Related Experiment Video

Updated: Aug 22, 2026

Activation of Apoptosis by Cytoplasmic Microinjection of Cytochrome c
07:42

Activation of Apoptosis by Cytoplasmic Microinjection of Cytochrome c

Published on: June 29, 2011

Diazoxide-mediated preconditioning against apoptosis involves activation of cAMP-response element-binding protein

Roman A Eliseev1, Beth Vanwinkle, Randy N Rosier

  • 1Musculo-Skeletal Research Unit, University of Rochester School of Medicine, Rochester, New York 14642, USA.

The Journal of Biological Chemistry
|August 25, 2004
PubMed

Insights

Diazoxide preconditioning protects cells from apoptosis by activating transcription factors CREB and NF-kappaB, leading to increased levels of survival proteins like cytochrome c and Bcl-xl.

Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Mitochondrial ATP-sensitive K+ channel openers, such as diazoxide, are known to precondition cells against injury and inhibit apoptosis.
  • The precise molecular mechanisms underlying diazoxide-mediated preconditioning remain incompletely understood.

Purpose of the Study:

  • To investigate the effects of diazoxide pretreatment on mitochondrial function and morphology in HL60 cells.
  • To elucidate the role of diazoxide in modulating cellular susceptibility to apoptosis.

Main Methods:

  • HL60 cells were pretreated with diazoxide, followed by exposure to etoposide to induce apoptosis.
  • Mitochondrial morphology, function (oxidative phosphorylation, membrane potential), and protein/mRNA levels (cytochrome c, Bcl-xl) were assessed.
  • Activation of transcription factors cAMP-response element-binding protein (CREB) and NF-kappaB was analyzed.
  • Specific inhibitors of mitochondrial K+ channels (5-hydroxydecanoate) and NF-kappaB (SN50) were used to confirm pathway specificity.

Main Results:

  • Diazoxide pretreatment inhibited etoposide-induced apoptosis and mitochondrial dysfunction.
  • Diazoxide induced moderate mitochondrial swelling and release of cytochrome c, alongside increased expression of cytochrome c and Bcl-xl.
  • Activation of CREB and NF-kappaB was observed, correlating with increased prosurvival protein levels and resistance to apoptosis.
  • Inhibitor studies confirmed the involvement of mitochondrial K+ channels and NF-kappaB in diazoxide's protective effects.

Conclusions:

  • Diazoxide-mediated preconditioning against apoptosis involves the activation of pro-survival transcription factors CREB and NF-kappaB.
  • This activation leads to the upregulation of key proteins like cytochrome c and Bcl-xl, conferring cellular resistance to apoptotic stimuli.
  • The observed effects are likely initiated by an increase in cytosolic calcium following diazoxide treatment.

More Related Videos

Lighting Up the Pathways to Caspase Activation Using Bimolecular Fluorescence Complementation08:47

Lighting Up the Pathways to Caspase Activation Using Bimolecular Fluorescence Complementation

Published on: March 5, 2018

Related Concept Videos

The Intrinsic Apoptotic Pathway01:31

The Intrinsic Apoptotic Pathway

Internal cellular stress, such as cellular injury or hypoxia, triggers intrinsic apoptosis. The B-cell lymphoma 2 (Bcl-2) family of proteins are the primary regulators of the intrinsic apoptotic pathway. For example, during DNA damage, checkpoint proteins, such as Ataxia Telangiectasia Mutated (ATM protein) and Checkpoints Factor-2 (Chk2) proteins, are activated. These proteins phosphorylate p53 which further activates pro-apoptotic proteins, such as Bax, Bak, PUMA, and Noxa, and inhibits...
Caspases01:24

Caspases

Caspase, a family of cysteine proteases, serve as effectors in apoptosis. The ced3 gene in C.elegans was first identified to be involved in apoptosis. This gene encodes the ced-3 caspase that is similar to the interleukin-1-beta converting enzyme or ICE in mammals. In addition to apoptosis, caspases also function in the inflammatory response. Inflammatory caspases are essential in activating pro-inflammatory cytokines that recruit immune cells and block the replication of pathogens inside cells.
The Extrinsic Apoptotic Pathway01:17

The Extrinsic Apoptotic Pathway

The extrinsic apoptotic pathway is initiated when extracellular death-inducing signals, such as specific cytokines, activate the death receptors expressed on the cell surface. The immune cells involved in this pathway are natural killer cells (NK cells) and cytotoxic T-lymphocytes. NK cells are critical in innate immune response, while cytotoxic T-lymphocytes are associated with adaptive immune response. These cells recognize specific receptors expressed on the altered cells and activate...
MAPK Signaling Cascades01:07

MAPK Signaling Cascades

Mitogen-activated protein kinase, or MAPK pathway, activates three sequential kinases to regulate cellular responses such as proliferation, differentiation, survival, and apoptosis. The canonical MAPK pathway starts with a mitogen or growth factor binding to an RTK. The activated RTKs stimulate Ras, which recruits Raf or MAP3 Kinase (MAPKKK), the first kinase of the MAPK signaling cascade. Raf further phosphorylates and activates MEK or MAP2 Kinases (MAPKK), which in turn phosphorylates MAP...
Apoptosis01:30

Apoptosis

Apoptosis is a combination of two Greek words, 'apo' and 'ptosis,' meaning separation and falling off, respectively. Hippocrates used this word to describe gangrene, which was caused due to bandaging of fractured bones. Apoptosis was distinguished from necrosis in 1970 when John Kerr reported observations of morphological changes occurring during apoptosis. During one experiment, he observed that the disruption of blood supply to the liver tissue resulted in a size reduction of the tissue.
cAMP-dependent Protein Kinase Pathways01:25

cAMP-dependent Protein Kinase Pathways

Cyclic Adenosine Monophosphate (cAMP) is an essential second messenger that activates protein kinase A (PKA) and regulates various biological processes. A single epinephrine molecule binds to GPCR and activates several heterotrimeric G proteins, each stimulating multiple adenylyl cyclase, amplifying the signal, and synthesizing large numbers of cAMP molecules. Small changes in cAMP concentration affect PKA activity. The binding of four cAMP molecules induces a conformational change in PKA,...