Autophagy in Stem Cells
Introduction:
Stem-cell:
Stem cells have been identified and characterized in several mammalian tissues. In addition, pluripotent embryonic stem cells have been derived from pre-implantation embryos in both mice and humans. Whereas the recent advances in stem-cell research have the potential to revolutionize medicine, the critical scientific challenge remains to elucidate the fundamental cellular and molecular controls of stem cells. An understanding of the molecular mechanisms that govern stem-cell fate and the identification of specific stem-cell markers is of fundamental significance in cell and developmental biology.
Stem cells have been identified and characterized in several mammalian tissues. In addition, pluripotent embryonic stem cells have been derived from pre-implantation embryos in both mice and humans. Whereas the recent advances in stem-cell research have the potential to revolutionize medicine, the critical scientific challenge remains to elucidate the fundamental cellular and molecular controls of stem cells. An understanding of the molecular mechanisms that govern stem-cell fate and the identification of specific stem-cell markers is of fundamental significance in cell and developmental biology.
+ add some more words such as induced pluripotent stem cells (called iPS cells), tzpes of steë=cells in tblem, etc
->--add some more descriptions, and integrate the picture here asw ell as the abreviations--)
Autophagy:
Autophagy or ‘‘self-eating’’ catabolism of cytoplasm, is a genetically programmed process that degrades long-lived cellular proteins and organelles and during nutrient deprivation, provides a supply of amino acids needed for cell survival. (Klionsky DJ, Emr SD. Autophagy as a regulated pathway of cellular degradation. Science 2000;290:1717–1721). Autophagy is important in normal development and response to changing environmental stimuli and autophagy has been found to be important in growth control and is defective in some tumor cells, bacterial and viral infections, neurodegenerative disorders, and cardiovascular disease (Shintani T, Klionsky DJ. Autophagy in health and disease: a double-edged sword. Science 2004;306:990–5.). The turnover of cytoplasmic constituents for energy is regulated by the endosomal/autophagic/lysosomal (EAL) system, complementing the degradation of most short-lived proteins by the ubiquitin-proteasome system. (Barry Boland etal Neuronal macroautophagy: From development to degeneration Molecular Aspects of Medicine 27 (2006) 503–519).
Though autophagy generally considered as a ‘self-eating’ process, this process occur by at least minimum 3 different routes:
Types of autophagy:
Macroautophagy(MA):
-this pathway proceeds the sequestration of cytosolic regions containing proteins, sugars, lipids, RNA, as well as organelles such as mitochondria, perixosomes into double-membraned vacuoles
that deliver their contents to late endosomal and lysosomal compartments for degradation with the help of the Apg12-Apg5 conjugate( Shintani T etal Autophagy in health and disease: a double-edged sword. Science 2004 306,990–995)
- this pathway activated during conditions of serum withdrawal in cell culture ,
-inhibitors for MA is 3-methyladenine, wortmannin, and LY294002
-protein degradation by MA is sensitive to microtubule inhibitors such as colcemide and vinblastine,
- Activators of MA is rapamycin,
chaperone-mediated autophagy (CMA):
-this pathway proceeds the selective targeting of proteins containing a KFERQ-like peptide motif to lysosomes for degradation ( Majeski, A.E etal Mechanisms of chaperone-mediated autophagy. Int. J. Biochem. Cell Biol 2004 .36, 2435–2444.)
- CMA are activated during short-term and prolonged starvation in organisms,
-CMA is inhibited by the protein synthesis inhibitors such as anisomycin and cycloheximide, partially inhibited when the p38 mitogen activated protein kinase is blocked,
- CMA activators are glucose-6-phophate dehydrogenase inhibitor, 6-aminonicotinamide, heat shock protein of 90 kilodaltons inhibitor, geldanamycin,
-CMA declines with age because of a decrease in the levels of lysosome-associated membrane protein (LAMP) type 2A, a lysosomal receptor for this pathway,
-CMA also induced cross-talk among different forms of autophagy,
microautophagy:
- this pathway involves the pinocytosis of small quantities of cytosol directly by lysosomes Muller, O etal Autophagic tubes: vacuolar invaginations involved in lateral membrane sorting and inverse vesicle budding. J. Cell Biol. 2000.151, 519–528.
xenophagy:
-when the autophagy machinery is also used to degrade foreign microbial invader
Crinophagy, pexophagy??
> add a picture as well as the abreviations----)
Recent studies have shown that a mammalian autophagy gene, beclin 1, is monoallelically deleted in 40–75% of breast tumors and can inhibit tumorigenesis when overexpressed in transformed cells (1). Other studies in Lamp2-deficient mice have also highlighted defective autophagy as a possible cause of cardiomyopathy and myopathy of Danon disease (2,3).
1. Liang XH, Jackson S, Seaman M, Brown K, Kempkes B, Hibshoosh H, Levine B. Induction of autophagy and inhibition of tumorigenesis
by beclin 1. Nature 1999;402:672–676.
2. Nishino I, Fu J, Tanji K, Yamada T, Shimojo S, Koori T, Mora M, Riggs JE, Oh SJ, Koga Y, Sue CM, Yamamoto A, Murakami N, Shanske S, Byrne E et al. Primary LAMP-2 deficiency causes X-linked vacuolar cardiomyopathy and myopathy (Danon disease). Nature 2000;
406:906–910.
3. Tanaka Y, Guhde G, Suter A, Eskelinen EL, Hartmann D, Lüllmann- Rauch R, Janssen PML, Blanz J, von Figura K, Saftig P. Accumulation
of autophagic vacuoles and cardiomyopathy in LAMP-2-deficient mice. Nature 2000;406:902–906.
Further evidence for the importance of autophagy in protecting against nutritional stress comes from studies where tumor cells aredeprived of growth/survival factors, leading to an increase in autophagy that prevents the cells from dying. Moreover, when autophagy is prevented under these conditions, the cells undergo apoptosis. Thus, when tumor cells are starved, autophagy stops them from dying by inhibiting apoptosis. In a tumor, this may mean that autophagy keeps tumor cells alive when limited angiogenesis leads to nutrient deprivation and hypoxia; therefore, we would expect that increased autophagy would promote the growth of solid tumors, whereas reduced autophagy might provide a useful way to limit tumor growth (1 and 2).
1. Boya P, Gonzalez-Polo RA, Casares N, et al. Inhibition of macroautophagy triggers apoptosis. Mol Cell Biol 2005;25:1025–40.
2. Lum JJ, Bauer DE, Kong M, et al. Growth factor regulation of autophagy and cell survival in the absence of apoptosis. Cell 2005;120:237–48.
Autophagy proceeds by nonselective uptake of cytoplasmic constituents into membrane-bound vesicles, termed autophagosomes, which are formed by a double- or multiple-membraned cisterna that wraps around and encloses a portion of the cytoplasm. Once formed, the autophagosomes mature rapidly by fusion with endo/lysosomes and acquire the proton pumps and lysosomal enzymes needed for degradation of the enclosed material.(1) (1. Kisen GO, Tessitore L, Costelli P, Gordon PB, Schwarze PE, Baccino FM, Seglen PO. Reduced autophagic activity in primary rat hepatocellular carcinoma and ascites hepatoma cells. Carcinogenesis 1993;14:2501–2505.)
