Cleavage to gastrulationCleavage to gastrulation

M.A.Kai-KaiM.A.Kai-Kai

Learning Objectives.Learning Objectives.

Define major Define major mechanismsmechanisms of development. of development.Describe the mammalian Describe the mammalian zygotezygote..Understand the process of mitotic division/Understand the process of mitotic division/cleavage cleavage of the of the zygote zygote

Understand process of Understand process of blastulation blastulation Understand the process of Understand the process of gastrulation gastrulation and the functions and the functions i.e. formation of the embryonic three i.e. formation of the embryonic three germ germ layerslayersectoderm, endoderm, mesoderm.ectoderm, endoderm, mesoderm.

Overview of the adult Overview of the adult derivativesderivatives of the germ layers. of the germ layers.

Mechanisms of DevelopmentMechanisms of Development1.1.MitosisMitosis and and growthgrowth differential mitosis and growth differential mitosis and growthbody formbody form--cell proliferation e.g.in cleavage--cell proliferation e.g.in cleavageincrease in number, cells smaller in size.increase in number, cells smaller in size.--growth by deposition of extracellular matrix,intracellular organelles and matrix.--growth by deposition of extracellular matrix,intracellular organelles and matrix.--spread of epithelial sheets e.g. extraembryonic membranes. --spread of epithelial sheets e.g. extraembryonic membranes. 2.2.RestrictionRestriction and and determinationdeterminationtotipotent blastomerestotipotent blastomeresrestricted potencyrestricted potency3.3.Gene activationGene activation and differential and differential expressionexpression of functional genes. of functional genes.4.4.DifferentiationDifferentiatione.g.cytodifferentiation into specific phenotype.e.g.cytodifferentiation into specific phenotype.5.5.Cell/tissue interactionCell/tissue interactionmediate embryonic signals e.g.primary induction-mediate embryonic signals e.g.primary induction-notochord.notochord.

6.6.CellCell movementmovementshort or long migrations, e.g. neural crest cells.short or long migrations, e.g. neural crest cells.7. 7. PatternPattern formationformationintrinsic blueprint directing development.intrinsic blueprint directing development.8.8.FoldinFoldingge.g.cephalic, caudal and lateral body foldse.g.cephalic, caudal and lateral body foldsbody formbody form9. 9. MorphogenesisMorphogenesiscumulative mechanisms transform internal and external body cumulative mechanisms transform internal and external body form.form.

EMBRYOLOGYCELL MOTILITY AND SHAPE CHANGE

CELL SIGNALLING

EXTRACELLULAR MATRIX

CELL-CELL AND CELL MATRIX INTERACTIONS

CONTROL OF GENE EXPRESSION

CONTROL OF CELL DIVISION

STEM CELLS

APOPTOSIS

LIMBS

NERVOUS SYSTEM

CARDIOVASCULAR SYSTEM RESPIRATORY

SYSTEM

URINARY SYSTEM

DIGESTIVE SYSTEM

REPRODUCTIVE SYSTEM

Cell movement, changing cell interactions, cell division, changing patterns of gene expression are all quite restricted in the adult but are rampant in the embryo.

The ZygoteThe Zygote The The zygotezygotea single cell formed at a single cell formed at fertilisation.fertilisation.Structure.Structure.

----diploiddiploid nucleus from both parents. nucleus from both parents. --cytoplasm is maternal.--cytoplasm is maternal.

--surrounded by zona pellucida--surrounded by zona pellucidaCleavageCleavagemitotic division at mitotic division at 12hours, 2days and 3days in pig and 12hours, 2days and 3days in pig and dog. Rate about one division/day.dog. Rate about one division/day.Zygote period lasts from fertilisation Zygote period lasts from fertilisation to hatching of the to hatching of the blastocystblastocyst..NutritionNutrition/embryotroph /embryotroph

--mammalian zygotes is provided by --mammalian zygotes is provided by uterine secretions/histiotroph and the uterine secretions/histiotroph and the zygotes own reserve. zygotes own reserve.

