Editing Transdifferentiation (section)

==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<ref name="ReferenceB"/> and functionally replace its natural counterpart. In one study, transdifferentiating tail-tip [[fibroblasts]] into hepatocyte-like cells using transcription factors [[GATA4|Gata4]], Hnf1α and [[FOXA3|Foxa3]], and inactivation of p19(Arf) restored hepatocyte-like liver functions in only half of the mice using survival as a means of evaluation.<ref>{{Cite journal 
| last1 = Huang | first1 = P. 
| last2 = He | first2 = Z. 
| last3 = Ji | first3 = S. 
| last4 = Sun | first4 = H. 
| last5 = Xiang | first5 = D. 
| last6 = Liu | first6 = C. 
| last7 = Hu | first7 = Y. 
| last8 = Wang | first8 = X. 
| last9 = Hui | first9 = L. 
| doi = 10.1038/nature10116 
| title = Induction of functional hepatocyte-like cells from mouse fibroblasts by defined factors 
| journal = Nature 
| volume = 475 
| issue = 7356 
| pages = 386–389 
| year = 2011 
| pmid = 21562492 
| s2cid = 1115749 
}}</ref>

===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|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.<ref>{{Cite journal 
| last1 = Pang | first1 = Z. P. 
| last2 = Yang | first2 = N. 
| last3 = Vierbuchen | first3 = T. 
| last4 = Ostermeier | first4 = A. 
| last5 = Fuentes | first5 = D. R. 
| last6 = Yang | first6 = T. Q. 
| last7 = Citri | first7 = A. 
| last8 = Sebastiano | first8 = V. 
| last9 = Marro | first9 = S. 
| last10 = Südhof 
| doi = 10.1038/nature10202 | first10 = T. C. 
| last11 = Wernig | first11 = M. 
| title = Induction of human neuronal cells by defined transcription factors 
| journal = Nature 
| volume = 476 
| issue = 7359 
| pages = 220–223 
| year = 2011 
| pmid = 21617644 
| pmc =3159048 
| bibcode = 2011Natur.476..220P 
}}</ref>

===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 progenitor]]s can differentiate into [[granulocyte]]/[[monocyte]] progenitor, [[eosinophil]], [[basophil]] or bipotent [[basophil]]/[[mast cell]] progenitor lineages.<ref>{{Cite journal 
| last1 = Iwasaki | first1 = H. 
| last2 = Mizuno | first2 = S. -I. 
| last3 = Arinobu | first3 = Y. 
| last4 = Ozawa | first4 = H. 
| last5 = Mori | first5 = Y. 
| last6 = Shigematsu | first6 = H. 
| last7 = Takatsu | first7 = K. 
| last8 = Tenen | first8 = D. G. 
| last9 = Akashi | first9 = K. 
| doi = 10.1101/gad.1493506 
| title = The order of expression of transcription factors directs hierarchical specification of hematopoietic lineages 
| journal = Genes & Development 
| volume = 20 
| issue = 21 
| pages = 3010–3021 
| year = 2006 
| pmid = 17079688 
| pmc =1620021 
}}</ref>

===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.<ref name="ReferenceA"/>

=== Method of transfection ===

In order to accomplish [[transfection]], one may use integrating [[viral vector]]s such as [[lentivirus]]es or [[retrovirus]]es, non-integrating vectors such as [[Sendai virus]]es or [[Adenoviridae|adenoviruses]], [[microRNA]]s and a variety of other methods including using proteins and [[plasmid]]s;<ref>{{Cite journal 
| last1 = Patel | first1 = M. 
| last2 = Yang | first2 = S. 
| doi = 10.1007/s12015-010-9123-8 
| title = Advances in Reprogramming Somatic Cells to Induced Pluripotent Stem Cells 
| journal = Stem Cell Reviews and Reports 
| volume = 6 
| issue = 3 
| pages = 367–380 
| year = 2010 
| pmid = 20336395 
| pmc =2924949 
}}</ref> one example is the non-viral delivery of transcription factor-encoding plasmids with a polymeric carrier to elicit neuronal transdifferentiation of fibroblasts.<ref>{{Cite journal 
| last1 = Adler | first1 = A. F. 
| last2 = Grigsby | first2 = C. L. 
| last3 = Kulangara | first3 = K. 
| last4 = Wang | first4 = H. 
| last5 = Yasuda | first5 = R. 
| last6 = Leong | first6 = K. W. 
| doi = 10.1038/mtna.2012.25 
| title = Nonviral Direct Conversion of Primary Mouse Embryonic Fibroblasts to Neuronal Cells 
| journal = Molecular Therapy: Nucleic Acids 
| volume = 1 
| issue = 7 
| pages = e32– 
| year = 2012 
| pmid = 23344148 
| pmc =3411320 
}}</ref> 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]]<ref>{{Cite journal 
| last1 = Sommer | first1 = C. A. 
| last2 = Sommer | first2 = A. 
| last3 = Longmire | first3 = T. A. 
| last4 = Christodoulou | first4 = C. 
| last5 = Thomas | first5 = D. D. 
| last6 = Gostissa | first6 = M. 
| last7 = Alt | first7 = F. W. 
| last8 = Murphy | first8 = G. J. 
| last9 = Kotton | first9 = D. N. 
| doi = 10.1002/stem.255 
| last10 = Mostoslavsky | first10 = G. 
| title = Excision of Reprogramming Transgenes Improves the Differentiation Potential of iPS Cells Generated with a Single Excisable Vector 
| journal = Stem Cells 
| volume = 28 
| issue = 1 
| pages = 64–74 
| year = 2009 
| pmid = 19904830 
| pmc =  4848036}}</ref> Non-integrating vectors have other issues concerning efficiency of reprogramming and also the removal of the vector.<ref>{{Cite journal 
| last1 = Zhou | first1 = W. 
| last2 = Freed | first2 = C. R. 
| doi = 10.1002/stem.201 
| title = Adenoviral Gene Delivery Can Reprogram Human Fibroblasts to Induced Pluripotent Stem Cells 
| journal = Stem Cells 
| volume = 27 
| issue = 11 
| pages = 2667–2674 
| year = 2009 
| pmid = 19697349 
| doi-access = free 
}}</ref> Other methods are relatively new fields and much remains to be discovered.