The formation of the double-membraned vesicle is a complex process involving 16autophag y-related proteins (1). Two ubiquitin-like conjugation systems are involved in
autophagy. These systems produce modified complexes of autophagy regulators (Atg8-PE and Atg5-Atg12-Atg16) that may determine the formation and size of the autophagosome. Nucleation, expansion, uncoating, and completion of the autophagosome then occurs, priming it to fuse with lysosomes (1). The initiating signal for autophagosome formation is poorly understood, but the mammalian target of rapamycin (mTOR) is a negative regulator, and the extent of autophagy is regulated by proteins upstream of mTOR signaling, including PTEN, PDK1, Akt, and TSC1/2 (2). For example, PTEN and TSC1/2 positively regulate autophagy, whereas Akt inhibits it. Downstream targets of mTOR, including elongation factor-2 kinase (3) and S6kinase (2), have been shown to regulate autophagy.
1. Yorimitsu T, Klionsky DJ. Autophagy: molecular machinery for self-eating. Cell Death Differ 2005;12 Suppl 2:1542–52.
2. Codogno P, Meijer AJ. Autophagy and signaling: their role in cell survival and cell death. Cell Death Differ 2005;12 Suppl 2:1509–18.
3. Wu H, Yang JM, Jin S, Zhang H, Hait WN. Elongation factor-2 kinase regulates autophagy in human glioblastoma cells. Cancer Res 2006;66:3015–23.
Behind to this potential cancer-promoting effect of autophagy, numerous lines of evidence indicate an anticancer role for autophagy. The autophagy gene Beclin 1 (the mammalian counterpart of the yeast Atg 6 gene), which is part of a type III phosphatidylinositol 3-kinase complex required for autophagic vesicle formation, is a haploinsufficient tumor suppressor in mice (1,2) and is monoallelically lost in human breast, ovarian, and other tumors (3). Moreover, p53 and PTEN, two of the most commonly mutated tumor suppressor genes, both induce autophagy (4,5). Conversely, the oncogenic protein Bcl-2 directly interacts with Beclin 1 to inhibit autophagy (6). Because oncogenes can inhibit autophagy and tumor suppressors induce autophagy whereas a bona fide autophagy regulator is itself a tumor suppressor, these data suggest that autophagy serves an anticancer role.
1. Yue Z, Jin S, Yang C, Levine AJ, Heintz N. Beclin 1, an autophagy gene essential for early embryonic development, is a haploinsufficient tumor suppressor. Proc Natl Acad Sci U S A 2003;100:15077–82.
2. Qu X, Yu J, Bhagat G, et al. Promotion of tumorigenesis by heterozygous disruption of the beclin 1 autophagy gene. J Clin Invest 2003;112:1809–20.
3. Kondo Y, Kanzawa T, Sawaya R, Kondo S. The role of autophagy in cancer development and response to therapy. Nat Rev Cancer 2005;5:726–34.
4. Feng Z, Zhang H, Levine AJ, Jin S. The coordinate regulation of the p53 and mTOR pathways in cells. Proc Natl Acad Sci U S A 2005;102:8204–9.
5. Arico S, Petiot A, Bauvy C, et al. The tumor suppressor PTEN positively regulates macroautophagy by inhibiting the phosphatidylinositol 3-kinase/protein kinase B pathway. J Biol Chem 2001;276:35243–6.
6. Pattingre S, Tassa A, Qu X, et al. Bcl-2 antiapoptotic proteins inhibit beclin 1-dependent autophagy. Cell 2005;122:927–39.
Alternatively, autophagy may kill developing tumor cells. In support of this idea, a cell death pathway that involves both autophagy and apoptosis is selectively inactivated when primary epithelial cells become immortal (14), and in model systems of mammary acini formation, both apoptosis and autophagy are involved in the removal of epithelial cells to form luminal structures (2), suggesting that autophagy prevents early steps in epithelial tumor development. Taken together, these data suggest that autophagy can both stimulate and prevent cancer
depending on the context.
1. Thorburn J, Moore F, Rao A, et al. Selective inactivation of a FADD-dependent apoptosis and autophagy pathway in immortal epithelial cells. Mol
Biol Cell 2005;16:1189–99.
2. Mills KR, Reginato M, Debnath J, Queenan B, Brugge JS. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is required for induction of autophagy during lumen formation in vitro . Proc Natl Acad Sci U S A 2004;101:3438–43.
->--add some more descriptions, and integrate the picture here asw ell as the abreviations--)
Autophagy:
Autophagy or ‘‘self-eating’’ catabolism of cytoplasm, is a genetically programmed process that degrades long-lived cellular proteins and organelles and during nutrient deprivation, provides a supply of amino acids needed for cell survival. (Klionsky DJ, Emr SD. Autophagy as a regulated pathway of cellular degradation. Science 2000;290:1717–1721). Autophagy is important in normal development and response to changing environmental stimuli and autophagy has been found to be important in growth control and is defective in some tumor cells, bacterial and viral infections, neurodegenerative disorders, and cardiovascular disease (Shintani T, Klionsky DJ. Autophagy in health and disease: a double-edged sword. Science 2004;306:990–5.). The turnover of cytoplasmic constituents for energy is regulated by the endosomal/autophagic/lysosomal (EAL) system, complementing the degradation of most short-lived proteins by the ubiquitin-proteasome system. (Barry Boland etal Neuronal macroautophagy: From development to degeneration Molecular Aspects of Medicine 27 (2006) 503–519).
Though autophagy generally considered as a ‘self-eating’ process, this process occur by at least minimum 3 different routes:
Types of autophagy:
Macroautophagy(MA):
-this pathway proceeds the sequestration of cytosolic regions containing proteins, sugars, lipids, RNA, as well as organelles such as mitochondria, perixosomes into double-membraned vacuoles
that deliver their contents to late endosomal and lysosomal compartments for degradation with the help of the Apg12-Apg5 conjugate( Shintani T etal Autophagy in health and disease: a double-edged sword. Science 2004 306,990–995)
- this pathway activated during conditions of serum withdrawal in cell culture ,
-inhibitors for MA is 3-methyladenine, wortmannin, and LY294002
-protein degradation by MA is sensitive to microtubule inhibitors such as colcemide and vinblastine,
- Activators of MA is rapamycin,
chaperone-mediated autophagy (CMA):
-this pathway proceeds the selective targeting of proteins containing a KFERQ-like peptide motif to lysosomes for degradation ( Majeski, A.E etal Mechanisms of chaperone-mediated autophagy. Int. J. Biochem. Cell Biol 2004 .36, 2435–2444.)
- CMA are activated during short-term and prolonged starvation in organisms,
-CMA is inhibited by the protein synthesis inhibitors such as anisomycin and cycloheximide, partially inhibited when the p38 mitogen activated protein kinase is blocked,
- CMA activators are glucose-6-phophate dehydrogenase inhibitor, 6-aminonicotinamide, heat shock protein of 90 kilodaltons inhibitor, geldanamycin,
-CMA declines with age because of a decrease in the levels of lysosome-associated membrane protein (LAMP) type 2A, a lysosomal receptor for this pathway,
-CMA also induced cross-talk among different forms of autophagy,
microautophagy:
- this pathway involves the pinocytosis of small quantities of cytosol directly by lysosomes Muller, O etal Autophagic tubes: vacuolar invaginations involved in lateral membrane sorting and inverse vesicle budding. J. Cell Biol. 2000.151, 519–528.
xenophagy:
-when the autophagy machinery is also used to degrade foreign microbial invader
Crinophagy, pexophagy??