--avian zygotes feed on the yolk.--avian zygotes feed on the yolk.

CleavageCleavageCleavage is series of Cleavage is series of mitotic divisionmitotic division of the zygote. of the zygote.Rapid cell divisionRapid cell divisionsubdivide large zygote into progressively subdivide large zygote into progressively smaller cellularsmaller cellular units--blastomeresunits--blastomeres

--cells--cells morphological morphological and and metabolicalmetabolical unspecialised unspecialisedFirst cleavage synchronous.First cleavage synchronous.Differences in pattern of cleavage dependent upon amount of yolk.Differences in pattern of cleavage dependent upon amount of yolk.TotipotentTotipotent early early blastomeresblastomeres(Hans Driesch(1893).(Hans Driesch(1893).

Unique Features of Mammalian CleavageUnique Features of Mammalian Cleavage11stst cleavage cleavagemeriodal(2-cells)meriodal(2-cells)22ndnd cleavage cleavageone one meriodionalmeriodional and one and one equatorialequatorialtermed rotational termed rotational cleavagecleavageSecond cleavage not synchronised in Second cleavage not synchronised in blastomeresblastomeresSlow rate, first cleavage last to 24hour, subsequent divisions 10-12h.Slow rate, first cleavage last to 24hour, subsequent divisions 10-12h.Zygote genome switched on early at 2-cell stageZygote genome switched on early at 2-cell stageCompactionCompaction occurs at 8-cell stage. occurs at 8-cell stage.--blastomeres flatten and form intercellular connections--blastomeres flatten and form intercellular connections--E-CAD(cadherin:glycoprotein) on cell surface--E-CAD(cadherin:glycoprotein) on cell surfaceadhesion.adhesion.--microvilli(actin) extend on surfaces of adjacent cells and anchor cells --microvilli(actin) extend on surfaces of adjacent cells and anchor cells together.together.--tight junctions prevent free exchange of fluid between the inside and outside --tight junctions prevent free exchange of fluid between the inside and outside allowing accumulation of fluid inside blastomeres.allowing accumulation of fluid inside blastomeres.--the gap junctions couple all the blastomeres of the compacted embryo and --the gap junctions couple all the blastomeres of the compacted embryo and permit exchange of ions and small molecules from one cell to the next.permit exchange of ions and small molecules from one cell to the next.

The MorulaThe MorulaMorula; Morula; 16-cell16-cell stage, embryo stage, embryo enclosed in the zona pellucida.enclosed in the zona pellucida.Late morula, first differentiation Late morula, first differentiation event in mammalian development. event in mammalian development.

--cells aggregate into internal --cells aggregate into internal inner inner massmass cells(ICM) and external cells(ICM) and external trophoblasttrophoblast..

--at --at 64-cell stage64-cell stage ICM and ICM and trophoblast form distinct populations.trophoblast form distinct populations.Trophoblast/trophoectodermTrophoblast/trophoectodermforms forms ectoderm of chorion/ placenta.ectoderm of chorion/ placenta.Inner mass cells:Inner mass cells:

--internal cells--internal cellspluripotentpluripotent; form ; form embryo embryo and partly and partly extraembryonicextraembryonic membranes. membranes. BlastogenesisBlastogenesis starts at 32-cell starts at 32-cell stage.stage.

0

10

20

30

40

SEGMENTATION/ SOMITE FORMATION

BLASTULATION

GASTRULATION/ NEURAL TUBE FORMATION

FORMATION OF BODY FOLDS AND EXTRA- EMBRYONIC MEMBRANES

EARLY ORGANO-GENESIS

FETAL PHASE

DAYS

IMPLANTATION

5 mm

10 mm

30 mm

50 mm

114

A TIMELINE - PIG DEVELOPMENT

Size of embryo

The embryonic phase contains both series and parallel components

In the mid to large size domestic mammals the embryonic phase lasts 30-40 days and the fetal phase varies with the size and maturity of the neonate

BlastulationBlastulation Formation of the blastocystFormation of the blastocyst--A series of rapid cell divisions produce

a blastula of 64 cells with an inner cell mass and an outer layer of trophoblasts.

