Craniofacial Development Part 1 Study Notes

Course Information and Textbook Reference

• Course Title: DENT 6500: Craniofacial development part 1.

• Lecturer: Brian L. Foster, PhD, Associate Professor, Biosciences Division.

• Primary Reference: Ten Cate’s Oral Histology, 9th edition, by Antonio Nanci.

  • Chapter 2 (General Embryology): Pages 12-22.

  • Chapter 3 (Embryology of the Head, Face, and Oral Cavity): Pages 23-28.

• Test Preparation/FAQ:

  • Understand important terms and concepts.

  • Be able to identify key structures in provided classroom images.

  • Memorize the order of developmental events and specified timelines.

  • Knowledge of specific diseases and genes discussed in "Research Minutes" and "Clinic Minutes" is required.

  • Class interaction was facilitated via Socrative.com (Room: TRAVERS9918).

Cell Biology Refresher: Differentiation and Induction

• Cell Differentiation: The process where a generalized cell (stem cell) becomes specialized to perform a specific job, manifesting as changes in size, shape, products, and activities.

  • The specialized state is known as "terminally differentiated."

  • It is a multi-step process: Stem cells $\rightarrow$ Multiple levels of progenitor cells $\rightarrow$ Specialized cells.

• Induction: The process that initiates differentiation.

  • Inducer: An agent/signal that provides cells with the instruction to differentiate.

• Competence: The inherent ability of a target cell to receive and respond to an inducing signal based on its internal and external hardware.

  • Receptors: Required to recognize the signal.

  • Internal Molecular Machinery: Required to respond to the signal.

• Examples of Inducers and Receptors:

  • Bone morphogenetic protein (BMP): Binds to cell surface BMP receptor type I or II ($BMPRI, BMPRII$).

  • Vitamin D: Binds to the intracellular vitamin D receptor ($VDR$).

  • Wnt signal: Binds to cell surface $LRP6$ AND Frizzled receptor.

Cell Signaling and Transcription Factors

• Cell Signaling Process: Transfer of information to cause changes in gene expression and function within a cell.

  • Path: External inducing signal $\rightarrow$ Competent receptor $\rightarrow$ Intracellular changes $\rightarrow$ Change in cell function.

• Types of Signaling:

  • Autocrine: A cell signals to itself.

  • Paracrine: A cell signals to adjacent/neighboring cells.

  • Endocrine: A cell signals to distant cells through the bloodstream.

• Signaling Effects: Affects cell differentiation, cell proliferation (division), and cell migration (movement).

• Transcription Factors (TFs): Proteins produced by cells that control whether DNA is transcribed into mRNA for protein translation.

  • TFs can activate or repress target genes.

  • Amplification: One single TF can control the expression of tens or hundreds of downstream genes.

Clinic Minute: DLX3 and TDO Syndrome

• DLX3 (Distal-less Homeobox 3): A transcription factor in the DLX family consisting of 214 amino acids.

• Tricho-dento-osseous (TDO) Syndrome: Caused by mutations in $DLX3$. It is characterized by pleiotropic effects involving hair, teeth, and bone.

  • Hair: Thin, kinky hair due to defects in follicle differentiation.

  • Teeth: Enamel defects because $DLX3$ regulates enamel genes such as Amelogenin, Enamelin, and Kallikrein 4.

  • Bone: Effects on BMP signaling, bone formation, resorption, and homeostasis.

General Embryology: Tissue Organization and Orientation

• Hierarchy of Organization: Genes/DNA $\rightarrow$ mRNA $\rightarrow$ Protein $\rightarrow$ Cells $\rightarrow$ Tissues $\rightarrow$ Organs $\rightarrow$ Fetus/Embryo.

• Anatomical Planes:

  • Coronal: Divides body into Front (Anterior) and Back (Posterior).

  • Sagittal: Divides body into Left and Right sections.

  • Transverse: Divides body into Top (Superior) and Bottom (Inferior).

