发育生物学_embryonic stem cell_F1.ppt

1、Stem Cell,By Wei-Jun Cai Dept. of Histology 292(5819): 154-6. Develop indefinitely in culture, normal karyotype,1995- from a non-human primate rhesus monkey embryos 1998 - first extract stem cells from human embryos,Embryonic Stem Cell Histoly,James Thomson,In 1998, James Thomson (University of Wisc

2、onsin-Madison) isolated cells from the inner cell mass of the early embryo, and developed the first human embryonic stem cell lines.,In 1998, John Gearhart (Johns Hopkins University) derived human embryonic germ cells from cells in fetal gonadal tissue (primordial germ cells). Pluripotent stem cell

3、“lines” were developed from both sources,What Are Embryonic Stem Cell?,Embryonic stem cells (ES cells) are pluripotent stem cells derived from the inner cell mass of the blastocyst, an early-stage embryo. Human embryos reach the blastocyst stage 45 days post fertilization, at which time they consist

4、 of 50150 cells,The Characteristics Of Embryonic Stem Cell,ESC are distinguished by two distinctive properties:,their ability to replicate indefinitely their pluripotency,able to differentiate into all derivatives of the three primary germ layers: ectoderm, endoderm, and mesoderm. These include each

5、 of the more than 220 cell types in the adult body.,The Characteristics Of Embryonic Stem Cell,Inner cell mass as a niche,ES cells can make tumors if in wrong environment,嵌合胚,An Overview of Early Development,Two kinds of stem cells from animals and humans:,In the 1960s, researchers discovered that t

6、he bone marrow contains at least two kinds of stem cells Hematopoietic stem cells, forms all the types of blood cells in the body Bone marrow stromal cells (bone, cartilage, fat, and fibrous connective tissue) In the 1990s - scientists agreed that the adult brain does contain stem cells that are abl

7、e to generate the brains three major cell types astrocytes and oligodendrocytes (non-neuronal cells) neurons, or nerve cells,An undifferentiated cell found among differentiated cells in a tissue or organ can renew itself can differentiate to yield the major specialized cell types of the tissue or or

8、gan The primary roles in a living organism are to maintain and repair the tissue Some scientists now use the term somatic stem cell instead of adult stem cell Unlike embryonic stem cells, which are defined by their origin, the origin of adult stem cells in mature tissues is unknown Brain, bone marro

9、w, peripheral blood, blood vessels, skeletal muscle, skin and liver,Labeling the cells in a living tissue with molecular markers and then determining the specialized cell types they generate Removing the cells from a living animal, labeling them in cell culture, and transplanting them back into anot

10、her animal to determine whether the cells repopulate their tissue of origin Isolating the cells, growing them in cell culture, and manipulating them, often by adding growth factors or introducing new genes, to determine what differentiated cells types they can become,Also, a single adult stem cell s

11、hould be able to generate a line of genetically identical cellsknown as a clonewhich then gives rise to all the appropriate differentiated cell types of the tissue,Cont.,Hematopoietic stem cells give rise to all the types of blood cells: red blood cells, B lymphocytes, T lymphocytes, natural killer

12、cells, neutrophils, basophils, eosinophils, monocytes, macrophages, and platelets Bone marrow stromal cells (mesenchymal stem cells) give rise to a variety of cell types: bone cells (osteocytes), cartilage cells (chondrocytes), fat cells (adipocytes), and other kinds of connective tissue cells such

13、as those in tendons,Neural stem cells in the brain give rise to its three major cell types: Neurons and two categories of non-neuronal cellsastrocytes and oligodendrocytes Epithelial stem cells in the lining of the digestive tract occur in deep crypts and give rise to several cell types: Absorptive

14、cells, goblet cells, Paneth cells, and enteroendocrine cells Skin stem cells occur in the basal layer of the epidermis and at the base of hair follicles. The epidermal stem cells give rise to keratinocytes, which migrate to the surface of the skin and form a protective layer The follicular stem cell

15、s can give rise to both the hair follicle and to the epidermis,Hematopoietic stem cells may differentiate into: Three major types of brain cells (neurons, oligodendrocytes, and astrocytes) Skeletal muscle cells Cardiac muscle cells and Liver cells Bone marrow stromal cells may differentiate into: Ca

16、rdiac muscle cells and Skeletal muscle cells Brain stem cells may differentiate into: Blood cells and skeletal muscle cells,Plasticity of adult stem cells,Adult stem cells,Embryonic stem cells,Typically reside in or near their tissue,Are capable of giving rise to functional cells in their tissue (bu

