Introduction to embryology III Petr Vaňhara LECTURE CONTENT 1. Revision of early embryogenesis 2. Trilaminar germ disc embryo 3. Folding of embryo 4. Pregnancy and fetal development 5. Prenatal diagnostics 1 2 3 4 5 8 6 7 Fertilization START! 9 10 11 12 • Adhesion of blastocyst • Differentiation of trofoblast Implantation • End of implantation Embryogenesis • Differentiation of embryoblast • Bilaminar germ disc • Amniotic cavity • Primary yolk sac • Extraembryonic mesoderm 13 14 15 16 17 18 1st week 2nd week Cleavage Blastogenesis 3rd week • Trilaminar germ disc • Differentiation of epiblast • Primitive node and primitive streak • Differentiation of mesoderm • Differentiation of ectoderm and neuroectoderm • Somitogenesis 19 20 21 FIRST EVENTS IN HUMAN LIFE 1st week „inside-outside“ „cell polarity“ • 16-cell embryo is still totipotent – later (32-cell), it loses the full developmental potential → cell are determined. FIRST EVENTS IN HUMAN LIFE DETERMINATION OF BLASTOMERES Mechanism of diferentiation 1st-2nd week Day 5 Day 6 Day 7 1/2 Day 8 Day 9 end of 2nd week hypoblast epiblast syncytiotrophoblast amnion primary yolk sac extraembryonic mesoderm extraembryonic coelom cytotrophoblast lacunae DEVELOPMENT OF GERM DISCFIRST EVENTS IN HUMAN LIFE BILAMINAR GERM DISC 15 16 17 18 19 20 21 22 23 24 25 26 27 28 3. týden Day 16 Primitive node Primitive streak Neural plate Day 18 3rd week begins 2nd week ends 14 NEW STRUCTURES TRILAMINAR GERM DISC PRIMITIVE STREAK AND PRIMITIVE NODE Prechordal plate Neural plate 3rd week TRILAMINAR GERM DISC Notochordal process PRIMITIVE STREAK AND PRIMITIVE NODE Day 17 Day 18 Day 19 3rd week TRILAMINAR GERM DISC PRIMITIVE STREAK AND PRIMITIVE NODE A new cell population appears - MESODERM 3rd week TRILAMINAR GERM DISC PRIMITIVE STREAK AND PRIMITIVE NODE NOTOCHORDDERIVATIVES OF MESODERM End of 2nd week End of 3rd week 3rd week Notochord induces differentiation of ectoderm – cellular basis of nerve system is established – NEUROECTODERM NEUROECTODERMNEURULATION 3rd week NEURAL TUBENEURULATION • Neural plate • Neural folds • Neural tube • Neural crest NEURAL TUBE AND NEURAL CRESTNEURULATION 3rd – 4th week NEURAL TUBE AND NEURAL CRESTNEURULATION NEURAL CREST AND ITS DERIVATIVES • Peripheral nerve system including glia • Enterochromaffin cells • Melanocytes • Craniofacial connective tissue • Odontoblasts • Adrenal medulla • ... NEURAL TUBE INTERVERTEBRAL DISCNOTOCHORD IN ADULT BODY 3rd week Chordomezodermový výběžek MESODERMGASTRULATION SOMITESDERIVATIVES OF MESODERM 3rd – 4th week SOMITESDERIVATIVES OF MESODERM OTHER DERIVATIVES OF MESODERM 3rd – 4th week • heart, cardiovascular system • urogenital system • muscle and skeletal system • hematopoietic and lymphatic systems • connective tissue, dermis • mesothelium DEVELOPMENTAL DISORDERS DURING GASTRULATION • Primitive streak is a temporary embryonic structure. Persistent primitive streak causes sacrococcgyeal teratoma. • Failure of primitive streak leads to absence of mesoderm in affected region - sirenomelia - limbs - urogenital system - GIT DEVELOPMENTAL DISORDERS DURING GASTRULATION • If two primitive streaks form, conjoined twins may develop FOLDING OF EMBRYONEW BODY PLAN 4th week FOLDING OF EMBRYONEW BODY PLAN extraembryonic coelom umbilical vein umbilical artery extraembryonic coelom FOLDING OF EMBRYONEW BODY PLAN Cytotrophoblast Extraembryonic somatopleura Extraembryonic splanchnopleura Connective stalk and allantois Extraembryonic coelom Chorion and chorionic villi Skin umbilicus Neural