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Role of gelatinase B and elastase in human polymorphonuclear neutrophil migration across basement membrane
C Delclaux1, C Delacourt, M P D'Ortho
1Département de Physiologie, Service de Réanimation Médicale, Créteil, France.
Abstract:
Polymorphonuclear neutrophil (PMN) migration across basement membrane is thought to be dependent on the degradation of membrane constituents. PMN gelatinase B, a metalloproteinase able to degrade type IV collagen, may be involved in this phenomenon. PMN gelatinase B is released in the extracellular medium as a latent proform and then activated, mainly by PMN elastase. We investigated the role of gelatinase B in PMN migration across a Matrigel basement membrane matrix coated onto a filter, in a Boyden chamber. The effects of gelatinase and elastase inhibitors on PMN migration in this system were tested. Chemokinesis of PMN was tested in the same Boyden chamber across a filter free of basement membrane. The agarose method was used to test the same inhibitors for effects on PMN chemotaxis. In both systems, FMLP 10(-7)M was used as a chemoattractant. Addition of 10(-8)M TIMP-1 (the preferential gelatinase B inhibitor) inhibited trans-basement membrane PMN migration by 52 +/- 6% (P<0.05), without affecting PMN chemokinesis, chemotaxis, or degranulation. Also, (Ala)(2) Pro Val chloromethyl ketone (AAPVCK) 100 micron, a specific elastase inhibitor, inhibited trans-basement membrane PMN migration by 51 +/- 8% (P<0.05), without affecting PMN chemokinesis, chemotaxis, or degranulation. The AAPVCK-TIMP combination led to a decrease in migration across Matrigel basement membrane (46 +/- 2%, P,0.05)similar to that seen with TIMP alone. AAPVCK was responsible for inhibition of gelatinase B activation, leading to a decrease in activated gelatinase from 14% to 2% of total gelatinase release (P<0.05). All these results strongly suggest that gelatinase B is a major factor of PMN migration across basement membrane and that elastase may contribute to this process by activating pro-gelatinase B.
Insights
Polymorphonuclear neutrophil (PMN) migration across basement membranes relies on gelatinase B. Inhibiting gelatinase B or elastase significantly reduced PMN migration, highlighting their crucial roles.
Area of Science:
- Cell Biology
- Biochemistry
- Immunology
Background:
- Polymorphonuclear neutrophil (PMN) migration through basement membranes is essential for immune responses.
- Degradation of basement membrane components is believed to facilitate PMN transmigration.
- PMN gelatinase B, a matrix metalloproteinase, degrades type IV collagen, a key basement membrane constituent.
Purpose of the Study:
- To investigate the role of PMN gelatinase B in PMN migration across a basement membrane matrix.
- To determine the effects of gelatinase B and elastase inhibitors on PMN migration.
- To elucidate the mechanism by which elastase influences gelatinase B activity during PMN migration.
Main Methods:
- Utilized a Boyden chamber model with Matrigel-coated filters to assess trans-basement membrane PMN migration.
- Administered specific inhibitors for gelatinase B (TIMP-1) and elastase (AAPVCK) to evaluate their impact on migration.
- Assessed PMN chemokinesis, chemotaxis, and degranulation in parallel experiments to isolate the effect on transmigration.
Main Results:
- TIMP-1, a gelatinase B inhibitor, reduced PMN migration across Matrigel by 52% (P<0.05) without affecting other PMN functions.
- AAPVCK, an elastase inhibitor, similarly inhibited trans-basement membrane migration by 51% (P<0.05).
- AAPVCK inhibited pro-gelatinase B activation, decreasing active gelatinase levels from 14% to 2% (P<0.05).
Conclusions:
- Gelatinase B is a critical enzyme mediating PMN migration across basement membranes.
- Elastase plays a significant role, likely by activating pro-gelatinase B, thus facilitating basement membrane degradation.
- Targeting gelatinase B and elastase represents a potential strategy to modulate inflammatory cell infiltration.