flow cytometry were used to detect MIP-1alpha mRNA and protein respectively. The nuclear protein NF-kappaB P65 subunit and the inhibitory protein IkappaB-alpha were analyzed by Western blotting.

RESULTSCompared with controls, HCY, at a concentration of 0.1 mmol/L, was able to enhance the expression of MIP-1alpha mRNA (up to 3.69-fold) and protein (1.16-fold) in THP-1 monocytes, as well as enhance NF-kappaB P65 transcription to nuclear proteins. These actions were significantly suppressed after pretreatment with 100 micromol/L PDTC for 30 minutes before HCY incubation; whereas incubation of THP-1 monocytes with PDTC only had no effect on both the expression of MIP-1alpha and nuclear transcription of NF-kappaB P65. Moreover, the level of IkappaB-alpha protein in THP-1 monocytes decreased after a 30-minute incubation with HCY, which gradually increased after 120 minutes.

CONCLUSIONSHomocysteine at a pathologic concentration stimulates MIP-1alpha expression in THP-1 monocytes, probably via NF-kappaB activation. Such activation may be caused by enhanced phosphorylation and degradation of the inhibitor protein IkappaB-alpha.

Cell Line, Tumor ; Chemokine CCL3 ; Chemokine CCL4 ; Homocysteine ; pharmacology ; Humans ; I-kappa B Proteins ; metabolism ; Leukemia, Monocytic, Acute ; metabolism ; pathology ; Macrophage Inflammatory Proteins ; biosynthesis ; genetics ; Monocytes ; metabolism ; NF-KappaB Inhibitor alpha ; NF-kappa B ; antagonists & inhibitors ; Phosphorylation ; Proline ; analogs & derivatives ; pharmacology ; RNA, Messenger ; biosynthesis ; genetics ; Thiocarbamates ; pharmacology ; Transcription Factor RelA ; biosynthesis ; genetics ; Transcription, Genetic

A previous report by this laboratory demonstrated that bacterial iron chelator (siderophore) triggers inflammatory signals, including the production of CXC chemokine IL-8, in human intestinal epithelial cells (IECs). Microarray-based gene expression profiling revealed that iron chelator also induces macrophage inflammatory protein 3 alpha (MIP-3alpha)/ CC chemokine-ligand 20 (CCL20). As CCL20 is chemotactic for the cells involved in host adaptive immunity, this suggests that iron chelator may stimulate IECs to have the capacity to link mucosal innate and adaptive immunity. The basal medium from iron chelator deferoxamine (DFO)-treated HT-29 monolayers was as chemotactic as recombinant human CCL20 at equivalent concentrations to attract CCR6+ cells. The increase of CCL20 protein secretion appeared to correspond to that of CCL20 mRNA levels, as determined by real-time quantitative RT-PCR. The efficacy of DFO at inducing CCL20 mRNA was also observed in human PBMCs and in THP-1 cells, but not in human umbilical vein endothelial cells. Interestingly, unlike other proinflammatory cytokines, such as TNF-alpha and IL-1beta, a time-dependent experiment revealed that DFO slowly induces CCL20, suggesting a novel mechanism of action. A pharmacologic study also revealed that multiple signaling pathways are differentially involved in CCL20 production by DFO, while some of those pathways are not involved in TNF-alpha-induced CCL20 production. Collectively, these results demonstrate that, in addition to some bacterial products known to induce host adaptive immune responses, direct chelation of host iron by infected bacteria may also contribute to the initiation of host adaptive immunity in the intestinal mucosa.
Calcium/metabolism ; Cell Movement/drug effects ; Chemokines, CC/genetics/*metabolism ; Deferoxamine/*pharmacology ; Egtazic Acid/analogs & derivatives/pharmacology ; HT29 Cells ; Humans ; Immunity, Mucosal/*drug effects ; Intestinal Mucosa/*drug effects/immunology/metabolism ; Iron Chelating Agents/*pharmacology ; Macrophage Inflammatory Proteins/genetics/*metabolism ; NF-kappa B/metabolism ; Phosphoprotein Phosphatase/physiology ; Protein Transport/drug effects ; Protein-Serine-Threonine Kinases/physiology ; RNA, Messenger/genetics/metabolism ; Receptors, Chemokine/metabolism ; Research Support, Non-U.S. Gov't

