Development of the

Thomas A. Marino, Ph.D. Competencies: Upon completion of this section of the course, you must be able to:

• Define the segments of the primitive gut tube. • Describe the embryological movements of the respiratory diverticulum as it develops into the , bronchi, and . • Explain the origin of cells that develop into the tissue. • Compare and contrast morphology of the lungs during the four stages of lung development. • Describe the primitive and how it becomes subdivided into pleural, pericardial and peritoneal cavities. Introduction

• Development of the lungs begins at 4 weeks. • The of the respiratory system develops from . • The , cartilage and muscle develop from splanchnic . Early Embryonic Morphology

• Early vertebrate body plan • At 26 days a small opening in the appears. • At 28 days it evaginates to form a laryngotracheal diverticulum. Separation of the Laryngotracheal Diverticulm • Longitudinal folds - tracheoesophageal ridges develop • Form

Notochord

Sox2 Nkx2-1 FGF10 RA Noggin Wnt2 Bmp Significance

• Lack of • Result – Shh – Trachoesophageal – Retinoic acid fistula receptors – Esophageal atresia – FGF10 – Sox2 – Nkx2-1 – Bmp4 – Noggin – Wnt Tracheoesophageal septum separates • Trachea and lung buds - ventral • Esophagus - dorsal Development of the

• Epithelium develops from endoderm of laryngotracheal tube. • Connective tissue and cartilage develops from splanchnic mesoderm. • Cartilages develop from neural crest cells. Development of the Trachea

• Epithelium develops from

endoderm of laryngo- Splanchnic tracheal tube Mesoderm • including glands Endoderm • Cartilage, connective tissue and muscle from Smooth Cartilage muscle splanchnic mesoderm Epithelial/Mesenchymal Interactions ! ! Endoderm Mesoderm In FGF10 deficient mice there are no lung buds.

FGF2

FGF10 Development of the Lungs

• 4th week the lung bud develops • divides into two lung buds Development of the Lungs

• Two lung buds divide: • The right one into three main bronchi • The left one into two main bronchi

https://syllabus.med.unc.edu/ courseware/embryo_images/ unit-digest/digest_htms/ digest012a.htm Development of the Lungs

• Bronchi continue to divide. • By 6 months there have been 17 generations of subdivisions. • After birth there are an additional 6 divisions of the bronchial tree. • As growth occurs there is a caudal development of the lungs. • At birth the tracheal bifurcation is at the level of the 4th thoracic vertebra. Maturation of the Lungs

• There are 3 Stages of Lung Maturation • 1. Pseudoglandular Period ( 5 - 16 weeks) • 2. Canalicular Period (16 - 26 weeks) • 3. Terminal Sac (Saccular) Period (26 weeks to birth) • There are 4(5) Stages of Lung Maturation 1. Embryonic ( 4 – 11 weeks) 2. Pseudoglandular Period ( 5 - 16 weeks) 3. Canalicular Period (16 - 26 weeks) 4. Saccular (Terminal Sac) Period (26 weeks to after birth) 5. Alveolar Period (late fetal period to childhood) Maturation of the Lungs

• Pseudoglandular Period - 5 to16 weeks • All elements of the lungs are developed except those elements involved in . • Branching morphogenesis is prominent. • Terminal Bronchioles present no respiratory bronchioles Branching morphogenesis

• Bud elongation FGF10 • Elongation stops FGF2 • Tip of the bud widens • Bifurcation Factors involved

• Retinoic acid forms gradient with highest levels proximally – RA inhibits FGF10 • SHH promotes BMP4 which inhibits FGF10 • Wnt3b regulates BMPs which promote proliferation of mesoderm. • What BMPs are regulating is not precisely known at the cellular level. Maturation of the Lungs

• Canalicular Period - 16 - 26 weeks • Overlap as cranial segments mature faster than caudal ones. • Lumen of bronchi and bronchioles become large relative to tissues • Bronchial tree branches become narrower. • Respiratory bronchioles and alveolar ducts develop. • Tissue becomes more vascular. Maturation of the Lungs

• Canalicular Period (16 - 26 weeks) – Note the cuboidal epithelium of the airway. – The blood vessels are not close to the epithelium Maturation of the Lungs

• Terminal Sac Period ( 26 weeks - birth) – Now the epithelium is much thinner. – The blood vessels abut the epithelium • What regulates the switch from pseudoglandular to canalicular an to terminal sac stage? – Alveolar sacs begin to form – Blood vessels become closely associated with the alveolar cells. Maturation of the Lungs

• Terminal Sac Period (26 weeks to birth) • Terminal sacs develop • Epithelium becomes very thin • bulge into the alveoli • Type I alveolar cells develop • network develops rapidly Formation of Alveoli

