Thoracic: Lung Transplant: Basic Science
Protective effect of necrosulfonamide on rat pulmonary ischemia-reperfusion injury via inhibition of necroptosis

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Abstract

Background

Necroptosis plays an important role in cell death during pulmonary ischemia-reperfusion injury (IRI). We hypothesized that therapy with necrosulfonamide (NSA), a mixed-lineage kinase domain-like protein inhibitor, would attenuate lung IRI.

Methods

Rats were assigned at random into the sham operation group (n = 6), vehicle group (n = 8), or NSA group (n = 8). In the NSA and vehicle groups, the animals were heparinized and underwent left thoracotomy, and the left hilum was clamped for 90 minutes, followed by reperfusion for 120 minutes. NSA (0.5 mg/body) and a solvent were administered i.p. in the NSA group and the vehicle group, respectively. The sham group underwent 210 minutes of perfusion without ischemia. After reperfusion, arterial blood gas analysis, physiologic data, lung wet-to-dry weight ratio, histologic changes, and cytokine levels were assessed. Fluorescence double immunostaining was performed to evaluate necroptosis and apoptosis.

Results

Arterial partial pressure of oxygen/fraction of inspired oxygen (PaO2/FiO2) was better, dynamic compliance was higher, and mean airway pressure and lung edema were lower in the NSA group compared with the vehicle group. Moreover, in the NSA group, lung injury was significantly alleviated, and the mean number of necroptotic cells (55.3 ± 4.06 vs 78.2 ± 6.87; P = .024), but not of apoptotic cells (P = .084), was significantly reduced compared with the vehicle group. Interleukin (IL)-1β and IL-6 levels were significantly lower with NSA administration.

Conclusions

In a rat model, our results suggest that NSA may have a potential protective role in lung IRI through the inhibition of necroptosis.

Key Words

ischemia-reperfusion injury
cell death
apoptosis
necrosis
necroptosis
necrosulfonamide
receptor-interacting protein kinase-1
receptor-interacting protein kinase-3
mixed lineage kinase domain-like protein

Abbreviations and Acronyms

DAMP
damage-associated molecular pattern
DAPI
4′,6-diamidino-2-phenylindole
DMSO
dimethyl sulfoxide
ELISA
enzyme-linked immunoassay
HMGB1
high-mobility group box protein 1
HPF
high-power field
IL
interleukin
IRI
ischemia-reperfusion injury
MLKL
mixed-lineage kinase domain-like protein
Nec-1
necrostatin-1
NSA
necrosulfonamide
RIPK1
receptor-interacting protein kinase-1
RIPK3
receptor-interacting protein kinase-3
TNF-α
tumor necrosis factor alpha
TUNEL
terminal deoxynucleotidyl transferase-dUTP nick-end labeling
TV
tidal volume

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