--trophoblast secrete fluid into the --trophoblast secrete fluid into the morula. Creates a cavity; the morula. Creates a cavity; the blastocoele(A)blastocoele(A)

--ICM/embryo proper lies to one side--ICM/embryo proper lies to one side Transition from morula to blastula Transition from morula to blastula

marked by:marked by:--rapid enlargement of blastocoele--rapid enlargement of blastocoele--differentiation of blastomeres into ICM --differentiation of blastomeres into ICM

and trophoblast cells(B).and trophoblast cells(B). Trophoblast cells induce special Trophoblast cells induce special

changes in the uterine lining at changes in the uterine lining at implantation.implantation.

Trophoblast cells preferentially Trophoblast cells preferentially express maternal genes, and express maternal genes, and inactivate paternal genes.inactivate paternal genes.

The blastula hatches from the zona pellucida and implants on the uterine epithelium

segregation

Trophoblast

ICM

Formation of hypoblastFormation of hypoblastand epiblastand epiblast..

Formation of hypoblast begins late in Formation of hypoblast begins late in blastulation blastulation

1.1. Segregation(A)Segregation(A)

2.2. Delamination Delamination of ICM cells.Cells expand of ICM cells.Cells expand beneath trophoblast form beneath trophoblast form hypoblasthypoblast(B)/(B)/extraembryonic endodermextraembryonic endoderm

Hypoblast tube(Hypoblast tube(blastocoeleblastocoele) inside tube ) inside tube of trophoblast(C)of trophoblast(C)

Formation of hypoblast results in two-Formation of hypoblast results in two-layered embryo(C) layered embryo(C)

Cells on surface ICM form Cells on surface ICM form epiblastepiblast(C)(C)

Blastula: Hatching and Implantation.Blastula: Hatching and Implantation.BlastulationBlastulation embryo arrives in uterus(4) embryo arrives in uterus(4) Blastula Blastula hatcheshatches from from zona pellucidazona pellucida and contact uterus. and contact uterus.Blastocyst surrounded by ZP prevents premature implantation and ectopic Blastocyst surrounded by ZP prevents premature implantation and ectopic pregnancy.pregnancy.Hatching involvesHatching involvesA trypsin-like protease lyses of ZPA trypsin-like protease lyses of ZP

--Trophoblast cells secrete proteases which degrades the endometrial wall and --Trophoblast cells secrete proteases which degrades the endometrial wall and blastocyst embeds(5).blastocyst embeds(5).

1

2

3 4 4

5

Stem cellsStem cellsProperties Properties of stem cells.of stem cells.Pluripotent Pluripotent embryonic stem adult stem cells.embryonic stem adult stem cells.EmbryonicEmbryonic stem cells(ESC) from inner mass cells, stem cells(ESC) from inner mass cells,totipotenttotipotent can give rise to can give rise to all embryonic cells except trophoblast.all embryonic cells except trophoblast.ectoderm, endoderm and ectoderm, endoderm and mesoderm.mesoderm.ESCESC form form pluripotent stem cellpluripotent stem cellcommittedcommitted stem cell then stem cell then progenitorprogenitor/precursor cells/precursor cellsdifferentiate differentiate into a cell lineage.into a cell lineage.Example Example haemangioblasthaemangioblastmultipotent haematopoietic stem cellmultipotent haematopoietic stem cellmyeloid myeloid progenitor cellprogenitor cellblood cells.blood cells.AdultAdult stem cells more lineage-restricted in their ability to differentiate stem cells more lineage-restricted in their ability to differentiateRestriction Restriction on the on the potencypotency of stem cells is determined by the of stem cells is determined by the microenvironmentmicroenvironmentAdult stem cells important for the continual production of tissues e.g Adult stem cells important for the continual production of tissues e.g intestinal epithelium. intestinal epithelium. Inner massInner mass cells also used for creating cells also used for creating chimericchimeric animals. animals. Expression of Expression of transcription factor transcription factor e.g Oct-4 is required for the maintenance e.g Oct-4 is required for the maintenance of pluripotency.of pluripotency.Murine embryonic stem cells useful models to elucidate the unique Murine embryonic stem cells useful models to elucidate the unique properties of mammalian stem cells. properties of mammalian stem cells. Valuable therapeutic potential in treatment of degenerative diseases. Valuable therapeutic potential in treatment of degenerative diseases.

