Thrombin stimulates albumin transcytosis in lung microvascular endothelial cells via activation of acid sphingomyelinase

Abstract

Transcellular albumin transport occurs via caveolae that are abundant in lung microvascular endothelial cells. Stimulation of albumin transcytosis by proinflammatory mediators may contribute to alveolar protein leak in lung injury, yet the regulation of albumin transport and its underlying molecular mechanisms are so far incompletely understood. Here we tested the hypothesis that thrombin may stimulate transcellular albumin transport across lung microvascular endothelial cells in an acid-sphingomyelinase dependent manner. Thrombin increased the transport of fluorescently labeled albumin across confluent human lung microvascular endothelial cell (HMVEC-L) monolayers to an extent that markedly exceeds the rate of passive diffusion. Thrombin activated acid sphingomyelinase (ASM) and increased ceramide production in HMVEC-L, but not in bovine pulmonary artery cells, which showed little albumin transport in response to thrombin. Thrombin increased total caveolin-1 (cav-1) content in both whole cell lysates and lipid rafts from HMVEC-L, and this effect was blocked by inhibition of ASM or de novo protein biosynthesis. Thrombin-induced uptake of albumin into lung microvascular endothelial cells was confirmed in isolated-perfused lungs by real-time fluorescence imaging and electron microscopy of gold-labeled albumin. Inhibition of ASM attenuated thrombin-induced albumin transport both in confluent HMVEC-L and in intact lungs, whereas HMVEC-L treatment with exogenous ASM increased albumin transport and enriched lipid rafts in cav-1. Our findings indicate that thrombin stimulates transcellular albumin transport in an acid sphingomyelinase-dependent manner by inducing de novo synthesis of cav-1 and its recruitment to membrane lipid rafts.

Platelet depletion limits the severity but does not prevent the occurrence of experimental transfusion‐related acute lung injury

“…Interestingly, in our hand, the platelet level detected in the BALs was significantly higher in the [LPS + anti‐MHC I + anti‐GPIbα] group than in the [LPS + anti‐MHC I] group, which itself presented significantly increased platelet infiltration compared to the control group. We cannot rule out the contribution of transcytosis/paracytosis to the increased BAL protein levels . Surprisingly, following platelet depletion, we noted a significant increase in the number of platelets with a low mean volume in the BAL.…”

“…We cannot rule out the contribution of transcytosis/paracytosis to the increased BAL protein levels. 43 Surprisingly, following platelet depletion, we noted a significant increase in the number of platelets with a low mean volume in the BAL. The latter result may be supported by the recent description of pulmonary thrombopoiesis.…”

Transcellular vesicular transport in epithelial and endothelial cells: Challenges and opportunities

“…Along the same lines, the contribution of transcytosis to overall endothelial permeability during inflammation is relatively understudied. A number of reports suggest that inflammatory states can induce transcytosis even in the absence of concomitant paracellular leak . However, how this contributes to bona fide pathology remains uncertain.…”

Dynamic interactions ofPlasmodiumspp. with vascular endothelium

“…The process of albumin transcytosis in resting and thrombin-stimulated human lung endothelial cells is regulated by acid sphingomyelinase production of ceramide which results in increases in caveolin-1, the major structural protein of caveolae. 7 Besides SFK, pathologic increases in transcytosis of albumin (transcellular leak) and paracellular leak are co-regulated by dynamin and Rac1. 8 In addition to albumin, transcytosis transports immunoglobulins, transferrin, aminopeptidase P and numerous other molecules.…”