Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2007 Sep;117(9):2692-701.
doi: 10.1172/JCI29134.

TNF provokes cardiomyocyte apoptosis and cardiac remodeling through activation of multiple cell death pathways

Sandra B Haudek  1 , George E Taffet, Michael D Schneider, Douglas L Mann
Affiliations

TNF provokes cardiomyocyte apoptosis and cardiac remodeling through activation of multiple cell death pathways

Sandra B Haudek et al. J Clin Invest. 2007 Sep.

Abstract

Transgenic mice with cardiac-restricted overexpression of secretable TNF (MHCsTNF) develop progressive LV wall thinning and dilation accompanied by an increase in cardiomyocyte apoptosis and a progressive loss of cytoprotective Bcl-2. To test whether cardiac-restricted overexpression of Bcl-2 would prevent adverse cardiac remodeling, we crossed MHCsTNF mice with transgenic mice harboring cardiac-restricted overexpression of Bcl-2. Sustained TNF signaling resulted in activation of the intrinsic cell death pathway, leading to increased cytosolic levels of cytochrome c, Smac/Diablo and Omi/HtrA2, and activation of caspases -3 and -9. Cardiac-restricted overexpression of Bcl-2 blunted activation of the intrinsic pathway and prevented LV wall thinning; however, Bcl-2 only partially attenuated cardiomyocyte apoptosis. Subsequent studies showed that c-FLIP was degraded, that caspase-8 was activated, and that Bid was cleaved to t-Bid, suggesting that the extrinsic pathway was activated concurrently in MHCsTNF hearts. As expected, cardiac Bcl-2 overexpression had no effect on extrinsic signaling. Thus, our results suggest that sustained inflammation leads to activation of multiple cell death pathways that contribute to progressive cardiomyocyte apoptosis; hence the extent of such programmed myocyte cell death is a critical determinant of adverse cardiac remodeling.

