Mitochondrial Death Throes

Knockout mice reveal a path to necrosis.

Andrea Gawrylewski
Mar 31, 2007
<figcaption>Scanning electron micrograph of apoptotic T-cells. Credit: © STEM JEMS / PHOTO RESEARCHERS, INC</figcaption>
Scanning electron micrograph of apoptotic T-cells. Credit: © STEM JEMS / PHOTO RESEARCHERS, INC

Apoptosis normally proceeds in an orderly manner as part of natural cell turnover and development, but that's not the only path to death. A quick influx of calcium ions, or extreme oxidative stress, as happens in cases of stroke, heart attack, or other ischemic events, can cause mitochondria to swell and burst, releasing necrotic mediators. The path that cells take appears intimately linked with the permeability of mitochondrial membranes.

Four groups, publishing in 2005, independently showed that the mitochondrial protein cyclophilin D (CyPD) can mediate the transition to mitochondrial permeability that leads to necrotic cell death.1-4 Two of these papers have been cited more than 100 times each.1, 2 The studies offered insight into the regulation of cell-death decisions and established a key protein that works with the long-sought after permeability pore. Though the...

Pointing to the Pore

These papers confirmed suspicions about the interaction of CyPD and the permeability transition pore that researchers had observed pharmacologically: Cyclosporin A targets cyclophilin proteins minimizing mitochondrial destruction. The groups demonstrated the connection by knocking out CyPD in mouse mitochondria and then inducing ischemia, while recording the response. In response to Ca2+ influx or oxidative stress, mitochondria lacking CyPD had limited pore opening and cell death.

Yoshihide Tsujimoto's group, at the Osaka Medical School in Japan, showed in their Hot Paper that in various cell types lacking CyPD, apoptosis still occurs, further linking the CyPD protein to necrosis.2 The work supports earlier hypotheses that pore opening is the result of a conformational change of the adenine nucleotide translocase (ANT), a protein on the inner mitochondrial membrane. Some researchers, including Tsujimoto, originally believed necrosis to be an outer-membrane-mediated process.

The picture isn't yet complete. While these papers showed that mitochondria lacking the CyPD protein were protected from reperfusion, very high Ca2+ levels could still induce pore opening. Also, the Molkentin group showed that the permeability transition pore could be involved in cases of apoptosis, given high levels of CyPD.

"The pore can open even in the absence of cyclophilin D or in the presence of cyclosporin A," says Paolo Bernardi, senior author on a fourth CyPD-knock out paper.3 Bernardi, at the University of Padova neuroscience department, remains skeptical about the exact mechanisms of the mitochondrial pore involved in programmed cell death. "Knock-out mice for cyclophilin D can only address the role of cyclophilin D in pore regulation; they cannot address the question of whether the pore is involved or not [in apoptosis]."

Pointing to Drug Targets

Speculation is widespread about the structure and activity of the pore, and the role that CyPD plays in stable, healthy cells remains undefined. Nonetheless, researchers still consider CyPD a strong target for mediating cell trauma after ischemic events such as stroke or heart attack. Indeed, in the 2005 paper that Stanley Korsmeyer's lab published, cerebral ischemia was used as the model to test protection against injury by CyPD-deficient mitochondria.4

Continued research has focused on both necrotic and apoptotic events, some of which can be mediated by cyclophilin inhibitors such as cyclosporin A. Alessia Angelin, in Bernardi's lab, has shown that cyclosporin A can mitigate fiber disintegration in muscular dystrophy.5 The group has worked with a compound called Debio-025 (Debio Pharmaceuticals developed it in the late 1990s), which has a 5 to 10 times stronger blocking effect on cyclophilins than cyclosporin A. Debio is currently testing Debio-025's efficacy as an antiviral against HIV and hepatitis C, while Novartis is working on a compound that would block CyPD during ischemic events.

Andrew Halestrap, from the University of Bristol, who was not involved in any of the studies, has worked with the Debio compound but doesn't see it having a future in protecting against ischemic trauma. "Our own work is to find better mimics of preconditioning to block the pore, rather than relying on the cyclosporin," Halestrap says. Preventing the entire chain of events that leads to mitochondrial explosion may be more practical than treating cells with necrotic mediators after ischemic injury has occurred, he adds.

Data derived from the Science Watch/Hot Papers database and the Web of Science (Thomson ISI) show that Hot Papers are cited 50 to 100 times more often than the average paper of the same type and age.
C.P. Baines et al., "Loss of cyclophilin D reveals a critical role for mitochondrial permeability transition in cell death," Nature, 434:658-62, 2005. (Cited in 120 papers) T. Nakagawa et al., "Cyclophilin D-dependent mitochondrial permeability transition regulates some necrotic but not apoptotic cell death," Nature, 434:652-8, 2005. (Cited in 112 papers)

Naturally, understanding cell death is vital to cancer studies as well. The two Hot Papers, and accompanying research, established a potential target for apoptosis induction in cancer therapy: Successful protection from reperfusion achieved by knocking out CyPD indicates an important role of the ANT protein. The CyPD mice experiments were also ideal in that they demonstrated natural cell function, even without the missing protein, a result that surprised other researchers such as Guido Kroemer from the Gustave Roussy Institute in France. This will help them locate the key protein targets for inducing cell death without ruining the entire function of the cell.

Mitochondria in cancer cells have shown strong resistance to calcium ions and therefore do not easily open their necrotic pores. "The challenge is to take advantage of this distinct [characteristic] and make drugs that target the cancer mitochondria and leave the regular mitochondria alone."

References

1. C.P. Baines et al., "Loss of cyclophilin D reveals a critical role for mitochondrial permeability transition in cell death," Nature, 434:658-62, 2005. (Cited in 120 papers) 2. T. Nakagawa et al., "Cyclophilin D-dependent mitochondrial permeability transition regulates some necrotic but not apoptotic cell death," Nature, 434:652-8, 2005. (Cited in 113 papers) 3. E. Basso et al., "Properties of the permeability transition pore in mitochondria devoid of cyclophilin D," J Biol Chem, 280:18558-61, 2005. (cited in 57 papers) 4. A. Schinzel et al., "Cyclophilin D is a component of mitochondrial permeability transition and mediates neuronal cell death after focal cerebral ischemia," Proc Natl Acad Sci, 102:12005-10, 2005. (cited in 26 papers) 5. A. Angelin et al., "Mitochondrial dysfunction in the pathogenesis of Ullrich congenital muscular dystrophy and prospective therapy with cyclosporins," Proc Natl Acad Sci, 104:991-6, Jan. 16, 2007.