Limits...
Autophagy and senescence in cancer-associated fibroblasts metabolically supports tumor growth and metastasis via glycolysis and ketone production.

Capparelli C, Guido C, Whitaker-Menezes D, Bonuccelli G, Balliet R, Pestell TG, Goldberg AF, Pestell RG, Howell A, Sneddon S, Birbe R, Tsirigos A, Martinez-Outschoorn U, Sotgia F, Lisanti MP - Cell Cycle (2012)

Bottom Line: An important clue comes from recent studies linking autophagy with the onset of senescence.Thus, we genetically validated the existence of the autophagy-senescence transition.Autophagic-senescent fibroblasts stimulated mitochondrial metabolism in adjacent cancer cells, when the two cell types were co-cultured, as visualized by MitoTracker staining.

View Article: PubMed Central - PubMed

Affiliation: The Jefferson Stem Cell Biology and Regenerative Medicine Center, Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA, USA.

ABSTRACT
Senescent fibroblasts are known to promote tumor growth. However, the exact mechanism remains largely unknown. An important clue comes from recent studies linking autophagy with the onset of senescence. Thus, autophagy and senescence may be part of the same physiological process, known as the autophagy-senescence transition (AST). To test this hypothesis, human fibroblasts immortalized with telomerase (hTERT-BJ1) were stably transfected with autophagy genes (BNIP3, CTSB or ATG16L1). Their overexpression was sufficient to induce a constitutive autophagic phenotype, with features of mitophagy, mitochondrial dysfunction and a shift toward aerobic glycolysis, resulting in L-lactate and ketone body production. Autophagic fibroblasts also showed features of senescence, with increased p21(WAF1/CIP1), a CDK inhibitor, cellular hypertrophy and increased β-galactosidase activity. Thus, we genetically validated the existence of the autophagy-senescence transition. Importantly, autophagic-senescent fibroblasts promoted tumor growth and metastasis, when co-injected with human breast cancer cells, independently of angiogenesis. Autophagic-senescent fibroblasts stimulated mitochondrial metabolism in adjacent cancer cells, when the two cell types were co-cultured, as visualized by MitoTracker staining. In particular, autophagic ATG16L1 fibroblasts, which produced large amounts of ketone bodies (3-hydroxy-butyrate), had the strongest effects and promoted metastasis by up to 11-fold. Conversely, expression of ATG16L1 in epithelial cancer cells inhibited tumor growth, indicating that the effects of autophagy are compartment-specific. Thus, autophagic-senescent fibroblasts metabolically promote tumor growth and metastasis, by paracrine production of high-energy mitochondrial fuels. Our current studies provide genetic support for the importance of "two-compartment tumor metabolism" in driving tumor growth and metastasis via a simple energy transfer mechanism. Finally, β-galactosidase, a known lysosomal enzyme and biomarker of senescence, was localized to the tumor stroma in human breast cancer tissues, providing in vivo support for our hypothesis. Bioinformatic analysis of genome-wide transcriptional profiles from tumor stroma, isolated from human breast cancers, also validated the onset of an autophagy-senescence transition. Taken together, these studies establish a new functional link between host aging, autophagy, the tumor microenvironment and cancer metabolism.

Show MeSH

Related in: MedlinePlus

Figure 9. BNIP3-, CTSB- and ATG16L1- fibroblasts all show mitochondrial dysfunction, with increased production of L-lactate or ketone bodies. (A) Loss of functional mitochondrial causes changes in cell metabolism, leading to the accumulation of L-lactate. Note that BNIP3- and CTSB fibroblasts both show significant increases in L-lactate production, between 25-to-37%. However, ATG16L1 fibroblasts did not show any increases in L-lactate accumulation. (B, C) Mitochondrial dysfunction can also activate ketone body production, resulting in the accumulation of 3-hydroxy-butyrate. Note that only ATG16L1 fibroblasts showed increases in ketone production, resulting in an up to 2.3-fold accumulation of 3-hydroxy-butyrate. The data were normalized either for cell number or for protein content per well. Thus, BNIP3- and CTSB fibroblasts produce L-lactate, while ATG16L1 fibroblasts produce ketone bodies, as a consequence of autophagy and the resulting mitochondrial dysfunction. (D) Autophagic fibroblasts were co-cultured with MDA-MB-231-GFP cells, and mitochondrial activity was visualized by MitoTracker staining. Note that all three autophagic fibroblast cell lines (BNIP3, CTSB, and ATG16L1) increased the MitoTracker staining (RED; Upper panels) in adjacent MDA-MB-231 cells (GREEN; Lower panels), during co-culture. This is consistent with the notion that autophagic fibroblasts provide mitochondrial fuels, such as L-lactate and ketone bodies, for the anabolic growth of cancer cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3383590&req=5

