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DB-Lec 12

DB-Lec 12 - Biology 442 Biology Developmental Biology...

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Unformatted text preview: Biology 442 Biology Developmental Biology Developmental Lecture 12 Gastrulation Gastrulation marks the onset of Morphogenesis. The cells of the embryo undergo movements that reorganize the 3 germ layers to generate the embryo’s organ rudiments the 5 questions central to morphogenesis that one should think about as we look at gastrulation and organogenesis. How are tissues formed from populations of cells ? How How are organs formed from tissues How are organs formed in particular locations and how How do migrating cells reach their destinations ? do How do organs and their cells grow and how is their How growth coordinated with development ? Or How do they know what size an organ or tissue should become ? know How do organs achieve polarity ? – An arm/hand contains the same tissues (bone, muscle, cartilage, skin, but the hand looks different from the arm. 2 Major type of cell arrangements in the embryo. Epithelial cells – cells tightly connected to each other in Epithelial sheets or tubes. Usually they rest on a basement membrane and form tight junctions at the apical or outer surface. Some epithelia consist of a single sheet others are stratified (consist of multiple layers). The outer layers are connected by tight junctions while the inner layers are connected by gap junctions. are Mesenchymal cells – Aggregates of single cells surrounded by large amounts of extracellular matrix material. material. Morphogenetic Movement of Epithelia Cells Morphogenetic Invagination – Inward “buckling” of an epithelium to create a depression. Involution – Inward Involution movement of an expanding epithelium around an edge. epithelium Convergent extension – Elongation of an epithelium in 1 dimension while shortening in the other. shortening Epiboly – spreading of an Epiboly epithelium to envelop the mass of cells or yolk. mass Delamination – splitting of 1 Delamination layre of cells into 2 parallel layers layers Passive Movements – epithelia pushed or dragged epithelia Migration – single cells Migration move over substrate. move Intercalation – cells move in between each other – lateral (in same layer) or lateral radial (between adjacent layers). layers). Ingression – transition of single cells from an epithelium into a cavity. epithelium Shape Changes – can cause layers to buckle or elongate. elongate. Cell division – orientation, size. size. Changes in Adhesion of cells. cells. Programmed cell death – can carve or sculpt organs. can Epithelia and mesenchyme cells can interconvert: epithelial-mesenchymal transitions are common . They transitions involve some of the following types of movements and can change the shape and organization of epithelial layers. The objective of these cell movements of gastrulation is to bring the endodermal cells to the interior, spread the endodermal ectodermal cells in an outer layer surrounding the embryo and ectodermal place the mesodermal cells in the middle between the mesodermal ectodermal and endodermal layers. ectodermal Gastrulation starts with the formation of the blastopore, an indentation or grove through which cells move inside to form the mesoderm and endoderm. The fate of the blastopore tissue is used as a basis for classifying animals into 2 groups: Protostomes (blastopore becomes mouth) and Deuterostomes Deuterostomes (blastopore becomes anus). We will look at gastrulation in 5 Dictyostelium model organisms: 1. A protostome – sea urchin 2. Deuterostomes – amphibians 3. Zebra fish 4. Chickens 5. Mammals Sea Urchin Gastrulation Sea Blastula is a single layered epithelia of ~1000 cells surrounding a fluid filled blastocoel cavity. The first step in gastrulation is a change in cell shape: blastmeres in cell the vegetal plate become tall and columnar. Next these cells change their adhesions, breaking the tight junctions that seal them to neighboring cells and they ingress into the blastocoel ingress and become primary mesenchyme. Epitheal to mesenchyme transition In changing their adhesion the ingressing mesenchyme cells decrease their adhesion to the hyaline layer and neighboring cells and increase their adhesion to the basement membrane on the inside of the blastula. As the primary mesenchyme cells migrate, the remaining vegetal plate cells start to invaginate to form the Archenteron. invaginate Archenteron Archentron = primative gut, opening is the blastopore that becomes the mouth. Invagination starts with the vegetal plate cells becoming bottle shaped. The hyaline layer buckles because of changes in the composition of one of the 2 lamina that make up the hyaline layer. At the time of invagination the vegetal plate cells secrete a chondroitin sulfate proteoglycan (CSPG) proteoglycan into the inner lamina of the hyaline layer directly underneath them. CSPG is hygroscopic (water absorbing) and swells the inner lamina but not the outer. This causes the vegetal region of the hyaline layer