> add a picture as well as the abreviations----)
Recent studies have shown that a mammalian autophagy gene, beclin 1, is monoallelically deleted in 40–75% of breast tumors and can inhibit tumorigenesis when overexpressed in transformed cells (1). Other studies in Lamp2-deficient mice have also highlighted defective autophagy as a possible cause of cardiomyopathy and myopathy of Danon disease (2,3).
1. Liang XH, Jackson S, Seaman M, Brown K, Kempkes B, Hibshoosh H, Levine B. Induction of autophagy and inhibition of tumorigenesis
by beclin 1. Nature 1999;402:672–676.
2. Nishino I, Fu J, Tanji K, Yamada T, Shimojo S, Koori T, Mora M, Riggs JE, Oh SJ, Koga Y, Sue CM, Yamamoto A, Murakami N, Shanske S, Byrne E et al. Primary LAMP-2 deficiency causes X-linked vacuolar cardiomyopathy and myopathy (Danon disease). Nature 2000;
406:906–910.
3. Tanaka Y, Guhde G, Suter A, Eskelinen EL, Hartmann D, Lüllmann- Rauch R, Janssen PML, Blanz J, von Figura K, Saftig P. Accumulation
of autophagic vacuoles and cardiomyopathy in LAMP-2-deficient mice. Nature 2000;406:902–906.
Further evidence for the importance of autophagy in protecting against nutritional stress comes from studies where tumor cells aredeprived of growth/survival factors, leading to an increase in autophagy that prevents the cells from dying. Moreover, when autophagy is prevented under these conditions, the cells undergo apoptosis. Thus, when tumor cells are starved, autophagy stops them from dying by inhibiting apoptosis. In a tumor, this may mean that autophagy keeps tumor cells alive when limited angiogenesis leads to nutrient deprivation and hypoxia; therefore, we would expect that increased autophagy would promote the growth of solid tumors, whereas reduced autophagy might provide a useful way to limit tumor growth (1 and 2).
1. Boya P, Gonzalez-Polo RA, Casares N, et al. Inhibition of macroautophagy triggers apoptosis. Mol Cell Biol 2005;25:1025–40.
2. Lum JJ, Bauer DE, Kong M, et al. Growth factor regulation of autophagy and cell survival in the absence of apoptosis. Cell 2005;120:237–48.
Autophagy proceeds by nonselective uptake of cytoplasmic constituents into membrane-bound vesicles, termed autophagosomes, which are formed by a double- or multiple-membraned cisterna that wraps around and encloses a portion of the cytoplasm. Once formed, the autophagosomes mature rapidly by fusion with endo/lysosomes and acquire the proton pumps and lysosomal enzymes needed for degradation of the enclosed material.(1) (1. Kisen GO, Tessitore L, Costelli P, Gordon PB, Schwarze PE, Baccino FM, Seglen PO. Reduced autophagic activity in primary rat hepatocellular carcinoma and ascites hepatoma cells. Carcinogenesis 1993;14:2501–2505.)
The formation of the double-membraned vesicle is a complex process involving 16autophag y-related proteins (1). Two ubiquitin-like conjugation systems are involved in
autophagy. These systems produce modified complexes of autophagy regulators (Atg8-PE and Atg5-Atg12-Atg16) that may determine the formation and size of the autophagosome. Nucleation, expansion, uncoating, and completion of the autophagosome then occurs, priming it to fuse with lysosomes (1). The initiating signal for autophagosome formation is poorly understood, but the mammalian target of rapamycin (mTOR) is a negative regulator, and the extent of autophagy is regulated by proteins upstream of mTOR signaling, including PTEN, PDK1, Akt, and TSC1/2 (2). For example, PTEN and TSC1/2 positively regulate autophagy, whereas Akt inhibits it. Downstream targets of mTOR, including elongation factor-2 kinase (3) and S6kinase (2), have been shown to regulate autophagy.
1. Yorimitsu T, Klionsky DJ. Autophagy: molecular machinery for self-eating. Cell Death Differ 2005;12 Suppl 2:1542–52.
2. Codogno P, Meijer AJ. Autophagy and signaling: their role in cell survival and cell death. Cell Death Differ 2005;12 Suppl 2:1509–18.
3. Wu H, Yang JM, Jin S, Zhang H, Hait WN. Elongation factor-2 kinase regulates autophagy in human glioblastoma cells. Cancer Res 2006;66:3015–23.
Behind to this potential cancer-promoting effect of autophagy, numerous lines of evidence indicate an anticancer role for autophagy. The autophagy gene Beclin 1 (the mammalian counterpart of the yeast Atg 6 gene), which is part of a type III phosphatidylinositol 3-kinase complex required for autophagic vesicle formation, is a haploinsufficient tumor suppressor in mice (1,2) and is monoallelically lost in human breast, ovarian, and other tumors (3). Moreover, p53 and PTEN, two of the most commonly mutated tumor suppressor genes, both induce autophagy (4,5). Conversely, the oncogenic protein Bcl-2 directly interacts with Beclin 1 to inhibit autophagy (6). Because oncogenes can inhibit autophagy and tumor suppressors induce autophagy whereas a bona fide autophagy regulator is itself a tumor suppressor, these data suggest that autophagy serves an anticancer role.
1. Yue Z, Jin S, Yang C, Levine AJ, Heintz N. Beclin 1, an autophagy gene essential for early embryonic development, is a haploinsufficient tumor suppressor. Proc Natl Acad Sci U S A 2003;100:15077–82.
2. Qu X, Yu J, Bhagat G, et al. Promotion of tumorigenesis by heterozygous disruption of the beclin 1 autophagy gene. J Clin Invest 2003;112:1809–20.
3. Kondo Y, Kanzawa T, Sawaya R, Kondo S. The role of autophagy in cancer development and response to therapy. Nat Rev Cancer 2005;5:726–34.
4. Feng Z, Zhang H, Levine AJ, Jin S. The coordinate regulation of the p53 and mTOR pathways in cells. Proc Natl Acad Sci U S A 2005;102:8204–9.
5. Arico S, Petiot A, Bauvy C, et al. The tumor suppressor PTEN positively regulates macroautophagy by inhibiting the phosphatidylinositol 3-kinase/protein kinase B pathway. J Biol Chem 2001;276:35243–6.
6. Pattingre S, Tassa A, Qu X, et al. Bcl-2 antiapoptotic proteins inhibit beclin 1-dependent autophagy. Cell 2005;122:927–39.
Alternatively, autophagy may kill developing tumor cells. In support of this idea, a cell death pathway that involves both autophagy and apoptosis is selectively inactivated when primary epithelial cells become immortal (14), and in model systems of mammary acini formation, both apoptosis and autophagy are involved in the removal of epithelial cells to form luminal structures (2), suggesting that autophagy prevents early steps in epithelial tumor development. Taken together, these data suggest that autophagy can both stimulate and prevent cancer
depending on the context.
1. Thorburn J, Moore F, Rao A, et al. Selective inactivation of a FADD-dependent apoptosis and autophagy pathway in immortal epithelial cells. Mol
Biol Cell 2005;16:1189–99.
2. Mills KR, Reginato M, Debnath J, Queenan B, Brugge JS. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is required for induction of autophagy during lumen formation in vitro . Proc Natl Acad Sci U S A 2004;101:3438–43.
