Derivatives of the Germ Layers

In the previous chapter we have seen how the embryonic disc transformed into a bilaminar disc as a result of the development of the epiblast and the hypoblast. This was followed by the process of gastrulation and formation of the primitive streak due to migration of the epiblast cells along the surface of the embryonic disc towards the midline and their ingression to form the primitive streak and the primitive pit. Ingression of the surface cells, their migration in the reverse direction, and their arrangement into two sheets of cells underneath the epiblast resulted in the formation of a trilaminar embryonic disc made up of an ectoderm, mesoderm and endoderm. The trilaminar embryonic disc is also known as the trilaminar blastoderm or the trilaminar germ disc.  

Fig. 7.1. The illustration shows the trilaminar embryonic disc that has resulted from the process of gastrulation. The three laminae are the ectoderm, mesoderm and endoderm. The notochord is part of the mesoderm constituting the axial mesoderm.

The Three Germ Layers

The three germ layers are the outcome of gastrulation that takes place in the third week of the embryonic development, when the blastocyst differentiates into a gastrula. This differentiation is a change from a homogenous single layered cells into distinct cell lineages, and also sets up the axis along which the embryo will continue to develop. The three germ layers are named according to their relative position in the embryonic disc as ectoderm, which is the outermost layer, the mesoderm which is the middle layer, and the endoderm which is the innermost layer. The three germ layers interact with each other and differentiate to form all tissues and organs of the body, as well as all extraembryonic tissues and structures. Normal embryonic development requires proper formation of the three germ layers and normal interactive signaling between them.

The Ectoderm

The ectoderm is the outermost of the three germ layers; it overrules the mesoderm. It is the last of the three germ layers to be established by cell of the epiblast that remain on the surface after formation of the mesoderm and the endoderm. After gastrulation, a process called neurulation commences. The ectoderm differentiates during neurulation into two parts, the surface ectoderm and the neural ectoderm (neuroectoderm). The surface ectoderm gives rise to tissues on the outer surface of the body like epidermis, hair, and nails, whereas the neuroectoderm gives rise to the nervous system. The neuroectoderm differentiates into the neural tube and the neural crest. The neural tube develops into the central nervous system and sense organs, whereas the neural crest gives rise to ganglia and connective tissues and bones of the head region.

Formation of the neural tube commences in the third and fourth weeks of embryonic development. This process, which is known as the primary neurulation, begins with induction of the notochord to the overlying ectoderm to proliferate and forming a narrow-thickened plate called the neural plate made of columnar cells. The neural plate transform into the neural tube via bending, convergence and closure.  By bending of the peripheries of the neural plate and sinking of its middle part, the plate gradually changes into a groove flanked by a fold on each side. The  groove is called the neural groove and the folds are known as the neural folds. The folds gradually approach each other and fuse to form a tube known as the neural tube. The cranial part of the tube gives rise to the grain, eye and inner ear, whereas the rest of the tube gives rise to the spinal cord,

Fig. 7.2. The illustration shows differentiation of epiblast into ectoderm, mesoderm and endoderm along with the derivatives of ectoderm.

When the neural folds fuse with each other, some cells are left behind not incorporated into the neural tube or the general ectoderm by a process known as delamination. These cells aggregate into ridgelike structures known as the neural crest. These cells are multipotent stem cells and migrate throughout the embryonic body giving rise to wide range of functional cells that include ganglionic cells, chemoreceptor cells, endocrine cells, melanocytes, Schwann cells, and chromaffin cells. The location to where they migrate provides a signaling aid for their differentiation.

Fig. 7.3. The illustrations show the process of development of the neural tube from the ectoderm as neural plate that transform into neural groove into a neural tube. It also shows the site of origin of the neural crest.

