Relationship between matrix metalloproteinases MMP-2, MMP-9, tissue inhibitor of matrix metalloproteinases-1 and IL-5, IL-8 in nasal polyps

BACKGROUND: Nasal polyps (NP), a subgroup of chronic rhinosinusitis, are characterized by interleukin 5 (IL-5) mediated infiltration of eosinophils in sinus mucosa, leading to pseudostratified ciliated columnar epithelium, thickening of the epithelial basement membrane and tissue edema. Matrix metalloproteinases (MMP) constitute a large group of Zn2+ dependent endopeptidases with the ability to degrade extracellular matrix and are possibly responsible for the development of tissue edema in chronic sinusitis. OBJECTIVE: The aim of this study was to determine the expression of MMP-2, MMP-9 and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) mRNA and to locate the distribution of MMP-2, MMP-9 and TIMP-1 by immunohistochemistry in ethmoid sinus mucosa in NP. Furthermore the correlation between IL-5 or IL-8 and MMP-2, MMP-9 or TIMP-1 is examined. METHODS: Nasal polyps of 33 patients and 18 specimens of inferior turbinate mucosa were examined by real time RT-PCR for MMP-2, MMP-9, TIMP-1, IL-5 and IL-8 mRNA expression. Immunohistochemical labeling for MMP-2, MMP-9 and TIMP-1 was performed. RESULTS: Differences between both locations were detectable for MMP-9 (P < 0.001) and IL-5 (P=0.003) but not for MMP-2 (P=0.278), TIMP-1 (P=0.515) and IL-8 (P=0.386). Correlation was detected only between TIMP-1 and IL-5 (r=0.422, P =0.014). Cytoplasmic staining of MMP-2 was present in the apical part of the ciliated cells, submucosal glands and in smooth muscle cells. Matrix metalloproteinase-9 was expressed in surface epithelium, in seromucous glands and in polymorphonuclear cells. CONCLUSIONS: Expression of MMP-9 and IL-5 mRNA are associated with NP. The correlation between IL-5 and TIMP-1 indicates the role of TIMP-1 in maintaining the homeostasis in NP.     

Different expression profiles of the lysyl oxidases and matrix metalloproteinases in human ACL fibroblasts after co-culture with synovial cells

Purposes The anterior cruciate ligament (ACL) has poor functional healing response. The synovial tissue surrounding ACL ligament might be a major regulator of the microenvironment in the joint cavity after ACL injury, thus affecting the repair process. Using transwell co-culture, this study explored the direct influence of human synovial cells (HSCs) on ACL fibroblasts (ACLfs) by characterizing the differential expression of the lysyl oxidase family (LOXs) and matrix metalloproteinases (MMP-1, ?2, ?3), which facilitate extracellular matrix (ECM) repair and degradation, respectively. Methods The mRNA expression levels of LOXs and MMP-1, ?2, ?3 were analyzed by semi-quantitative PCR and quantitative real-time PCR. The protein expression levels of LOXs and MMP-1, ?2, ?3 were detected by western blot. Results We found that co-culture resulted in an increase in the mRNAs of LOXs in normal ACLfs and differentially regulated the expression of MMPs. Then we applied 12% mechanical stretch on ACLfs to induce injury and found the mRNA expression levels of LOXs in injured ACLfs were decreased in the co-culture group relative to the mono-culture group. Conversely, the mRNA expression levels of MMPs in injured ACLfs were promoted in the co-culture group compared with the mono-culture group. At translational level, we found that LOXs were lower while MMPs were highly expressed in the co-culture group compared to the mono-culture group. Conclusions The co-culture of ACLfs and HSCs, which mimicked the cell-to-cell contact in a micro-environment, could contribute to protein modulators for wound healing, inferring the potential reason for the poor self-healing of injured ACL.     

