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Lecture 25

Part Ib. Stem Cells - Therapy and Medical Research

Issues in Biotechnology:The Way We Work With Life

Dr. Albert P. Kausch

life edu.us

Medical Biotechnology

Issues in Biotechnology:Biotechnology, Our Society and Our Future

OnCampus LiveOnCampus LiveBCH 190, MIC 190, AFS 190, NRS 190, PLS 190BCH 190, MIC 190, AFS 190, NRS 190, PLS 190

OnLine BCH 190OnLine BCH 190

A Sweeping General Survey on Life and BiotechnologyA Public Access College Course

The University of Rhode Island

Kimberly Nelson

Issues in Biotechnology:The Way We Work With Life

Dr. Albert P. Kausch

life edu.us

© life_edu

A Sweeping General Survey on Life and Biotechnology

The University of Rhode Island

Issues in Biotechnology:The Way We Work With Life

Dr. Albert P. Kausch

life edu.us

BCH 190BCH 190 Section II.

The Applications of Biotechnology

Stem Cell ResearchWhat is involved?

What is the potential?

What are the downsides?

Who decides?

What, for you, divides our thinking about stem cell research?

(A) support for the research that may result in human cloning(B) uncertain research, not near clinical trials(C) lack of application, still unproven results(D) right to life issues(E) lack of information on the topic

Do you support federal funding for embryonic stem cell research?

(A) yes(B) no(C) yes, but with restrictions

Therapeuticvs

Reproductive Cloning

Embryonic Stem Cellsvs

Adult Stem Cellsvs

iPS Cells

Stem Cell Research

Obama lifts the ban of Federal funding for Stem Cell Research

New research will bring innovation

Stem Cell Basics

Embryonic, Adult and

Induced Pluripotent Stem Cells

Stem Cell BasicsWhat is a Stem Cell?

What Are the Different Types of Stem Cells?

Stem Cell Therapy: How will this be used?What is the progress?What are the controversies?

Characteristics of Stem Cells:

A capacity for extensive self-renewal

Developmental potential (The ability to differentiate)

The type of stem cell is defined both by its origin (site, species, age) and by its differentiation abilities

What is a stem cell?

Examples of types of stem cells based on age of organism at time of derivation

• Embryonic• Fetal• Newborn• Adult• iPSc

Stem cells give rise to specialized cells residing in organs

Stem Cells

Stem Cell Self-renewal

Symmetric Cell Division

(Proliferation)

1 stem cell

2 stem cells

Stem Cell Differentiation

Asymmetric Cell Division

(Proliferation)

1 stem cell

1 stem cell +1 differentiatedcell

Stem Cells

Stem Cells

Totipotency

Types of Stem Cells

Embryonic Stem Cells (ES Cells)--Derived from early embryos (blastocysts) ES cells are totipotent--they can form all the cell-types in an adult

ES cells are embryonic stem cells and immortal (unlimited proliferation in dishes)

hES cells are Human embryonic stem cells Adult Stem Cells - Derived from pluripotent adult tissues (i.e. bone

marrow transplants) IPS Cells - Induced pluripotent cells are derived from adult tissues

and genetically modified with genes that override the cell division pathways to become stem cells capable of then differentiating

Embryonic Stem Cells

(ES cells)

Preimplantation Development

Morula

Blastocyst

Zygote

ICM

Extraembryonicprogenitors

Preimplantation Development

Morula

Blastocyst

Zygote

ICM

Extraembryonicprogenitors

Derivation of Embryonic Stem Cells

Morula

Zygote

Blastocyst

ES cells

Totipotency of ES cells

Zygote

Morula

Blastocyst

ES cells

How embryonic stem cell lines are produced

Dissociate blastocyst into single cells

Grow in the presence of growth factors

Only rarely does a single cell continue to grow indefinitely

How embryonic stem cell lines are validated

EctodermSkin

Neurons

MesodermMuscleBloodBone

EndodermPancreas

LungLiver

Grow cells under different conditions to stimulate differentiation into multiple different cell types

Immuno-rejection

Solutions to immuno-rejectionPersonalized Stem Cells

Somatic cell nuclear transfer

Unfertilized egg cell

Remove nucleus from the egg cell and replace it with that ofthe patient

Use newly formed stem cells for therapeutic purposesSpecific to the patientIn addition, ES cells canbe genetically engineered