Autophagy in developmental cell death:
the first wave of PCD occurs during cavitation of the early embryo; the solid embryonic ectoderm undergoes apoptosis to form the proamniotic cavity (1). Autophagy is most pronounced in developmental processes that involve massive cell elimination, such as embryogenesis, insect metamorphosis, glandular atresia,and lumen or cavitation formation (2 ).
In an invitro study by culturing mouse embryonic stem cells (ES), Xueping Qu group were the first one demostrated the novel role of autophagy, which removes the cells that die during normal embryonic development. In their study they have found that the ES cells form undifferentiated cell aggregates that develop into simple embryoid bodies (EBs), containing an outer layer of endodermal cells and an inner solid core of ectodermal cells. The inner ectodermal cells of the simple EBs undergo PCD to form cystic EBs. ES cells lacking either atg5 or beclin 1 are defective in autophagosome formation (3).
Further their study proved that the two different autophagy genes, atg5 (autophagy 5 ) and beclin 1, are required for the clearance of dead cells during EB cavitation. In the absence of autophagy genes, the cells didn’t expose phosphatidylserine and secreted low levels of lysophosphatidylcholine and as a result the amount of cell death in the inner ectodermal cells fail to be removed due to absent engulfment by phagocytic cells and consequently, cavitation is impaired. Thus, autophagy genes are required for the phagocytic removal of corpses during PCD both in the in vitro EB model as well as in vivo during mouse development. This study shows that autophagy-induced signals are essential for normal development.
Furthermore they have also found that the cells of the autophagy-deficient mouse embryos had low levels of ATP, a vital energy source for many cellular functions. Autophagy is known to generate amino and fatty acids utilized in ATP production. Treatment with an alternative fuel, methylpyruvate, restored normal levels of ATP in autophagy-deficient mouse embryonic bodies and bypassed the bodies’ failure to prompt signals needed for the healthy cells to engulf the dead ones.
1 Coucouvanis, E., and Martin, G.R. (1995). Signals for death and survival: a two-step mechanism for cavitation in the vertebrate embryo. Cell 83, 279–287.
2) Xueping Qu, Etal Autophagy Gene-Dependent Clearance of Apoptotic Cells during Embryonic Development Cell 128, 931–946, March 9, 2007
3) Mizushima, N., Yamamoto, A., Hatano, M., Kobayashi, Y., Kabeya, Y., Suzuki, K., Tokuhisa, T., Ohsumi, Y., and Yoshimori, T. (2001). Dissection of autophagosome formation using Apg5-deficient mouse embryonic stem cells. J. Cell Biol. 152, 657–668.
Within this energy derivation, interestingly Yang Y etal reported that the neurite degeneration was associated with a decrease in neuritic ATP levels and was caused by energy failure, because an exogenous supply of nicotinamide adenine dinucleotide (NAD) or its precursor nicotinamide suppressed the degeneration by delaying axonal ATP reduction caused by Zn2+ depletion. As a result, the cellular Zn2+ depletion induces a "dying-back" degeneration characterized by an NAD- and autophagy-dependent process, independently of neurite elongation dynamics(Yang Y etal Cellular Zn2+ chelators cause "dying-back" neurite degeneration associated with energy impairment. Neurosci Res. 2007 Oct;85(13):2844-55).Interestingly Yue etal also dedmostrated that beclin 1, a critical component of mammalian autophagy, is a haploinsufficient tumor suppressor gene. Their study supported the role of autophagy in tumor suppression function and it’s suggest that mutations in other genes operating in this pathway may contribute to tumor formation through deregulation of autophagy.(Yue Z, Beclin 1, an autophagy gene essential for early embryonic development, is a haploinsufficient tumor suppressor. Proc Natl Acad Sci U S A. 2003 Dec 9;100(25):15077-82.)
Apart from the Beclin1, a principal regulator in autophagosome formation, deficiency results in early embryonic lethality (Xueping Qu (2 refer above ref*), Very recently Fimia etal demonstrated an another novel protein Ambra1 (activating molecule in Beclin1-regulated autophagy), a large, previously unknown protein bearing a WD40 domain at its amino terminus, regulates autophagy and has a crucial role in embryogenesis. By using RNA interference experiments in vitro, they found that Ambra1 is a positive regulator of the Becn1-dependent programme of autophagy. Further studies proved that the Ambra1 functional deficiency in mouse embryos leads to various developmental diseases such as severe neural tube defects associated with autophagy impairment, accumulation of ubiquitinated proteins, unbalanced cell proliferation and excessive apoptotic cell death. By proving this novel role and the complex system , they also proved in vivo evidence that there is an existence of a complex interplay between autophagy, cell growth and cell death, which is required for neural development in mammals(Fimia GM etal Ambra1 regulates autophagy and development of the nervous system.Nature. 2007 Jun 28;447(7148):1121-5. Epub 2007 Jun 24).
In connection to this, the deficiency of upstream regulators of the mitochondrial death pathway has been recently shown to trigger in vitro a cellular process of self-clearance with features of autophagy. Within the context, Moreno S group demostrated that when there is an impairement on the apotosome during cellular development, then there would be an activation of an autophagy program in cerebral cortex, which might activated by a depletion in growth factors in the cells' microenvironment. By utilising the neural precursor cell line as a model system, they proved the brain development. It would be ineterstign if the system develops further any inflammation and any neurodegenration diseases or diseases which might be associated with the neural cancer stem cells(Moreno S etal Apoptosome impairment during development results in activation of an autophagy program in cerebral cortex. Apoptosis. 2006 Sep;11(9):1595-602)
In extending the microenvironment role, Cárdenas-Aguayo Mdel C etal indentified that growth factor deprivation induces an alternative non-apoptotic, autophagy-like death mechanism that is inhibited by the Bcl2 in cells derived from neural precursor cells. By utilsign the mesencephalic neural-progenitor-cell cultures the authors found that the anti-apoptotic Bcl2 family members are key molecules controlling death activation which independent to the cell degeneration mechanism???(Cárdenas-Aguayo Mdel C etal Growth factor deprivation induces an alternative non-apoptotic death mechanism that is inhibited by Bcl2 in cells derived from neural precursor cells. J Hematother Stem Cell Res. 2003 Dec;12(6):735-48)
Adult stem cells and autophagy:
Neural:
While transiting from the embryonic state into adult stem cells stage, the area which have studied most is the neural system.
Jiang H group from the University of Texas M. D. Anderson Cancer Center, have focused first time on the therapeutic side by utilising the adenovirus system as a mediator to induce the autophagic cell death in cancer stem-cells.
While transiting from the embryonic state into adult stem cells stage, the area which have studied most is the neural system.
Jiang H group from the University of Texas M. D. Anderson Cancer Center, have focused first time on the therapeutic side by utilising the adenovirus system as a mediator to induce the autophagic cell death in cancer stem-cells.
Having enhanced tropism to glioma cells and selective replication in cancer cells with an abnormal Rb pathway, ability to completely overcome the molecular machinery of the infected cell, prevent cancer stem cells from developing resistance to other forms of therapy they pursued the Delta- 24-RGD, an oncolytic adenovirus as an therapeutic agent, targeting to the abnormal p16INK4/Rb pathway in brain tumor stem cells that are resistant to radiation and chemotherapy.