Complement activation triggers metalloproteinases release inducing cervical remodeling and preterm birth in mice

Inflammation is frequently linked to preterm delivery (PTD). Here, we tested the hypothesis that complement activation plays a role in cervical remodeling and PTD. We studied two mouse models of inflammation-induced PTD. The first model was induced by vaginal administration of lipopolysaccharide (LPS) and the second one by administration of progesterone antagonist RU486. Increased cervical C3 deposition and macrophages infiltration and increased serum C3adesArg and C5adesArg levels were observed in both models when compared to gestational age matched controls. A significant increase in collagen degradation, matrix metalloproteinase 9 (MMP-9) activity and tissue distensibility was observed in the cervix in both models. Mice deficient in complement receptor C5a did not show increased MMP-9 activity and cervical remodeling and did not deliver preterm in response to LPS or RU486, suggesting a role for C5aR in the cervical changes that precede PTD. In vitro studies show that macrophages release MMP-9 in response to C5a. Progesterone diminished the amount of C5aR on the macrophages surface, inhibited the release of MMP-9 and prevented PTD. In addition, macrophages depletion also prevented cervical remodeling and PTD in LPS-treated mice. Our studies show that C5a-C5aR interaction is required for MMP-9 release from macrophages, and the cervical remodeling that leads to PTD. Complement inhibition and supplementation with progesterone may be good therapeutic options to prevent this serious pregnancy complication.     

TGF-beta1 antisense therapy modulates expression of matrix metalloproteinases in keloid-derived fibroblasts

Transforming growth factor-beta1 (TGF-beta1) has been identified as an important regulator of wound healing. Recent developments in molecular therapy offer exciting prospects for the modulation of wound healing, specifically those targeting TGF-beta1. The purpose of this study was to analyze the effect of TGF-beta1 targeting on the expression of matrix metalloproteinases (MMPs) in fibroblasts cultured from earlobe keloids. The expression of MMP-2 and -9 in tissue samples from keloids was investigated by immunohistochemistry. The effect of TGF-beta1 targeting using antisense oligonucleotides on the expression of MMPs in keloid-derived fibroblasts was analysed by ELISA and multiplex RT-PCR. Immunohistochemical studies demonstrated an increased expression of MMP protein in tissue samples from keloids compared to normal human skin. Antisense TGF-beta1 oligonucleotide treatment significantly downregulated MMP-9 secretion in vitro. In conclusion, TGF-beta1 antisense oligonucleotide technology may be a potential therapeutic option for the inhibition of proteolytic tissue destruction in keloids.     

Acute stress results in skin corticotropin-releasing hormone secretion,mast cell activation and vascular permeability,an effect mimicked by intradermal corticotropin-releasing hormone and inhibited by histamine-1 receptor antagonists

BACKGROUND: Mast cells play an important role in allergic inflammation by releasing vasoactive molecules, proteases and cytokines. Corticotropin-releasing hormone (CRH) and its structural analogue urocortin (Ucn) were shown to trigger skin mast cell activation and vascular permeability. We investigated the effect of acute stress on rat skin vascular permeability and CRH secretion, as well as the effect of intradermal CRH, and that of two histamine-1 receptor antagonists, azelastine and olopatadine, on vascular permeability. METHODS: Rats were stressed by restraint and vascular permeability was assessed by extravasation of (99)Tc-gluceptate, while mast cell activation was determined by skin rat mast cell protease-1 (RMCP-1) content. Skin CRH content was evaluated by ELISA. The effect of intradermal injection of CRH and Ucn, as well as that of two histamine-1 receptor antagonists, azelastine and olopatadine, was assessed by Evan's blue extravasation. Purified rat peritoneal mast cells (RPMCs) were also pretreated with azelastine (24 microM) or olopatadine (133 microM) for 5 min before challenge with compound 48/80 (0.5 microg/ml) for 30 min. Histamine secretion was measured fluorometrically. Intracellular Ca(2+) ions were evaluated in RPMCs loaded with calcium crimson and stimulated with compound 48/80. RESULTS: Acute stress increased skin vascular permeability and CRH content, while it decreased RMCP-1. Intradermal injection of CRH or Ucn induced substantial Evan's blue extravasation that was inhibited by pretreatment with azelastine (24 microM) and olopatadine (133 microM). Both antihistamines also inhibited histamine release and intracellular increase of Ca(2+) ions from RPMCs stimulated by compound 48/80. CONCLUSIONS: These results indicate that acute stress increases skin CRH that can trigger mast cell-dependent vascular permeability, effects inhibited by certain histamine-1 receptor antagonists, possibly acting to reduce intracellular Ca(2+) ion levels.     

