# Heme toxicity and vaso-occlusion in sickle cell disease

> **NIH NIH R01** · UNIVERSITY OF MINNESOTA · 2020 · $494,306

## Abstract

Hemolysis and the intravascular release of hemoglobin S are central to the pathophysiology of sickle
cell disease (SCD). During the current funding period, we showed that heme derived from sickle red
blood cells acts as a damage-associated molecular pattern that can activate toll-like receptor 4
(TLR4) of the innate immune system, independently of its cognate ligand lipopolysaccharide, leading
to oxidant production, rapid P-selectin and von Willebrand factor expression on endothelium, and
vaso-occlusion (VO) in SCD mice. We hypothesize that the innate immune system, including TLR4
and complement, is fundamental to understanding hemolysis-driven inflammation, coagulation, VO,
and the cumulative organ pathology in SCD. The first aim of our proposal is to identify the heme-
binding site on the myeloid differentiation factor 2 (MD-2)/TLR4 complex. To date, no studies have
examined the heme-binding site on the MD-2/TLR4 complex. Identifying the heme-binding site on
MD-2/TLR4 is vital for developing therapies to interrupt heme-driven inflammation and VO. The
second aim will examine the impact of global Tlr4 deficiency and the specific contribution of TLR4 in
leukocytes, platelets, and the vessel wall to SCD pathogenesis. Recent studies by our group and
others used Tlr4-/-, non-sickle cell mice transplanted with Tlr4+/+ sickle bone marrow to underscore the
importance of TLR4 in the vessel wall in promoting VO and acute chest syndrome in SCD. However,
the impact of Tlr4 deficiency on inflammation, coagulation, and cumulative organ pathology has not
been tested in a global Tlr4-/- SCD mouse model. We will test the hypothesis that TLR4 is a key
signaling pathway that translates hemolysis into inflammation, VO, and organ pathology in SCD. Our
third aim will examine the role of complement activation in SCD hemolysis, inflammation, and VO.
The alternative complement pathway is abnormally activated in SCD and is amplified by
phosphatidylserine on the outer leaflet of sickle red blood cells and microparticles. We will test the
hypothesis that complement activation on the surface of sickle red blood cells and microparticles
stimulates inflammation, coagulation, hemolysis, and VO in SCD. Using state-of-the-art molecular
techniques including site-directed mutagenesis, bone marrow transplants, cellular/biochemical
studies, breeding a unique sickle/Tlr4-deficient mouse model, and in vivo vascular imaging in murine
models of SCD, we will provide the foundation for the development of new therapies targeting
multiple pathways to interrupt SCD pathophysiology. Understanding TLR4 and complement activation
in SCD is expected to lead to therapies targeting the heme-binding site on the MD-2/TLR4 complex
and complement inhibitors that will improve the quality of life of SCD patients.

## Key facts

- **NIH application ID:** 9905547
- **Project number:** 5R01HL114567-08
- **Recipient organization:** UNIVERSITY OF MINNESOTA
- **Principal Investigator:** Gregory M Vercellotti
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $494,306
- **Award type:** 5
- **Project period:** 2017-02-01 → 2022-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9905547

## Citation

> US National Institutes of Health, RePORTER application 9905547, Heme toxicity and vaso-occlusion in sickle cell disease (5R01HL114567-08). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/9905547. Licensed CC0.

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