In this study, they isolated neurosphere forming potential brain tumor stem cell w ith marker of CD133, from four fresh surgical specimens of glioblastoma multiforme, which exhibited the in vitro stem cell characteristics of extensive self-renewal (for more than fi ve passages in culture), ability to differentiate to neurons and astrocytes, expressed high levels of adenoviral receptors and allowed for efficient viral infection, replication, and oncolysis in an Rb-dependent manner.
The four cell lines were efficiently infected with an oncolytic adenovirus Delta-24- RGD, which induced autophagic cell death as indicated by accumulation of Atg5 and LC3-II protein and autophagic vacuoles. Further analyses of treated tumors showed that Atg5 expression colocalized with viral fiber protein and delineated a wave front of autophagic cells that circumscribed areas of virally induced necrosis.
Moreover, transplanted into the basal ganglia of immunodeficient mice carrying xenograft tumors that were derived from one of the cell lines survived longer after treatment with Delta-24-RGD than with an inactivated form of the virus.
By demonstrating this novel therapeutic approach, they have also proved that the brain tumor stem cells are susceptible to adenovirus-mediated cell death via autophagy in vitro and in vivo.
However, as the paper indicated, it is unclear how similar the cell lines developed in this study are to the brain tumor stem cells that exist in human brain cancer or whether the oncolytic adenovirus developed in this study would be efficacious and safe in humans.
Jiang H etal Examination of the therapeutic potential of Delta-24-RGD in brain tumor stem cells: role of autophagic cell death J Natl Cancer Inst. 2007 Sep 19;99(18):1410-4.
Interestingly, Ito H etal also developed an another novel cancer therapy approach by using the conditionally replicating adenoviruses (CRAds) which engineered to be replicate selectively in cancer cells and cause cancer-specific cell lysis. In this study the authors developed four different cell lines initially, infected by the conditionally replicating adenoviruses (CRAds) system which regulated by the human telomerase reverse transcriptase promoter (hTERT-Ad) or control nonreplicating adenoviruses (Ad-GFP). The author found that the hTERT-Ad induced tumor-specific autophagic cell death in tumor cells and in subcutaneous gliomas as well as the hTERT-Ad may kill telomerase-positive cancer cells by inducing autophagic cell death by suppressing the mTOR signaling pathway. (????? Relationship??) (Ito H etal Autophagic cell death of malignant glioma cells induced by a conditionally replicating adenovirus. J Natl Cancer Inst. 2006 May 3;98(9):625-36)
Osteo :
In this study, they isolated neurosphere forming potential brain tumor stem cell w ith marker of CD133, from four fresh surgical specimens of glioblastoma multiforme, which exhibited the in vitro stem cell characteristics of extensive self-renewal (for more than fi ve passages in culture), ability to differentiate to neurons and astrocytes, expressed high levels of adenoviral receptors and allowed for efficient viral infection, replication, and oncolysis in an Rb-dependent manner.
The four cell lines were efficiently infected with an oncolytic adenovirus Delta-24- RGD, which induced autophagic cell death as indicated by accumulation of Atg5 and LC3-II protein and autophagic vacuoles. Further analyses of treated tumors showed that Atg5 expression colocalized with viral fiber protein and delineated a wave front of autophagic cells that circumscribed areas of virally induced necrosis.
Moreover, transplanted into the basal ganglia of immunodeficient mice carrying xenograft tumors that were derived from one of the cell lines survived longer after treatment with Delta-24-RGD than with an inactivated form of the virus.
By demonstrating this novel therapeutic approach, they have also proved that the brain tumor stem cells are susceptible to adenovirus-mediated cell death via autophagy in vitro and in vivo.
However, as the paper indicated, it is unclear how similar the cell lines developed in this study are to the brain tumor stem cells that exist in human brain cancer or whether the oncolytic adenovirus developed in this study would be efficacious and safe in humans.
Jiang H etal Examination of the therapeutic potential of Delta-24-RGD in brain tumor stem cells: role of autophagic cell death J Natl Cancer Inst. 2007 Sep 19;99(18):1410-4.
Interestingly, Ito H etal also developed an another novel cancer therapy approach by using the conditionally replicating adenoviruses (CRAds) which engineered to be replicate selectively in cancer cells and cause cancer-specific cell lysis. In this study the authors developed four different cell lines initially, infected by the conditionally replicating adenoviruses (CRAds) system which regulated by the human telomerase reverse transcriptase promoter (hTERT-Ad) or control nonreplicating adenoviruses (Ad-GFP). The author found that the hTERT-Ad induced tumor-specific autophagic cell death in tumor cells and in subcutaneous gliomas as well as the hTERT-Ad may kill telomerase-positive cancer cells by inducing autophagic cell death by suppressing the mTOR signaling pathway. (????? Relationship??) (Ito H etal Autophagic cell death of malignant glioma cells induced by a conditionally replicating adenovirus. J Natl Cancer Inst. 2006 May 3;98(9):625-36)
Osteo :
Chondrocytes are the only cells found in cartilage, which produce and maintain the cartilaginous matrix, which consists mainly of collagen and proteoglycans. During the lineage, the chondrocytic undergoes ulmost 4 types of difefrentiation process namely, Colony-forming unit-fibroblast (CFU-F), Mesenchymal stem cell / marrow stromal cell (MSC), Chondrocyte anf finally Hypertrophic chondrocyte.
Recently it has been demonstrated that the mesenchymal stem cells ( or multipotent adult progenitor cells ) are commonly known as osteochondrogenic (or osteogenic, chondrogenic, osteoprogenitor, etc.) cells has shown the ability to differentiate into chondrocytes or osteoblasts, depending on the medium. In vivo, differentiation of an MSC in a vascularized area (such as bone) yields an osteoblast, whereas differentiation of an MSC in a non-vascularized area (such as cartilage) yields a chondrocyte. Chondrocytes undergo terminal differentiation when they become hypertrophic during endochondral ossification. This last stage is characterized by major phenotypic changes in the cell (1,2,3)
-Yuehua Jiang etal Pluripotency of mesenchymal stem cells derived from adult marrow Nature 418, 41-49 (4 July 2002)
-Dominici M etal Bone marrow mesenchymal cells: biological properties and clinical applications. J Biol Regul Homeost Agents 2001 Jan-Mar;15(1):28-37
-Bianco P etal Bone marrow stromal stem cells: nature, biology, and potential applications Stem Cells 19 (3): (2001). 180-92.
-Yuehua Jiang etal Pluripotency of mesenchymal stem cells derived from adult marrow Nature 418, 41-49 (4 July 2002)
-Dominici M etal Bone marrow mesenchymal cells: biological properties and clinical applications. J Biol Regul Homeost Agents 2001 Jan-Mar;15(1):28-37
-Bianco P etal Bone marrow stromal stem cells: nature, biology, and potential applications Stem Cells 19 (3): (2001). 180-92.
Recently HEGERT Claudia etal has demonstrated that the embryonic stem cells differentiate into chondrocytes, which progressively develop into hypertrophic and calcifying cells. At a terminal differentiation stage, cells expressing an osteoblast-like phenotype appeared either by transdifferentiation from hypertrophic chondrocytes or directly from osteoblast precursor cells. They also showed that the chondrocytic cells are able to transdifferentiate into other mesenchymal cells such as osteogenic and adipogenic cell types.(HEGERT Claudia Differentiation plasticity of chondrocytes derived from mouse embryonic stem cells Journal of Cell Science 2002, vol. 115 (23), pp. 4617-4628).