Hypoxia promotes the proliferation of cervical carcinoma cells through stimulating the secretion of IL-8

To explore whether hypoxia and interleukin 8 (IL-8) regulate the viability and apoptosis of cervical carcinomas cells and the possible mechanism. We evaluated the expression of hypoxia inducible factor-1α (HIF-1α), IL-8 and its receptors (CXCR1 and CXCR2) in cervical cancer and cervicitis tissues by immunohistochemistry. Then the effects of hypoxia and IL-8 on the viability and apoptosis of HeLa and SiHa cells were detected by the SRB and apoptosis assays. Here we observed that the expression of HIF-1α, IL-8 and CXCR1 in cervical cancer tissues was significantly higher than that in cervicitis tissues. Hypoxic condition stimulated the secretion of IL-8 and the expression of CXCR1 and CXCR2 on HeLa and SiHa cells. Recombinant human IL-8 enhanced the viability and reduced the apoptosis in HeLa and SiHa cells. HeLa and SiHa cells cultured in 1% oxygen showed the increased viability and apoptosis, and the former effect could be partly reversed by anti-human IL-8 neutralizing antibody. This data suggested that IL-8 secreted by cervical carcinomas cells induced by hypoxia can stimulate the viability of cervical carcinomas cells in an autocrine dependent manner, and contribute to the pathogenesis of cervical cancer.     

Cell-associated interleukin-8 in cord blood of term and preterm infants

To assess the effect of gestational age and labor on the interleukin-8 (IL-8) concentration in whole cord blood and serum, IL-8 levels were determined simultaneously in cord blood serum and lysate in 134 infants. Following the elimination of some of the samples due to exclusion criteria, the data for 99 uninfected infants (71 term and 28 preterm) and 9 infants with neonatal bacterial infection delivered either vaginally or by elective or emergency cesarean section were analyzed. The effects of labor and gestational age were tested by analysis of variance. IL-8 was not detectable in the serum of 25 infants, whereas IL-8 levels in whole blood were measurable in all of the samples. The median IL-8 conncentrations in whole cord blood lysate were 106 pg/ml (range, 20 to 415 pg/ml) in preterm infants and 176 pg/ml (range, 34 to 1,667 pg/ml) in term infants. In contrast to the IL-8 levels in serum, IL-8 levels in whole blood were reduced after ECS. Gestational age had no independent effect on the IL-8 concentrations in either serum or whole blood; these concentrations increased in infected infants after labor. We conclude that the neonatal proinflammatory response to labor stress was more evident in the concentrations of IL-8 in whole blood than in serum. The levels of IL-8 in whole-blood lysate reflect proinflammatory stimulation in neonates and may be a useful diagnostic tool for the early diagnosis of neonatal infection.     

Surface-adsorbed alpha1-microglobulin modulation of human fibroblasts spreading and matrix metalloproteinases release

The lipocalin alpha1-microglobulin (alpha1-m), an immunoregulatory protein produced by human hepatocytes and distributed in various organs and fluids, is physiologically adsorbed onto polymer surfaces from both serum and urine, and its adsorption correlated to the degree of surface hydrophobicity. Starting from the hypothesis that alpha1-m holds a modulatory role at the biomaterials-tissue interface, we have observed a dose-dependent reduction in adhesion of human fibroblasts (cell line MRC-5) seeded onto polystyrene (PS) in a serum-free medium in the presence of adsorbed alpha(1)-m (2.1+/-0.2 x 10(4) cells/cm2 at 200 ng/ml alpha1-m ) compared to cells seeded onto cell grade PS (2.9+/-0.05 x 10(4) cells/cm2) after 72 h. Moreover, in the presence of alpha1-m, adherent MRC-5 cells exhibit an altered shape due to inhibition of cell spreading, and release of matrix metalloproteinase -2 (gelatinase A, MMP-2) by fibroblasts was also increased by 1.6-1.9-fold after 72 h of incubation. These data extend the known spectrum of alpha1-m activities, suggesting a possible role of this protein in the complex series of events occurring at the tissue-biomaterial interface.     

