Transdifferentiation

Transdifferentiation, also known as lineage reprogramming,[1] is a process where one mature somatic cell transforms into another mature somatic cell without undergoing an intermediate pluripotent state or progenitor cell type.[2] It is a type of metaplasia, which includes all cell fate switches, including the interconversion of stem cells. Current uses of transdifferentiation include disease modeling and drug discovery and in the future may include gene therapy and regenerative medicine.[3] The term 'transdifferentiation' was originally coined by Selman and Kafatos in 1974 to describe a change in cell properties as cuticle producing cells became salt-secreting cells in silk moths undergoing metamorphosis.[4]

Discovery

Davis et al. 1987 reported the first instance of transdifferentiation where a cell changed from one adult cell type to another. Forcing mouse embryonic fibroblasts to express MyoD was found to be sufficient to turn those cells into myoblasts.[5]

Natural examples

There are no known instances where adult cells change directly from one lineage to another except Turritopsis dohrnii. Rather, cells dedifferentiate and then redifferentiate into the cell type of interest. In newts when the eye lens is removed, pigmented epithelial cells de-differentiate and then redifferentiate into the lens cells.[6]

While it was previously believed that oesophageal cells were developed from the transdifferentiation of smooth muscle cells, that has been shown to be false.[7]

Induced and therapeutic examples

The first example of functional transdifferentiation has been provided by Ferber et al.[8] by induce a shift in the developmental fate of cells in liver and convert them into 'pancreatic beta-cell-like' cells. The cells induced a wide, functional and long-lasting transdifferentiation process that reduced the effects of hyperglycemia in diabetic mice.[9] Moreover, the trans-differentiated beta-like cells were found to be resistant to the autoimmune attack that characterizes type 1 diabetes.[10]

The second step was to undergo transdifferentiation in human specimens. By transducing liver cells with a single gene, Sapir et al. were able to induce human liver cells to transdifferentiate into human beta cells.[11]

This approach has been demonstrated in mice, rat, xenopus and human tissues (Al-Hasani et al., 2013).

Schematic model of the hepatocyte-to-beta cell transdifferentiation process. Hepatocytes are obtained by liver biopsy from diabetic patient, cultured and expanded ex vivo, transduced with a PDX1 virus, transdifferentiated into functional insulin-producing beta cells, and transplanted back into the patient.[12]

Methods

Lineage-instructive approach

In this approach, transcription factors from progenitor cells of the target cell type are transfected into a somatic cell to induce transdifferentiation.[13] There exists two different means of determining which transcription factors to use: by starting with a large pool and narrowing down factors one by one[14] or by starting with one or two and adding more.[15] One theory to explain the exact specifics is that ectopic TFs directs the cell to an earlier progenitor state and then redirects it towards a new cell type. Rearrangement of the chromatin structure via DNA methylation or histone modification may play a role as well.[16] Here is a list of in vitro examples and in vivo examples. In vivo methods of transfecting specific mouse cells utilize the same kinds of vectors as in vitro experiments, except that the vector is injected into a specific organ. Zhou et al. (2008) injected Ngn3, Pdx1 and Mafa into the dorsal splenic lobe (pancreas) of mice to reprogram pancreatic exocrine cells into β-cells in order to ameliorate hyperglycaemia.[17]

Initial epigenetic activation phase approach

Somatic cells are first transfected with pluripotent reprogramming factors temporarily (Oct4, Sox2, Nanog, etc.) before being transfected with the desired inhibitory or activating factors.[18] Here is a list of examples in vitro.