Bone growth takes place through the activities of chondrocytes embedded in a cartilaginous structure called the epiphyseal growth plate. Growth-plate chondrocytes differentiate from resting cells via proliferating to hypertrophic cells, and eventually deleted from the cartilage through the induction of apoptosis when the potent chondrocyte apoptogens stimulated the solubilization of mineral and hydrolysis of organic matrix constituents by septoclasts generates high local concentrations of ions, peptides, and glycans, and secreted matrix metalloproteins (Shapiro IM Fate of the hypertrophic chondrocyte: microenvironmental perspectives on apoptosis and survival in the epiphyseal growth plate Birth Defects Res C Embryo Today. 2005 Dec;75(4):330-9). This differentiation process is controlled by a variety of growth factors. (René C.C. van Donkelaar, UNLOADING IS ESSENTIAL FOR GROWTH PLATE DEVELOPMENT 2005 Summer Bioengineering Conference, June 22-26, Vail Cascade Resort & Spa, Vail, Colorado).
Since the life history of the growth plate chondrocyte is very short (1–2 days) even minor disturbances in the metabolic state can result in gross impairment of growth. In addition, the authors contend that the induction of the autophagic response permits the terminally differentiated cells to survive the brief rigors of the harsh local microenvironment (Srinivas V etal Chondrocytes embedded in the epiphyseal growth plates of long bones undergo autophagy prior to the induction of osteogenesis Autophagy. 2006 Jul-Sep;2(3):215-6). Furthermore, when the autophagy was suppressed, chondrocytes promoted caspase-8 activation and activated BID which ruether regulate the apoptosis (Bohensky J HIF-1 regulation of chondrocyte apoptosis: induction of the autophagic pathway. Autophagy. 2007 May-Jun;3(3):207-14)
Since the life history of the growth plate chondrocyte is very short (1–2 days) even minor disturbances in the metabolic state can result in gross impairment of growth. In addition, the authors contend that the induction of the autophagic response permits the terminally differentiated cells to survive the brief rigors of the harsh local microenvironment (Srinivas V etal Chondrocytes embedded in the epiphyseal growth plates of long bones undergo autophagy prior to the induction of osteogenesis Autophagy. 2006 Jul-Sep;2(3):215-6). Furthermore, when the autophagy was suppressed, chondrocytes promoted caspase-8 activation and activated BID which ruether regulate the apoptosis (Bohensky J HIF-1 regulation of chondrocyte apoptosis: induction of the autophagic pathway. Autophagy. 2007 May-Jun;3(3):207-14)
At the microenvironment and it’s association level, Shapiro group also proposed a novel concept that the hypertrophic cells die through the induction of autophagy. In the cartilage microenvironment, combinations of local factors cause chondrocytes to express an initial survival phenotype and oxidize their own structural macromolecules to generate ATP. While delaying death, autophagy leads to a state in which cells are further sensitized to changes in the local microenvironment. One such change is similar to ischemia reperfusion injury, a condition that leads to tissue damage and cell death. In the growth cartilage, an immediate effect of this type of injury is sensitization to local apoptogens. These two concepts (type II programmed cell death and ischemia reperfusion injury) emphasize the importance of the local microenvironment, in particular pO(2), in directing chondrocyte survival and apoptosis(Shapiro IM Fate of the hypertrophic chondrocyte: microenvironmental perspectives on apoptosis and survival in the epiphyseal growth plate Birth Defects Res C Embryo Today. 2005 Dec;75(4):330-9).
Further more they have also demonstrated the involement of PIM-2 (a serine/threonine protein kinases) in the growth plate modulated the activity of a key regulator of apoptosis, BAD. Since BAD inhibition and Beclin-1 expression activated autophagy, it is likely that induction of the autophagic pathway would serve to inhibit apoptosis and preserve the life of the terminally differentiated chondrocyte. As a result the authors concluded that the PIM-2 regulates a new intermediate stage in the differentiation pathway, the induction of autophagy. It’s would also be more interesting if this PIM-2 have any role in the self-renewal of the chondrocyte (stem-cells) because recently Pim-1 and Pim-3 proved to play role on the self-renewal function in the murine model (Irène Aksoy etal Self-Renewal of Murine Embryonic Stem Cells Is Supported by the Serine/Threonine Kinases Pim-1 and Pim-3. Stem Cells (December 2007; Vol. 25, No. 12)) + (Bohensky J etal PIM-2 is an independent regulator of chondrocyte survival and autophagy in the epiphyseal growth plate. J Cell Physiol. 2007 Oct;213(1):246-51)
(+ J Kramer etal Ultrastructural analysis of mouse embryonic stem cell-derived chondrocytes Anatomy and Embryology 2005 Oct;210(3):175-85. Abstract Pluripotent embryonic stem (ES) cells cultivated as cellular aggregates, so called embryoid bodies (EBs), differentiate spontaneously into different cell types of all three germ layers in vitro resembling processes of cellular differentiation during embryonic development. Regarding chondrogenic differentiation, murine ES cells differentiate into progenitor cells, which form pre-cartilaginous condensations in the EB-outgrowths and express marker molecules characteristic for mesenchymal cell types such as Sox5 and Sox6. Later, mature chondrocytes appear which express collagen type II, and the collagen fibers show a typical morphology as demonstrated by electron-microscopical analysis. These mature chondrogenic cells are organized in cartilage nodules and produce large amounts of extracellular proteoglycans as revealed by staining with cupromeronic blue. Finally, cells organized in nodules express collagen type X, indicating the hypertrophic stage. In conclusion, differentiation of murine ES cells into chondrocytes proceeds from the undifferentiated stem cell via progenitor cells up to mature chondrogenic cells, which then undergo hypertrophy. Furthermore, because the ES-cell-derived chondrocytes did not express elastin, a marker for elastic cartilage tissue, we suggest the cartilage nodules to resemble hyaline cartilage tissue)
Mesenchymal:
Catherine Verfaillie group recently opened a new therapeutic avenue by developiong a novel mesenchymal stem cells also termed multipotent adult progenitor cells or MAPCs, which differentiate, at the single cell level, not only into mesenchymal cells, but also cells with visceral mesoderm, neuroectoderm and endoderm characteristics in vitro. Followed by in-vitro study when injected into an early blastocyst, single MAPCs contribute to most, if not all, somatic cell types. On transplantation into a non-irradiated host, MAPCs engraft and differentiate to the haematopoietic lineage, in addition to the epithelium of liver, lung and gut, proliferate extensively without obvious senescence or loss of differentiation potential(Yuehua Jiang etal Pluripotency of mesenchymal stem cells derived from adult marrow Nature 418, 41-49 (4 July 2002))
This underlines the importance of the population of mesenchymal stem cells (MSCs) with stem cell properties similar to embryonic stem (ES) cells. These cells can be cultured and expanded in vitro without losing their stem cell potential making them an attractive target for cell therapy. Finally, it is still not clear if stem cells for various tissues are present in peripheral blood, or bone marrow and thus can be directly purified from these sources.