PubMed Disclaimer

Figures

Figure 1
Figure 1. Characterization of mouse models.
Representative Western blots showing (A) levels of exogenous human Bcl-2 (hBcl-2), endogenous mouse Bcl-2 (mBcl-2), and total (human and mouse) Bcl-2 (ttBcl-2) as well as (B) levels of mouse TNF in cytosolic and mitochondrial protein extracts of 12-week-old littermate control (LM), MHCsTNF, MHCsTNF/Bcl-2, and Bcl-2 mouse hearts. GAPDH (cytosol) and COX IV (mitochondria) served as loading normalization. (C) Myocardial TNF protein levels were measured by ELISA. **P < 0.05 compared with LM.
Figure 2
Figure 2. Effect of Bcl-2 on LV remodeling.
(A) Representative M-mode echocardiograms for littermate control, MHCsTNF, MHCsTNF/Bcl-2, and Bcl-2 mouse hearts. Group data for (B) LVEDD, (C) posterior wall thickness (PWth), (D) r/h, and (E) percent fractional shortening (%FS) for LM, MHCsTNF, MHCsTNF/Bcl-2, and Bcl-2 mouse hearts. *P < 0.05 between 4 and 12 weeks of age within the same transgenic mouse line; **P < 0.05 between MHCsTNF and MHCsTNF/Bcl-2 at 12 weeks of age.
Figure 3
Figure 3. Cardiomyocyte apoptosis.
The prevalence of cardiomyocyte apoptosis was determined by the in situ DNA ligation method in littermate control, MHCsTNF, MHCsTNF/Bcl-2, and Bcl-2 mouse hearts at 12 weeks of age. *P < 0.05 between MHCsTNF and MHCsTNF/Bcl-2 mice; **P < 0.05 compared with LM.
Figure 4
Figure 4. Myocardial cytochrome c release.
The release of cytochrome c from (A) mitochondria to (B) the cytosol was determined by Western blotting in 12-week-old littermate control, MHCsTNF, MHCsTNF/Bcl-2, and Bcl-2 mouse hearts. Shown are representative Western blots next to the group data for mitochondrial cytochrome c levels (normalized to COX IV) and cytosolic cytochrome c levels (normalized to GAPDH). Group data represent the ratio of experimental (transgenic) to control (wild-type) mouse groups. *P < 0.05 between MHCsTNF and MHCsTNF/Bcl-2 groups; **P < 0.05 compared with LM.
Figure 5
Figure 5. Myocardial levels of proapoptotic proteins.
(A) The release of mitochondrial Smac/Diablo, Omi/HtrA2, apoptosis-inducing factor (AIF), and Endo G into the cytosol was determined by Western blotting in littermate control, MHCsTNF, MHCsTNF/Bcl-2, and Bcl-2 mouse hearts at 12 weeks of age. (B) Group data for the corresponding protein levels (normalized to GAPDH) are expressed as the ratio of experimental (transgenic) to control (wild-type) mouse groups. *P < 0.05 between MHCsTNF and MHCsTNF/Bcl-2 groups; **P < 0.05 compared with LM.
Figure 6
Figure 6. Myocardial caspase-3 and -9 activation.
(A) Caspase-3–like and (B) caspase-9–like activities were determined in 12-week-old littermate control, MHCsTNF, MHCsTNF/Bcl-2, and Bcl-2 mouse hearts using a fluorogenic assay. *P < 0.05 between MHCsTNF and MHCsTNF/Bcl-2 groups; **P < 0.05 compared with LM.
Figure 7
Figure 7. Myocardial caspase-8 activation, Bid, and IAPs.
(A) Caspase-8–like activity was determined in 12-week-old littermate control, MHCsTNF, MHCsTNF/Bcl-2, and Bcl-2 mouse hearts using a fluorogenic assay. Cytosolic levels of (B) full-length Bid (23 kDa) and t-Bid (15 kDa) and of (C) c–IAP-1 and c–IAP-2 as determined by Western blotting. Group data for the corresponding levels (normalized to GAPDH) are shown as the ratio of experimental (transgenic) to control (wild-type) mouse groups. x, empty lane. *P < 0.05 compared with LM.
Figure 8
Figure 8. Myocardial c-FLIPL and C-FLIPS levels.
(A) Cytosolic protein levels of c-FLIPL and c-FLIPS were determined in 12-week-old littermate control, MHCsTNF, MHCsTNF/Bcl-2, and Bcl-2 mouse hearts by Western blotting. Group data for the corresponding protein levels (normalized to GAPDH) are shown as the ratio of experimental (transgenic) to control (wild-type) mouse groups. (B) c-FLIPL mRNA levels were determined by RNase protection assay, and representative results as well as group data (normalized to L32) are shown. **P < 0.05 compared with LM.
Figure 9
Figure 9. Pro- and anti-apoptotic proteins that govern the extrinsic and intrinsic cell death pathways.
Proapoptotic proteins are shown in italic, antiapoptotic proteins in bold. TNF-induced signaling involves the formation of 2 sequential signaling complexes, complex I and complex II. Complex I is bound to TNF receptor superfamily, member 1 (TNFR1) and contains TNFR1-associated death domain (TRADD), TNFR-interacting serine-threonine kinase (RIP), and TNFR-associated factor 2 (TRAF2). Complex II is located in the cytosol and contains TRADD, RIP1, Fas-associated death domain (FADD), and caspase-8. See Discussion for further details.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Sivasubramanian N., et al. Left ventricular remodeling in transgenic mice with cardiac restricted overexpression of tumor necrosis factor. Circulation. 2001;104:826–831. - PubMed
    1. Bryant D., et al. Cardiac failure in transgenic mice with myocardial expression of tumor necrosis factor-α (TNF). Circulation. 1998;97:1375–1381. - PubMed
    1. Kubota T., et al. Dilated cardiomyopathy in transgenic mice with cardiac specific overexpression of tumor necrosis factor-alpha. Circ. Res. 1997;81:627–635. - PubMed
    1. Engel D., Peshock R., Armstrong R.C., Sivasubramanian N., Mann D.L. Cardiac myocyte apoptosis provokes adverse cardiac remodeling in transgenic mice with targeted TNF overexpression. Am. J. Physiol. Heart Circ. Physiol. 2004;287:H1303–H1311. - PubMed
    1. Bishopric N.H., Andreka P., Slepak T., Webster K.A. Molecular mechanisms of apoptosis in the cardiac myocyte. Curr. Opin. Pharmacol. 2001;1:141–150. - PubMed

Publication types