Pharmacological agents

The DNA methylation inhibitor, 5-azacytidine is also known to promote phenotypic transdifferentiation of cardiac cells to skeletal myoblasts.[19]

Mechanism of action

The transcription factors serve as a short term trigger to an irreversible process. The transdifferentiation liver cells observed 8 months after one single injection of pdx1.[20]

The ectopic transcription factors turn off the host repertoire of gene expression in each of the cells. However, the alternate desired repertoire is being turned on only in a subpopulation of predisposed cells.[21] Despite the massive dedifferentiation – lineage tracing approach indeed demonstrates that transdifferentiation originates in adult cells.[22]

Mogrify algorithm

Determining the unique set of cellular factors that is needed to be manipulated for each cell conversion is a long and costly process that involved much trial and error. As a result, this first step of identifying the key set of cellular factors for cell conversion is the major obstacle researchers face in the field of cell reprogramming. An international team of researchers have developed an algorithm, called Mogrify(1), that can predict the optimal set of cellular factors required to convert one human cell type to another. When tested, Mogrify was able to accurately predict the set of cellular factors required for previously published cell conversions correctly. To further validate Mogrify's predictive ability, the team conducted two novel cell conversions in the laboratory using human cells, and these were successful in both attempts solely using the predictions of Mogrify.[23][24] Mogrify has been made available online for other researchers and scientists.

Issues

Evaluation

When examining transdifferentiated cells, it is important to look for markers of the target cell type and the absence of donor cell markers which can be accomplished using green fluorescent protein or immunodetection. It is also important to examine the cell function, epigenome, transcriptome, and proteome profiles. Cells can also be evaluated based upon their ability to integrate into the corresponding tissue in vivo[25] and functionally replace its natural counterpart. In one study, transdifferentiating tail-tip fibroblasts into hepatocyte-like cells using transcription factors Gata4, Hnf1α and Foxa3, and inactivation of p19(Arf) restored hepatocyte-like liver functions in only half of the mice using survival as a means of evaluation.[26]

Transition from mouse to human cells

Generally transdifferentiation that occurs in mouse cells does not translate in effectiveness or speediness in human cells. Pang et al. found that while transcription factors Ascl1, Brn2 and Myt1l turned mouse cells into mature neurons, the same set of factors only turned human cells into immature neurons. However, the addition of NeuroD1 was able to increase efficiency and help cells reach maturity.[27]

Order of transcription factor expression

The order of expression of transcription factors can direct the fate of the cell. Iwasaki et al. (2006) showed that in hematopoietic lineages, the expression timing of Gata-2 and (C/EBPalpha) can change whether or not a lymphoid-committed progenitors can differentiate into granulocyte/monocyte progenitor, eosinophil, basophil or bipotent basophil/mast cell progenitor lineages.[28]

Immunogenicity

It has been found for induced pluripotent stem cells that when injected into mice, the immune system of the synergeic mouse rejected the teratomas forming. Part of this may be because the immune system recognized epigenetic markers of specific sequences of the injected cells. However, when embryonic stem cells were injected, the immune response was much lower. Whether or not this will occur within transdifferentiated cells remains to be researched.[29]

Method of transfection

In order to accomplish transfection, one may use integrating viral vectors such as lentiviruses or retroviruses, non-integrating vectors such as Sendai viruses or adenoviruses, microRNAs and a variety of other methods including using proteins and plasmids;[30] one example is the non-viral delivery of transcription factor-encoding plasmids with a polymeric carrier to elicit neuronal transdifferentiation of fibroblasts.[31] When foreign molecules enter cells, one must take into account the possible drawbacks and potential to cause tumorous growth. Integrating viral vectors have the chance to cause mutations when inserted into the genome. One method of going around this is to excise the viral vector once reprogramming has occurred, an example being Cre-Lox recombination[32] Non-integrating vectors have other issues concerning efficiency of reprogramming and also the removal of the vector.[33] Other methods are relatively new fields and much remains to be discovered.

