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Abstract Background Inflammatory cells are believed to play a prominent role during tissue repair and remodeling. Since repair processes develop and mature over extended time frames, the present study was designed to evaluate the effect of monocytes and fibroblasts in prolonged culture in three-dimensional collagen gels. Methods Blood monocytes from healthy donors and human fetal lung fibroblasts were cast into type I collagen gels and maintained in floating cultures for three weeks. Results Fibroblast-mediated gel contraction was initially inhibited by the presence of monocytes (P < 0.01). However, with extended co-culture, contraction of the collagen gels was greatly augmented (P < 0.01). In addition, with extended co-culture, degradation of collagen in the gels occurred. The addition of neutrophil elastase to the medium augmented both contraction and degradation (P < 0.01). Prostaglandin E2 production was significantly increased by co-culture and its presence attenuated collagen degradation. Conclusion The current study, therefore, demonstrates that interaction between monocytes and fibroblasts can contract and degrade extracellular matrix in extended culture.

Abstract Background Extended culture of monocytes and fibroblasts in three-dimensional collagen gels leads to degradation of the gels (see linked study in this issue, "Fibroblasts and monocytes contract and degrade three-dimensional collagen gels in extended co-culture"). The current study, therefore, was designed to evaluate production of matrix-degrading metalloproteinases by these cells in co-culture and to determine if neutrophil elastase could collaborate in the activation of these enzymes. Since co-cultures produce prostaglandin E2 (PGE2), the role of PGE2 in this process was also evaluated. Methods Blood monocytes from healthy donors and human fetal lung fibroblasts were cast into type I collagen gels and maintained in floating cultures for three weeks. Matrix metalloproteinases (MMPs) were assessed by gelatin zymography (MMPs 2 and 9) and immunoblotting (MMPs 1 and 3). The role of PGE2 was explored by direct quantification, and by the addition of exogenous indomethacin and/or PGE2. Results Gelatin zymography and immunoblots revealed that MMPs 1, 2, 3 and 9 were induced by co-cultures of fibroblasts and monocytes. Neutrophil elastase added to the medium resulted in marked conversion of latent MMPs to lower molecular weight forms consistent with active MMPs, and was associated with augmentation of both contraction and degradation (P < 0.01). PGE2 appeared to decrease both MMP production and activation. Conclusion The current study demonstrates that interactions between monocytes and fibroblasts can mediate tissue remodeling.

Fibrotic diseases are characterized by the accumulation of extracellular matrix together with distortion and disruption of tissue architecture. Phosphodiesterase (PDE)4 inhibitors, by preventing the breakdown of cAMP, can inhibit fibroblast functions and may be able to mitigate tissue remodeling. Transforming growth factor (TGF)-[beta]1, a mediator of fibrosis, can potentially modulate cAMP by altering [PGE.sub.2] metabolism. The present study assessed whether PDE4 inhibitors functionally antagonize the profibrotic activity of fibroblasts stimulated by TGF-[beta]1. The PDE4 inhibitors roflumilast and rolipram both inhibited fibroblast-mediated contraction of three-dimensional collagen gels and fibroblast chemotaxis toward fibronectin in the widely studied human fetal lung fibroblast strain HFL-1 and several strains of fibroblasts from adult human lung. Roflumilast was ~10-fold more potent than rolipram. There was a trend for PDE4 inhibitors to inhibit more in the presence of TGF-[beta]1 (0.05 < P < 0.08). The effect of the PDE4 inhibitors was mediated through camp-stimulated protein kinase A (PKA), although a PKA-independent effect on gel contraction was also observed. The effect of PDE4 inhibitors depended on fibroblast production of [PGE.sub.2] and TGF[beta]-induced [PGE.sub.2] production. PDE4 inhibitors together with TGF-[beta]1 resulted in augmented [PGE.sub.2] production together with increased expression of COX mRNA and protein. The present study supports the concept that PDE4 inhibitors may attenuate fibroblast activities that can lead to fibrosis and that PDE4 inhibitors may be particularly effective in the presence of TGF-[beta]1-induced fibroblast stimulation. phosphodiesterase 4; chemotaxis; collagen gel contraction; transforming growth factor-[beta]1; prostaglandin [E.sub.2]