Figure 9: Figure 9. BNIP3-, CTSB- and ATG16L1- fibroblasts all show mitochondrial dysfunction, with increased production of L-lactate or ketone bodies. (A) Loss of functional mitochondrial causes changes in cell metabolism, leading to the accumulation of L-lactate. Note that BNIP3- and CTSB fibroblasts both show significant increases in L-lactate production, between 25-to-37%. However, ATG16L1 fibroblasts did not show any increases in L-lactate accumulation. (B, C) Mitochondrial dysfunction can also activate ketone body production, resulting in the accumulation of 3-hydroxy-butyrate. Note that only ATG16L1 fibroblasts showed increases in ketone production, resulting in an up to 2.3-fold accumulation of 3-hydroxy-butyrate. The data were normalized either for cell number or for protein content per well. Thus, BNIP3- and CTSB fibroblasts produce L-lactate, while ATG16L1 fibroblasts produce ketone bodies, as a consequence of autophagy and the resulting mitochondrial dysfunction. (D) Autophagic fibroblasts were co-cultured with MDA-MB-231-GFP cells, and mitochondrial activity was visualized by MitoTracker staining. Note that all three autophagic fibroblast cell lines (BNIP3, CTSB, and ATG16L1) increased the MitoTracker staining (RED; Upper panels) in adjacent MDA-MB-231 cells (GREEN; Lower panels), during co-culture. This is consistent with the notion that autophagic fibroblasts provide mitochondrial fuels, such as L-lactate and ketone bodies, for the anabolic growth of cancer cells.

Mentions: Loss of functional mitochondria is associated with key changes in cell metabolism, leading to the accumulation of L-lactate and/or ketone bodies as end products in the tissue culture media. Figure 9A shows that BNIP3- and CTSB fibroblasts both increase L-lactate production, between 25%-to-37%. However, ATG16L1 fibroblasts did not show any increase in L-lactate accumulation.


Autophagy and senescence in cancer-associated fibroblasts metabolically supports tumor growth and metastasis via glycolysis and ketone production.

Capparelli C, Guido C, Whitaker-Menezes D, Bonuccelli G, Balliet R, Pestell TG, Goldberg AF, Pestell RG, Howell A, Sneddon S, Birbe R, Tsirigos A, Martinez-Outschoorn U, Sotgia F, Lisanti MP - Cell Cycle (2012)

Figure 9. BNIP3-, CTSB- and ATG16L1- fibroblasts all show mitochondrial dysfunction, with increased production of L-lactate or ketone bodies. (A) Loss of functional mitochondrial causes changes in cell metabolism, leading to the accumulation of L-lactate. Note that BNIP3- and CTSB fibroblasts both show significant increases in L-lactate production, between 25-to-37%. However, ATG16L1 fibroblasts did not show any increases in L-lactate accumulation. (B, C) Mitochondrial dysfunction can also activate ketone body production, resulting in the accumulation of 3-hydroxy-butyrate. Note that only ATG16L1 fibroblasts showed increases in ketone production, resulting in an up to 2.3-fold accumulation of 3-hydroxy-butyrate. The data were normalized either for cell number or for protein content per well. Thus, BNIP3- and CTSB fibroblasts produce L-lactate, while ATG16L1 fibroblasts produce ketone bodies, as a consequence of autophagy and the resulting mitochondrial dysfunction. (D) Autophagic fibroblasts were co-cultured with MDA-MB-231-GFP cells, and mitochondrial activity was visualized by MitoTracker staining. Note that all three autophagic fibroblast cell lines (BNIP3, CTSB, and ATG16L1) increased the MitoTracker staining (RED; Upper panels) in adjacent MDA-MB-231 cells (GREEN; Lower panels), during co-culture. This is consistent with the notion that autophagic fibroblasts provide mitochondrial fuels, such as L-lactate and ketone bodies, for the anabolic growth of cancer cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3383590&req=5