Cell from patient

These cells can also be genetically engineered

Promoter Coding Sequence Terminator

Stem Cells can be Genetically ModifiedGenetic constructs could be used in stem

cell therapy

Controlled expressionCell and developmentally

specific

Your favorite gene

InsulinAnti-cancerTay-sachsmany others

Stop transcriptionMessage stability

Nuclear Transfer Stem Cell Therapy

Advantages:

No Rejection (Complete tissue-type match)Complete CuresVersatile-Genetic Modification

Disadvantages:

Controversial ExpensiveRequires supply of oocytes

ES Cell Technology is Well Established in Mice

Morula

Blastocyst

Zygote

ICM

Extraembryonicprogenitors

Use of ES technology is well established in the mouse model

ES cells from mice can be made genetically engineered and established in recipients

The same Technology Could be Applied to Humans

Morula

Blastocyst

Zygote

ICM

Extraembryonicprogenitors

The same technology could be used to clone a human

Therapeuticvs

Reproductive Cloning

Embryonic Stem Cells

vsAdult Stem Cells

vsiPS Cells

Adult Stem Cells

Types of Stem Cells:

Adult Stem Cells--A small subpopulation of specialized stem cells present within some adult organs capable of self renewal

Adult stem cells are pleuripotent--They have more limited potential

Adult stem cells cannot divide indefinitely

Adult Stem Cells

Adult Stem Cells

pluripotency

• Hematopoietic stem cells are capable of differentiation into all mature peripheral blood cells

• Liver stem cells are capable of differentiation into hepatocytes and cholangiocytes

• Neural stem cells can differentiate into neurons, astroglia and oligodendrocytes

Some examples of types of stem cells based on their differentiation abilities

Adult Stem Cell Therapy

Steps:

1. Biopsy

2. In vitro culture of Adult Stem Cells

3. In vitro expansion or differentiation (If needed)

4. Genetic modification (as needed)

5. Engraftment

Bone Marrow Transplantation

Plasticity: The ability of cells to change developmental fate

GFP

Donor Bone Marrow(Contains HematopoeticStem Cells)

Engraftment

Repopulation of “Lethally” IrradiatedHost

(Lymphoid + Myeloid)

Genetic Modification

Adult stem cell differentiation has been demonstrated

Adult Stem Cell Therapy

Advantages:No Immuno-Rejection (if from self)Not Controversial

Disadvantages:Many organs do not contain adult stem cellsStem cells may be absent or defective in the affected organsUse of Plasticity and transdifferentiation is unproven

Adult Stem Cells

Adult Stem Cells

Self Renewal(symmetric cell division)

Adult Stem Cells

Adult Stem Cells

Self Renewal

Activation,Proliferation(asymmetric cell division)

Adult Organs

Bone Marrow/ Blood Skin Liver Pancreas Intestine Testes Brain ( but limited)

Organs With Adult Stem Cells

More to come?

Probably!

And these can be genetically modified to suit a specific disease or disorder

Question about Plasticity

Plasticity: The ability of cells to change developmental fate

GFP

Donor Bone Marrow(Contains HematopoeticStem Cells)

Engraftment

Repopulation of “Lethally” IrradiatedHost

(Lymphoid + Myeloid)

+

Neurons(a few!)

True Plasticity

GFP+

Donor Bone Marrow(Contains HematopoeticStem Cells)

EngraftedCells

(Lymphoid + Myeloid)

Neurons(a few!)

Induced Pluripotent Stem Cells (iPS)

Induced Pluripotent Stem Cells (iPS)

Induced pluripotent stem cells, commonly abbreviated as iPS cells or iPSc, are a type of pluripotent stem cell artificially derived from a non-pluripotent cell, typically an adult somatic cell, by inducing a “forced” expression of certain genes that regulate the cell division cycle thus making them immortal.

Induced Pluripotent Stem Cells are believed to be identical to natural pluripotent stem cells, such as embryonic stem cells in many respects, such as the expression of certain stem cell genes and proteins, chromatin methylation patterns, doubling time, embryo formation, teratoma formation, viable chimera formation, and potency and differentiability, but the full extent of their relation to natural pluripotent stem cells is still being assessed.