The controlled accumulation of fibroblasts to sites of inflammation is crucial to effective tissue repair after injury. Either inadequate or excessive accumulation of fibroblasts could result in abnormal tissue function. Prostacyclin (PG[I.sub.2]) is a potent mediator in the coagulation and inflammatory processes. The aim of this study was to investigate the effect of PG[I.sup.2] on chemotaxis of human fetal lung fibroblasts (HFL-1). Using the blind well chamber technique, we found that the PG[I.sub.2] analog carbaprostacyclin ([10.sup.-6] M) inhibited HFL-1 chemotaxis to human plasma fibronectin (20 [micro]g/ml) 58.0 [+ or -] 13.2% (P < 0.05) and to platelet-derived growth factor (PDGF)-BB (10 ng/ml) 48.7 [+ or -] 4.6% (P < 0.05). Checkerboard analysis demonstrated that carbaprostacyclin inhibits both directed and undirected migration. The inhibitory effect of the carbaprostacyclin was concentration dependent and blocked by the cAMP-dependent protein kinase (PKA) inhibitor KT-5720, suggesting that a cAMP-PKA pathway may be involved in the process. Two other PG[I.sub.2] analogs, ciprostene and dehydro-15-cyclohexyl carbaprostacyclin (both [10.sup.-6] M), significantly inhibited fibroblast migration to fibronectin. In summary, PG[I.sub.2] appears to inhibit fibroblast chemotaxis to fibronectin and PDGF-BB. Such an effect may contribute to the regulation of fibroblasts in wound healing and could contribute to the pathogenesis of diseases characterized by abnormal tissue repair remodeling. prostaglandin [I.sub.2]; protein kinase; tissue repair; inflammation

Byline: Fu-Qiang Wen (1,2), C. Magnus Skold (3), Xiang-Der Liu (1,2), Ronald F. Ertl (1,2), Yun Kui Zhu (1,2), Tadashi Kohyama (1,2), Hangjun Wang (1,2), Stephen I. Rennard (1,2) Keywords: glucocorticoids; transforming growth factor-[beta] (TGF-[beta]); prostaglandin; collagen gel contraction; fibroblast Abstract: TGF-[beta] plays a central role in the initiation and progression of pulmonary fibrosis. Glucocorticoids are frequently used to treat fibrotic diseases, but beneficial effects are often modest. Both TGF-[beta] and glucocorticoids have been reported to increase fibroblast contraction of native collagen gels, a model of fibrotic tissue remodeling. Therefore, we sought to determine how glucocorticoids interact with TGF-[beta] in this system. In this study, human fetal lung fibroblasts (HFL-1) were pretreated with or without TGF-[beta] for 72 h before they were cast into type I collagen gels. Various concentrations of glucocorticoids (budesonide or hydrocortisone) were added at the time of casting. Gel size was then monitored at different times after gel release. The surrounding media were collected for the assay of prostaglandin E.sub.2 (PGE.sub.2) and the cell lysates were analyzed for cyclooxygenase (COX) expression by immunoblot. Glucocorticoids alone significantly enhanced fibroblast-mediated contraction of collagen gels (P &lt 0.01) and dose-dependently inhibited PGE.sub.2 release by HFL-1 fibroblasts. TGF-[beta] significantly augmented gel contraction but also induced a 30% increase in PGE.sub.2 release and increased the expression of COX-1. Glucocorticoids inhibited TGF-[beta]1 induced-PGE.sub.2 release, and enhanced TGF-[beta] augmented gel contraction without significantly affecting TGF-[beta] augmented COX-1 expression. Indomethacin, a COX inhibitor, increased TGF-[beta] augmented gel contraction but had no further effect when added together with glucocorticoids. Thus, glucocorticoids can synergize with TGF-[beta] in augmenting fibroblast mediated collagen gel contraction through the inhibition of PGE.sub.2 production. Such interactions between glucocorticoids and TGF-[beta] may account, in part, for the lack of response of fibrotic diseases to glucocorticoids. Author Affiliation: (1) Pulmonary and Critical Care Medicine Section, Department of Internal Medicine, University of Nebraska Medical Center, 985125, Nebraska (2) Nebraska Medical Center, Omaha, Nebraska, 68198-5125 (3) Department of Medicine, Karolinska Hospital, Stockholm, Sweden Article History: Registration Date: 17/10/2004

An analysis of the theory that epithelial cells lining the airway regulate the development of peribronchial fibrosis and fixed airway obstruction by directing fibroblast proliferation reveals that airway epithelial cells modify fibroblast proliferation. They also play a role in peribronchial fibrosis in chronic bronchitis, which causes irreversible airway blockade. The modifications of airway architecture with abnormal airway connective tissue is believed to play a crucial role in the process.