Pluripotent reprogramming

See also

References

  1. ↑ Orkin, S. H.; Zon, L. I. (2008). "Hematopoiesis: An Evolving Paradigm for Stem Cell Biology". Cell. 132 (4): 631–644. doi:10.1016/j.cell.2008.01.025. PMC 2628169Freely accessible. PMID 18295580.
  2. ↑ Graf, T.; Enver, T. (2009). "Forcing cells to change lineages". Nature. 462 (7273): 587–594. doi:10.1038/nature08533. PMID 19956253.
  3. ↑ Pournasr, B.; Khaloughi, K.; Salekdeh, G. H.; Totonchi, M.; Shahbazi, E.; Baharvand, H. (2011). "Concise Review: Alchemy of Biology: Generating Desired Cell Types from Abundant and Accessible Cells". Stem Cells. 29 (12): 1933–1941. doi:10.1002/stem.760. PMID 21997905.
  4. ↑ Selman, K.; Kafatos, F. C. (2013). "Transdifferentiation in the labial gland of silk moths: Is DNA required for cellular metamorphosis?". Cell differentiation. 3 (2): 81–94. doi:10.1016/0045-6039(74)90030-x. PMID 4277742.
  5. ↑ Davis, R. L.; Weintraub, H.; Lassar, A. B. (1987). "Expression of a single transfected cDNA converts fibroblasts to myoblasts". Cell. 51 (6): 987–1000. doi:10.1016/0092-8674(87)90585-x. PMID 3690668.
  6. ↑ Jopling, C.; Boue, S.; Belmonte, J. C. I. (2011). "Dedifferentiation, transdifferentiation and reprogramming: Three routes to regeneration". Nature Reviews Molecular Cell Biology. 12 (2): 79–89. doi:10.1038/nrm3043. PMID 21252997.
  7. ↑ Rishniw, M.; Xin, H. B.; Deng, K. Y.; Kotlikoff, M. I. (2003). "Skeletal myogenesis in the mouse esophagus does not occur through transdifferentiation". Genesis. 36 (2): 81–82. doi:10.1002/gene.10198. PMID 12820168.
  8. ↑ Ferber S, Halkin A, Cohen H, Ber I, Einav Y, Goldberg I, Barshack I, Seijffers R, Kopolovic J, Kaiser N, Karasik A (2000) Pancreatic and duodenal homeobox gene 1 induces expression of insulin genes in liver and ameliorates streptozotocin-induced hyperglycemia. http://www.nature.com/nm/journal/v6/n5/full/nm0500_568.html
  9. ↑ Sarah Ferber, Amir Halkin, Hofit Cohen, Idit Ber, Yulia Einav, Iris Goldberg, Iris Barshack, Rhona Seijffers, Juri Kopolovic, Nurit Kaiser & Avraham Karasik (2000) - "Pancreatic and duodenal homeobox gene 1 induces expression of insulin genes in liver and ameliorates streptozotocin-induced hyperglycemia"
  10. ↑ Shternhall-Ron K et al., Ectopic PDX-1 expression in liver ameliorates type 1 diabetes, Journal of Autoimmunity (2007), doi:10.1016/j.jaut.2007.02.010. http://www.orgenesis.com/uploads/default/files/shternhall-jai-2007.pdf
  11. ↑ Tamar Sapir, Keren Shternhall, Irit Meivar-Levy, Tamar Blumenfeld, Hamutal Cohen, Ehud Skutelsky, Smadar Eventov-Friedman, Iris Barshack, Iris Goldberg, Sarah Pri-Chen, Lya Ben-Dor, Sylvie Polak-Charcon, Avraham Karasik, Ilan Shimon, Eytan Mor, and Sarah Ferber (2005) Cell-replacement therapy for diabetes: Generating functional insulin-producing tissue from adult human liver cells
  12. ↑ Tamar Sapir, Keren Shternhall, Irit Meivar-Levy, Tamar Blumenfeld, Hamutal Cohen, Ehud Skutelsky, Smadar Eventov-Friedman, Iris Barshack, Iris Goldberg, Sarah Pri-Chen, Lya Ben-Dor, Sylvie Polak-Charcon, Avraham Karasik, Ilan Shimon, Eytan Mor, and Sarah Ferber (2005) Cell-replacement therapy for diabetes: Generating functional insulin-producing tissue from adult human liver cells
