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Keratinocyte stimulation of matrix metalloproteinase-1 production and proliferation in fibroblasts: regulation through mitogen-activated protein kinase signalling events.

Moon SE, Bhagavathula N, Varani J - Br. J. Cancer (2002)

Bottom Line: In fibroblasts exposed to keratinocyte-conditioned medium, mitogen-activated protein kinase signalling through both the extracellular signal-related kinase pathway and p38 pathway occurred.A p38 signalling inhibitor (SB203580) blocked matrix metalloproteinase-1 elaboration induced by keratinocyte-conditioned medium or interleukin-1beta, but did not inhibit matrix metalloproteinase-1 elaboration or cell growth induced by epidermal growth factor.These data indicate that keratinocyte-fibroblast interactions are mediated by multiple stimulating agents acting on specific receptors to induce signalling through different mitogen-activated protein kinase pathways leading to altered expression of key biological functions.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, The University of Michigan Medical School, 1301 Catherine Road, Box 0602, Ann Arbor, Michigan, MI 48109, USA.

ABSTRACT
Incubation of human dermal fibroblasts in keratinocyte-conditioned culture medium led to a 5.7-fold increase in the level of matrix metalloproteinase-1. Virtually all of the matrix metalloproteinase-1 - inducing activity could be related to agonists acting through members of the epidermal growth factor receptor family or to agonists acting through the interleukin-1 receptor. The same keratinocyte-conditioned medium also induced a modest increase in fibroblast proliferation (approximately 1.8-fold). Growth-stimulating activity could be attributed to epidermal growth factor receptor (but not interleukin-1 receptor) function. In fibroblasts exposed to keratinocyte-conditioned medium, mitogen-activated protein kinase signalling through both the extracellular signal-related kinase pathway and p38 pathway occurred. When recombinant epidermal growth factor or recombinant interleukin-1beta were used as a control, they induced mitogen-activated protein kinase signalling consistent with the combined effects of epidermal growth factor receptor - specific and interleukin-1 receptor - specific agonists in keratinocyte-conditioned medium. Recombinant epidermal growth factor stimulated both matrix metalloproteinase-1 induction and proliferation while recombinant interleukin-1beta stimulated matrix metalloproteinase-1 elaboration but not fibroblast growth. An inhibitor of extracellular signal-related kinase pathway signalling (U0126) blocked induction of matrix metalloproteinase-1 production induced by keratinocyte-conditioned medium (as well as by epidermal growth factor or interleukin-1beta), and also inhibited proliferation. A p38 signalling inhibitor (SB203580) blocked matrix metalloproteinase-1 elaboration induced by keratinocyte-conditioned medium or interleukin-1beta, but did not inhibit matrix metalloproteinase-1 elaboration or cell growth induced by epidermal growth factor. These data indicate that keratinocyte-fibroblast interactions are mediated by multiple stimulating agents acting on specific receptors to induce signalling through different mitogen-activated protein kinase pathways leading to altered expression of key biological functions.

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Fibroblast production of MMP-1 in response to stimulation by (A) keratinocyte-conditioned medium, (B) EGF and (C) IL-1β. Fibroblasts were exposed to culture medium alone or to a 50 : 50 mixture of culture medium and keratinocyte-conditioned medium. U0126 (10 μM) or SB203580 (15 μM) was included as indicated. At the end of the 2-day incubation period, MMP-1 was assessed by β-casein zymography. Quantitation was accomplished by densitometry scanning as described in the Materials and Methods section. Values shown are means and standard deviations based on four separate experiments. The insert shows a β-casein zymogram from one experiment. Lane 1 demonstrates MMP-1 levels (54 kD) in control fibroblast culture fluid; Lane 2 shows increased enzyme produced by fibroblasts exposed to keratinocyte-conditioned medium. Lanes 3 and 4 show the effects of U0126 and SB203580, respectively, on up-regulated enzyme production. A gelatin zymogram is shown for control. Neither inhibitor influenced expression of MMP-2 (72 kD).
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fig1: Fibroblast production of MMP-1 in response to stimulation by (A) keratinocyte-conditioned medium, (B) EGF and (C) IL-1β. Fibroblasts were exposed to culture medium alone or to a 50 : 50 mixture of culture medium and keratinocyte-conditioned medium. U0126 (10 μM) or SB203580 (15 μM) was included as indicated. At the end of the 2-day incubation period, MMP-1 was assessed by β-casein zymography. Quantitation was accomplished by densitometry scanning as described in the Materials and Methods section. Values shown are means and standard deviations based on four separate experiments. The insert shows a β-casein zymogram from one experiment. Lane 1 demonstrates MMP-1 levels (54 kD) in control fibroblast culture fluid; Lane 2 shows increased enzyme produced by fibroblasts exposed to keratinocyte-conditioned medium. Lanes 3 and 4 show the effects of U0126 and SB203580, respectively, on up-regulated enzyme production. A gelatin zymogram is shown for control. Neither inhibitor influenced expression of MMP-2 (72 kD).

Mentions: Human dermal fibroblasts were exposed to 72-h keratinocyte-conditioned medium and incubated for 2 days. At the end of the incubation period, MMP-1 production and proliferation were assessed. Consistent with our recent findings (Moon et al, 2001), there was a greater than five-fold stimulation of MMP-1 production and an approximately two-fold increase in growth. Effects of two MAPK inhibitors – i.e., U0126 and SB203580 – on MMP-1 elaboration and growth were assessed. In the presence of 10 μM U0126, MMP-1 production was reduced by 88% and fibroblast growth was reduced by 95% (Figures 1AFigure 1


Keratinocyte stimulation of matrix metalloproteinase-1 production and proliferation in fibroblasts: regulation through mitogen-activated protein kinase signalling events.