Figure 9: Figure 9. BNIP3-, CTSB- and ATG16L1- fibroblasts all show mitochondrial dysfunction, with increased production of L-lactate or ketone bodies. (A) Loss of functional mitochondrial causes changes in cell metabolism, leading to the accumulation of L-lactate. Note that BNIP3- and CTSB fibroblasts both show significant increases in L-lactate production, between 25-to-37%. However, ATG16L1 fibroblasts did not show any increases in L-lactate accumulation. (B, C) Mitochondrial dysfunction can also activate ketone body production, resulting in the accumulation of 3-hydroxy-butyrate. Note that only ATG16L1 fibroblasts showed increases in ketone production, resulting in an up to 2.3-fold accumulation of 3-hydroxy-butyrate. The data were normalized either for cell number or for protein content per well. Thus, BNIP3- and CTSB fibroblasts produce L-lactate, while ATG16L1 fibroblasts produce ketone bodies, as a consequence of autophagy and the resulting mitochondrial dysfunction. (D) Autophagic fibroblasts were co-cultured with MDA-MB-231-GFP cells, and mitochondrial activity was visualized by MitoTracker staining. Note that all three autophagic fibroblast cell lines (BNIP3, CTSB, and ATG16L1) increased the MitoTracker staining (RED; Upper panels) in adjacent MDA-MB-231 cells (GREEN; Lower panels), during co-culture. This is consistent with the notion that autophagic fibroblasts provide mitochondrial fuels, such as L-lactate and ketone bodies, for the anabolic growth of cancer cells.
Mentions: Loss of functional mitochondria is associated with key changes in cell metabolism, leading to the accumulation of L-lactate and/or ketone bodies as end products in the tissue culture media. Figure 9A shows that BNIP3- and CTSB fibroblasts both increase L-lactate production, between 25%-to-37%. However, ATG16L1 fibroblasts did not show any increase in L-lactate accumulation.

Bottom Line: An important clue comes from recent studies linking autophagy with the onset of senescence.Thus, we genetically validated the existence of the autophagy-senescence transition.Autophagic-senescent fibroblasts stimulated mitochondrial metabolism in adjacent cancer cells, when the two cell types were co-cultured, as visualized by MitoTracker staining.

View Article: PubMed Central - PubMed

Affiliation: The Jefferson Stem Cell Biology and Regenerative Medicine Center, Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA, USA.

ABSTRACT
Senescent fibroblasts are known to promote tumor growth. However, the exact mechanism remains largely unknown. An important clue comes from recent studies linking autophagy with the onset of senescence. Thus, autophagy and senescence may be part of the same physiological process, known as the autophagy-senescence transition (AST). To test this hypothesis, human fibroblasts immortalized with telomerase (hTERT-BJ1) were stably transfected with autophagy genes (BNIP3, CTSB or ATG16L1). Their overexpression was sufficient to induce a constitutive autophagic phenotype, with features of mitophagy, mitochondrial dysfunction and a shift toward aerobic glycolysis, resulting in L-lactate and ketone body production. Autophagic fibroblasts also showed features of senescence, with increased p21(WAF1/CIP1), a CDK inhibitor, cellular hypertrophy and increased β-galactosidase activity. Thus, we genetically validated the existence of the autophagy-senescence transition. Importantly, autophagic-senescent fibroblasts promoted tumor growth and metastasis, when co-injected with human breast cancer cells, independently of angiogenesis. Autophagic-senescent fibroblasts stimulated mitochondrial metabolism in adjacent cancer cells, when the two cell types were co-cultured, as visualized by MitoTracker staining. In particular, autophagic ATG16L1 fibroblasts, which produced large amounts of ketone bodies (3-hydroxy-butyrate), had the strongest effects and promoted metastasis by up to 11-fold. Conversely, expression of ATG16L1 in epithelial cancer cells inhibited tumor growth, indicating that the effects of autophagy are compartment-specific. Thus, autophagic-senescent fibroblasts metabolically promote tumor growth and metastasis, by paracrine production of high-energy mitochondrial fuels. Our current studies provide genetic support for the importance of "two-compartment tumor metabolism" in driving tumor growth and metastasis via a simple energy transfer mechanism. Finally, β-galactosidase, a known lysosomal enzyme and biomarker of senescence, was localized to the tumor stroma in human breast cancer tissues, providing in vivo support for our hypothesis. Bioinformatic analysis of genome-wide transcriptional profiles from tumor stroma, isolated from human breast cancers, also validated the onset of an autophagy-senescence transition. Taken together, these studies establish a new functional link between host aging, autophagy, the tumor microenvironment and cancer metabolism.

Show MeSH
Related in: MedlinePlus