In this journal club I intend to discuss a review article discussing iPSC, which will broaden our knowledge about contemporary research in anatomy and cell biology.At the end of session one should be able to write at least a short note in iPSC.
To start with…Before we proceed to the article of discussion itself.. A brief note on how how came across this article and why we should know about iPSC .Baker, Monya (2007-12-06). "Adult cells reprogrammed to pluripotency, without tumors". Nature Reports Stem Cells. doi:10.1038/stemcells.2007.124
Chinese? Nepalese? Naaa Japanese. He had a very interesting and unique career.
Born in 1962, Osaka , JapanHe was regarded as a lousy surgeon by his colleagues and was nicknamed "Jamanaka” a Japanese word for ‘obstacle’ after he took more than an hour to remove a benign tumor which usually takes not more than 10 minutes. Interestingly, one reason he was admitted into his first stem cell research because of his can do attitude.
Like anybody else, I was more curious than before and wanted to know more about iPSC. I turned to PubMed.
Within 4-5 years around 5 thousand articles were published on iPSC.
I came across what I was looking for..
A review article written by the master himselfBefore we discuss the article it would be a good idea to go through some terms that we need to understand.
Alexander Maksimov 1908 postulated hematopoetic stem cellsJoseph Altman and Gopal Das1960 Vs CajalTill and McCulloch 1963, hematopoeticSo there was this need for stem cells that were not antigenic and the best candidate for this was stem cell derived from the patient or subject himself.Another term is transcription factor.
These are inserted into cell and then into nucleus using vectors most common of which are retrovirus. Retroviruses remain in cell thereafter (Integrated)With this background, now lets proceed to the article.
First we will discuss the past? What made scientists to think that somatic or terminally differentiated cells could be reverted back to stem cells .There were 3 streams of researches which cumulated into scientists believe that iPSC were possible.
ESC: extra embryonic stem cellsDonar cell taken from mammary gland.Dolly Parton
iPSC are being generated and researched by labs throughout the world owing to its …..but has been criticized for ,,,,due to not very clearly known reasons.
What will the audience be able to do after this training is complete?Briefly describe each objective and how the audience will benefit from this presentation.
If somatic cells can be converted to stem cells why cant one somatic cell type converted to another somatic cell type e.g. hepatocyte to cardiomyocyte?
Use a section header for each of the topics, so there is a clear transition to the audience.
7 ipsc 3:5:13
I-PS CELLSCapt Rishi PokhrelJOURNAL CLUB
iPS cell‘Induced pluripotent stemcell is a type of pluripotentstem cell artificially derivedfrom a non-pluripotent cell -typically an adult somaticcell - by inducing a "forced"expression of specific genes’.Baker, Monya (2007-12-06). "Adult cells reprogrammed to pluripotency, withouttumors". Nature Reports Stem Cells. doi:10.1038/stemcells.2007.124
President of the International Societyfor Stem Cell Research (ISSCR).MBBSMD (1987)Ph. D (1993)Residency in Orthopedic surgeryPost doctoral fellowship in Cardiovascular diseaseProfessor of anatomyUniversity of California,San Francisco, USA
Nobel prize awarded for• Generation of induced pluripotent stem cells from adult mousefibroblasts (2006)• Closedly resembled embryonic stem cells (in vivo equivalent ofblastocyst)• iPSC were pluripotent – could generate whole iPSC mice• iPSC cells from human adult fibroblasts for the first time (2007)• Initially used 24 transcription factors for inducing pluri-potency• Successful in narrowing down the number of factors to just 4Sox2, Oct4, Klf 4 and c-Myc
Yamanaka SCell stem cellJune 14, 2012Volume 10, No 6Page no 678-684
Literature review• Stem cells• Transcription factors
How was iPSC possible?• Reprogramming by nuclear transfer• Tadpoles from unfertilized eggs thatreceived nucleus from intestinal cells ofadult frogs (Gurdon J 1962)• Cloning of Dolly (Wilmut W 1997)• Adult somatic cells contain all geneticinformation• Oocyte contain factors that canreprogram somatic cell nuclei, so do ESC(Tada T 2001)1Past
How was iPSC possible?• Discovery of transcription factors– Genes of drosophila coding for antenna couldform legs when ‘antennapeda’ was introduced(Schneuwly 1987)– Mammalian fibroblasts converted to myocyteusing MyoD (Davis 1987)2
How was iPSC possible?• Generation of ESC, mouse (Evans 1981),human (Thomson 1988) and culture media• Long term maintenance of pluripotency usingLIF (Smith 1988)• Optimal cultural conditions with bFGF3
• iPSC : simplicity and reproducibility• Poor efficacy: success rate 1% (?)• Integrated vectors used for introducingtranscription factors -> retroviruses, can causemutagenesis & other adverse effects• Use of non-integrated vectors: plasmid, Sendaivirus, adenovirus, synthetic RNA and proteins• Technology development -> applicationsPresent
Current works focused inRegenerative medicine–Parkinsons disease–Platelet deficiency–Spinal cord injury–Macular degenerationFuture
Disease models• Patient derived iPSC used for testing of drugs &toxins• Found useful for creating models of late onsetdiseases like Parkinson’s, Alzheimer’s,Schizophrenia• Analysis of disease mechanisms
Use in animals• Genetic engineering• Production of deficient proteins e.g.enzymes• Preservation or recreation of endangeredor extinct animals
Direct reprogramming• In vivo conversion of exocrine pancreatic cells toendocrine using 3 transcription factors (Zhou2008)• In vitro conversion of adult mature fibroblasts toneural cells, hepatocyte, cardiomyocyte orhematopoietic progenitor cells• Problem: source of cells?A step ahead
iPSC Vs ESC• Similar and different Source of tissue– Culture medium– Source of clone e.g. labs– Vectors used– Both are basically artificial cells
Dark side• Variation in– gene expression– DNA methylation– Pluripotent potential– Somatic mutations– Copy number variations– Immunogenicity
So is it just another hoax?DARK SIDEUNDER ATTACKFLAWEDTROUBLESOMEGROWINGPAINS
Not really• Genetic defects preexistedin source cells• Cloning magnified thedefects• Immunogenicity is veryweak - its effects nil inanimal experiments• ESC not gold standard forcomparisons of iPSC
Conclusions• iPSC technology ready for applications• Necessity of establishment of in-advancestocks of clones• Source of tissue: healthy donors, cord blood,HLA homozygous donors