Various reports are also proving the involment of mesenchymals stem-cells in the formation of cancer (Galiè M etal Mesenchymal stem cells share molecular signature with mesenchymal tumor cells and favor early tumor growth in syngeneic mice.Oncogene. 2007 Nov 12). In this series, Shima Y gropu recently developed a novel method which demonstrated morphological features of autophagy, could also serve as as a material with which to analyze the tumorigenic and differentiation-modifying effects of candidate oncogenes involved in the development of sarcomas (for details, refer the ‘methods’ table**)
** add further stem-cell systems
Methods:
A) Detection of Autophagy: (make it as table*)
I Morphological methods:
i)Electron microscopy:
-Immunoelectron microscopy using antibodies against autophagosomal marker (Mizushima, N. etal Dissection of autophagosome formation using Apg5-deficient mouse embryonic stem cells. Journal of Cell Biology, (2001). 152, 657–667)
-Quantification of autophagic activity by EM: the area or volume of autophagic vacuoles is
calculated and expressed as the ratio to the total cytoplasmic area or volume.
ii)Monodansylcadaverine (MDC) staining:
- a fluorescent compound, monodansylcadaverine (MDC) has been proposed as a tracer for autophagic
vacuoles (Biederbick, A. etal Monodansylcadaverine (MDC) is a specific in vivo marker for autophagic vacuoles. European Journal of Cell Biology, (1995).66, 3–14)
2. Biochemical methods
i) Bulk degradation of long-lived proteins:
-measurement of bulk degradation of long-lived proteins is often used to monitor
autophagic activity by incubating the cells with [14C] (or [3H])-valine or leucine to label all cellular
proteins. Since leucine is the most effective suppressor of autophagy , valine may be a preferred
tracer. Mortimore, G. E etal Intracellular protein catabolism and its control during nutrient deprivation and supply. Annual Review of Nutrition, (1987). 7, 539–564.
ii)Delivery of cytoplasmic componentsto lysosome:
- an ideal method to assess autophagic activity by measuring the delivery of cytosolic material to lysosomes.
- the autophagic activity can be determine by simple measurement of ALP activity using such genetically
modified strains.
- the cytosolic enzymes such as lactate dehydrogenase (LDH) were used as endogenous probes. The accumulation of these enzymes in sedimented vacuoles in the presence of lysosomal proteinase inhibitors was measured Seglen, P. O etal Autophagy and other vacuolar protein degradation mechanisms. Experientia, (1992). 48, 158– 172.
3. Specific markers for autophagy
i) GFP-LC3 localization:
LC3 associates with the isolation membrane in an Atg5-dependent manner and remains on the membrane even after
spherical autophagosomes are completely formed.These molecules are very good marker proteins for autophagic membranes. Atg12–Atg5 and Atg16L are specific markers for the isolation membrane, and LC3 is a general marker for autophagic membranes. Mizushima, N etal Dissection of autophagosome formation using Apg5-deficient mouse embryonic stem cells.
Journal of Cell Biology, (2001). 152, 657–667.
ii)Conversion of LC3-I to LC3-II:
- measurement of LC3-II by immunoblotting is a simple and quantitative method for determining the autophagic activity of mammalian cells.
B) Detection of cell-death in in-vivo: (make it as table*)
a) Recombinant luciferase reporter molecule: for the noninvasive monitoring of caspase-3 activity in living animals using bioluminescence-imaging techniques.
Applications/ Advantages: prove useful for rapid and dynamic screening aswell as validation of proapoptotic and antiapoptotic experimental therapeutic agents in animal models
b) Real-time imaging method: to visualize apoptotic membrane changes of single cardiomyocytes in the injured heart of the living mouse, by using fluorescent-labelled annexin V. Annexin V binds selectively to PS, a recognition signal on the surface of apoptotic
Applications/ Advantages: can be used in vivo as a noninvasive means to detect and serially image tissues and organs undergoing apoptosis. Moreover the annexin V-based method was also adapted for the measurement of tumour apoptosis in living animals , so this would makes further advantage to measure the antiproliferative effects of cancer chemotherapeutic regimens in patients.
c) Cytochrome c method: cytochrome c is release from the dying cell into the extracellular compartment was suggested to occur after induction of apoptosis, but not during necrosis, and elevated cytochrome c levels can be observed in the serum of patients with haematological malignancies.
Applications/ Advantages: Monitoring of serum levels of cytochrome c in cancer patients might, therefore, serve as a useful clinical marker reflecting therapyinduced
cell death in vivo
d) Circulating apoptotic microparticles: have been captured in the peripheral blood of individuals with HIV-1 infection
Applications/ Advantages: assessment of circulating microparticles was used to monitor apoptosis in patients with congestive heart failure
e) Radiolabelled annexin V: the most widely studied agent for the in vivo study of
apoptosis in patients
Applications/ Advantages:
i) Promising results have been obtained for cardiac disease patients, for instance. Heart transplant rejection is characterized pathologically by myocyte necrosis and apoptosis, and the recent administration of technetium-99m-labelled annexin V to cardiac allograft recipients has revealed the clinical feasibility and safety of annexin V imaging
for noninvasive detection of transplant rejection.
ii) Radiolabelled annexin V is now entering clinical trials for the assessment of therapeutic efficacy in cancer patients, the extent and severity of myocardial infarction, and the screening for acute rejection in heart transplant recipients, and could perhaps serve to obviate invasive biopsies in some cases
f) Magnetic resonance (MR) imaging: a promise methods to monitor apoptotic cell death. The detection of apoptotic cells by MR, using a targeted contrast agent (synaptotagmin) that binds to PS, was recently demonstrated in vivo, in mice bearing a tumour treated with chemotherapeutic drugs
Applications/ Advantages: recognition of cell death with relatively high spatial resolution, and further developments in this area are sure to provide exciting new opportunities for the noninvasive evaluation of the clinical condition of patients as well as the effects of various treatments.
C) Assays for Heamatopoietic Stem Cells: (make it as table*)
¨[Combine details from the previous posts] ++ add the very recent one- novel method for study of neural stem-cell niche by utilsign the HSC cells, which funtioning without adding any exogenous growth factors, etc +
Conclusions:
Abbreviations / explanations:
Abbreviations / explanations:
Totipotent:
- having unlimited capability. A totipotent cell has the capacity to form an entire organism,
- approximately four days after fertilization and after several cycles of cell division, these totipotent cells begin to specialize.
- totipotent specialize into pluripotent cells that can give rise to most of the tissues necessary for fetal development. Pluripotent cells undergo further specialization into multipotent cells that are committed to give rise to cells that have a particular function. For example, multipotent blood stem cells give rise to the red cells, white cells and platelets in the blood.
Pluripotent:
-the definition of pluripotency has come to refer to a stem cell that has the potential to differentiate into any of the three germ layers: endoderm (interior stomach lining, gastrointestinal tract, the lungs), mesoderm (muscle, bone, blood, urogenital), or ectoderm (epidermal tissues and nervous system). Pluripotent stem cells can give rise to any fetal or adult cell type. However, alone they cannot develop into a fetal or adult animal because they lack the potential to contribute to extraembryonic tissue, such as the placenta. In contrast, many progenitor cells are multipotent, i.e. they are capable of differentiating into a limited number of cell fates.
-the definition of pluripotency has come to refer to a stem cell that has the potential to differentiate into any of the three germ layers: endoderm (interior stomach lining, gastrointestinal tract, the lungs), mesoderm (muscle, bone, blood, urogenital), or ectoderm (epidermal tissues and nervous system). Pluripotent stem cells can give rise to any fetal or adult cell type. However, alone they cannot develop into a fetal or adult animal because they lack the potential to contribute to extraembryonic tissue, such as the placenta. In contrast, many progenitor cells are multipotent, i.e. they are capable of differentiating into a limited number of cell fates.