tube 1 Primitive streak 2 Neural folds 3 Amnion Day 25 1.5-2.5 mm Carnegie stage 9 1 Yolk sac 2 Primitive streak 3 Primitive node 4 Germ disc Day 19 0.4 mm Carnegie stage 7 Day 28 2-3.5 mm Carnegie stage 10 1 Amnion 2a Neural ridge 2b Neural tube 2c Caudal neuroporus 2d Rostral neuroporus 3 Neural folds 4 Somites 5 Yolk sac http://www.embryology.ch/anglais/iperiodembry/carnegie03.html#st710 Dorsal Ventral • bilaminar → trilaminar germ disc • cephalocaudal and lateral flexion of embryo FOLDING OF EMBRYONEW BODY PLAN 6W 6W5W4W week 4-6EMBRYONIC DEVELOPMENT • Mesoderm segmentation • Primitive gut • Esophagotracheal diverticulum • Heart (starts beating day 22-23) • Limb buds • Primary brain vesicles, closing of neuropores • Differentiation of neural crest • Origin of thyroid and anterior pituitary • Ectodermal placodes, optic vesicle • Liver diverticulum • Septum transversum • Segmentation of mesoderm continues • Posterior pituitary • Heart septation begins • Lung buds branch - pseudoglandular stage of lung development • Cochlea grows • Lens vesicle, nasal placodes • Fourth brain ventricle forms • Pharyngeal arches, ridges and pouches • Limb buds grow • Hematopoiesis in liver • Retinal pigment • Derivatives of endodermal pharyngeal pouches (parathyroid, thymus) • Adrenal gland • Heart and lungs descended to thorax • Innervation of limbs, differentiation of myoblasts • Face development – maxillary and mandibulary processes, palatine, choans • Telencephalon stratifies – archicortex, paleocortex and neocortex. Choroid plexus • Rotation of stomach • Pancreatic diverticula fuse week 8 7-8W 8-9W • Secretion from endocrine pancreas • Growth of liver, growth and luminization of bile ducts • Ossification of limbs begin • Development of brain nuclei • Joints of upper and later lower limbs allow rotation • Fingers grow • Stratification of cerebellar cortex • Perforation of anal membrane • Herniation of intestinal loops • Testes produce testosterone • Nose, meatus, eyelids, developer, external ears start to grow • Backbone - 33-34 cartilaginous vertebrae • Embryonic tail diminished EMBRYONIC DEVELOPMENT 280 days (= 40 weeks = 10 lunar months) from the first day of the last menstruation 266 days (= 38 weeks) from ovulation (gestation age) Calculation of term: First day of the last menstruation + 1 year– 3 months + 7 days LENGTH OF PREGNANCY Conception age Menstrual age Fertilization Beginning of the last menstruation 38 weeks 40 weeks DELIVERY 0 3 8 weeks preembryo embryo fetus 3M month 4-5 • Fetus swallows amniotic fluid – necessary for GIT development • Rapid growth of head (non-proportional to rest of body) • Eyelids fuse • Ossification centers visible by ultrasound examination • Development of external genitalia • Kidneys produce urine, other organs start to work • Skeletal muscles innervated • Physiological umbilical hernia, in 12th weeks reposition of intestinal loops 60-70mm, 150 g FETAL DEVELOPMENT 4M 130mm, 450 g • Rapid growth of fetus • Ossification of skeleton • Face growths, mandible visible • Apparent external genitalia month 5-9 5M • Limbs growth • Mother feels fetal movements • Vernix caseosa, lanugo • Short hairs and eyelashes • Fetus reacts to sound and later to light • Lungs start to produce surfactant • Limit of viability 160-200mm, 500-600 g • Eyelids open • Wrinkled skin with visible capillaries • Subcutaneous fat • Hairs grow • Maturing of organ systems 300mm, 800-900 g 6-7M 8-9M • Subcutaneous fat accumulates in limbs • Smooth, red skin • Hallmarks of