Role of the leucine-rich repeat domain of cryopyrin/NALP3 in monosodium urate crystal-induced inflammation in mice.
OBJECTIVE
The mechanism by which monosodium urate monohydrate (MSU) crystals intracellularly activate the cryopyrin inflammasome is unknown. The aim of this study was to use a mouse molecular genetics-based approach to test whether the leucine-rich repeat (LRR) domain of cryopyrin is required for MSU crystal-induced inflammation.


METHODS
Cryopyrin-knockout lacZ (Cryo(-Z/-Z)) mice and mice with the cryopyrin LRR domain deleted and fused to the lacZ reporter (Cryo(DeltaLRR Z/DeltaLRR Z)) were generated using bacterial artificial chromosome-based targeting vectors, which allow for large genomic deletions. Bone marrow-derived macrophages from Cryo(DeltaLRR Z/DeltaLRR Z) mice, Cryo(-Z/-Z) mice, and congenic wild-type (WT) mice were challenged with endotoxin-free MSU crystals under serum-free conditions. Phagocytosis and cytokine expression were assessed by flow cytometry and enzyme-linked immunosorbent assay. MSU crystals also were injected into mouse synovial-like subcutaneous air pouches. The in vivo inflammatory responses were examined.


RESULTS
Release of interleukin-1beta (IL-1beta), but not CXCL1 and tumor necrosis factor alpha, was impaired in Cryo(DeltaLRR Z/DeltaLRR Z) and Cryo(-Z/-Z) mouse bone marrow-derived macrophages compared with WT mouse bone marrow-derived macrophages in response to not only MSU crystals but also other known stimuli that activate the cryopyrin inflammasome. In addition, a comparable percentage of MSU crystals taken up by each type of bone marrow-derived macrophage was observed. Moreover, total leukocyte infiltration in the air pouch and IL-1beta production were attenuated in Cryo(-Z/-Z) and Cryo(DeltaLRR Z/DeltaLRR Z) mice at 6 hours postinjection of MSU crystals compared with WT mice.


CONCLUSIONS
MSU crystal-induced inflammatory responses were comparably attenuated both in vitro and in vivo in Cryo(DeltaLRR Z/DeltaLRR Z) and Cryo(-Z/-Z) mice. Hence, the LRR domain of cryopyrin plays a role in mediating MSU crystal-induced inflammation in this model.
Hoffman
Hal M
HM
University of California, San Diego, USA.
Scott
Peter
P
Mueller
James L
JL
Misaghi
Amir
A
Stevens
Sean
S
Yancopoulos
George D
GD
Murphy
Andrew
A
Valenzuela
David M
DM
Liu-Bryan
Ru
R
D000818:Animals; D015210:Arthritis, Gouty / Q000276:immunology / Q000378:metabolism*; D001854:Bone Marrow Cells; D002352:Carrier Proteins / Q000276:immunology / Q000378:metabolism*; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism; D005260:Female; D000090263:Leucine-Rich Repeat Proteins; D008262:Macrophage Activation; D008264:Macrophages / Q000187:drug effects / Q000276:immunology; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; D010587:Phagocytosis / Q000187:drug effects; D011506:Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D020449:Repetitive Sequences, Amino Acid; D012867:Skin / Q000187:drug effects / Q000473:pathology; D014527:Uric Acid / Q000276:immunology / Q000633:toxicity
D002352:Carrier Proteins; D016207:Cytokines; D000090263:Leucine-Rich Repeat Proteins; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; C492716:Nlrp3 protein, mouse; D011506:Proteins; D014527:Uric Acid