Extraembryonic Ectoderm

The three germ layers extend beyond the limits. To begin with, the boundaries of the embryonic body is clear and the embryonic and extraembryonic membranes are discernible. Extraembryonic membranes and tissues are derived from the zygote but do not contribute to the body of the fetus. Embryonic tissues are those whose fate is to contribute to structures retained within the fetus and become part of the newborn baby. Extraembryonic tissue on the other hand, are those whose fate is to give rise to structures that support the embryo during its development – such as the placenta and extraembryonic membranes - and are not retained as part of the newborn baby.

The trophoblast which is in direct contact with the embryonic ectoderm form the extraembryonic ectoderm, which is also known as the trophectoderm. This occurs during early gastrulation.  After splitting of the lateral plate mesoderm of mesoderm into splanchnic and somatic mesoderm, the ectoderm fuses with the somatic mesoderm together forming what is known as the somatopleure. The somatopleure forma the wall of the amnion and the chorion.

Fig. 7.4. The illustration shows formation of the somatopleure.

The Mesoderm

The mesoderm is the middle layer of the three germ layers that is formed during the process of gastrulation by migration of epiblast cells towards the primitive streak and ingression to form a new layer beneath the epiblast. This new layer is formed in the third week of embryonic development and is called the mesoderm. During the process of formation of the mesoderm, the columnar cells of the epiblast dissociate from each other, lose their epithelial characteristics and assume mesenchymal cell characteristics and the ingress down the primitive streak. To begin with, these precursor cells have the potential to become either mesoderm or endoderm and accordingly are called mesoendodermal cells. The process that gives rise to the mesoderm creates a dorso-ventral pattern within the mesoderm that organizes cells in specific locations along the dorso-ventral axis. Location of the cell determines its  kind and the fate of its progeny. Mesoderm is the mother germ layer of the body’s cells and tissues. Several bimolecular signaling factors induce and regulate development of the mesoderm; these include activin, FGF, TBx6, Sox2, Brachyury, and BMP-4. After its formation, the mesoderm differentiates into four parts, namely the axial mesoderm, the paraxial mesoderm, the intermediate mesoderm and the lateral mesoderm.

The Axial Mesoderm

The axial mesoderm, which is also known as the chordamesoderm, is that part of the mesoderm that lies along the central axis of the embryonic disc. It gives rise to a number of structures including the notochord. Its formation begins with the epiblast cells migrating towards the cranial end of the primitive streak where they ingress and migrate cranially along the midline beneath the epiblast at the same level as the rest of the mesoderm. It differentiates into the prechordal plate and the notochord. The notochord provides physical support to the developing embryo and as well produces signaling factors that induce differentiation of cells in the neighborhood. After attaining full size, it regresses and forms the nucleus pulposus of intervertebral discs. Formation of the axial mesoderm is induced and maintained by signaling proteins that include NODAL, FOXH1, TBXT, and SMAD2. The notochord plays a significant role in the development of the nervous system. It induces the overlying ectoderm to differentiate into the neural plate and then the neural groove to form the neural tube and the neural crest.

The prechordal plate is a median mass of cells, located anterior to the cranial end of the notochord; it appears during gastrulation. The prechordal plate differentiates into the oropharyngeal membrane that in future will give rise to the oropharyngeal membrane, which separates the embryonic mouth (stomodeum) from the embryonic pharynx. On the other end, there is the cloacal membrane which develops similarly as a circular structure located caudal to the caudal end of the primitive streak. It covers the embryonic cloaca during development of the reproductive and urinary systems.

Fig, 7.5. The illustration shows the derivatives of the axial mesoderm.

Fig, 7.6. The illustration shows migration of epiblast cells (curved green arrows), their ingression in the primitive pit, and their migration cranially (straight green arrow) forming the notochord.

The Paraxial Mesoderm

This is also known as the somitic mesoderm. It is called paraxial because it parallels the axial mesoderm flanking it on both sides and is called the somitic mesoderm because its cells aggregate into masses known as the somites. Somites are a characteristic feature of the embryo disc at the early stage of development and distinctively arranged in pairs of discrete spherical cell masses. The paraxial mesoderm develops by migration of cells from the primitive streak at  the time when the neural plate develops from the ectoderm.