17、t not other tissue types) Are typically found only in tissues that undergo regular turnover.,Decrease in both number and activity as one ages.,Created from inner cell mass cells that exist only in very early embryos,Are capable of giving rise to all of the cell types in the body including non-regene

18、rative,Can be divided many times (possibly indefinitely) in culture to make many cells.,Embryonic or Adult Stem Cells for Cell Replacement Therapy Advantages and Disadvantages,Embryonic SC “Pluripotent” Stable. Can undergo many cell divisions. Easy to obtain but blastocyst is destroyed (Ethics) Poss

19、ibility of immune rejection High potential for tumours,Adult SC “Multipotent” Less Stable. Capacity for self-renewal is limited. No ethical concerns Difficult to isolate in adult tissue. Host rejection minimized or absent Less tumorigenic potential,For those who believe that the embryo is a human pe

20、rson from the moment of conception, destruction of the embryo is equivalent to murder But the stem cells obtained from human embryos offer great hope for curing diseases Question: Is there any way of obtaining human embryonic stem cell without destroying or harming human embryos?,The other sources o

21、f stem cells,Therapeutic Cloning,Clone baby,A sexual reproduction,History of Animal Cloning,Amphibian (pollywog) (1962),1973年，获第一批carp(鲤鱼)和鲫鱼(crucian)核移植鱼 （童第周等，动物学报）；,History of Animal Cloning,1996年7月5日，乳腺上皮细胞克隆绵羊“Dolly”诞生， 哺乳动物体细胞克隆里程碑,History of Animal Cloning,Since then, 13 species of animals in

22、cluding mice (1998), cows (1998), pigs (2000), cats (2001), and rabbits (2002) were successfully cloned.,RABBIT,Cattle,horse,How Successful Was Animal Cloning? Very low (1-3%),自1997年“多莉”羊问世之后，全球陆续有几个实验室开展人类体细胞核移 植研究(陆长富等，2000；Cibelli et al., 2001；Lavoir et al., 2005；Heindrychx et al., 2007； Hall et

23、al., 2007；Fan et al., 2009 )。 2000年，中南大学生殖与干细胞工程研究所首次报道人类体细胞核移植胚胎可发育至桑椹胚； 2001年，美国先进细胞公司 Ciblelli 等报道人核移植胚胎可发育至6-c； 2003年，南大学生殖与干细胞工程研究所再次报道人类体细胞核移植胚胎能发育至囊胚； 后来陆续有几个实验室发表文章获得人类核移植囊胚，但获得囊胚数量极少（1-5枚）（Lu et al, 2003, Strojkovic et al., 2005, French et al., 2008, Li et al., 2009, Yu et al., 2009 , Xu et

24、 al, 2011）。,人类体细胞核移植技术进展？,The other sources of stem cells,Alternative sources of pluripotent stem cells from amnitotic fluid,The process of deriving stem cell lines from amniotic fluid,Proposed by Landry and Zucker Landry, D. W. and H. A. Zucker, “Embryonic death and the creation of human embryonic

25、stem cells,” The Journal of Clinical Investigation 114, 1184-1186 (2004). “organismic death for the early-stage human embryo: the irreversible loss of the capacity for continued and integrated cellular division, growth and differentiation.” Extracting stem cells from an embryo that is organismically

26、 dead would not harm the embryo, since it would already be dead.,Howard Zucker,The other sources of embryonic stem cells,Dead embryos,Such entities in fact sometimes occur in naturethey are know as teratomasand they can never develop into full human beings even though the teratomas may show evidence

27、 of certain human characteristics such as fingernails. Typically these are viewed as freakish and some critics have maintained that this solution is morally and aesthetically repulsive.,Teratomas,The other sources of embryonic stem cells,Currently, to perform preimplantation genetic diagnosis (PGD),

28、 doctors extract a single cell from the early embryo. Can we do the same in order to then use the cell to develop hESCs?,Removing a single Cell,The other sources of embryonic stem cells,The Induced Pluripotent Stem (iPS) Cell,iPS : Induced pluripotent stem (iPS) cells, which are functionally compara

29、ble to embryonic stem (ES) cells. iPS cells are generated from somatic cells.,iPS cells generation in patient fibroblasts,Parkinsons disease (Wernig and Jaenisch, 2008, Maehr and Melton PNAS 2009). Amyopathic Lateral Sclerosis, (Dimos and Eggan Science 2008) Type I diabetes (Maehr and Melton PNAS 20