IPScs were first produced in 2006 from mouse cells and in 2007 from human cells. This has been cited as an important advancement in stem cell research, as it may allow researchers to obtain pluripotent stem cells, which are important in research and potentially have therapeutic uses, without the controversial use of embryos.

Induced Pluripotent Stem Cell Basics

A scheme of the generation of induced pluripotent stem (iPS) cells. (1) Isolate and culture donor cells. (2) Transfect stem cell-associated genes into the cells by viral vectors. Red cells indicate the cells expressing the exogenous genes. (3) Harvest and culture the cells according to ES cell culture, using mitotically inactivated feeder cells (4) A small subset of the transfected cells become iPS cells and generate ES-like colonies.

INDUCED PLURIPOTENT Stem Cell Lines Derived from Human Somatic Cells Junying Yu 1*, Maxim A. Vodyanik 2, Kim Smuga-Otto 1, Jessica Antosiewicz-Bourget 1, Jennifer L Frane 3, Shulan Tian 4, Jeff Nie 4, Gudrun A. Jonsdottir 4, Victor Ruotti 4, Ron Stewart 4, Igor I. Slukvin 5, James A. Thomson 6 Science DOI: 10.1126/science.1151526

Stem Cell Therapy:Recent Progress

Stem Cell Therapy

Human Degenerative Disorders:

Many diseases are characterized by organ failure or cell loss:

Degenerative disease states (unknown etiology or autoimmune disease)

Age-related degenerative disease

Cell or organ destruction due to genetic or infectious disease

Stem Cell Therapy:

Stem Cell Therapy is the replenishment of cells or organs lost to degenerative disease or loss of function with new cells derived from stem cells

Stem Cell Therapy is Experimental (Except Bone Marrow Transplantation)

Stem Cell therapy should be free of pharmaceutical toxicity

Cellular Pharmaceutical

Stem Cell Therapy

Stem Cell Therapy offers a method to treat degenerative disorders

A Cellular Pharmaceutical

Diseases amenable to Stem Cell Therapy Diabetes Spinal chord injury Alzheimer’s Disease Parkinson’s Disease Heart Disease Many more…..but not overnight

Stem Cell Therapy

Established Stem Cell Therapies:

Bone Marrow Transplantation--Restores blood system with hematopoetic stem cells (1950s)

(many variations)

Simplest Diseases to cure next involve secretory cells: Parkinson’s Disease--Dopamine Secretion Type I Diabetes--Insulin Secretion (Type II is difficult due to insulin insensitivity)

ES cell Therapy(Parkinson’s Disease)

ES cells

Dopaminergic Neurons

Engraftment

Dopamine Production Restored

in vitro:

in vivo:

Parkinson’s Disease-Experimental Therapy

From Lindvall et al., Nat. Med. 2004, S42-S50.

Stem Cell TherapyCriticisms and Controversies

Embryogenic Stem Cell

CriticismsI. Destruction of human embryos

II. A slippery slope to:

• Human Genetic Engineering

• Human cloning

Somatic cell nuclear transfer techniques are the same as those that could be applied to

cloning

Unfertilized egg cell

Remove nucleus from the egg cell and replace it with that ofthe patient

Cell from patient

These cells can also be genetically engineered

Use newly formed stem cells for therapeutic purposesSpecific to the patientIn addition, ES cells canbe genetically engineered

Where is DNA in a cell?

• All of the information for each protein in a cell is in it’s DNA• All of the information that codes for a complete organism is in DNA

7. Induced Pluripotent Stem Cells (iPCs) are made by the 7. Induced Pluripotent Stem Cells (iPCs) are made by the introduction of genes that cause the normal cell cycle to introduction of genes that cause the normal cell cycle to remain in the cell division mode, ensuring that they will divide remain in the cell division mode, ensuring that they will divide in culture indefinitely. Two or more cloned genes are inserted in culture indefinitely. Two or more cloned genes are inserted into the genome of adult differentiated cells; these genes are into the genome of adult differentiated cells; these genes are similar to those involved with tumor genesis that override similar to those involved with tumor genesis that override normal cell division. These cells can then be triggered to normal cell division. These cells can then be triggered to become any adult cell type, thereby obviating the controversy become any adult cell type, thereby obviating the controversy about using cells derived from human embryos. The process about using cells derived from human embryos. The process of inserting genes into cells is called:of inserting genes into cells is called:

