Stem Cell Biology Presentors: Ayesha Bano, Allyn Bryan, and Caleb

History Timeline Of Stem Cell Research  1868 – The words "stem cell" first used by Ernst Haeckel in literature to describe embryological development  1909 – Russian-American scientist Alexander Maximow theorizes that all blood cells are derived from the same ancestral cell thus introducing the idea of blood stem cells  1963 – Evidence of blood stem cells gathered by Canadian scientists Ernest McCulloch and James Till who performed experiments on bone marrow of mice  First time that pluripotent stem cells were isolated from mouse embryo, work done independently by Martin Evans and Gail Martin  1989 – First gene knockout mouse produced using embryonic stem cells and homologous recombination  The first batch of human embryonic stem cells were isolated by team of researchers at the University of Wisconsin, Madison.  2006-Shinya Yamanaka of Kyoto University in Japan developed "induced pluripotent stem cells" (Jointly awarded Nobel Prize for this work with John Gurdon in 2012).

Introduction Stem cells can divide and create copies of themselves as well as differentiate into specialized cell types Zygote is the origin of all stem cells

Embryonic Stem Cells  The zygote contains "totipotent" stem cells which can develop into any cell including those that make up the human embryo  Once the zygote has developed into a blastocyst, the inner cell mass will contain embryonic stem cells  Pluripotent -> limited capabilities but can still further create several specialized cell types

Adult Stem Cells  Undifferentiated cells present throughout the body  Found among other differentiated cells in a tissue or organ  Maintain and repair the tissue they are found in  Can divide, replicate and renew themselves  Multipotent -> Specialization is limited to one or more cell types  Mesenchymal stem cell can only give rise to bone, cartilage, or connective tissue  Unipotent -> Stem cells that generate only one cell type  Spermatagonial stem cell can only give rise to sperm cells

Adult Stem Cells - Bone Marrow Stemcells.nih.gov

Inducible Pluripotent Stem Cells (iPSCs) Yamanaka, first produced iPSCs in 2006 from a mouse and in 2007 from human fibroblasts through genetic reprogramming. iPSCs are artificially derived from an adult somatic cell by inducing a "forced expression" of certain genes. Specifically Oct3/4, Sox2, Nanog, Klf4, and c-Myc These four genes are introduced using retrovirus and encode transcription factors involved in cell development Takes 3-4 weeks to convert differentiated cell to pluripotent stem cells Artificially generated iPSCs are remarkably similar to naturally isolated pluripotent cells iPSCs is the answer to ethical concerns of wasting embryos.

Inducible Pluripotent Stem Cells (iPSCs) (Photo: iPS Academia Japan, Inc.) Regular path VS induced Path

Inducible Pluripotent Stem Cells (iPSCs) Nuclear transferViral reprogramming

(iPSCs) Applications  iPSCs are regarded as "holy grail of stem cell research"  Help understand disease models in vitro  Drug screening  Possibility of personalized iPSCs in specific diseases Deng, W. (2010). Induced pluripotent stem cells: paths to new medicines.EMBO Reports, 11(3), 161–165.

Stem Cell Therapies  Bone marrow transplantation  Cancer  Engineering skin tissue  Burn patients and skin grafts  Potential for organ tissue repair and regrowth  Heart, brain, spinal cord, limbs, teeth, eyes, ears  Myocardial infarction  Neurodegenerative diseases  Parkinson's disease  Treatment of Diabetes  Stem cells -> beta cells  Fertility Treatment

Visual illustration

Challenges of Stem Cell Therapies  Technical hurdles  Treat humans without giving them mutations and cancers  Controlling and mimicking conditions for drug modeling is essential .Ethical concerns  Treat suffering while respecting human/animal life  Moral status of embryo? 14 day vs 40 day cut off period?  Role of religion in determining status of embryo as "human" varies  Accessibility  need for transplantable tissues and organs far outweighs the available supply  Cost  Unpredictibility in funding due to politics 

Transdifferentiation or Lineage reprogramming  Definition:  Particular somatic cell is switched from one lineage to an entirely different identity.  Direct conversion of one type to another bypassing pluripotency  Routes to switch cell lineage  Using either transcription factors or microRNAs  Two major types of routes: 1. Direct Route: via a predefined set of transcription factors and/or microRNAs 2.Plastic State: via an indirect route reprogramming factors and followed by adding growth factors into culture medium

Transdifferentiation Direct Route

Transdifferentiation Timeline

Transdifferentiation Timeline

How to identify transcription factors

Pluripotency vs Transdifferentiation Pluripotency  Removal of all epigenetic marks  IPSCs must be differentiated before use  Induced pluripotency is riskier due to potential teratomas in vivo  Pluripotent cells have unlimited potential  In vivo regeneration hasn’t been accomplished yet  All transcription factors have been identified Transdifferentiation  Removal of some epigenetic marks  Final products can be used clinically  Transdifferentiated cell lineages may be safer due to fewer cell passages and less mutation  Transdifferentiation is focused towards conversion between similar lineages  In vivo regeneration has been achieved  TFs still under research

Direct Differentiation using small chemical molecules  The idea is to use small chemical molecules instead of transcription factors Oct4, Sox2, Klf4, c-Myc  Use of chemical molecules can increase efficiency and quality of reprogramming up to 100 folds  CiPSCs utlize XEN-like intermediate step which is closer to pluripotent state and more stable

Small Chemical Molecules that can replace TFs

Video Illustration on iPSCs Generation of Induced Pluripotent Stem Cells by Reprogramming Mouse Embryonic Fibroblasts with a Four Transcription Factor, Doxycycline Inducible Lentiviral Transduction System stem-cells-reprogramming-mouse

References agreement-for-induced-pluripotent-stemips-cell-patent-tec/ agreement-for-induced-pluripotent-stemips-cell-patent-tec/ Deng, W. (2010). Induced pluripotent stem cells: paths to new medicines.EMBO Reports, 11(3), 161– Takahashi, K.; Yamanaka, S. Induced pluripotent stem cells in medicine and biology. Development 2013, 140, 2457–2461. [Google Scholar] [CrossRef] [PubMed]Google ScholarCrossRefPubMed Extreme Makeover: Converting One Cell into Another:Zhou, Qiao et al.Cell Stem Cell, Volume 3, Issue 4, 382 – 388 Cohen, D. E., & Melton, D. (2011). Turning straw into gold: directing cell fate for regenerative medicine. Nature Reviews Genetics, 12(4), doi: /nrg2938