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Updated: Feb 24, 2026
Characterization of MLKL-mediated Plasma Membrane Rupture in Necroptosis
Published on: August 7, 2018
RIP3 AND pMLKL promote necroptosis-induced inflammation and alter membrane permeability in intestinal epithelial
Anna Negroni1, Eleonora Colantoni2, Maria Pierdomenico2
1Division of Health Protection Technologies, Territorial and Production Systems Sustainability Department, ENEA, Rome, Italy.
Background:
Necroptosis is an inflammatory form of programmed cell death requiring receptor-interacting protein kinase 3 (RIP3) and mixed lineage kinase domain-like protein (MLKL).
Aims:
The aim of this study is to examine in depth in vitro and ex vivo the contribution of necroptosis to intestinal inflammation.
Methods:
In vitro: we used an intestinal cell line, HCT116RIP3, produced in our laboratory and overexpressing RIP3. Ex vivo: intestinal mucosal biopsies were taken from patients with inflammatory bowel disease (IBD) (20 with Crohn's disease; 20 with ulcerative colitis) and from 20 controls.
Results:
RIP3-induced necroptosis triggers MLKL activation, increases cytokine/alarmin expression (IL-8, IL-1β, IL-33, HMGB1), NF-kBp65 translocation and NALP3 inflammasome assembly. It also affects membrane permeability by altering cell-cell junctional proteins (E-cadherin, Occludin, Zonulin-1). Targeting necroptosis through Necrostatin-1 significantly reduces intestinal inflammation in vitro and in cultured intestinal explants from IBD.
Conclusion:
We show for the first time in vitro and ex vivo that RIP3-driven necroptosis seriously affects intestinal inflammation by increasing pMLKL, activating different cytokines and alarmins, and altering epithelial permeability. The inhibition of necroptosis causes a significant decrease of all these effects. These data strongly support the view that targeting necroptosis may represent a promising new option for the treatment of inflammatory enteropathies.
Insights
Necroptosis, a form of programmed cell death involving RIP3 and MLKL, significantly drives intestinal inflammation by increasing cytokines and altering gut barrier function. Inhibiting necroptosis reduces these inflammatory markers, offering a potential therapeutic strategy for inflammatory enteropathies.
Area of Science:
- Cell Biology
- Immunology
- Gastroenterology
Background:
- Necroptosis is an inflammatory programmed cell death pathway.
- It requires receptor-interacting protein kinase 3 (RIP3) and mixed lineage kinase domain-like protein (MLKL).
Purpose of the Study:
- To investigate the role of necroptosis in intestinal inflammation.
- To assess necroptosis in vitro and ex vivo using cell lines and patient biopsies.
Main Methods:
- Utilized an in vitro intestinal cell line (HCT116RIP3) overexpressing RIP3.
- Examined ex vivo intestinal mucosal biopsies from inflammatory bowel disease (IBD) patients and controls.
- Assessed markers of necroptosis, inflammation, and epithelial barrier function.
Main Results:
- RIP3-induced necroptosis activates MLKL, increases pro-inflammatory cytokines (IL-8, IL-1β, IL-33, HMGB1), and promotes NF-kB p65 translocation.
- Necroptosis alters epithelial barrier integrity by affecting cell-cell junction proteins (E-cadherin, Occludin, Zonulin-1).
- Necrostatin-1, a necroptosis inhibitor, significantly reduced intestinal inflammation in vitro and in cultured IBD explants.
Conclusions:
- RIP3-driven necroptosis exacerbates intestinal inflammation through cytokine release and epithelial barrier disruption.
- Inhibition of necroptosis effectively mitigates these inflammatory effects.
- Targeting necroptosis presents a promising therapeutic avenue for inflammatory enteropathies.
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