成骨细胞旁分泌影响软骨细胞中MMPs与TIMPs的表达

背景：细胞之间体外共培养能最大限度的模拟体内真实的微环境,细胞划痕实验及炎症因子白细胞介素1β刺激后基质金属蛋白酶及基质金属蛋白酶抑制剂之间的平衡可能破坏,从而导致关节软骨细胞外基质的降解,软骨细胞功能的失调,关节软骨的退变。目的：在成骨细胞上清液与软骨细胞体外共培养下,观察炎症因子白细胞介素1β对体外培养的软骨细胞的迁移、基质金属蛋白酶及组织金属蛋白酶抑制剂表达的影响。方法：实验分为软骨细胞单培养组﹑软骨细胞与成骨细胞上清液共培养组和软骨细胞与成骨细胞上清液共培养+白细胞介素1β组,划痕实验观察3组24 h软骨细胞的迁移变化;半定量PCR实验分析以上3组24 h软骨细胞中基质金属蛋白酶1,2,3,9及组织金属蛋白酶抑制剂1,2,3,4的变化情况。结果与结论：与单培养组比较,共培养组和共培养+白细胞介素1β组细胞迁移率显著增加(P < 0.01);与单培养组比较,共培养组中基质金属蛋白酶1,2,3,9基因表达明显增高(P < 0.05),共培养+白细胞介素1β组基质金属蛋白酶1,3,9基因表达明显增高(P < 0.01);与单培养组比较,共培养组和共培养+白细胞介素1β组中组织金属蛋白酶抑制剂1基因表达明显升高(P < 0.01),组织金属蛋白酶抑制剂3,4基因表达明显下降(P < 0.05)。提示成骨细胞上清液与软骨细胞共培养促进软骨细胞的迁移,增强软骨细胞中基质金属蛋白酶1,2,3,9的基因表达且调节组织金属蛋白酶抑制剂家族的基因表达。白细胞介素1β抑制共培养的软骨细胞迁移及组织金属蛋白酶抑制剂家族的基因表达。        中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程     

Inhibitory effect of water-soluble chitosan on TNF-alpha and IL-8 secretion from HMC-1 cells

Mast cells are known to play an active role as effector cells in allergic inflammation and in diverse immunological and pathological processes. Activated mast cell-derived pro-inflammatory cytokines are important pathologic factors of progression of allergic inflammation. In this study, we investigated whether pro-inflammatory cytokines (TNF-alpha and IL-8) can be induced by calcium stimulation in HMC-1 cells, and high molecular weight water-soluble chitosan (WSC) can inhibit the production of these cytokines. We provided evidence that the secretion of TNF-alpha and IL-8 from HMC-1 cells was induced by Ca2+-ionophore A23187 or Ca2+-ATPase inhibitor TSG. Treatment of WSC (10 microg/ml) prior to stimulation with calcium agonists significantly blocked the secretion of TNF-alpha by 65.1% for A23187 and 87.7% for TSG. IL-8 secretion in response to A23187 or TSG was inhibited by 49.2% for A23187 and 34.1% for TSG, respectively, compared to absence of WSC. These results suggest that WSC has potential regulatory effects on allergic inflammatory diseases by down-modulating Ca2+-induced mast cell activation.     