(A) eugenics (A) eugenics (B)(B) carcinogenics carcinogenics (C)(C) transgenics transgenics (D)(D) genomics genomics (E) idiomics(E) idiomics

8. Stem cell research in humans is controversial, in part, 8. Stem cell research in humans is controversial, in part, because of the so-called ‘slippery slope’ argument. It is because of the so-called ‘slippery slope’ argument. It is suggested that the same technology that is developed for hES suggested that the same technology that is developed for hES stem cells and therapeutic cloning could easily be extended to:stem cells and therapeutic cloning could easily be extended to:

(A) determining who can have children(A) determining who can have children(B)(B) reproductive cloning of humans reproductive cloning of humans (C)(C) increased abortions increased abortions (D)(D) a policy of ethnic cleansinga policy of ethnic cleansing(E) the creation of bioweapons (E) the creation of bioweapons

9. The defining characteristic(s) of adult stem cells are that:9. The defining characteristic(s) of adult stem cells are that:  (A) they are pluripotent - they have a limited potential to (A) they are pluripotent - they have a limited potential to differentiate to other cell types differentiate to other cell types (B) all of these answers are appropriate (B) all of these answers are appropriate (C) they are seen as a potential source for therapeutic (C) they are seen as a potential source for therapeutic purposes that will not be controversial purposes that will not be controversial (D) they are unable to divide indefinitely (D) they are unable to divide indefinitely (E) they can be genetically engineered to carry and express (E) they can be genetically engineered to carry and express foreign genesforeign genes

10. Adult Stem Cells are:10. Adult Stem Cells are:  (A) outlawed in every country in the world as part of a wide (A) outlawed in every country in the world as part of a wide ban on cloning ban on cloning (B) from a small subpopulation of specialized stem cells (B) from a small subpopulation of specialized stem cells present within some adult organs and capable of self renewal present within some adult organs and capable of self renewal and limited capacity to differentiateand limited capacity to differentiate(C) an artifact that was famous for being fraudulently (C) an artifact that was famous for being fraudulently reported reported (D) unable to differentiate into any other cell type (D) unable to differentiate into any other cell type (E) unable to be genetically altered the way hES cells are(E) unable to be genetically altered the way hES cells are

11. Degenerative diseases that result in cell death or function, 11. Degenerative diseases that result in cell death or function, such as Parkinson’s and Alzheimer’s, injuries that may result such as Parkinson’s and Alzheimer’s, injuries that may result in damages cells and tissues, such as spinal cord injuries, and in damages cells and tissues, such as spinal cord injuries, and the lack of available human donor organs for transplant has the lack of available human donor organs for transplant has motivated scientists to investigate new ways to replace the motivated scientists to investigate new ways to replace the functions of diseased organs. Promising approaches to these functions of diseased organs. Promising approaches to these problems that have received considerable research are:problems that have received considerable research are:  (A) xenotransplantation and tissue engineering (A) xenotransplantation and tissue engineering (B) all of these approaches (B) all of these approaches (C) gene therapy (C) gene therapy (D) creation of artificial biomedical devices(D) creation of artificial biomedical devices(E) stem cells (E) stem cells

12. Human Embryonic Stem Cells (hES Cells) are:12. Human Embryonic Stem Cells (hES Cells) are:  (A) derived from adult bone marrow as leukemia treatments(A) derived from adult bone marrow as leukemia treatments(B) derived from early (five day old) embryos (blastocysts) (B) derived from early (five day old) embryos (blastocysts) (C) outlawed in every country in the world (C) outlawed in every country in the world (D) made with great ease at the Seoul National University in (D) made with great ease at the Seoul National University in South KoreaSouth Korea(E) were funded through vast amount of Federal dollars in the (E) were funded through vast amount of Federal dollars in the US from 2001-2008 US from 2001-2008

13. The history behind the development and therapeutic 13. The history behind the development and therapeutic applications for adult stem cells includes that they:applications for adult stem cells includes that they:

