PhD Scientific Days 2024

Poster Session K - Theoretical and Translational Medicine 2.

A cell-based screening platform for testing inflammasome inhibitors

Előadó neve

Dr. Onódi, Zsófia, PhD

Előadó munkahelye

Department of Pharmacology and Pharmacotherapy

Előadó telefonszáma

06308771710

Előadó e-mail címe

onodi.zsofia@semmelweis.hu

Az előadás címe

A cell-based screening platform for testing inflammasome inhibitors

Szerző(k) neve és munkahelye

Zsófia Onódi¹, Márton Kocsis¹, Gergely Kozma², Márta Szabó¹, Zoltán V. Varga¹

1: Department of Pharmacology and Pharmacotherapy, Semmelweis University, Budapest, Hungary
2: Center of Translational Medicine, Semmelweis University, Budapest, Hungary

Bemutatás módja

Poszter

Szekció

Poster Session K - Theoretical and Translational Medicine 2.

Language of the presentation

Hungarian

Preferred session

Theoretical and Translational Medicine

Összefoglaló szövege

Introduction: Inflammasome activation has been shown to be an important player in inflammation-related conditions such as autoimmune or cardiovascular diseases; thus, targeting inflammasomes may be a promising therapeutic approach for these conditions. Several candidates were identified as potential inflammasome inhibitors; however, there is a lack of evidence on direct inhibitory effects of these agents.
Aims: to optimize a cell culture-based model for testing inflammasome inhibitors.
Method: THP1 ASC-GFP cell line was used for examining inflammasomes. For NLRP3 inflammasome induction, the cells were primed with lipopolysaccharide (LPS) for 24h to induce expression of inflammasome proteins, then activated by nigericin. For AIM2 induction, cells were incubated with poly(dA:dT) for 24h. During the 24-hour priming, drug candidates were used to inhibit inflammasomes including probenecid (an uricosuric), sotagliflozin (an SGLT1/2 inhibitor) and montelukast (a leukotriene receptor antagonist). NLRP3 inhibitor MCC950 and caspase-1 inhibitor VX-765 were used as positive controls. Treated cells were harvested for Western blot (WB) and confocal microscopy to assess inflammasome priming and activation. Inflammasome oligomerization was visualized and assessed by imaging flow cytometry (IFC). The ratio of ASC diffuse (primed) and ASC speck+ (activated) cells were calculated.
Results: NLRP3 priming and activation was successfully induced with LPS and nigericin, as increased expression and cleavage of inflammasome proteins (e.g. interleukin 1 beta (IL-1b), caspase-1, cleaved IL-1b) was detected by WB. AIM2 activation was induced in THP-1 cells; however, markers showed milder changes compared to NLRP3 model. MCC950 reduced the amount of cleaved IL-1b and ASC speck+ cells. Probenecid attenuated cleaved IL-1b secretion suggesting its modulatory effect on inflammasomes. Sotagliflozin did not alter inflammasome priming or activation. Surprisingly, montelukast significantly reduced the viability of THP-1 cells.
Conclusion: Our screening platform successfully detected inhibition of inflammasomes in THP-1 cells. Some of the tested agents did not show clear inhibition in our system suggesting that reported anti-inflammatory effect might be mediated by other pathways.
Funding: The work was supported by the MTA [LP2021-14], NKFIH [ÚNKP-23-4-II-SE-26], NCL (RRF-2.3.1-21-2022-00003).

University

Semmelweis University

Supervisor

n/a

Publication of my abstract

I do not give consent to the publication of my abstract on the website of the congress.

Kind

Szabad

Status

elfogadva

Accepted presentation method

poszter

Előadás fájl jóváhagyás

nem rendelkezett róla

Előadó

2853

Start

16:10

End

16:13