OBJECTIVE: To explore the microvessel counts and the expressions of interleukin-8 (IL-8), monocyte chemoattractant protein-1 (MCP-1), and macrophage inflammatory protein-1 ( MIP-1) alpha mRNA in the pathological scar tissues. METHODS: Immunohistochemical method of avidin-biotin complex was used for microvessel counts on the routinely formalin-fixed and paraffin-embedded sections of specimens of hypertrophic scars, keloids, normal skin, and surgical scar, and in situ hybridization for the expressions of IL-8, MCP-1, MIP-1alpha mRNA. RESULTS: The microvessel counts as well as the positive rates and the scorings of IL-8, MCP-1, and MIP-1alpha mRNA were significantly higher in pathological scars than those in the normal skin and surgical scar (all P < 0.05). The microvessel counts were significantly higher in the positive cases of IL-8, MCP-1 and MIP-1alpha mRNA than those in the negative ones (P < 0.05). The close positive correlations were found among the microvessel counts and the expressive scorings of 3 factors (P < 0.05). The close positive correlations were also found among the expressive scorings of IL-8, MCP-1, and MIP-1alpha mRNA in pathological scars. Microvessel counts were significantly higher in hypertrophic scars with the course less than 1 year than those with the course more than 1 year. CONCLUSION IL-8, MCP-1 and MIP-1alpha play important roles in promoting the neovascularization of pathological scars.
Adolescent ; Adult ; Burns ; complications ; Capillaries ; metabolism ; Chemokine CCL2 ; biosynthesis ; genetics ; Chemokine CCL3 ; Chemokine CCL4 ; Cicatrix ; etiology ; metabolism ; Female ; Humans ; Interleukin-8 ; biosynthesis ; genetics ; Macrophage Inflammatory Proteins ; biosynthesis ; genetics ; Male ; Middle Aged ; RNA, Messenger ; biosynthesis ; genetics ; Skin ; blood supply

OBJECTIVETo study the effect of macrophage inflammatory protein-1alpha(MIP-1alpha) and its mRNA on airway inflammation of mouse with induced asthma.

METHODSSeventy male BALB/C mice were randomly divided into the control group and asthma group (including 7 subgroups, 10 mice each). The control group included group A(24) (the lavaging subgroup was sacrificed 24 h after the last challenge) and group A(0) (the non-lavaging subgroup was sacrificed from 18 h to 24 h after the last challenge); asthma group included group B(3) (the lavaging subgroup was sacrificed 3 h after the last challenge), group B(8) (the lavaging subgroup was sacrificed 8 h after the last challenge), group B(24) (the lavaging subgroup was sacrificed 24 h after the last challenge), group B(36) (the lavaging subgroup was sacrificed 36 h after the last challenge) and group B(0) (the non-lavaging subgroup was sacrificed from 18 h to 24 h after the last challenge). In the experiment, the mice model of asthma was established by the ovalbumin (OVA) challenge methods. Eosinophils (EOS) numbers and differentiated cell numbers in bronchoalveolar lavage fluid (BALF) were counted; the concentrations of MIP-1alpha in serum and BALF were measured by sandwich enzyme-linked immunosorbent assay (sandwich ELISA); the protein expressions of MIP-1alpha were detected by immunohistochemical techniques; the mRNA expressions of MIP-1alpha were determined by in situ hybridization technique.

RESULTS(1) The concentrations of MIP-1alpha in BALF and serum of group B(3) [(30.2 +/- 4.2) pg/ml, (30.8 +/- 4.6) pg/ml], group B(8) [(35.3 +/- 4.9) pg/ml, (34.9 +/- 5.1) pg/ml], group B(24) [(42.9 +/- 5.8) pg/ml, (41.7 +/- 6.3) pg/ml] and group B(36) [(37.8 +/- 4.7) pg/ml, (35.7 +/- 4.9) pg/ml] were significantly higher than those of group A(24) [(20.9 +/- 3.8) pg/ml, (22.4 +/- 4.3) pg/ml] (P < 0.01); the concentrations of MIP-1alpha in BALF and serum went up at 3 h, reached peak at 24 h, and had descended at 36 h. (2) Immunohistochemistry showed that the protein expressions of MIP-1alpha around the bronchus of group B(0) [(26.4 +/- 6.2)%] were significantly elevated as compared to those of group A(0) [(10.3 +/- 2.5)%] (P < 0.01), the epithelial cell was the chief expression cell. (3) In situ hybridization showed that the mRNA expressions of MIP-1alpha around the bronchus of group B(0) [(23.9 +/- 4.2)%] were significantly increased when compared to those of group A(0) [(8.7 +/- 1.8)%] (P < 0.01), the epithelial cell was the chief expression cell. (4) There was a significant correlation between the concentrations of MIP-1alpha and the numbers of EOS in BALF and between the concentrations of MIP-1alpha and the percentage of EOS numbers in the total cell numbers (EOS%) in BALF.

CONCLUSIONSMIP-1alpha protein and MIP-1alpha mRNA were found strongly expressed in mouse asthma model, the epithelial cell was the chief expression cell; the kinetic characteristic of MIP-1alpha showed that its level increased at 3 h, reached peak at 24 h and declined at 36 h; MIP-1alpha and EOS gathering had a significant correlation.