  13. ↑ Graf, T.; Enver, T. (2009). "Forcing cells to change lineages". Nature. 462 (7273): 587–594. doi:10.1038/nature08533. PMID 19956253.
  14. ↑ Ieda, M.; Fu, J. D.; Delgado-Olguin, P.; Vedantham, V.; Hayashi, Y.; Bruneau, B. G.; Srivastava, D. (2010). "Direct Reprogramming of Fibroblasts into Functional Cardiomyocytes by Defined Factors". Cell. 142 (3): 375–386. doi:10.1016/j.cell.2010.07.002. PMC 2919844Freely accessible. PMID 20691899.
  15. ↑ Vierbuchen, T.; Ostermeier, A.; Pang, Z. P.; Kokubu, Y.; Südhof, T. C.; Wernig, M. (2010). "Direct conversion of fibroblasts to functional neurons by defined factors". Nature. 463 (7284): 1035–1041. doi:10.1038/nature08797. PMC 2829121Freely accessible. PMID 20107439.
  16. ↑ Ang, Y. S.; Gaspar-Maia, A.; Lemischka, I. R.; Bernstein, E. (2011). "Stem cells and reprogramming: Breaking the epigenetic barrier?". Trends in Pharmacological Sciences. 32 (7): 394–401. doi:10.1016/j.tips.2011.03.002. PMC 3128683Freely accessible. PMID 21621281.
  17. ↑ Zhou, Q.; Brown, J.; Kanarek, A.; Rajagopal, J.; Melton, D. A. (2008). "In vivo reprogramming of adult pancreatic exocrine cells to β-cells". Nature. 455 (7213): 627–632. doi:10.1038/nature07314. PMID 18754011.
  18. ↑ Efe, J. A.; Hilcove, S.; Kim, J.; Zhou, H.; Ouyang, K.; Wang, G.; Chen, J.; Ding, S. (2011). "Conversion of mouse fibroblasts into cardiomyocytes using a direct reprogramming strategy". Nature Cell Biology. 13 (3): 215–222. doi:10.1038/ncb2164. PMID 21278734.
  19. ↑ kaur, keerat; yang, jinpu; eisenberg, carol; eisenberg, leonard (2014). "5-azacytidine promotes the transdifferentiation of cardiac cells to skeletal myocytes." (PDF). cellular reprogramming. 16: 324–30. doi:10.1089/cell.2014.0021. PMID 25090621.
  20. ↑ Sarah Ferber, Amir Halkin, Hofit Cohen, Idit Ber, Yulia Einav, Iris Goldberg, Iris Barshack, Rhona Seijffers, Juri Kopolovic, Nurit Kaiser & Avraham Karasik (2000) - "Pancreatic and duodenal homeobox gene 1 induces expression of insulin genes in liver and ameliorates streptozotocin-induced hyperglycemia"
  21. ↑ Meivar-Levy, I.; Sapir, T.; Gefen-Halevi, S.; Aviv, V.; Barshack, I.; Onaca, N.; Mor, E.; Ferber, S. (2007). "Pancreatic and duodenal homeobox gene 1 induces hepatic dedifferentiation by suppressing the expression of CCAAT/enhancer-binding protein β". Hepatology. 46 (3): 898–905. doi:10.1002/hep.21766. PMID 17705277.
  22. ↑ Mauda-Havakuk, M.; Litichever, N.; Chernichovski, E.; Nakar, O.; Winkler, E.; Mazkereth, R.; Orenstein, A.; Bar-Meir, E.; Ravassard, P.; Meivar-Levy, I.; Ferber, S. (2011). Linden, Rafael, ed. "Ectopic PDX-1 Expression Directly Reprograms Human Keratinocytes along Pancreatic Insulin-Producing Cells Fate". PLoS ONE. 6 (10): e26298. doi:10.1371/journal.pone.0026298. PMC 3196540Freely accessible. PMID 22028850.
  23. ↑ Mapping out cell conversion
  24. ↑ Owen, Rackham; Gough, Julian (2016). "A predictive computational framework for direct reprogramming between human cell types". Nature Genetics. doi:10.1038/ng.3487.
