Background/Purpose:
MicroRNAs (miRNAs)
have been shown to serve as important regulators for inflammatory and immune
responses and are implicated in several immune disorders including gouty
arthritis. The expression of miR-146a is upregulated in peripheral blood
mononuclear cells in patients with intercritical gout
compared to normouricaemic and hyperuricaemic
controls and those with acute gout flares. However, the role of miR-146a in
gout development remains unknown. Here, we use miR-146a knockout (KO) mice to
test miR-146a function in monosodium urate (MSU) -induced
gouty arthritis.  

Methods: miR-146a KO  and WT control 
mice were injected with MSU suspension into the foot pad (1mg/40ul) or
ankle (0.5mg/20ul), respectively to induce 
the acute gouty arthritis. The bone marrow-derived macrophages (BMDM) were
stimulated with 250ug/ml MSU and miR-146a, interleukin 1 beta (IL-1β),
tumor necrosis factor-α (TNF-α) and NACHT, LRR and PYD
domains-containing protein 3 (NALP3) inflammasome gene
expression were evaluated by qRT-PCR.
The TNF-α and IL-1β protein level in BMDM
were detected by FACS staining and western blot. Gene and protein levels of TRAF-6
and IRAK-1, the targets of miR-146a, were measured with qRT-PCR
and western blot.

Results:  MiR-146a
expression in BMDM from C57/B6 mice was dramatically upregulated (160-folds
compared to unstimulated cells) at 4 hours post MSU crystal stimulation.
Significantly increased  paw
swelling index was observed in miR-146aKO mice compared to WT controls at 6h
and 24h after MSU treatment (37.18±6.47% vs 27.03±8.03 at the 6h, p<0.05;
61.87±6.50% vs 37.78±3.38% at 24h, p<0.05, N=10 for each group). Consistent
with increased paw swelling, miR-146aKO mice showed more severe ankle joint
swelling compared to WT mice (40.38±2.19% vs 15.14±2.54% at 6h, P<0.01;
32.69 ±2.85% vs 24.75±3.96% at 24h and 8.74±3.27% vs 3.36±2.98% at 48h,
p<0.05, N=10 for each group).  The expression
of IL-1β, TNF-α and NALP3 inflammatory, including of NALP3, ASC and
caspase-1 were upregulated in BMDM after exposed to MSU crystals for 4 hours,
compared to wild type mice. Consistent with the gene expression, the IL-1β
and TNF-α protein were up-regulated in miR-146aKO mice. Finally, the qRT-PCR and western blot results showed that miR-146a
targets, TRAF-6 and IRAK-1, were dramatically upregulated in BMDMs from miR-146
KO mice compared to that from WT.

Conclusion: Collectively, our data suggests that
miR-146a provides
feedback regulation of gout arthritis development and lack of miR-146a enhances
gouty arthritis through upregulating TRAK-6/IRAK-1 and NALP3 inflammsome function (figure 1).