The role of IL-18 and IL-12 in the modulation of matrix metalloproteinases and their tissue inhibitors in monocytic cells

The matrix metalloproteinases (MMP) are enzymes crucial for the physiological patrol as well as pathological chemotaxis of immune cells to target tissues. The present study examined differential effects of pro-inflammatory [IL-18, IL-12 and tumor necrosis factor (TNF)-alpha] versus anti-inflammatory (IL-4) cytokines on the modulation of MMP and their endogenous tissue inhibitors (TIMP) expression in the U937 cell line. IL-18 and IL-12 separately and synergistically enhanced MMP-2, while TNF-alpha led to the elevation of MMP-9. All pro-inflammatory cytokines enhanced MT1-MMP expression and IL-4 suppressed TNF-alpha-induced MMP-9 expression. This study demonstrated that elevated IL-18 and IL-12, and related pro-inflammatory activity, may be associated with aberrant MMP activity, suggesting modulation of MMP expression using IL-12 and IL-18 antagonists as future therapeutic strategies to attenuate inflammatory and autoimmune disorders.     

Variations in cervical IL-10 and IL-8 concentrations throughout gestation in normal pregnancies

PROBLEM: Data regarding cervical interleukin 18 (IL-8) and IL-10 concentrations during pregnancy is limited. METHOD OF STUDY: This was a cross sectional study of healthy pregnant women. Specimens were collected from the cervical os secretions. IL-8 and IL-10 levels were measured by using enzyme-linked immunosorbent assay. Median (range) cytokine concentrations were derived for each trimester and compared across trimesters. The relationship between gestational age and cytokine levels was assessed by regression analysis. The mean of the ratios of IL-8 to IL-10 was compared in each trimester using anova. RESULTS: The median (range) IL-8 concentrations in cervical secretions were in pg/mL: 1562 (1210-4100), 2460 (1047-4688), 3660 (1451-4748) (P < 0.0021); the median (range) IL-10 concentrations in cervical secretions were in pg/mL: 38.3 (6.8-227.9), 10.9 (0-263.3), 9.5 (0-35.6); the mean IL10/IL-8 x 100 (+/- standard deviation) concentrations were: 3.33 +/- 0.65, 1.47 +/- 0.41, 0.38 +/- 0.52 (P = 0.0035) during the first, second and third trimesters, respectively. CONCLUSION: The patterns of cervical IL-8 concentration is inversely related to gestational age, and the ratio of IL-10/IL-8 decreases with advancing gestation.     

Expression and activities of matrix metalloproteinases under oscillatory shear in IL-1-stimulated synovial cells

Matrix metalloproteinases (MMPs) are known to play a critical role in tissue disintegration, and an elevated level of MMPs is observed in synovium and synovial fluid of joints with rheumatoid arthritis. During joint movement, synovial tissue receives various mechanical stimuli, but effects of mechanical challenges on regulation of MMPs in rheumatic synovium are poorly understood. Focusing on cellular responses to oscillatory fluid shear in human synovial cells, we determined the expression of MMP-1 and MMP-13 by polymerase chain reaction and immunoblotting as well as proteolytic activities of total MMPs by a fibril degradation assay and zymography. The results revealed that approximately 0.5 dyn/cm2 oscillatory shear at 1 Hz not only reduced an mRNA level and a protein level of MMP-1 and MMP-13, but it also decreased collagenase and gelatinase activities of total MMPs. Furthermore, the induction of the MMP expression and activities by interleukin-1 was suppressed by the oscillatory shear. Interestingly, the oscillatory shear upregulated the mRNA expression of TIMP-1 and TIMP-2. Our results support a potential role of oscillatory shear in regulating expression and activities of MMPs in the presence and the absence of proinflammatory cytokine.     

Activity patterns assessed throughout 24-hour recordings in preterm and near term infants

The motility of 10 low-risk infants, aged between 34 and 40 weeks of postmenstrual age, has been continuously recorded for 24 h. Four codes were distinguished: code 1 (absence of motility or occasional occurrence of startles), code 2 (presence of small general or isolated body movements, startles, smiles, grimaces, and other facial activity), code 3 (forceful and prolonged general movements, startles, and stretches), code 4 (vigorous and abrupt general body movements accompanied by crying). Changes with age concern mainly the increase of the duration of code 1 (quiescence) episodes. Confrontation between day and night showed higher levels of motility during the night than during the day. The last weeks before term represent a time for increase in the ability to sustain a quiet behavior and to reorganize day-night motility distribution.     