Fig. 7.7. The illustration shows the axial and paraxial mesoderm. The paraxial mesoderm is made of a series of distinct pairs of somites.

Several differentiation factors are involved in the development of the paraxial mesoderm; these include BMPs (bone morphogenetic proteins) VV (ventral signal), DV (dorsal signal), FGF (fibroblast growth factor), Brachyury (T-box signal), Wnt and noggin factors. 

The paraxial mesoderm differentiates in a craniocaudal direction on each side of the axial mesoderm (notochord) forming pairs of spherical cellular masses known as somites; the cranial most pair is the oldest whereas the most caudal one is the youngest. The first pair of somites appears in the 20^th day of embryonic development. Thereafter new pairs are added caudal to the existing ones, at rate of about 3 pairs / day, until 44 pairs are added. As new pairs of somites are added caudally, the oldest ones present cranially may further differentiate and disappear. The age of embryo at this stage is often related to the number of somites present. In the fourth week of embryonic development, the somites begin to differentiate into dermatomes, myotomes, and sclerotomes. The dermatomes and myotomes are collectively referred to as dermomyotomes.

The mesoderm as a whole develops from the epiblast by gastrulation via the primitive streak. After ingression in the primitive streak to form a new layer beneath the epiblast, the presumptive mesodermal cells migrate laterally and cranially and arrange them into two sheet, one on either side of the primitive streak and the notochord, assuming the shape of a butterfly wings.

The sheet of mesoderm gradually differentiates into three components, the paraxial mesoderm, the intermediate mesoderm. The paraxial mesoderm thicken and transform into somites, which transit cuboidal or spherical structures that are arranged in pairs in craniocaudal direction alongside the neural tube. They are regularly produced by budding off from the unsegmented paraxial mesoderm. The somitic cells differentiate into three masses known as the dermatome, the myotome and the sclerotome. The sclerotome develops into vertebrae and proximal parts of ribs, whereas the dermatome gives rise the dermis of the body back. On the other hand, the myotome differentiates into an epaxial myotome and a hypaxial myotome. The epaxial myotome gives rise to musculature of the back, whereas the hypaxial myotome gives rise to musculature of the body wall and the musculature of the limbs.

Differentiation of the somites is regulated by number of biomolecules which include the Hairy protein, cadherin, fibronectin, Sonic hedgehog protein produced by the notochord, neurotrophin-3 secreted by the neural tube, Wnt protein by the neural tube, and BMP-4 by the lateral mesoderm.

Fig. 7.8. The illustration on the left shows parts of the mesoderm, whereas the one on the right shows the somite is differentiated into myotome, dermatome and sclerotome.

Fig. 7.9.  The illustration shows main derivative of the paraxial mesoderm.

 The Intermediate Mesoderm

The intermediate mesoderm is defined as that part of the mesoderm which lies between the paraxial mesoderm and the lateral mesoderm. It maintains its sheet-like shape but ultimately differentiates into the kidneys, reproductive organs, and adrenal cortex.

Several biomolecules initiate regulate morphogenesis of the intermediate mesoderm. Early intermediate mesoderm transcription factors include Lim1, Lhx1, Osr1, Pax2, and Sim1. All these markers are expressed before epithelial duct formation and respond to signals from adjacent mesoderm and overlying ectoderm. Most of these factors are not specific for the intermediate mesoderm, yet Lhx1 and Pax2/8 are critical for early differentiation of the intermediate mesoderm.

The paired kidney structures develop in a rostral-to-caudal sequence during embryogenesis via specific stages, the pronephros, mesonephros, and metanephros stages. Thus, the development of the human kidney progresses through three major stages. The first two stages are transient; only the third one, the metanephros, persists as a functional kidney. All kidney progenitors originate from the intermediate mesoderm, which forms as bilateral stripes of mesoderm in the caudal trunk of the embryo between the somites and the lateral plate mesoderm. The pronephros appears at about the 22^nd day of human embryonic development and disappears four days later and is made of a set of tubules. Development of the pronephros is regulated by Pasx2/8. The mesonephros is set of new tubules that arise caudal to the pronephros close to the gonadial ridge. It is a functional kidney that carries on its functions between the 8^th and 10^th week of embryonic development. The human mesonephros regresses completely by the 16^th week of embryonic development. The metanephros which is the permanent develops further caudally. It appears in the 10^th week of human embryonic development. It has a dual origin partly developing from the ureteric bud and partly from the metanephrogenic tissue (mesenchyme). BMP7, FGF2, and GDNF factors participate in the morphogenesis of the metanephros.

