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Abstract
Plasma membrane disruptions occur in mechanically active tissues such as the epidermis and can lead to cell death if the damage remains unrepaired. Repair occurs through fusion of vesicle patches to the damaged membrane region. The enzyme phospholipase D (PLD) is involved in membrane traffickiing; therefore, the role of PLD in membrane repair was investigated. Generation of membrane disruptions by lifting epidermal keratinocytes from the substratum induced PLD activation, whereas removal of cells from the substratum via trypsinization had no effect. Pretreatment with 1,25-dihydroxyvitamin D₃, previously shown to increase PLD1 expression and activity, had no effect on, and a PLD2-selective (but not a PLD1-selective) inhibitor decreased, cell lifting-induced PLD activation, suggesting PLD2 as the isoform activated. PLD2 interacts functionally with the glycerol channel aquaporin-3 (AQP3) to produce phosphatidylglycerol (PG); however, wounding resulted in decreased PG production, suggesting a potential PG deficiency in wounded cells. Cell lifting-induced PLD activation was transient, consistent with a possible role in membrane repair, and PLD inhibitors inhibited membrane resealing upon laser injury. In an in vivo full-thickness mouse skin wound model, PG accelerated wound healing. These results suggest that PLD and the PLD2/AQP3 signaling module may be involved in membrane repair and wound healing.
Highlights
Plasma membrane disruptions occur in mechanically active tissues such as the epidermis and can lead to cell death if the damage remains unrepaired
We demonstrate that plasma membrane disruptions, as induced by cell lifting from the substratum, resulted in the activation of phospholipase D (PLD) in a time-dependent manner (Figs. 1, 2)
This activation did not appear to be the result of specific factors in the medium, inasmuch as similar activation was observed in Hank’s buffered salt solution (HBSS) (Fig. 1B) or serum-free keratinocyte medium (SFKM) (Fig. 1A)
Summary
Plasma membrane disruptions occur in mechanically active tissues such as the epidermis and can lead to cell death if the damage remains unrepaired. As keratinocytes migrate up from the stratum basale, they undergo a distinct pattern of differentiation that is essential for the function of the skin as a protective barrier This pattern is characterized by growth arrest and expression of the mature keratins 1 and 10 in the first differentiated layer of the epidermis, the spinous layer. Many tissues of the body in addition to the skin are exposed to mechanical stresses that result in tearing, or disrupting, the plasma membrane of the constituent cells. It is critical that cells in these mechanically active tissues be able to repair membrane
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