30、09) ADA-SCID, SBDS, Gaucher disease, Duchenne and Becker Muscular dystrophi, Huntington disease, JDM, Down syndrome, Lesch-Nyhan syndrome. (Park and Daley Cell 2008).,iPS cells generation from other cell types,Blood cells (Loh and Daley 2009). B-cells (Hanna and Jaenisch Cell 2008) Blood stem cells

31、(Emiinli and Hochedlinger Nat Genet 2009) Pancreatic b-cells (Stadtfeld and Hochedlinger Cell Stem Cell2008) Hepatic and gastric endoderm (Aoi and Yamanaka Science 2008) Neural stem cells (Kim and Scholar, Nature 2008),iPS cell reprogramming: Problems,Use of viral vectors for induction Low efficienc

32、y of reprogramming Risk of tumour formation Efficient differentiation protocols required,To test new drugs. Tested for safety on differentiated cells generated from human pluripotent cell lines Cancer cell lines, for example, are used to screen potential anti-tumor drugs The generation of cells and

33、tissues which can be used for cell-based therapies Replacement cells and tissues to treat diseases Parkinsons and Alzheimers diseases Spinal cord injury Stroke, burns Heart disease, Diabetes Osteoarthritis, and Rheumatoid arthritis,Approximately one billion heart cells are lost during a serious hear

34、t attack. If one survives the initial event, a progressive chronic heart failure often occurs due to the replacement of functional beating heart cells with a tough fibrous scar,Substantia Nigra and Parkinson Disease,Parkinson disease vs,normal,Alzheimers Disease & the Brain Loss of nerve cells withi

35、n brain “shrunken, shriveled” Hippocampus, basal forebrain and cortex are affected,AD,Normal,Barriers to bringing hESCs to clinic,Changes in their epigenetic profiles,Chromosomal aberrations during their establishment and maintenance,Post transplantation challenges like risk of tumors,Immune-rejecti

36、ons,Fate decision,How do stem cells develop into a functional population of specialized cells ?,50,Thank YOU!,Essentially involves tricking a human egg into thinking it had been fertilized when it had not The egg would then develop to the 50-100 cell stage, at which point hESCs could be extracted Ar

37、e these really embryos? Could they actually develop as human beings? There is no way to answer that question without implanting the embryo, and this is itself a morally dangerous step,The other sources of embryonic stem cells,Paethenogenosis,Can we know that the IVF embryos are really dead? Will the

38、 screening to find dead embryos itself harm some embryos?,Will this be an incentive to produce even more embryos than necessary for IVF? Will this in fact yield stem cells of sufficient quality?,Will this harm the embryo? Are the removed cells not themselves equivalent to embryos? Can one research t

39、his issue without harming embryos?,Pluripotent Stem Cells Derived from Organismically Dead Embryos (Landry-Zucker Proposal) Pluripotent Stem Cells via Blastomere Extraction from Living Embryos Pluripotent Stem Cells Derived from Biological Artifacts Pluripotent Stem Cells via “Parthenogenesis.” Amni

40、otic Fluid that contains embryotic stem cells Pluripotent Stem Cells Derived from “cloning”,Six Possible Sources for ESC,Takahashi and Yamanaka, Cell, Aug 25, 2006,therapeutic cloning,Goal is to make genetically matched ES cells Proof-of-concept in mice allows somatic (not germline) gene/cell therap

41、y,Rideout WM et al. 2002. Correction of a genetic defect by nuclear transplantation and combined cell and gene therapy. Cell 109: 17-27.,A list of animal injury and disease models where hESCs have been shown to be effective,Risk of tumors,Transplantation of hES cell based therapies involves the risk

42、 of tumor formation arising from undifferentiated population of the transplanted cells. Studies with both ESCs and ES derived differentiated cells have shown that they can form teratocarcinomas in adult mice if injected subcutaneously, intramuscularly or into the testis. Presence of even one undiffe

43、rentiated cell may potentially lead to teratomas, a cancerous tumor which is derived from germ cells and can from all the three germ layers.,Genetic instability,Questions on the suitability of ESCs for transplantation purpose is raised because of the observed genetic instability of cloned cells and extreme inefficiency of the process. Cloned animals like Dolly give the outward appearance of full health, but the probability of their having numerous genetic defects is very high. Hochedlinger and Rudolf Jaenisch (2002) showed that in mice, the reprogramming of the inserted genetic