Trends in Biotechnology Stem Cells

References Understanding Stem Cells National Academy Science dels.nas.edu/dels/rpt_briefs/Understanding_Stem_Cells.pdf eBioscience – Stem Cell Booklet http://www.ebioscience.com/ebioscience/whatsnew/pdf/Stem_Cell_Booklet.pdf

Possible fix for type I diabetes. Beta cells are destroyed by the patient’s overactive immune system. The cells of the body cannot take up glucose and they starve. Patients need insulin injections several times a day. Transform embryonic stem cells into insulin-producing cells. Combine beta-cell transplants with methods to “fix” the patient’s immune system.

http://sciencestage.com/v/882/stem-cell-primer.html Stem Cell Primer

http://www.youtube.com/watch?v=mUcE1Y_bOQE Stem Cells

In Vitro Fertilization • can produce all cell types • relatively easy to identify, isolate, maintain, and grow in the laboratory • large source of “excess” blastocysts from IVF clinics But - risk of creating teratomas (tumors) from implanting undifferentiated stem cells

Nuclear Transfer • can produce all cell types • relatively easy to identify, isolate, maintain, and grow in the laboratory • stem cells may be genetically matched to patient • not yet achieved with human cells • risk of creating teratomas (tumors)

Adult Tissues • demonstrated success in some treatments • stem cells may be genetically matched to patient • produce limited number of cell types • not found in all tissues • difficult to identify, isolate, maintain, and grow in the laboratory

Identifying Stem Cells Every cell displays an array of proteins on its surface. Different cell types have different proteins. Use these surface proteins as “markers” that characterize individual cell types—a type of “molecular ID.” Also identify stem cells by observing their behavior in the laboratory: stem cells must be able to remain unspecialized and self-renew for long periods of time.

Test whether embryonic stem cells have differentiated by labeling for specific marker proteins found in specialized cells.

Fluorescent markers can be used to identify stem cells hidden among ordinary adult cells. Here, human embryonic stem cells are recognized by the marker proteins they express (green). Courtesy of Paul J. Tesar, Laboratory of Molecular Biology, NINDS and the NIH Stem Cell Unit.

eg Blood Stem Cells The body replenishes the blood cells through a small number of semi-specialized hematopoietic (heem-AT-oh-poh-EH-tik) stem cells contained in the blood and bone marrow. This type of adult stem cell is used to treat diseases such as leukemia, sickle cell anemia, bone marrow damage, and some metabolic disorders and immunodeficiencies where the body has lost its ability to replenish its own set of healthy blood cells. Hematopoietic stem cells give rise to all the blood cell types, from infection-fighting white blood cells to blood-clotting platelets. Preliminary results have suggested that they may also be able to produce other cell types not found in blood, but this is not yet proven.

CD = Cluster of Differentiation

See a more complete poster of stem cell types and markers at http://eee.ebioscience.com/ebioscience/whatsnew/pdf/Stem_Cell_Poster.pdf

©2008 Terese Winslow

Stem cell research provides a useful tool for studying the molecular mechanisms that determine the differentiation fate of a pluripotent cell and for understanding the gene expression properties and epigenetic modifications needed to maintain the pluripotent state. In the future, stem cells may be made for transplantation therapies, where a specific cell population affected by disease is replaced with new, functional cells. Differentiated derivatives of human pluripotent cells may be useful as models for understanding the biology of disease and developing new drugs, particularly when there is no animal model for the disease.

Recent results Overexpression of a small number of transcription factors can reprogram differentiated cells into induced pluripotent stem (iPS) cells that resemble embryonic stem (ES) cells.

A drug-inducible lentiviral reprogramming strategy to achieve tight control of transgene expression in iPS cells and their derivatives.

Genome map of a lentivirus by C. Büchen-Osmond and J. Whitehead

Packaging and infection by a lentiviral vector http://en.wikipedia.org/wiki/File:Lentiviral_vector.png Peter Znamenskiy public domain

The four original reprogramming factors (Oct4 (also known as Pou5f1), Sox2, Klf4 and c-Myc) were placed under control of the tetO promoter, which is activated by the reverse tetracycline transactivator (rtTA) protein in the presence of the tetracycline analogue doxycycline (dox).

Exposed MEFs to the histone deacetylase inhibitor valproic acid (VPA), which has been reported to enhance reprogramming efficiency and to select against incompletely reprogrammed cells by inhibiting cell division.

http://video. google. com/videoplay http://video.google.com/videoplay?docid=8370692532177471184&hl=en# What are induced pluripotent stem cells?

A scheme of the generation of induced pluripotent stem (iPS) cells. Isolate and culture host cells. e.g. mouse embryonic fibroblasts and adult human dermal fibroblasts. Introduce the ES specific genes (iPS factors) into the cells by using retrovirus vector. Red cells indicate the cells expressing the exogenous genes. Harvest and culture the cells according to the method for ES cell culture using feeder cells (gray). A subset of the cells generates ES-like colonies, that is, iPS cells. http://en.wikipedia.org/wiki/File:Induction_of_iPS_cells.svg Y tambe GNU Free Documentation License

Assessed lines for similarity to ES cells by morphology, proliferation rate, expression of pluripotency markers and ability to generate embryoid bodies

Adult mice generated from induced pluripotent stem cells (2009) Michael J. Boland, Jennifer L. Hazen, Kristopher L. Nazor, Alberto R. Rodriguez, Wesley Gifford, Greg Martin, Sergey Kupriyanov & Kristin K. Baldwin Letters to Nature 461:91-94. 3 September 2009

Other videos available on the internet: http://www.youtube.com/watch?v=WaRnVcwZ0i8&feature=related What's the difference between embyonic and adult stem cells? http://www.youtube.com/watch?v=eBpTPhGaHYE&NR=1 Adult Stem Cells and Regeneration

Or you can watch a complete lecture series at http://www. hhmi Lecture 1 — Understanding Embryonic Stem Cells, Lecture 2 — Adult Stem Cells and Regeneration, Lecture 3 — Coaxing Embryonic Stem Cells, Lecture 4 — Stem Cells and the End of Aging, Discussion Session — Stem Cell Research: Policies and Ethics