Objective:

2

To evaluate the protective effects of gentiopicroside (GPS) against reactive oxygen species (ROS)-induced NOD-like receptor family

pyrin domain containing 3 (NLRP3) inflammasome activation in endothelial cells

aiming to reduce atherosclerosis.

Methods:

2

Eight-week-old male ApoE-deficient mice were randomly divided into 2 groups (

n

=10 per group): the vehicle group and the GPS treatment group. Both groups were fed a high-fat diet for 16 weeks. GPS (40 mg/kg per day) was administered by oral gavage to the GPS group

while the vehicle group received an equivalent volume of the vehicle solution. At the end of the treatment

blood and aortic tissues were collected for assessments of atherosclerosis

lipid profiles

oxidative stress

and molecular expressions related to NLRP3 inflammasome activation

ROS production

and apoptosis. Additionally

in vitro

experiments on human aortic endothelial cells treated with oxidized low-density lipoprotein (ox-LDL) were conducted to evaluate the effects of GPS on NLRP3 inflammasome activation

pyroptosis

apoptosis

and ROS production

specifically examining the role of the sirtuin 1 (SIRT1)/nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. SIRT1 and Nrf2 inhibitors were used to confirm the pathway's role.

Results:

2

GPS treatment significantly reduced atherosclerotic lesions in the en face aorta (

P

<

0.01)

as well as in the thoracic and abdominal aortic regions

and markedly decreased sinus lesions within the aortic root (

P

<

0.05 or

P

<

0.01). Additionally

GPS reduced oxidative stress markers and proinflammatory cytokines

includi

ng interleukin (IL)-1β and IL-18

in lesion areas (

P

<

0.05

P

<

0.01).

In vitro

GPS inhibited ox-LDL-induced NLRP3 activation

as evidenced by reduced NLRP3 (

P

<

0.01)

apoptosis-associated speck-like protein containing a CARD

cleaved-caspase-1

and cleaved-gasdermin D expressions (all

P

<

0.01). GPS also decreased ROS production

apoptosis

and pyroptosis

with the beneficial effects being significantly reversed by SIRT1 or Nrf2 inhibitors.

Conclusion:

2

GPS exerts an anti-atherogenic effect by inhibiting ROS-dependent NLRP3 inflammasome activation via the SIRT1/Nrf2 pathway.