to buckle and start the invagination of the archentron 2 forces drive continued extension of the archenteron into the interior of the blastocoel. Convergent Extension occurs by intercalation of cells to form a longer, narrower tissue Secondary mesenchyme cells form at the tip of the archenteron and extend filopodia which attach to the inner surface of the blastocoel wall and pull the archenteron further into the blastocoel. With Sea Urchin Gastrulation we have seen a number of different types of cell motility and adhesion at work bringing about the reorganization of the embryo. Changes in cell shape Changes in cell adhesion strength Changes Ingression of cells Invagination of cell layers Convergent Extension A situation where individual cells pull a cell situation layer. layer. Gastrulation in Amphibians Gastrulation Gastrulation initiates in the cells derived from the grey crescent. These cells invaginate to form a slit like blastopore The cells undergo a major shape change – they form bottle shaped cells with a constriction at one end. The second phase of gastrulation involves: 1) involution of the marginal cells while involution 2) the animal cells undergo epiboly spreading animal over the entire embryo and converging at the blastopore. 2 types of marginal cells 1) superficial and 2) deep marginal cells. The migrating marginal zone cells reach the dorsal lip of the blastopore and turn inward and migrate along the inner surface of the outer animal hemisphere cells. As these marginal zone cells (mesoderm) enter, the archenteron (primative gut) forms and the blastocoel cavity is displaced. This means that the cells that occupy the position of dorsal lip of the blastopore (the organizer) are constantly changing. the A detailed look at the mechanism of involution: 1) Bottle cells initiate movement but are not necessary for continued involution: can remove them after involution starts and no problem ! 2) Active migration of the deep marginal zone cells drives involution. Replacing superficial cells has no effect but replacing deep cells stops involution. 3) Migration of the deep cells requires the ECM protein fibronectin. Cells along the path secrete fibronectin. Antibodies to fibronectin or integrin block migration. Yellow =fibronectin, Red =cells The involuting marginal zone is initially several layers thick. Before involution through the DBL these cells intercalate radially to form a thin broad layer. After the deep cell involute through the DBL they initiate a 2ond intercalation that causes convergent extension along the mediolateral axis. Differential cell adhesion drives convergent extension. Cells undergoing convergent extension express protocadherins. Dominant negative protocadherin blocks convergent extension. Epiboly of the Ectoderm (spreading of the ectodermal cells Epiboly over the entire embryo) Is Accomplished by Cell Division and Cell Intercalation. Cell Antibody to Phosphorylated Histone Antibody H3 marks cells undergoing mitosis H3 Here we see gastrulation driven by Here Changes in cell shape Involution of a tissue layer Intercalation leading to convergent extension Epiboly (spreading of a sheet of cells) And we begin to see a role for cell-cell adhesion mediated by cadherins and cell substrate adhesion mediated by integrin receptors binding cells to fibronectin. End of Gastrulation Red = mesoderm Blue = ectoderm Yellow = endoderm Green = cells from bottle cells Gastrulation in Fish, Birds and Reptiles (very similar) Superficial cleavage leaves cluster of cells on top of dense yolk. egg is laid at 20,000 cells and gastrulation starts Space between blastoderm and yolk is subgerminal space space First step is ingression of individual cells from single layer ingression into subgerminal space – these meet up with another group of cells migrating from posterior end of blastoderm and migrating form second layer of cells under the first – hypoblast layer – second hypoblast space between them is blastocoel blastocoel Embryo comes form the Epiblast and the Hypoblast Epiblast Hypoblast generates the Extra embryonic tissues (yolk embryonic sac) and germ cells. germ The primitive streak, a major structure characteristic of birds, primitive reptiles and mammals forms next. It starts as a thickening of epiblast layer at the posterior marginal zone caused by an increase in height of the cells. Convergent extension is responsible for the extension of the primitive streak. During this process a depression, the primitive grove forms. primitive It is the equivalent of the amphibian blastopore and serves as It an opening for cells to enter the blastocoel. an The most anterior end of the primitive streak is Hensen’s node. It is the equivalent of the dorsal blastopore lip or in fish node. embryos, the embryonic shield – It acts as the organizer. embryos, Epiblast cells undergo an epithelial to mesenchyme transition, with the underlying basal lamina breaking down as they migrate as single cells into migrate the primitive grove. This is different from amphibians where cells migrate into the blastocoel as sheets of cells. blastocoel