  25. ↑ Ieda, M.; Fu, J. D.; Delgado-Olguin, P.; Vedantham, V.; Hayashi, Y.; Bruneau, B. G.; Srivastava, D. (2010). "Direct Reprogramming of Fibroblasts into Functional Cardiomyocytes by Defined Factors". Cell. 142 (3): 375–386. doi:10.1016/j.cell.2010.07.002. PMC 2919844Freely accessible. PMID 20691899.
  26. ↑ Huang, P.; He, Z.; Ji, S.; Sun, H.; Xiang, D.; Liu, C.; Hu, Y.; Wang, X.; Hui, L. (2011). "Induction of functional hepatocyte-like cells from mouse fibroblasts by defined factors". Nature. 475 (7356): 386–389. doi:10.1038/nature10116. PMID 21562492.
  27. ↑ Pang, Z. P.; Yang, N.; Vierbuchen, T.; Ostermeier, A.; Fuentes, D. R.; Yang, T. Q.; Citri, A.; Sebastiano, V.; Marro, S.; Südhof, T. C.; Wernig, M. (2011). "Induction of human neuronal cells by defined transcription factors". Nature. 476 (7359): 220–223. doi:10.1038/nature10202. PMC 3159048Freely accessible. PMID 21617644.
  28. ↑ Iwasaki, H.; Mizuno, S. -I.; Arinobu, Y.; Ozawa, H.; Mori, Y.; Shigematsu, H.; Takatsu, K.; Tenen, D. G.; Akashi, K. (2006). "The order of expression of transcription factors directs hierarchical specification of hematopoietic lineages". Genes & Development. 20 (21): 3010–3021. doi:10.1101/gad.1493506. PMC 1620021Freely accessible. PMID 17079688.
  29. ↑ Pournasr, B.; Khaloughi, K.; Salekdeh, G. H.; Totonchi, M.; Shahbazi, E.; Baharvand, H. (2011). "Concise Review: Alchemy of Biology: Generating Desired Cell Types from Abundant and Accessible Cells". Stem Cells. 29 (12): 1933–1941. doi:10.1002/stem.760. PMID 21997905.
  30. ↑ Patel, M.; Yang, S. (2010). "Advances in Reprogramming Somatic Cells to Induced Pluripotent Stem Cells". Stem Cell Reviews and Reports. 6 (3): 367–380. doi:10.1007/s12015-010-9123-8. PMC 2924949Freely accessible. PMID 20336395.
  31. ↑ Adler, A. F.; Grigsby, C. L.; Kulangara, K.; Wang, H.; Yasuda, R.; Leong, K. W. (2012). "Nonviral Direct Conversion of Primary Mouse Embryonic Fibroblasts to Neuronal Cells". Molecular Therapy — Nucleic Acids. 1 (7): e32–. doi:10.1038/mtna.2012.25. PMC 3411320Freely accessible. PMID 23344148.
  32. ↑ Sommer, C. A.; Sommer, A.; Longmire, T. A.; Christodoulou, C.; Thomas, D. D.; Gostissa, M.; Alt, F. W.; Murphy, G. J.; Kotton, D. N.; Mostoslavsky, G. (2009). "Excision of Reprogramming Transgenes Improves the Differentiation Potential of iPS Cells Generated with a Single Excisable Vector". Stem Cells. 28 (1): 64–74. doi:10.1002/stem.255. PMID 19904830.
  33. ↑ Zhou, W.; Freed, C. R. (2009). "Adenoviral Gene Delivery Can Reprogram Human Fibroblasts to Induced Pluripotent Stem Cells". Stem Cells. 27 (11): 2667–2674. doi:10.1002/stem.201. PMID 19697349.
  34. 1 2 3 4 5 6 Zhou, Q.; Melton, D. A. (2008). "Extreme Makeover: Converting One Cell into Another". Cell Stem Cell. 3 (4): 382–388. doi:10.1016/j.stem.2008.09.015. PMID 18940730.
  35. ↑ Passier, R.; Mummery, C. (2010). "Getting to the Heart of the Matter: Direct Reprogramming to Cardiomyocytes". Cell Stem Cell. 7 (2): 139–141. doi:10.1016/j.stem.2010.07.004. PMID 20682439.
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