News and Publications

Programmed Cell Death

R.A. Gottlieb

Programmed cell death, or apoptosis, refers to a process in which a cell responds to an external stimulus by committing suicide in a highly stereotyped manner. Events in apoptosis include cell shrinkage, degradation of the cell's chromatin and destruction of its genome, appearance of new antigens on the cell surface that mark the cell as a target for phagocytosis, cross-linking of cell proteins to form a leathery envelope at the cell surface, and eventually disintegration of the cell by blebbing and elimination of the resulting fragments by phagocytes. Apoptosis is 1 of the 3 general types of behavior of cells (the other 2 types are replication and differentiation), and it plays an indispensable role in development, tissue homeostasis, and response to injury.

This rapidly expanding field can be broken down into several areas: the conditions under which apoptosis is induced (many different diseases manifest disorders of apoptosis, including myocardial infarction, neurodegenerative diseases, and cancer), the genetic control of cell death (many transcripts are induced in cells subjected to an apoptotic stimulus), and the biochemical mechanisms of cell destruction (death proteases, mitochondrial alterations, and the role of Bcl-2 and its relatives are currently the focus of intense scrutiny).

Our earlier work showed that neutrophils acidify their cytoplasm during apoptosis and that the cytokine granulocyte colony-stimulating factor protected against both acidification and apoptosis by upregulating a membrane-associated proton pump. We have shown that components of the proton pump are stabilized in the presence of granulocyte colony-stimulating factor and that cytosolic components translocate to the membrane to form active pumps.

Similar studies in cardiomyocytes from adult rabbits showed that the stress response known as preconditioning appears to require activity of the proton pump for protection against injury mediated by ischemia and reperfusion. The proton pump spares activity of the sodium and hydrogen exchanger and secondary accumulation of calcium through the sodium and calcium exchanger. Previously, we showed that ischemia-reperfusion injury resulted in apoptosis of cardiomyocytes. Caspase activation is a general and essential feature of apoptosis; we have been able to show that caspase inhibitors are highly protective against ischemia-reperfusion injury. We are now cloning the cardiac caspases.

Acidification may be necessary for activation of the endonuclease responsible for degradation of DNA during apoptosis. In a cell culture model of cystic fibrosis, the elevated resting pH of the cells interferes with cytoplasmic acidification and apoptosis. Studies of apoptosis in the mouse model of cystic fibrosis (deletion of the gene that encodes the cystic fibrosis transmembrane conductance regulator [CFTR]) suggest that apoptosis is defective in exocrine tissues. In studies with cultures of mouse small intestinal crypt cells, intestinal cells that lacked CFTR were highly resistant to the induction of apoptosis caused by removal of extracellular matrix (Fig. 1).

Mitochondrial function is greatly impaired during apoptosis. Apoptosis results in defunctionalization of cytochrome c. This defunctionalization is characterized by dissociation of the cytochrome from the electron transport chain and is due to the action of a cytosolic factor known as cytochrome c inactivating factor of apoptosis. Bcl-2 opposes the action of this factor. Purification of the factor is under way, in collaboration with B. Babior, Department of Molecular and Experimental Medicine.

Mitochondrial inactivation is a feature of the response of neuronlike PC12 cells exposed to the amyloidogenic peptide derived from amyloid precursor protein, which is elevated in patients with Alzheimer's disease. Mitochondrial abnormalities are a general feature of neurodegenerative diseases, and cytochrome oxidase defects are implicated in Alzheimer's disease. Amyloid peptide appears to result in production of cytochrome c inactivating factor of apoptosis, with subsequent mitochondrial inactivation.

PUBLICATIONS

Adachi, S., Gottlieb, R.A., Babior, B.M. Lack of release of cytochrome c from mitochondria into cytosol early in the course of Fas-mediated apoptosis of Jurkat cells. J. Biol. Chem. 273:19892, 1998.

Karwatowska-Prokpczuk, E., Nordberg, J.A., Li, H.-L., Engler, R.L., Gottlieb,R.A. Effect of vacuolar proton ATPase on intracellular pH, calcium, and on apoptosis in neonatal cardiomyocytes during metabolic inhibition/recovery. Circ. Res. 82:1139, 1998.

Niessen, H., Meisenholder, G.W., Li, H.-L., Gluck, S.L., Lee, B.S., Forgac, M., Bowman, B., Engler, R.L., Babior, B.M., Gottlieb, R.A. G-CSF upregulates the vacuolar proton ATPase in human neutrophils. Blood 90:4598, 1997.