(A) have been used to reproductively clone primates and in (A) have been used to reproductively clone primates and in one case an adult human beingone case an adult human being(B) have been derived from every tissue type in the human (B) have been derived from every tissue type in the human body and have been shown to have equal developmental body and have been shown to have equal developmental plasticity compared with hES cells plasticity compared with hES cells (C) have been shown to be more useful than embryonic stem (C) have been shown to be more useful than embryonic stem cells for any applications, demonstrating the importance of cells for any applications, demonstrating the importance of hES cells hES cells (D) have been used to treat leukemia using bone marrow since (D) have been used to treat leukemia using bone marrow since 1956 and have been used successfully without controversy for 1956 and have been used successfully without controversy for many years many years (E) in spite of their great promise, because of serious errors (E) in spite of their great promise, because of serious errors will never be successful in clinical trials to treat any disease will never be successful in clinical trials to treat any disease and have resulted in many fatalitiesand have resulted in many fatalities

14. Many diseases are characterized by organ failure or cell 14. Many diseases are characterized by organ failure or cell loss. These types of disease are attractive candidates for stem loss. These types of disease are attractive candidates for stem cell therapy and include:cell therapy and include:  (A) disease states that require gene therapy(A) disease states that require gene therapy(B) only degenerative disease of unknown etiology or (B) only degenerative disease of unknown etiology or autoimmune disease autoimmune disease (C) only age-related degenerative diseases, such as (C) only age-related degenerative diseases, such as Parkinson’s and Alzheimer’s Parkinson’s and Alzheimer’s (D) only cell or organ destruction due to genetic, (D) only cell or organ destruction due to genetic, environmental or infectious disease, such as diabetes environmental or infectious disease, such as diabetes (E) all of the disease states described in the other answers (E) all of the disease states described in the other answers

15. Stem cells give rise to specialized cells residing in organs. 15. Stem cells give rise to specialized cells residing in organs. One of the hallmark characteristics of stem cells is a prolonged One of the hallmark characteristics of stem cells is a prolonged capacity for self-renewal. Another hallmark characteristic of capacity for self-renewal. Another hallmark characteristic of stem cells is their:stem cells is their:  (A) origin exclusively in embryos that are five days old (A) origin exclusively in embryos that are five days old (B) developmental potential (their ability to differentiate in (B) developmental potential (their ability to differentiate in any cell type) any cell type) (C) inability to be genetically engineered (C) inability to be genetically engineered (D) inability to grow in culture (D) inability to grow in culture (E) ability to be cultured back to cloned human beings(E) ability to be cultured back to cloned human beings

16. The production of human embryonic stem cells is 16. The production of human embryonic stem cells is controversial because:controversial because:  (A) they have been used to clone President George W. Bush (A) they have been used to clone President George W. Bush (B) a three month old fetus must be aborted in order to (B) a three month old fetus must be aborted in order to produce them produce them (C) a blastocyst stage embryo usually must be ‘dissociated’ to (C) a blastocyst stage embryo usually must be ‘dissociated’ to recovery ICM (Inner Cell Mass) cells in order to produce them recovery ICM (Inner Cell Mass) cells in order to produce them in culture in culture (D) these types of cells can be used only for reproductive (D) these types of cells can be used only for reproductive cloning cloning (E) they have been shown to cause delusions in NIH (E) they have been shown to cause delusions in NIH researchers about the prospects of receiving federal funding researchers about the prospects of receiving federal funding for this important workfor this important work

17. Stem Cell Research:17. Stem Cell Research:  (A) was banned from receiving federal funding in the US in (A) was banned from receiving federal funding in the US in August 2001 under the Bush Administration until recently but August 2001 under the Bush Administration until recently but is still in contention by many public opponentsis still in contention by many public opponents(B) has resulted in human cloning by a group in the (B) has resulted in human cloning by a group in the Netherlands as reported on the internetNetherlands as reported on the internet(C) is an established science since first applied to a dog skull (C) is an established science since first applied to a dog skull bone in the 1600sbone in the 1600s(D) has no scientific basis and because of this should not (D) has no scientific basis and because of this should not receive federal funding receive federal funding (E) is a scientific ruse developed by a fraudulent researcher in (E) is a scientific ruse developed by a fraudulent researcher in South Korea named Dr. Woo Suk WongSouth Korea named Dr. Woo Suk Wong