Stem CellsThe Basics

BackgroundCells of the Human Body

The human body is composed of many different types of cells

e.g. muscle cells, skin cells, liver cells, cardiovascular cells, etc.

Not all cells have the same potential

Some cells remain immature (blank slates)these are stem cells When stem cells mature, they turn into the different cells of the body

Embryonic development
You know how this goes

What does this remind you of? Morula, latin for Mulberry

Not exactly a blackberry

In Vitro Fertilizationcurrent method of deriving an hESC line

Eggs and sperm donated and fused to create a fertilized egg in a petri dish Fertilized egg matures into a blastocyst Embryonic stem cells extracted from blastocyst Cells replated on another petri dish and grown in culture

Embryonic development

What is a blastocyst?
Trophoblasta hollow sphere of cells that implants in the uterus and develops into the extra-embryonic membranes such as the placenta, umbilical cord, and amnion. Inner cell mass (ICM)embryonic stem cells are the ICM
=

Special Characteristics of ALL Stem Cells

Self-renewal (proliferation)- the ability of a stem cell to clone itself indefinitely by cell division. Asymmetric cell division Relocation and Differentiation are abilities of stem cells to migrate to where theyre needed in the body and specialize into a particular type of mature cell

Can form the entire human being

Generates every cell in the body including the placenta and extraembryonic tissues

Cannot form the entire human being

Can generate every cell in the body except placenta and extraembryonic tissues

Become specific cell types; may or may not have plasticity

There are two different types of Stem CellsAdult & Embryonic Stem Cells
Adult stem cells HELP YOU HEAL. They are cells found in post-natal tissue that can yield only the specialized cell types of the tissue from which they originated.
hematopoietic stem cells give rise to blood cells mesenchymal stem cells give rise to cells of connective tissues and bones umbilical cord stem cells a rich source of hematopoietic stem cells stem cells found in amniotic fluid might be more flexible than adult stem cells

Current Adult Stem Cell Therapies: Bone Marrow Transplant

Hematopoietic Stem Cell

There are two different types of Stem CellsAdult & Embryonic Stem Cells

Embryonic stem cells are derived from the inner cell mass of a blastocyst
Can self-renew indefinitely in culture Embryonic stem cells used for research are made in a Petri dish, not a woman's body They hold great potential for alleviating the symptoms of or even curing: Paralysis Diabetes Alzheimers

Real Mouse Embryonic Stem cells

What is Stem Cell Research?

Experimental model systems, understanding more about development Cell-based therapies Pharmaceutical research and testing

Experimental model systemCardiomyocytes

START

Cell-based therapySpinal Cord Injury

Clinical trials starting for treatment of spinal cord injury* in humans (after much data gathered using rats as an animal model)
Differentiate (+ growth factors) time

Oligodendrocytes
markedly recovered injured

Drug DevelopmentCancer Stem Cells

Reya, T., et al. Nature, 2001

Stem Cell Culture

Artificially directing stem cell fate
Why do you think this is useful?

Mouse feeder-layer

What problems do you foresee in trying to transplant this tissue into a human? Source: NIH

Somatic Cell Nuclear Transfer

Future Method of Deriving Stem Cells

Transcription Factor
Transcription factor glossary Transcription copying of DNA by RNA polymerase into RNA Factor a substance, such as a protein, that contributes to the cause of a specific biochemical reaction or bodily process Transcriptional regulation controlling the rate of gene transcription for example by helping or hindering RNA polymerase binding to DNA Upregulation, activation, or promotion increase the rate of gene transcription Downregulation, repression, or suppression decrease the rate of gene transcription Coactivator a protein that works with transcription factors to increase the rate of gene transcription

Corepressor a protein that works with transcription factors to decrease the rate of gene transcription

Mechanism of Action: TF
Transcription factors bind to either enhancer or promoter regions of DNA adjacent to the genes that they regulate. Depending on the transcription factor, the transcription of the adjacent gene is either up- or downregulated. Transcription factors use a variety of mechanisms for the regulation of gene expression.These mechanisms include: Stabilize or block the binding of RNA polymerase to DNA Catalyze the acetylation or deacetylation of histone proteins. The transcription factor can either do this directly or recruit other proteins with this catalytic activity. Many transcription factors use one or the other of two opposing mechanisms to regulate transcription:
histone acetyltransferase (HAT) activity acetylates histone proteins, which weakens the association of DNA with histones, which make the DNA more accessible to transcription, thereby up-regulating transcription histone deacetylase (HDAC) activity deacetylates histone proteins, which strengthens the association of DNA with histones, which make the DNA less accessible to transcription, thereby down-regulating transcription

Recruit coactivator or corepressor proteins to the transcription factor DNA complex

The induction of pluripotency

Krppel-like factor 4 (KlF4) octamer-binding transcription factor 4 (Oct-4 ) SOX (SRY-like HMG-box gene) transcription factor family

Not all genes are active at all times. DNA methylation is one of several epigenetic mechanisms that cells use to control gene expression.

Natural and artificial transdifferentiation

Nature Reviews Molecular Cell Biology 12, 79-89 (February 2011)

Reprogramming, Dedifferentiation, Transdifferentiation

The Nobel Prize in Physiology or Medicine 2007

Transcriptional proling of stem cells

Miguel Ramalho-Santos, et al., 2002, Science

Overlaps between genes enriched in stem cells

Miguel Ramalho-Santos, et al., 2002, Science

The 216 genes enriched in all three stem cells

Miguel Ramalho-Santos, et al., 2002, Science

Chromosomal locations of stem cell enriched genes

Miguel Ramalho-Santos, et al., 2002, Science

Stemness: Essential Contributes

Active JAK/STAT (Janus kinase/signal transducers and activators of transcription), TGF- (transforming growth factor), Yes (Yamaguchi sarcoma) kinase, and Notch signaling Capacity to sense growth hormone and thrombin Interaction with the extracellular matrix via integrin 6/1, Adam9, and bystin Engagement in the cell cycle, either arrested in G1 or cycling High resistance to stress, with upregulated DNA repair, protein folding, ubiquitin system, and detoxifier systems A remodeled chromatin, acted upon by DNA helicases, DNA methylases, and histone deacetylases Translation regulated by RNA helicases of the Vasa type. Only four of the 216 genes enriched in all SCs had absolute calls of absent in all the differentiated cell samples and present in all the SC samples, namely Uridine phosphorylase, Suppressor of Lec15, and two ESTs

Signaling factors

Transcription factors

Pluripotency
DNA methylation Cell cycle regulators

MicroRNA & genes involving in chromosome stability

Factors involved in pluripotency

Ras/Raf/ERK Wnt Signaling PI3K

BMP

LIF/STAT

Secondary Messengers Transcription factors

(Oct4, Nanog, Sox2)

PLURIPOTENCY GENES

DIFFERENTIATION GENES

Regulation of proliferation of mouse ES cells

Regulation of proliferation of mouse ES cells. (A) Pluripotent transcription factors activate the expression of (B) certain effectors that drive ES cell proliferation. Among these, Eras and Tcl1 stimulate the (C) phosphoinositide3-kinase (PI3K)/Akt signaling pathway to promote the cell cycle, whereas b-Myb and c-Myc activate the progression of the cell cycle directly. How Utf1 and Sall4 affect ES cell proliferation remains unknown.

 Questions?