Moon SE, Bhagavathula N, Varani J - Br. J. Cancer (2002)

Fibroblast production of MMP-1 in response to stimulation by (A) keratinocyte-conditioned medium, (B) EGF and (C) IL-1β. Fibroblasts were exposed to culture medium alone or to a 50 : 50 mixture of culture medium and keratinocyte-conditioned medium. U0126 (10 μM) or SB203580 (15 μM) was included as indicated. At the end of the 2-day incubation period, MMP-1 was assessed by β-casein zymography. Quantitation was accomplished by densitometry scanning as described in the Materials and Methods section. Values shown are means and standard deviations based on four separate experiments. The insert shows a β-casein zymogram from one experiment. Lane 1 demonstrates MMP-1 levels (54 kD) in control fibroblast culture fluid; Lane 2 shows increased enzyme produced by fibroblasts exposed to keratinocyte-conditioned medium. Lanes 3 and 4 show the effects of U0126 and SB203580, respectively, on up-regulated enzyme production. A gelatin zymogram is shown for control. Neither inhibitor influenced expression of MMP-2 (72 kD).
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2376127&req=5

fig1: Fibroblast production of MMP-1 in response to stimulation by (A) keratinocyte-conditioned medium, (B) EGF and (C) IL-1β. Fibroblasts were exposed to culture medium alone or to a 50 : 50 mixture of culture medium and keratinocyte-conditioned medium. U0126 (10 μM) or SB203580 (15 μM) was included as indicated. At the end of the 2-day incubation period, MMP-1 was assessed by β-casein zymography. Quantitation was accomplished by densitometry scanning as described in the Materials and Methods section. Values shown are means and standard deviations based on four separate experiments. The insert shows a β-casein zymogram from one experiment. Lane 1 demonstrates MMP-1 levels (54 kD) in control fibroblast culture fluid; Lane 2 shows increased enzyme produced by fibroblasts exposed to keratinocyte-conditioned medium. Lanes 3 and 4 show the effects of U0126 and SB203580, respectively, on up-regulated enzyme production. A gelatin zymogram is shown for control. Neither inhibitor influenced expression of MMP-2 (72 kD).
Mentions: Human dermal fibroblasts were exposed to 72-h keratinocyte-conditioned medium and incubated for 2 days. At the end of the incubation period, MMP-1 production and proliferation were assessed. Consistent with our recent findings (Moon et al, 2001), there was a greater than five-fold stimulation of MMP-1 production and an approximately two-fold increase in growth. Effects of two MAPK inhibitors – i.e., U0126 and SB203580 – on MMP-1 elaboration and growth were assessed. In the presence of 10 μM U0126, MMP-1 production was reduced by 88% and fibroblast growth was reduced by 95% (Figures 1AFigure 1

Bottom Line: In fibroblasts exposed to keratinocyte-conditioned medium, mitogen-activated protein kinase signalling through both the extracellular signal-related kinase pathway and p38 pathway occurred.A p38 signalling inhibitor (SB203580) blocked matrix metalloproteinase-1 elaboration induced by keratinocyte-conditioned medium or interleukin-1beta, but did not inhibit matrix metalloproteinase-1 elaboration or cell growth induced by epidermal growth factor.These data indicate that keratinocyte-fibroblast interactions are mediated by multiple stimulating agents acting on specific receptors to induce signalling through different mitogen-activated protein kinase pathways leading to altered expression of key biological functions.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, The University of Michigan Medical School, 1301 Catherine Road, Box 0602, Ann Arbor, Michigan, MI 48109, USA.

ABSTRACT
Incubation of human dermal fibroblasts in keratinocyte-conditioned culture medium led to a 5.7-fold increase in the level of matrix metalloproteinase-1. Virtually all of the matrix metalloproteinase-1 - inducing activity could be related to agonists acting through members of the epidermal growth factor receptor family or to agonists acting through the interleukin-1 receptor. The same keratinocyte-conditioned medium also induced a modest increase in fibroblast proliferation (approximately 1.8-fold). Growth-stimulating activity could be attributed to epidermal growth factor receptor (but not interleukin-1 receptor) function. In fibroblasts exposed to keratinocyte-conditioned medium, mitogen-activated protein kinase signalling through both the extracellular signal-related kinase pathway and p38 pathway occurred. When recombinant epidermal growth factor or recombinant interleukin-1beta were used as a control, they induced mitogen-activated protein kinase signalling consistent with the combined effects of epidermal growth factor receptor - specific and interleukin-1 receptor - specific agonists in keratinocyte-conditioned medium. Recombinant epidermal growth factor stimulated both matrix metalloproteinase-1 induction and proliferation while recombinant interleukin-1beta stimulated matrix metalloproteinase-1 elaboration but not fibroblast growth. An inhibitor of extracellular signal-related kinase pathway signalling (U0126) blocked induction of matrix metalloproteinase-1 production induced by keratinocyte-conditioned medium (as well as by epidermal growth factor or interleukin-1beta), and also inhibited proliferation. A p38 signalling inhibitor (SB203580) blocked matrix metalloproteinase-1 elaboration induced by keratinocyte-conditioned medium or interleukin-1beta, but did not inhibit matrix metalloproteinase-1 elaboration or cell growth induced by epidermal growth factor. These data indicate that keratinocyte-fibroblast interactions are mediated by multiple stimulating agents acting on specific receptors to induce signalling through different mitogen-activated protein kinase pathways leading to altered expression of key biological functions.

Show MeSH
Related in: MedlinePlus