Animals ; Asthma ; blood ; genetics ; Bronchoalveolar Lavage Fluid ; chemistry ; Chemokine CCL3 ; Chemokine CCL4 ; Disease Models, Animal ; Enzyme-Linked Immunosorbent Assay ; In Situ Hybridization ; Macrophage Inflammatory Proteins ; blood ; genetics ; Male ; Mice ; Mice, Inbred BALB C ; RNA, Messenger ; genetics ; metabolism

In order to explore the possible role of CC chemokine ligand 20 (CCL20) and its receptor CC chemokine receptor 6 (CCR6) in the pathogenesis of psoriasis, the expression levels of mRNA of them in psoriatic lesions were investigated. The skin biopsies were collected from skin lesions in 35 cases of psoriasis vulgaris and 18 normal controls. RT-PCR was used to semi-quantitatively analyze the mRNA expression of CCL20 and CCR6 in the psoriatic lesions and the normal skin tissues. The results showed that the mRNA of CCL20 and CCR6 was present in every specimen. The expression levels of CCL20 mRNA in skin lesions were 1.1397 +/- 0.0521, which were greatly higher than those in normal controls (0.8681 +/- 0.0308) (P<0.001). The expression levels of CCR6 mRNA in skin lesions were 1.1103 +/- 0.0538, significantly higher than in the controls (0.9131 +/- 0.0433, P<0.001). These findings indicate that up-regulated expression of CCL20 and CCR6 mRNA might be related to the pathogenesis of psoriasis.
Adolescent ; Adult ; Aged ; Chemokine CCL20 ; Chemokines, CC ; biosynthesis ; genetics ; Female ; Humans ; Macrophage Inflammatory Proteins ; biosynthesis ; genetics ; Male ; Middle Aged ; Psoriasis ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Receptors, CCR6 ; Receptors, Chemokine ; biosynthesis ; genetics

To investigate the role of Interleukin-17 (IL-17), Interferon-gamma (IFN-gamma), and macrophage inflammatory protein-3 alpha (MIP-3alpha) in the pathogenesis of psoriasis, reverse transcriptase-polymerase chain reaction (RT-PCR) was used to semi-quantitatively analyze the mRNA expression of IL-17, IFN-gamma, and MIP-3alpha in 31 psoriatic lesions and 16 normal skin tissues. The results showed that the mRNA of the three cytokines was present in all specimens. And the expression level of IL-17 mRNA in skin lesions was 1.1416 +/- 0.0591, which was significantly higher than that in normal controls (0.8788 +/- 0.0344, P<0.001). The expression levels of IFN-gamma mRNA were 1.1142 +/- 0.0561 and 0. 9050 +/- 0.0263, respectively, with significant difference (P<0.001). And the expression levels of MIP-3alpha mRNA in psoriatic lesions was 1.1397 +/- 0.0521, which was markedly higher than that in normal controls (0.8681 +/- 0.0308, P<0.001). These findings indicate that up-regulated expression of IL-17, IFN-gamma, and MIP-3alpha might be involved in the pathogenesis of psoriasis.
Adolescent ; Adult ; Aged ; Chemokine CCL20 ; Chemokines, CC ; biosynthesis ; genetics ; Female ; Humans ; Interferon-gamma ; biosynthesis ; genetics ; Interleukin-17 ; biosynthesis ; genetics ; Macrophage Inflammatory Proteins ; biosynthesis ; genetics ; Male ; Middle Aged ; Psoriasis ; metabolism ; RNA, Messenger ; biosynthesis ; genetics

OBJECTIVETo discuss the correlation between acute rejection episodes and expression of regulated upon activation normal T cell expressed and secreted (RANTES) and macrophage inflammatory protein-1 alpha (MIP-1 alpha) gene following kidney transplantation.

METHODSA total of 76 kidney biopsies (episode biopsy) were performed on both 57 patients with allograft dysfunction following transplantation and 19 patients without rejection, and the latter were served as controls. All acute rejections were confirmed by histological examination. The expressions of RANTES and MIP-1 alpha mRNA in all samples were assayed by reverse transcription polymerase chain reaction.

RESULTSThirty-eight (66.7%) of 57 cases with acute rejection had strong expression of RANTES, and 41 (72%) had expression of MIP-1 alpha. Only four (21%) of 19 controls had mild expression of RANTES, and 5 (26%) had expression of MIP-1 alpha, which evoked a statistical significance (P < 0.001) when compared with those with acute rejections.

CONCLUSIONSThe expressions of chemokines RANTES and MIP-1 alpha are important in the process of immune reaction in the rejection of transplanted kidney, which may imply a potential way for early diagnosis and treatment of acute rejection after further research.

Acute Disease ; Adolescent ; Adult ; Biomarkers ; Biopsy ; Chemokine CCL4 ; Chemokine CCL5 ; biosynthesis ; genetics ; Female ; Graft Rejection ; metabolism ; pathology ; Humans ; Kidney ; metabolism ; pathology ; Kidney Transplantation ; Macrophage Inflammatory Proteins ; biosynthesis ; genetics ; Male ; Middle Aged ; Reverse Transcriptase Polymerase Chain Reaction