The development of the testis and the ovary (gonadal differentiation) begin in the indifferent stage of embryonic development when the gonad is undifferentiated with a potentiality to develop into either a testis or an ovary. At about the 4^th week of embryonic development, the gonadal primordium appears differentiates into the urogenital ridge. The urogenital ridge contributes to the formation of the urinary system, the genital system and the adrenal cortex. It differentiate into a urinary component, and adrenal component and a gonadal component. The gonadal component differentiates into finger like projections called the gonadal cords. Until this stage the gonad is indifferent, made of a cortex and a medulla, and cannot be identified as a testis or an ovary. Primordial germ cells migrate from the yolk sac vesicle and settle between cells the gonadal cords. If the embryo is chromosomally male (46XY), the gonad develops into a testis under the influence of the SRY gene present in the Y chromosome. If the embryo is chromosomally female (46XY), the gonad develops into an ovary influenced by genes of the X chromosome. The ovary becomes identifiable in the 10^th week of embryonic development. 

Fig. 7.10. The illustration shows derivatives of the intermediate mesoderm.

Fig. 7.11. The illustration shows development of the pronephros, mesonephros and metanephros,

The Lateral Mesoderm

The lateral mesoderm is the peripheral most parts of the mesoderm, lying peripheral to the intermediate mesoderm on either side of the embryo. It is often referred to as the lateral plate mesoderm. It gives rise to  the circulatory system, body wall, some neck muscles, muscles of and the limbs except its muscles. During the third week of embryonic development, the lateral mesoderm splits into a splanchnic (visceral) mesoderm and a somatic (parietal) mesoderm; each has an embryonic part and extraembryonic part. The space between the somatic and the splanchnic is the coelom; the coelom later differentiates into the pleural cavity, the pericardial cavity, and peritoneal cavity. The splanchnic mesoderm fuses with the endoderm to form the splanchnopleure, which gives rise to the viscera and the heart. The somatic mesoderm on the other hand fuses with the ectoderm to form the somatopleure which contributes to the formation of the body walls and the dermis.

Fig. 7.12.  The illustration shows the derivatives of the lateral mesoderm.

Fig. 7.13. The illustration shows the mesoderm (in red colour) and how the somatic mesoderm and the splanchnic mesoderm fuse with ectoderm and endoderm respectively to form somatopleure and splanchnopleure.

The Endoderm

Endoderm is the innermost of the three primary germ layers. To begin with, the endoderm is made flat low cuboidal cells that may develop into columnar cells. It has an embryonic and an extraembryonic component. The embryonic endoderm differentiates into the epithelium of alimentary tract, the respiratory passage and associated glands, the liver, pancreas, the pharynx, eustachian tube, and tympanic cavity, the thyroid gland, the parathyroid gland and the thymus. Several growth factors are involved in the development of the endoderm; these include Activin A / Nodal, TGF-β, FGF, Wnt, BMP,  CXCR4, CD117, EPCAM and SOX17

The endoderm folds its cephalic and caudal ends forming blind-ended tubes called the foregut and the hindgut. In between the two is the midgut that overlies the yolk; it has no floor and accordingly is confluent with the yolk sac via the yolk stalk. The rostral end of the foregut dilates to form the primitive pharynx.

Fig. 7.14.  The shows the endoderm in yellow colour as the innermost layer of the developing embryo. By folding of its rostral part, it forms the foregut and folding of its caudal part it forms the hind gut. It extends beyond the limits of the embryonic body to line the yolk sac and the allantoic.