• PDGF • Fgf – Fgf2 and Fgf18 important for late alveolar development • Retinoic acid – High and low levels can disrupt lung development. Multipotential cell

Bronchiolar cells Alveolar cells

Non neuroendocrine Neuroendocrine Type II cells cells cells ?? ?? Ciliated cells Goblet cells Type I cells Maturation of the Lungs

• By 20 weeks Type II alveolar cells begin producing surfactant. • Surfactant permits expansion of terminal sacs. • Fetus needs to weigh 1000 gm and be between 26 and 28 weeks before enough surfactant is produced. • Surfactant and enough capillaries are necessary. Maturation of the Lungs

• Alveolar Period ( late fetal period to childhood) • Squamous epithelium forms. • During this period respiratory bronchioles end as terminal sacs. • Terminal sacs become alveolar ducts. • Alveoli form after birth. • From 3rd to 8th year alveoli continue to develop. Maturation of the Lungs

• Alveolar Period – Now there are: • Type I alveolar cells • Type II alveolar cells • Macrophages • Fibroblasts Lungs at Birth

• At birth the lungs are filled with fluid. • Fluid is replaced by air. • Fluid cleared through: • Mouth and nose • Pulmonary capillaries • Pulmonary arteries, veins and lymphatics • After birth most growth is in the number of respiratory bronchioles and alveoli and not an increase in the size of alveoli. Formation of blood vessels

• Angiogenesis – Angiogenesis new blood vessels from pre- existing blood vessels • Vasculogenesis – angioblasts develop into endothelial cells and new blood vessels form • Vegf helps regulate this along with ephrinB2 and B4 The pulmonary vasculature develops in the absence of lung specification.

• Cardiac outflow tract and pulmonary vasculature come from cardiopulmonary mesoderm progenitors that lie in the posterior splanchnic mesoderm. ! • Lung develops separately from these progenitors.

T Peng et al. Nature 000, 1-4 (2013) doi:10.1038/nature12358 Clonal analysis reveals that CPPs generate related lineages within the cardiopulmonary system.

T Peng et al. Nature 000, 1-4 (2013) doi:10.1038/nature12358 Hedgehog signaling is required in CPPs to coordinate the vascular connection between the and lung.

T Peng et al. Nature 000, 1-4 (2013) doi:10.1038/nature12358 Development of Body Cavities Development of Horseshoe- Shaped Pericardial Cavity Lateral body folding occurs as well as head folding. Development of Body Cavities • In the fourth week the embryo has: – large pericardial cavity – left and right pericardioperitoneal canals – large peritoneal cavity Embryonic Circulation Common Cardinal Vein Dorsal Aorta and Posterior Anterior Cardinal Vein Cardinal Vein Umbilical Artery

Umbilical Vein Yolk Aortic Arches Sac

Ventricle Vitelline Artery Atria & Vein

BODY CAVITY Division of Body Cavities

• Pericardioperitoneal canal is dorsal to septum transversum. • pericardioperitoneal canal is lateral to the foregut.

Septum Transversum Division of Body Cavities

• As lung bud grows a membrane develops between the lungs and the heart. • Ridge of tissue grows into the pericardioperitoneal canals. • Ridges grow from the lateral walls of each canal. • Ridges called PLEUROPERICARDIAL FOLDS Division of Body Cavities

• Pleuropericardial membranes separate pericardial cavity from pleural cavities. • Pleuropericardial membranes contain the common cardinal veins which drain in the primitive heart. • The internal layer of the pleuropericardial membrane becomes the fibrous . Division of Body Cavities

• Pleuropericardial Membranes Division of Body Cavities • Pleuroperitoneal membranes separate the Pleuroperitoneal from the membranes peritoneal cavity. • Attachment to the to the dorsolateral abdominal wall. • Project into the pericardioperitoneal canal. • Fuse with the: • dorsal mesentary of the esophagus • septum transversum • Lateral body wall mesoderm Development of the Diaphragm

• The diaphragm develops from: • Septum transversum • Pleuroperitoneal membrane • Dorsal mesentary of the esophagus • Lateral body walls (cervical somite myotomes). Development of the Diaphragm

• At week 4 the septum transversum lies opposite the 3rd, 4th, and 5th cervical somites. • Myoblasts from these somites migrate into the diaphragm. • Phrenic nerve comes from cervical nerves 3, 4, and 5. Development of the Diaphragm

• As body grows diaphragm appears to migrate caudally. • By 6 weeks diaphragm lies opposite thoracic somites. • Phrenic nerve passes through the pleuropericardial membrane. • Phrenic nerve comes to lie in fibrous pericardium.