More great More great ……..Student questions of the dayStudent questions of the day……..Q: How does the embryo know WHERE to form a tube or other structure?A: There is a connection between positional informationA: There is a connection between positional informationand the formation of specific structures. and the formation of specific structures. The details are poorly understood.The details are poorly understood.Q: What is the relationship between ECM ligandand cellular receptor interaction and focal adhesion?A: The A: The ligand/receptor ligand/receptor interaction causes the interaction causes the focal adhesion.focal adhesion.7.0134.09.07How-to 2Stem CellsSTARTSTARTFOUNDATIONSFOUNDATIONSHow-to 1How-to 1FORMATIONFORMATIONHow-to 2How-to 2SYSTEMSSYSTEMSPROBLEMSPROBLEMSFuture ofFuture ofBIOCHEM GENETICSCELL BIO.MOL. BIOSTEM CELLSREC. DNACELLTYPEFERTIL-IZATIONPRIONSBACTERIAVIRUSESCANCERLIFELIFEIMMUNENERVOUSVISUAL3DSTRUCTUREPurves 48.12:Lung tubules17Lung tubule branching: initial steps18An epithelial sheet can extend to form a tubeExample: primary tracheal tubules19change surface/vol ratio (epiboly)intercalation (convergent extension)Stretching out anepithelial sheetH. Sive MIT 200719aSingle cells can roll or hollow into tubesExamples: secondary and terminal tracheal tubules20epitheliumFGF (ligand)= branchlessFGF inhibitor = sproutygenesPrimary tracheal outgrowth and branching (Drosophila)See Purves 48.5FGF receptor = breathless21O2 stressPrimary tubule Secondary Terminal What are stem cells?Uncommitted cells(undecided)Committed/determined cells(decided)Differentiated cells(final function)Stem cellsH. Sive MIT 2007timeActivation of a fewregulatory genesActivation of manyeffector genesInput(s) 1Input(s) 21“stem cell”(somewhat committed)differentiatedcell type(s)(number measuresstem cell potency)Stem Cell Lineageself-renewingasymmetricdivision“progenitor”(committed)stem cell plus2H. Sive MIT 2007 DiscoveryCell populationPulse/Chase analysis of cell turnover timeAdd BrdU for short time = PULSE(incorporated into DNA) Labeled cell populationBrdU =Bromodeoxyuridine= thymine analogFollow cells over long time= CHASEAssay half-life oflabeled population3H. Sive MIT 2007Cell populationBrdU pulseLabeled cell populationDuring chase…BrdU labeling implies differentiated cellsare derived from progenitor or stem cellsAssay differentiated fate of cells4H. Sive MIT 2007Hematopoeitic stem cells give rise to all blood and immune cell types Blood cellsImmune cells5HSCPlanarian regenerationPlanarian stem cells (neoblasts)6 Isolation and Assay +-Cells labelled with fluorescent antibodiesFluorescence ActivatedCell Sorteris activated by specific fluorescenceCells with specific charge are deflected and collected7Theoretical result of FACS analysis with red and green labeled cellsRed labelGreen labelBoth red and greenNo label8Irradiationdestroys bone marrowand mouse lives!!irradiated mouse would dieBUTNormal bone marrowis injectedBone Marrow Transplant: repopulation assayIsolate morestem cells?9H. Sive MIT 2007In vitro assays for stem cell potencypure-ish stem cellsplate as single cells in culture mediumH. Sive MIT 200710add inducer #2culture (days)add inducer #1culture (days)assay spectrum of differentiated cell typesControl of stem cell fateby the “niche”(surrounding cells)Quiescent stem cellsSurrounding cells(“niche”)maintain stem cellsEnvironmental inputActivated stem cellsdivide to form… Progenitor cellschanges surrounding cellsSurrounding cellsinduce stem cellactivation(and renew stem cells)11H. Sive MIT 2007Hair follicle: the bulge is a source of stem cells12Dr. E. FuchsBulge cells transplantedinto a “nude” mouse strainform hair tufts13Dr. E. Fuchsgrowthregressioninductionbegin newgrowthHair follicle “niche”: dermal papilla induces bulge (stem cell)Wnt signaling(!-catenin!)14Dr. E. FuchsEmbryonic stem cellsBlastula stage mammalian embryo:inner cell mass (ICM) gives rise to whole embryoand is used as embryonic stem cell sourceICM15Purves 19.6:Making ES cellsEach colony may forma stem cell line:can have different potencies16Purves 19.6 A stem cell line may be induced to form various cell typesthat can repair different organs17Stem cell
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