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Polymorphonuclear leukocyte migration through human amnion membrane
Abstract:
A new in vitro model has been developed for studying migration of human polymorphonuclear leukocytes (PMN) through living native cellular and matrix barriers. Human amnion membrane consists of a single layer of epithelium bound to a continuous basement membrane interfacing an avascular collagenous stroma. Living amnion was placed in plastic chambers with separate compartments on each side of the membrane. PMN were introduced on the epithelial side of the amnion, and a Millipore filter (Millipore Corp., Bedford, Mass.) was placed against the stromal side. In response to N-formylmethionyl-leucyl- phenylanlanine (FMLP) chemoattractant, PMN penetrated the full thickness of the amnion and were collected and counted on the filter. The rate of PMN traversal of the amnion was dependent on the concentration of FMLP (optimal at 10(-8)M) as well as the slope of the FMLP gradient across the amnion. The route of PMN migration was studied by transmission electron microscopy. PMN first attached to the epithelial surface, then infiltrated between intercellular junctions. PMN migrated around or through tight junction and hemidesmosome attachments. The PMN then penetrated the basement membrane and migrated through the dense collagenous stroma. The present amnion migration system has characteristics of the in vivo inflammatory state not described in any previous method for monitoring PMN migration in vitro. Prior methods have not used native epithelium, whole basement membrane, or collagenous stroma. PMN penetration of these barriers occurs in the normal inflammatory response and probably involves biochemical mechanisms not required for simple migration through the pores of an artificial filter. The amnion system can be useful for future biochemical and morphological studies of PMN penetration of these barriers and possible repair processes that may follow.
Insights
A new in vitro model using human amnion membrane allows scientists to study polymorphonuclear leukocyte (PMN) migration through native barriers. This advanced system mimics in vivo inflammation, offering insights into cell movement and potential repair mechanisms.
Area of Science:
- Cell Biology
- Immunology
- Biomedical Engineering
Background:
- Studying polymorphonuclear leukocyte (PMN) migration is crucial for understanding inflammatory responses.
- Previous in vitro models lack the complexity of native cellular and matrix barriers found in vivo.
- A need exists for a more physiologically relevant model to investigate PMN transmigration.
Purpose of the Study:
- To develop and validate a novel in vitro model for studying human PMN migration through living native barriers.
- To analyze the factors influencing PMN migration rate and route using this new model.
- To provide a platform for future studies on PMN-matrix interactions and inflammatory processes.
Main Methods:
- Utilized human amnion membrane, comprising native epithelium, basement membrane, and collagenous stroma, as the barrier.
- Developed a two-compartment chamber system to simulate physiological conditions for PMN migration.
- Employed N-formylmethionyl-leucyl-phenylanlanine (FMLP) as a chemoattractant and transmission electron microscopy for route analysis.
Main Results:
- PMN successfully migrated through the full thickness of the native amnion barrier in response to FMLP.
- Migration rate was dependent on FMLP concentration and gradient slope, with optimal chemoattraction at 10(-8)M.
- Transmission electron microscopy revealed PMN infiltration between epithelial cells, through junctions, basement membrane, and collagenous stroma.
Conclusions:
- The developed amnion migration system accurately models in vivo inflammatory cell migration through complex native barriers.
- This model recapitulates key aspects of PMN transmigration, including interaction with epithelium, basement membrane, and stroma.
- The system offers a valuable tool for investigating the biochemical and morphological mechanisms underlying PMN penetration and potential tissue repair.