full term fetus 400mm, 2000g FETAL DEVELOPMENT Major: • length (50 – 51 cm), • weight (around 3500 g, physiological range 2500 - 4000g), • head sizes • boys - testes in scrotum, girls - labia majora over labia minora Minor: • eutrophic fetus, subcutaneous fat • skin is not blue (no cyanosis), lanugo remains on shoulders and back, • eyelashes, hairs several cm long, nails over fingertips • cranial bones hard, anterior and posterior fontanelle are palpable, and separated • newborn cries and moves (Apgar score) HALLMARKS OF FETAL MATURITY Full term – related to length of pregnancy (menstrual age) • preterm (<37 weeks) • full term (38 – 40 weeks) • after term (>42 weeks) (meconium in amniotic fluid) Fetal maturity – development of fetus: mature X immature LENGTH OF PREGNANCY  FETAL MATURITYFULLTERM NEWBORN Diameter bitemporalis – 8,00 cm (join of the most distant points on sutura coronaria) Diameter biparietalis – 9,5 cm (join of midpoints of tubera parietalia) Diameter bitemporalis Diameter biparietalis HEAD SIZEFULLTERM NEWBORN Oblique sizes: • Diameter frontooccipitalis – 12.0 cm (join of forehead midpoint and most distant point of occiput) • Circumferentia frontooccipitalis – 34.0 cm • Diameter suboccipitobregmatica – 9.5 cm (join of protuberantia occipitalis externa and midpoint of large fontanelle) • Circumferentia suboccipitobregmatica – 32.0 cm • Diameter mentooccipitalis – 13.5 cm (join of chin midpoint and most distant point of occiput) • circumferentia mentoocipitalis – 35 - 36 cm • Diameter biacromialis – 12.0 cm, circumferentia biacromialis – 35 cm • (join of acromion – acromion) Diameter frontooccipitalis Diameter suboccipitobregmatica Diameter mentoooccipitalis HEAD SIZEFULLTERM NEWBORN RELATIONSHIP BETWEEN AGE AND LENGTH OF FETUSRULE OF HAASE RULE OF HASSE • forensic medicne 3. – 5. lunar month: length in cm = square of month 6. – 10. lunar month: length in cm = months multiplied by 5 Lunar month Length of fetus[cm] 3 9 4 16 5 25 6 30 7 35 8 40 9 45 10 50 1. LIE (SITUS) = relationship of the long axis of the fetus to that of the mother • longitudinal: (99 %) • transverse: (1 %) perpendicular axes • oblique: unstable → longitudinal or transverse position 2. POSITION (POSITIO) = fetal backbone relative to uterus ridge • first= left (back to the left) • second= right (back to the right) • first/second common/less common 3. FETAL HABITUS (HABITUS) = relationship of one fetal part to another • regular = head and limbs in flexion • irregular = everything else 4. PRESENTATION (PRAESENTATIO) = that part of the fetus lying over the pelvic inlet; the presenting body part of the fetus. • occiput (most common) • vertex, forehead, face (1%) • breech • trunk, shoulder FETUS IN UTERUS SITUS longitudinal - occiput longitudinal - breech transverse oblique POSITIO 1. COMMON2. COMMON „LEFT“„RIGHT“ 2. LESS COMMON 1. LESS COMMON HABITUS AND PRESENTATION regular vertex forehead faceocciput PRESENTATION HABITUS irregular (any other) PHYSIOLOGICAL IMPOSITION OF FETUS IN UTERUS • LIE LONGITUDINAL - HEAD FIRST • POSITION FIRST COMMON • HABITUS REGULAR • PRESENTATION - OCCIPUT INTRODUCTION TO TERATOLOGY • Congenital disorders – due to abnormal developmental events • Genetic (inherited) or nongenetic (external) causes • Teratogens • Critical developmental periods • Life style (alcohol, smoking, drugs) • Infections (rubeola, HIV, toxoplasmosis) • Lack or abundance of key substances (folic acid  retinoids) • Chronic diseases (medical treatment) INTRODUCTION TO PRENATAL DIAGNOSTICS • Genetic counselling • Biochemical and ultrasound screening • Karyotyping and DNA diagnostics • Clinical diagnostics • Revealing high risk pregnancies, access to preventive and therapeutic care • Preventing delivery of fetuses with severe congenital malformations • Support of delivery of genetically high-risk babies • Planning and providing clinical care • Interdisciplinary care – biochemistry, genetics, gynecology and obstetrics, neonatology – parts of fetal medicine • Indication: - congenital disorders in family - positive screening in 1st or 2nd trimester - abnormal finding by ultrasound - maternal age (over 35 years) ČR 1994-2008 GENETIC COUNSELING • Anamnesis (case history) • Preconception counselling • Explaining of examination results, causes, clinical symptoms, therapeutic options • Minimization of risk of repeated disease • Providing diagnosis and information for free choice • Providing precise diagnosis and risk estimation • Providing care during pregnancy and later • NONDIRECTIVE • ALL EXAMINATIONS AND PROCEDURES ARE VOLUNTARY BIOCHEMICAL SCREENING • Non invasive - Revealing high risk pregnancies – chromosomal aberrations and clefts - Screening is not a diagnostics → further examinations (amniocentesis, karyotype, US) • Tests between weeks 14-16 („TRIPLE test“) - low sensitivity and specificity (50-60%), high false positivity (70%) - AFP, E3, hCG - chromosomal aberrations, abnormal closing of neural tube, defects of body walls • Combined screening in week 11-13 - chromosomal aberrations – Down: 47,XY,+21, Edwards: 47,XY,+18, Patau 47,XY,+18 - US –nuchal translucence, NT - PAPP-A, hCG (multiplies of median, MoM) - age included in algorithm - output: screening positive vs. negative (limit 1:100) INVASIVE DIAGNOSTICS • Amniocentesis - 16th-20th week - US controlled amniotic fluid aspiration - Cell culture, karyotype - Risk of miscarriage 0.5-1% • Chorion villus biopsy - 10th-13th week - Karyotype, molecular genetic examination - Risk 0.5-1% • Cordocentesis - 22nd week - Sampling of venous umbilical blood - Now diagnostics and therapy of blood diseases (anemia, infections), or diagnostics in multiple pregnancies - Risk 1% • Fetoscopy - Transabdominally (earlier transcervically) - Visualization and fetal biopsy - Risk 3-10%, done rarely ULTRASOUND DIAGNOSTICS • 6-8th week - confirmation of pregnancy, heart action - number of fetuses • 13-14th week - nuchal translucence (risk > 3 mm) - nasal bone (present  absent), minor markers (omphalocele, tricuspidal regurgitation, abnormality in ductus venosus flow, enlargement of urinary bladder-megavesica) - fetal size • 20-22nd week - detailed screening - fetal biometry (biparietal diameter, head circumference, length of femur) - head and CNS (shape, cavity in septum pellucidum, ventricles, cerebellum, cisterna magna), face (lip, jaws, nose, orbits, profile), heart (action, size, axis, 4-chamber projection, outflow tracts, ...), thorax (pathological structures), abdominal cavity (stomach, intestine, kidneys, urinary bladder, umbilicus and umbilical vessels), backbone, limbs, palms, feet - placenta, volume of amniotic fluid • 30th week - fetal size - volume of amniotic fluid - placenta (exclusion of placenta praevia) ADVANCEMENTS IN MOLECULAR GENETICS • Analysis of cell-free fetal DNA in maternal blood • Since 12th week • Massive paralell sequencing (Next-Gen Sequencing) • Common aneuploidies (trisomy 21,13,18) • Monogeneous disorders pvanhara@med.muni.cz http://www.med.muni.cz/histology THANK YOU FOR ATTENTION