The primitive pharynx differentiates to give to the thyroid gland, the parathyroid glands, lining of the middle ear, lining of the Eustachian tube, the epithelial lining of the definitive pharynx, and the epithelial components of the thymus. Several growth factors are involved in the differentiation of the pharynx; these include AX9, FOXA2, SOX2, EPCAM, and HOXA3. The rest of the foregut differentiates into the epithelium of esophagus, stomach, proximal duodenum, larynx, trachea, bronchi, lungs, and gallbladder. It also yields the liver, the pancreas and intramural glands of the respiratory passages and the digestive tract. The midgut gives rise to the distal duodenum, jejunum, ileum, cecum, appendix, ascending colon, and proximal parts of transverse colon. The hindgut becomes the distal part of the transverse colon, descending colon, sigmoid colon and the upper anal canal.

The endoderm has an embryonic component and an extraembryonic component. The extraembryonic component fuses with the splanchnic mesoderm to form the splanchnopleure which makes up the walls of the yok sac and the allantoic.

Fig. 7.15. The shows the derivatives of the endoderm.

Mesenchyme

Mesenchyme originates mostly from the mesoderm, yet a substantial amount of it develops from the ectoderm and specifically from the neural crest. Mesenchyme is made of cells and an intercellular matrix. Mesenchymal cells multipotent or possibly pluripotent stem cells that can differentiate into a wide array of functional cells including bone cells, cartilage cells and muscle cells. They are small spindle-shaped cells with large pale nuclei and prominent nucleoli and scant cytoplasm. They could appear stellate with multiple large processes. They give rise to all connective cells, cells of connective tissue proper such as fibroblasts and cells of specialized connective tissue such as bone cells, blood cells, fat cells, and cartilage cells.  In embryos mesenchyme is present in several locations occupying the spaces between the ectoderm and the endoderm. As the embryo and fetus grow the amount of mesenchyme decreases by differentiation in other types of tissues. In adults, mesenchyme is found in small quantities as part of the loose areolar connective tissue. Mesenchyme is also present in Wharton’s jelly of the umbilical cord, which represents a good source of mesenchymal stem cells.

Fig. 7.16. The illustration shows the multipotecy of mesechymal cells; they capable of differentiating into a wide asortment of functional cells including fat cell, connective tissue fibroblasts, skeletal muscle cells, cardiac muscle cells, smooth muscle cells, cartilage cells and bone cells.

 Mesenchymal Cells and Regenerative Medicine

The great capabilities of mesenchymal cells or the mesenchymal stem cells (MSCs) to proliferate, self-regenerate and differentiate into a wide array of functional cells, makes a good candidate for regenerative medicine. MSCs can modulate the immune system, reduce inflammation and promote tissue repair. Moreover, they can migrate to damaged tissues and contribute to repair and regeneration. They utilized for regenerative medicine in a number of medical fields that include orthopedics, cardiovascular diseases, neurological diseases and wound healing procedures. They are used for the treatment of bone and cartilage defects, such as osteoarthritis, bone fractures, myocardial infarction and heart failure, multiple sclerosis Parkinson's disease and are also enhance wound healing and tissue repair.

Mesenchymal stem cells are obtained from embryo or adults. Utilization of embryonic mesenchymal stem cells is accompanied by some ethical issues, however, MSCs can be obtained from the umbilical cord of newborns without such issues. Moreover, immunological issues can also be faced when grafting embryonic mesenchymal cells. this problem of immunology is overcome by utilize adult mesenchymal stem cells from the patient himself (autologous adult MSCs).  

Adult MSCs can be isolated easily from various body tissues including the bone marrow and adipose tissue. One important advantage MSC therapy is that it is carried out by minimally invasive procedures.

Fig. 7.17. The illustration shows the major steps in MSC therapy technique, that includes harvesting MSCs from the bone marrow by needle aspiration, induction of differentiation into specific functional cells, culturing of functional cells and their implantation into the patient's body. ss

Take a quiz