Multipotent progenitor cells:
-can give rise to several other cell types, but those types are limited in number. An example of a multipotent stem cell is a hematopoietic cell — a blood stem cell that can develop into several types of blood cells, but cannot develop into brain cells or other types of cells. At the end of the long series of cell divisions that form the embryo are cells that are terminally differentiated, or that are considered to be permanently committed to a specific function.
Embryonic stem cells:
-are stem cells derived from the inner cell mass of an early stage embryo known as a blastocyst. Human embryos reach the blastocyst stage 4-5 days post fertilization, at which time they consist of 50-150 cells.
-ES cells are pluripotent and can able to differentiate into all derivatives of the three primary germ layers: ectoderm, endoderm, and mesoderm.
Cord blood:
-Cord blood, which is also called "placental blood," is the blood that remains in the umbilical cord and placenta following birth and after the cord is cut. Cord blood is routinely discarded with the placenta and umbilical cord, which has valuable stem-cells.
Embryonal carcinoma (EC):
-cells are pluripotent stem cells derived from teratocarcinomas and are considered the malignant counterparts of human embryonic stem (ES) cells.
Transdifferentiation and redifferentiation: examples??
Cavitation:
-(embryoid bodies) cavitation or lumen formation, which derived from mouse embryonic stem cells??
Haploinsufficiency:
-"haploid" (one instead of two chromosomes or two genes) + "insufficiency." Haploinsufficiency is related to hemizygosity.
A situation in which the total level of a gene product (a particular protein) produced by the cell is about half of the normal level and that is not sufficient to permit the cell to function normally.
Haploinsufficiency can be due to a number of problems. One of the two copies of the gene may be missing due to a deletion. A mutation (change) in the gene may have wiped out production of message. Or the message or the protein produced by the cell may be unstable or degraded by the cell.
-ES cells are pluripotent and can able to differentiate into all derivatives of the three primary germ layers: ectoderm, endoderm, and mesoderm.
Cord blood:
-Cord blood, which is also called "placental blood," is the blood that remains in the umbilical cord and placenta following birth and after the cord is cut. Cord blood is routinely discarded with the placenta and umbilical cord, which has valuable stem-cells.
Embryonal carcinoma (EC):
-cells are pluripotent stem cells derived from teratocarcinomas and are considered the malignant counterparts of human embryonic stem (ES) cells.
Transdifferentiation and redifferentiation: examples??
Cavitation:
-(embryoid bodies) cavitation or lumen formation, which derived from mouse embryonic stem cells??
Haploinsufficiency:
-"haploid" (one instead of two chromosomes or two genes) + "insufficiency." Haploinsufficiency is related to hemizygosity.
A situation in which the total level of a gene product (a particular protein) produced by the cell is about half of the normal level and that is not sufficient to permit the cell to function normally.
Haploinsufficiency can be due to a number of problems. One of the two copies of the gene may be missing due to a deletion. A mutation (change) in the gene may have wiped out production of message. Or the message or the protein produced by the cell may be unstable or degraded by the cell.
Cancer stem cells
- expression of stem cell markers
- - capacity for self-renewal
- - multilineage differentiation
- - reestablishment of tumors after transplantation
- resistant to radiation and chemotherapy and may therefore be responsible for tumor recurrence
The non-obese diabetic-severe combined immunodeficient (NOD-SCID) mouse model:
-transplantation of human hematopoietic stem cells is the only true test of their long-term repopulation potential. Models are readily available to evaluate murine hematopoietic stem cells, but few exist that allow reliable quantification of human stem cells. The non-obese diabetic-severe combined immunodeficient (NOD-SCID) mouse model enables quantification of human hematopoietic stem cells.
Examples: Primitive human hematopoietic cells can be assayed on the basis of their ability to repopulate immune-deficient NOD/SCID mice and have been termed SCID repopulating cells (SRCs). The in vivo biological fate of individual SRCs can be tracked by following the unique retroviral insertion site in the progency of transduced SRCs. Distinct human SRCs were identified that differ in the proliferative and self-renewal capacity indicating that the primitive cell compartment is functionally heterogeneous.
mTOR signaling pathway:
-mTOR (mammalian target of rapamycin) appears to play a central role in signaling caused by nutrients and mitogens such as growth factors to regulate translation,
-the drug rapamycin acts on mammalian cells through the mTOR protein kinase,
-mTOR was also found to act as an ATP sensor to regulate cell growth,
-mTOR is a large class IV PI-3 kinase family member with protein kinase activity, but lacks any lipid kinase activity,
-biomarkers indicate that the mTOR pathway is hyperactive in certain types of cancers, suggesting that mTOR could be an attractive target for cancer therapy,
-activated mTOR may provide tumor cells with a growth advantage by promoting protein synthesis, which is the best-described physiological function of mTOR signaling. mTOR regulates Akt activity, a crucial downstream effector in the PI-3K–PTEN pathway, which controls cell proliferation and survival. Targeting this function of mTOR may also have therapeutic potential.
During nutrient deprivation, autophagy provides a supply of amino acids needed for cell survival (1). does qutophagy plqz role in cancer stem cells ???
- expression of stem cell markers
- - capacity for self-renewal
- - multilineage differentiation
- - reestablishment of tumors after transplantation
- resistant to radiation and chemotherapy and may therefore be responsible for tumor recurrence
The non-obese diabetic-severe combined immunodeficient (NOD-SCID) mouse model:
-transplantation of human hematopoietic stem cells is the only true test of their long-term repopulation potential. Models are readily available to evaluate murine hematopoietic stem cells, but few exist that allow reliable quantification of human stem cells. The non-obese diabetic-severe combined immunodeficient (NOD-SCID) mouse model enables quantification of human hematopoietic stem cells.
Examples: Primitive human hematopoietic cells can be assayed on the basis of their ability to repopulate immune-deficient NOD/SCID mice and have been termed SCID repopulating cells (SRCs). The in vivo biological fate of individual SRCs can be tracked by following the unique retroviral insertion site in the progency of transduced SRCs. Distinct human SRCs were identified that differ in the proliferative and self-renewal capacity indicating that the primitive cell compartment is functionally heterogeneous.
mTOR signaling pathway:
-mTOR (mammalian target of rapamycin) appears to play a central role in signaling caused by nutrients and mitogens such as growth factors to regulate translation,
-the drug rapamycin acts on mammalian cells through the mTOR protein kinase,
-mTOR was also found to act as an ATP sensor to regulate cell growth,
-mTOR is a large class IV PI-3 kinase family member with protein kinase activity, but lacks any lipid kinase activity,
-biomarkers indicate that the mTOR pathway is hyperactive in certain types of cancers, suggesting that mTOR could be an attractive target for cancer therapy,
-activated mTOR may provide tumor cells with a growth advantage by promoting protein synthesis, which is the best-described physiological function of mTOR signaling. mTOR regulates Akt activity, a crucial downstream effector in the PI-3K–PTEN pathway, which controls cell proliferation and survival. Targeting this function of mTOR may also have therapeutic potential.
During nutrient deprivation, autophagy provides a supply of amino acids needed for cell survival (1). does qutophagy plqz role in cancer stem cells ???