Gastrulation(1)Gastrulation(1)Gastrulation transforms flat two-layered Gastrulation transforms flat two-layered blastula(epiblast,hypoblast) into three-layered blastula(epiblast,hypoblast) into three-layered gastrulagastrulaectoderm, endoderm, mesoderm.ectoderm, endoderm, mesoderm.

Mechanism of gastrulation consists of:Mechanism of gastrulation consists of:

11..FormationFormation of of primitive streakprimitive streak, marked by:, marked by:

----expansion expansion of of epiblast epiblast cells and caudal cells and caudal convergenceconvergence(A).(A).

--Primitive streak(PS) forms as longitudinal ridge --Primitive streak(PS) forms as longitudinal ridge in midline(B)in midline(B)

--PS elongates, cranial tip widened as primitive --PS elongates, cranial tip widened as primitive node(C)node(C)

Gastrulation(2)Gastrulation(2)

22. . InvolutionInvolution of primitive streak(PS) of primitive streak(PS)

--epiblast cells leave PS --epiblast cells leave PS

--primitive groove in midline--primitive groove in midline

----firstfirst group of cells form intraembryonic group of cells form intraembryonic endoderm(endoderm(AA))

-- --second group of epiblast cells -- --second group of epiblast cells intercalateintercalate between endoderm and between endoderm and ectodermectodermform form mesodermmesoderm

33. PS . PS regressesregresses caudally caudally

4. Enlarged tip Of PS4. Enlarged tip Of PSHensen’s Hensen’s nodenode/primitive knot, moves to posterior /primitive knot, moves to posterior region(C).region(C).

44.Function of PS is to form three germ .Function of PS is to form three germ layerslayers

A

B

A

B

C

AA

B

C Hensen’s node

Cranial

Caudal

Formation of the NotochordFormation of the NotochordThe The NotochordNotochord--a rod-shaped aggregate --a rod-shaped aggregate of of chordamesodermchordamesoderm cells cells extending along entire extending along entire length of embryo.length of embryo.--notochord cells formed of --notochord cells formed of migrating cells from migrating cells from Hensen’s nodeHensen’s nodeFunctionsFunctions of notochord. of notochord.1.Defines1.Defines cranial-caudal cranial-caudal axis of embryo.axis of embryo.2. Serves as 2. Serves as primary primary inducerinducer at at neurulation.Induces neurulation.Induces formation of the neural tube formation of the neural tube and somitogenesis.and somitogenesis.3. 3. TransientTransient, remnants in , remnants in intervertebral disc as intervertebral disc as nucleus pulposus.nucleus pulposus.