Prostaglandin [E.sub.2] inhibits fibroblast chemotaxis. Am J Physiol Lung Cell Mol Physiol 281: L1257-L1263, 2001.--Fibroblasts are the major source of extracellular connective tissue matrix, and the recruitment, accumulation, and stimulation of these cells are thought to play important roles in both normal healing and the development of fibrosis. Prostaglandin [E.sub.2] (PG[E.sub.2]) can inhibit this process by blocking fibroblast proliferation and collagen production. The aim of this study was to investigate the inhibitory effect of PG[E.sub.2] on human plasma fibronectin (hFN)- and bovine bronchial epithelial cell-conditioned medium (BBECCM)-induced chemotaxis of human fetal lung fibroblasts (HFL1). Using the Boyden blind well chamber technique, PG[E.sub.2] ([10.sup.-7] M) inhibited chemotaxis to hFN 40.8 [+ or -] 5.3% (P < 0.05) and to BBEC-CM 49.7 [+ or -] 11.7% (P < 0.05). Checkerboard analysis demonstrated inhibition of both chemotaxis and chemokinesis. The effect of PG[E.sub.2] was concentration dependent, and the inhibitory effect diminished with time. Other agents that increased fibroblast cAMP levels, including isoproterenol ([10.sub.-5] M), dibutyryl cAMP ([10.sub.-5] M), and forskolin (3 x [10.sub.-5] M) had similar effects and inhibited chemotaxis 54.1, 95.3, and 87.0%, respectively. The inhibitory effect of PG[E.sub.2] on HFL1 cell chemotaxis was inhibited by the cAMP-dependent protein kinase (PKA) inhibitor KT-5720, which suggests a cAMP-dependent effect mediated by PKA. In summary, PG[E.sub.2] appears to inhibit fibroblast chemotaxis, perhaps by modulating the rate of fibroblast migration. Such an effect may contribute to regulation of the wound healing response after injury. eicosanoids; adenosine 3',5'-cyclic monophosphate; fibronectin; fibrosis; repair

Evaluates the effect of prostaglandin E[sub 2] on fibroblast chemotaxis. use of purified chemoattractant human plasma fibronectin for chemoattractant; Mechanism of PGE[sub 2]; Effect of PGE[sub 2] on human fetal lung fibroblasts cell chemotaxis.

Background: Extended culture of monocytes and fibroblasts in three-dimensional collagen gels leads to degradation of the gels (see linked study in this issue, "Fibroblasts and monocytes contract and degrade three-dimensional collagen gels in extended co-culture"). The current study, therefore, was designed to evaluate production of matrix-degrading metalloproteinases by these cells in co-culture and to determine if neutrophil elastase could collaborate in the activation of these enzymes. Since cocultures produce prostaglandin E2 (PGE2), the role of PGE2 in this process was also evaluated. Methods: Blood monocytes from healthy donors and human fetal lung fibroblasts were cast into type I collagen gels and maintained in floating cultures for three weeks. Matrix metalloproteinases (MMPs) were assessed by gelatin zymography (MMPs 2 and 9) and immunoblotting (MMPs 1 and 3). The role of PGE2 was explored by direct quantification, and by the addition of exogenous indomethacin and/or PGE2. Results: Gelatin zymography and immunoblots revealed that MMPs 1, 2, 3 and 9 were induced by cocultures of fibroblasts and monocytes. Neutrophil elastase added to the medium resulted in marked conversion of latent MMPs to lower molecular weight forms consistent with active MMPs, and was associated with augmentation of both contraction and degradation (P < 0.01). PGE2 appeared to decrease both MMP production and activation. Conclusion: The current study demonstrates that interactions between monocytes and fibroblasts can mediate tissue remodeling. [ABSTRACT FROM AUTHOR]

Background: Inflammatory cells are believed to play a prominent role during tissue repair and remodeling. Since repair processes develop and mature over extended time frames, the present study was designed to evaluate the effect of monocytes and fibroblasts in prolonged culture in three-dimensional collagen gels. Methods: Blood monocytes from healthy donors and human fetal lung fibroblasts were cast into type I collagen gels and maintained in floating cultures for three weeks. Results: Fibroblast-mediated gel contraction was initially inhibited by the presence of monocytes (P < 0.01). However, with extended co-culture, contraction of the collagen gels was greatly augmented (P < 0.01). In addition, with extended co-culture, degradation of collagen in the gels occurred. The addition of neutrophil elastase to the medium augmented both contraction and degradation (P < 0.01). Prostaglandin E2 production was significantly increased by co-culture and its presence attenuated collagen degradation. Conclusion: The current study, therefore, demonstrates that interaction between monocytes and fibroblasts can contract and degrade extracellular matrix in extended culture. [ABSTRACT FROM AUTHOR]