# Molecular regulation of the capillary barrier in acute critical illness

> **NIH NIH R01** · YALE UNIVERSITY · 2024 · $573,389

## Abstract

Project Summary
Capillary leak is common in acutely critically ill children. Although no gold standard definition exists, it is clinically
recognized as new or worsening organ failure despite appropriate cardiovascular resuscitation. Unfortunately,
little has been learned of the pathophysiologic processes despite decades of struggling at the bedside of volume
overloaded children with multiple organ dysfunction syndromes. Treatment is limited to intensive supportive care
for failing organ systems. Less confusion exists in vitro, where leak around cultured human microvascular
endothelial cells (EC) is identified as disruption of intercellular tight junctions (TJs) with functional changes in
monolayer permselectivity. Such changes may be modeled in the EC response to cytokines, including those
known to be elevated in the plasma of critically ill children. However, targeting specific cytokines has repeatedly
failed to improve patient outcomes. Our overarching hypothesis is that while a great many leak-producing
cytokines may be elevated in acute critical illness, there are only limited EC responses and that final
common signaling pathways result in leak either between (para-) or through (trans-) ECs are therapeutic
targets. However, the relative contributions of trans- and paracellular leak to the clinical manifestations of leak
(i.e., organ dysfunction) and the pathways that cause them are incompletely understood. We will focus on targets
we identified upregulated in ECs isolated from critically ill children (collected from vascular access insertion
equipment and immediately analyzed by single-cell RNA-sequencing). This unique data set has identified
candidate regulatory molecules associated with paracellular leak and oncostatin M (OSM) as a novel mediator
of transcellular leak. We will test the contribution of these targets to leak in our culture models of TJ-forming
human microvascular ECs from a healthy donor (both male and female) skin and lung using trans-endothelial
electrical resistance, macromolecular flux assays, morphological analyses, molecular engineering, and immuno-
chemical tools. Aim 1 will utilize tumor necrosis factor (TNF) to model paracellular leak to test the hypotheses
that ArhGEF15, ArhGAP21, and -26 regulate RhoB activity, promoting junctional disassembly via downstream
kinases directly acting on TJs and amplified new gene transcription. We have also discovered that formoterol,
but not other β2-adrenergic agonists, inhibits TNF-induced leak and will investigate potential mechanisms. Aim
2 will test the hypothesis that OSM induces vesicle-associated JAK3/STAT3 signaling resulting in transcellular
leak, which also depends on new gene expression, and investigate how formoterol reduces OSM-induced leak.
Finally, in Aim 3, we will determine if the specific pathways identified in Aims 1 and 2 are recapitulated in intact
human capillaries in vivo using human skin xenografts in mice and ex vivo using machine-perfused human lungs.
The...

## Key facts

- **NIH application ID:** 10875434
- **Project number:** 5R01HL170164-02
- **Recipient organization:** YALE UNIVERSITY
- **Principal Investigator:** RICHARD W PIERCE
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $573,389
- **Award type:** 5
- **Project period:** 2023-09-01 → 2027-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10875434

## Citation

> US National Institutes of Health, RePORTER application 10875434, Molecular regulation of the capillary barrier in acute critical illness (5R01HL170164-02). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10875434. Licensed CC0.

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