Polarized secretion of IL-6 and IL-8 by human retinal pigment epithelial cells

A number of cell types situated along interfaces of various tissues and organs such as the peritoneum and the intestine have been shown to secrete inflammatory cytokines in a polarized fashion. Retinal pigment epithelial (RPE) cells are positioned at the interface between the vascularized choroid and the avascular retina, forming part of the blood–retina barrier. These cells are potent producers of inflammatory cytokines and are therefore considered to play an important role in the pathogenesis of ocular inflammation. Whether cytokine secretion by these cells also follows a vectorial pattern is not yet known, and was therefore the subject of this study. Monolayers of human RPE cells (primary cultures and the ARPE-19 cell line) cultured on transwell filters were stimulated to produce IL-6 and IL-8 by adding IL-1β (100 U/ml) to either the upper or the lower compartment. After stimulation, the human RPE cell lines showed polarized secretion of IL-6 and IL-8 towards the basal side, irrespective of the side of stimulation. The ARPE-19 cell line also secreted IL-6 and IL-8 in a polarized fashion towards the basal side after basal stimulation; polarized secretion was, however, not apparent after apical stimulation. The observation that human RPE cells secrete IL-6 and IL-8 in a polarized fashion towards the choroid may represent a mechanism to prevent damage to the adjacent fragile retinal tissue.     

IL-10 secretion increases signal persistence of HEMA-MMA-microencapsulated luciferase-modified CHO fibroblasts in mice

Microencapsulation of cells in a polymer membrane [e.g., poly(hydroxyethyl methacrylate-co-methyl methacrylate) (HEMA-MMA)] has been proposed as a vehicle for the delivery of therapeutic biomolecules, but cells (especially xenogeneic cells) survive only for short times, limiting the utility of this approach. Murine interleukin-10 (mIL-10) has been shown to downregulate the xenogeneic immune response, and we tested the hypothesis that mIL-10 produced by microencapsulated Chinese hamster ovary (CHO) cells would modulate the transplant-site environment leading to prolonged cell function in a xenogeneic model without other immunomodulatory agents. Prior to encapsulation, CHO cells were genetically engineered to express mIL-10 and a firefly bioluminescence protein, luciferase, which allowed for noninvasive tracking of transplanted cells in vivo with the Xenogen IVIS Imaging System. This nondestructive imaging system was sufficiently sensitive to detect photon signal emitted by a single capsule containing around 800 luciferase-transduced CHO (CHO(LUC)) cells in vitro, and to track changes in luciferase expression in vivo over time. Effective modulation of the transplantation-site environment with mIL-10 secreted from capsules was evident by greater luciferase expression at 10 and 21 days after transplantation relative to encapsulated luciferase-transfected cells that did not produce mIL-10. Longer duration effects require further investigation to extend this proof-of-concept study.     

Expression of matrix metalloproteinases and tissue inhibitors of metalloproteinases in human chondrosarcomas

The aim of the present study was to characterise the ability of malignant chondrosarcomas to invade normal bone by analysing their production of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). For this purpose 12 chondrosarcomas were investigated for the expression of mRNAs for several MMPs and all 4 TIMPs by Northern hybridisation, and for immunohistochemical localisation of the proteins. A characteristic finding of these analyses was increased expression of MMP-13, MMP-14 and TIMP-2 mRNAs in chondrosarcomas when compared with nonmalignant control samples. Individual chondrosarcomas also exhibited elevated levels of MMP-1, MMP-7 and MMP-9 mRNAs. The results of Northern hybridisations were supported by immunohistochemical stainings of the corresponding tumour areas for MMP-2, MMP-14 and TIMP-2, further suggesting that these may have prognostic value for determining whether individual chondrosarcomas are locally aggressive or have a probability of recurrence. Another finding of the present study was a marked heterogeneity in histologic appearance and gene expression of the chondrosarcomas, emphasising the importance of analysing several areas of these tumours to get representative results. These findings suggest that analysis of MMPs could be a useful diagnostic indicator in patients with cartilaginous tumours and could help in differentiating between a low-grade malignant chondrosarcoma and a benign growing enchondroma.