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Mitochondrial dysfunction links ceramide activated HRK expression and cell death

Farhan Rizvi1, Tom Heimann, Anja Herrnreiter

  • 1Department of Ophthalmology, Medical College of Wisconsin, Milwaukee, Wisconsin, United States of America. frizvi@mcw.edu

Plos One
|April 13, 2011
PubMed
Abstract

Insights

Ceramide induces human corneal stromal fibroblast death by causing mitochondrial dysfunction and upregulating the harikari gene (HRK). HRK knockdown reduces ceramide-induced cell death, highlighting its role in corneal wound healing.

Area of Science:

  • Ophthalmology
  • Cell Biology
  • Molecular Biology

Background:

  • Cell death is crucial for corneal wound healing following injury.
  • The precise molecular mechanisms of corneal stromal cell death after injury, such as following LASIK surgery, are not fully understood.
  • Ceramide has been implicated in stromal cell death post-LASIK.

Purpose of the Study:

  • To investigate the mechanism by which C6 ceramide induces cell death in human corneal stromal fibroblasts (HCSF).
  • To elucidate the role of ceramide in corneal stromal cell death and its potential implications for wound healing.

Main Methods:

  • HCSF cultures were treated with C6 ceramide or a control.
  • Cell death was assessed using Live/Dead staining, Annexin V, caspase activation, and TUNEL assays.
  • Mitochondrial function, gene expression (qPCR, Western blotting), and protein interactions were analyzed.

Main Results:

  • Ceramide induced mitochondrial dysfunction, evidenced by reduced MTT, cytochrome c release, increased ROS, and loss of mitochondrial membrane potential.
  • Ceramide triggered the expression of the harikari gene (HRK) and JNK phosphorylation, leading to HRK translocation to mitochondria.
  • HRK interacted with p32 and BAD in mitochondria, and HRK knockdown significantly reduced ceramide-induced cell death.

Conclusions:

  • Ceramide induces human corneal stromal fibroblast death via HRK-mediated mitochondrial dysfunction.
  • This pathway is a key mechanism contributing to corneal stromal cell death and potentially corneal wound healing.

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