Embryo proper

Transverse through primitive streak

FORMATION OF THE MAMMALIAN GASTRULA - 3

Migration through the primitive streak forms a three-layered embryo

Primitive streak

Primitive gut

Ectoderm

Mesoderm

Endoderm

The 3 germ layers

Extra-embryonic

Splanchnopleure

Somatopleure

Transverse section through a mammalian embryo(14days in pig)

Splanchnopleure

Trophoectoderm

Mesoderm

Endoderm/hypoblast

Extraembryonic coelom

Junctional line

Periods of Development of ConceptusPeriods of Development of Conceptus Embryo implants by end of Embryo implants by end of blastogenesisblastogenesis

EmbryogenesisEmbryogenesis is period of differentiation and is period of differentiation and organogenesisorganogenesis

Major organs and systems formed, morphogenesis is later.Major organs and systems formed, morphogenesis is later.

Period ends with formation of the primordial nervous system, circulatory Period ends with formation of the primordial nervous system, circulatory system digestive and excretory systems and limb buds.system digestive and excretory systems and limb buds.

The embryo is a miniature adult of the species.The embryo is a miniature adult of the species.

Period is most vulnerable to Period is most vulnerable to teratogenic teratogenic malformationsmalformations

High mortality rate due to nutritional deficiency, excessive ambient High mortality rate due to nutritional deficiency, excessive ambient temperature temperature

and infections.and infections.

Embryonic deaths occur, about 15% in the queen cat, bitch and horses Embryonic deaths occur, about 15% in the queen cat, bitch and horses

SummarySummaryThe zygote goes through different stages of development by specific The zygote goes through different stages of development by specific morphogenetic processes.morphogenetic processes.Cleavage, a series of mitotic divisions produces a morulaCleavage, a series of mitotic divisions produces a morulaBlastulation forms the blastula which hatches from the zona pellucida and Blastulation forms the blastula which hatches from the zona pellucida and implants in the wall of the uterus. implants in the wall of the uterus. Gastrulation transforms the flat blastula into a three-layered gastrulaGastrulation transforms the flat blastula into a three-layered gastrulaThe process of gastrulation involves formation of the primitive streak by The process of gastrulation involves formation of the primitive streak by convergence of epiblast cells towards the midline and involution of the streak convergence of epiblast cells towards the midline and involution of the streak to form the endoderm and mesoderm. The surface epiblast cells form the to form the endoderm and mesoderm. The surface epiblast cells form the ectoderm. ectoderm. The notochord forms from mesoderm in Hensen’s node and acts as primary The notochord forms from mesoderm in Hensen’s node and acts as primary inducer of neurulation.inducer of neurulation.Organogenesis starts after neurulation.All adult organs are formed from the Organogenesis starts after neurulation.All adult organs are formed from the three germ layers.three germ layers.Periods of development arePeriods of development areblastogenesis, embryogenesis and the fetal blastogenesis, embryogenesis and the fetal stages.The entrance and duration of each stages in the species.stages.The entrance and duration of each stages in the species.

ReferencesReferences1.1. Carlson, B. M., Foundations of Embryology (6Carlson, B. M., Foundations of Embryology (6thth.Edition) 1996..Edition) 1996.

McGraw-Hill inc. London. Page 151 – 226McGraw-Hill inc. London. Page 151 – 226

2.2. Gilbert, S.F., Developmental Biology (8Gilbert, S.F., Developmental Biology (8thth. Edition) 2006. Sinauer . Edition) 2006. Sinauer Associates Inc. Sunderland, Massachuetts. USA. Page 348 - 354Associates Inc. Sunderland, Massachuetts. USA. Page 348 - 354

3.3. McGeady, T.A., Quinn, P.J., Fitzpatrick, E.S., & Rayan, M.T., McGeady, T.A., Quinn, P.J., Fitzpatrick, E.S., & Rayan, M.T., (2006).(2006).

Veterinary Embryology. Page 25 – 38.Veterinary Embryology. Page 25 – 38.

4.4. Noden, D.M., DeLaHunta, A., The Embryology of Domestic Animals.Noden, D.M., DeLaHunta, A., The Embryology of Domestic Animals.

1985, Williams & Wilkins. London. Page25 – 27, 33-40.1985, Williams & Wilkins. London. Page25 – 27, 33-40.