The epithelial to mesenchyme transition is induced by a 190Kd protein (scatter factor), secreted by the cells as they enter the streak. It down regulates cadherin and blocks cadherin its synthesis. The primitive streak extends posterior to anterior defining the axes of the embryo. Cells enter through the dorsal side, move to the ventral side. The first set of cells to ingress through the primitive streak are endodermal precursors. They migrate anteriorly and displace the hypoblast cells in the ventral layer. The second set of cells to ingress also move anteriorly but they remain between the epiblast and the endoderm. These cells will form the mesoderm. While the mesodermal and endodermal cells are migrating inward the ectodermal precursors proliferate and migrate to surround the yolk by epiboly. The enclosure of the entire yolk by the ectodermal cells takes 4 days! At the end of gastrulation – ectodermal cells outside, endodermal cells are inside and mesodermal cells are in the middle. Gastrulation in Mammals Gastrulation Birds and mammals are both descendents of reptilians and mammalian gastrulation is very similar to gastrulation in birds and reptiles which is adapted for yolky eggs. It is surprising that mammals have retained this adaptation even in the absence of large amounts of yolk ! At the end of blastula stage 1) Inner cell mass (ICM) – will form 1) Inner embryo, yolk sac, allantois and amnion (embryonic stem cells are obtained from here at day 5) obtained 2) Trophoblast cells or Trophoblast trophectoderm – these cells will not contribute to the embryo Trophoblast cells Trophoblast 1)secrete protease to hatch the blastula out of the Zona Pellucida so that it can implant in the uterine wall. 2) Secrete proteases to make a hole in the uterine lining to implant the embryo at about 7 days after fertilization 3) make chorion which is the embryonic part of the chorion placenta 1 step in Gastrulation A layer of cells delaminates from ICM and is called hypoblast – this gives two layers of cells like birds The hypoblast does not hypoblast contribute to the embryo but spreads out around inside of blastocoel and forms yolk sac under embryo – yolk sac structure but with no yolk. structure The cells of ICM left in top layer are called epiblast. This epiblast layer splits off a layer on top – these cells form the lining of the cavity above the embryo called the amnion. When it is amnion formed it is filled with amniotic fluid – this serves as shock amniotic absorber for embryo The embryonic epiblast makes all the embryonic structures structures Gastrulation proceeds like in birds – A thickening forms at the posterior end of epiblast layer and forms the primitive streak Epiblast cells migrate through the primitive streak and displace hypoblast and form endoderm. These are followed by cells that will form the mesoderm. This leaves only ectoderm on top – so same three-layered structure as in birds As in birds the ingression of the endodermal and mesodermal cells involves an epithelial to mesenchym transition. The cells loose E-cadherin, detach from their neighbors and migrate through the streak as single cells. Cell migration and specification are coordinated by FGF (fibroblast growth factor – ligands for tyrosine kinase receptors). Cells of the primitive streak synthesize and receptors). respond to FGF. In Embryos homozygous for the loss of the FGF-8 gene, cells fail to migrate through the primitive streak and no mesoderm or endoderm is formed. Changes are also happening in the Trophoblast cells – Represent an evolutionary adaptation so mammals can have internal development Trophoblast cells divide without cell division to from syncytiotrophoblast - this structure extends further into uterus and maternal blood vessels move towards it and into it A stalk of mesodermal cells extends out from embryo and forms embryonic blood vessels – this stalk become umbilical cord The trophoblast and extraembryonic mesoderm together is called chorion. It fuses with the uterine wall to become It placenta. The placenta contains both maternal and embryonic tissues. Functions of chorion 1) Deliver O2 and food from the mother and remove wastes (CO2 and urea) - fetal and maternal circulatory systems never merge. 2) Endocrine organ – secretes hormones a) chorionic gonadotropin (in EP tests) b) Which leads to progesterone secretion by chorion. It b) takes over from corpus luteum and keeps uterine lining rich with blood vessels with c) placental lactogen – breast development for milk c) production production 3) Inhibits immune system rejection of embryo by mother At the end of 3 weeks an embryo with 3 germ layers is ready for organogenesis. People think this works so well because they only see successful or mostly successful outcomes, BUT BUT ■ 75% of human embryos die before week 8 75% ■ 25% in 1st week before implantation (chromosome abnormalities) abnormalities) ■ 25% in second week 25% ■ 25% in next six weeks 25% ■ after 8 weeks, most hard development done, and lots of growth growth ...
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