• Anatomical Orientation Terms:

  • Rostral / Anterior: Toward the head.

  • Caudal / Posterior: Toward the tail/rear.

  • Dorsal / Superior: Toward the back/top.

  • Ventral / Inferior: Toward the belly/bottom.

Gestational Timeline

• Total Gestation: Approximately 40 weeks.

• Developmental Stages:

  1. Weeks 0 to 4: Primarily involved in proliferation and migration.

  2. Weeks 4 to 8: Morphogenesis stage involving cell differentiation and formation of major external/internal structures (the "Embryo").

  3. Weeks 8 to 40: Growth and maturation stage (the "Fetus").

• Time References:

  • Week 1: Days 0-7.

  • Week 2: Days 8-14.

  • Week 3: Days 15-21.

Early Embryonic Milestones: Weeks 1 to 3

• First Week:

  • Pre-implantation: Days 0-6 after fertilization.

  • Morula: A solid ball of cells formed around Days 3-4 via proliferation.

  • Blastocyst: A hollow, fluid-filled ball of cells formed around Day 5 via differentiation.

    • Embryoblast: The Inner Cell Mass (ICM). These are embryonic stem cells that form ALL tissues of the embryo.

    • Trophoblast: The outer layer of cells.

  • Implantation: Occurs into the uterine wall around Day 7.

• Second Week:

  • Bilaminar Embryo: Formed by the end of the 2nd week (~Day 13).

  • Germ Layers: Primary layers derived from the ICM.

    1. Ectoderm: Dorsal layer.

    2. Endoderm: Ventral layer.

  • Axes Establishment: Thickening of the rostral prochordal (prechordal) plate marks the union between germ layers.

• Third Week:

  • Gastrulation: The conversion from a bilaminar disk to a trilaminar disk (middle of 3rd week).

  • Primitive Streak: Formed as ectodermal cells converge toward the midline to migrate between the ectoderm and endoderm.

  • Mesoderm: The third germ layer newly formed during gastrulation.

  • Notochord: Formed by cephalic migrating cells to support the embryo.

  • Buccopharyngeal Membrane: Remains a bilayer of ectoderm and endoderm only, with NO intervening mesoderm.

Embryonic Folding: Week 4

• Rostro-caudal (Front-Back) Folding:

  • Occurs during the 4th week (commences around Day 21, complete by Day 28).

  • Consequence: Formation of the Stomatodeum (Primitive Oral Cavity).

• Lateral (Side-to-Side) Folding:

  • Transforms the disk into a 3D embryo.

  • Results in the formation of the neural tube, ectoderm epithelium, endoderm gut, and differentiation of mesoderm.

• Stomatodeum Boundaries:

  • Superior: Frontal prominence.

  • Inferior: Cardiac bulge (later replaced by the first branchial arch).

  • Posterior: Buccopharyngeal membrane.

  • Membrane Rupture: In the 4th week, the buccopharyngeal membrane ruptures, establishing communication between the stomatodeum and the foregut.

Neural Crest Cells (NCCs)

• Origin: Ectoderm; specifically adjacent to the neural tube.

• Formation: They separate from the neural plate when the neural tube closes (~Day 22/end of 3rd week).

• Characteristics: Highly migratory and have extensive capacity to differentiate into various tissues (atypical for ectoderm cells).

• Ectomesenchyme: NCCs undergo an epithelial-mesenchymal transformation. These ectomesenchymal cells form most of the connective tissues of the head.

• Craniofacial Contributions:

  • Dentin/Odontoblasts.

  • Dental pulp.

  • Dental follicle.

  • Cementum/Cementoblasts.

  • Periodontal ligament (PDL) fibroblasts.

  • Alveolar bone/Osteoblasts.

  • Note: Enamel/Ameloblasts are derived from traditional Ectoderm, NOT NCC-derived ectomesenchyme.

Brain Patterning and NCC Migration

• Brain Regions: Forebrain, midbrain, and hindbrain expand from the neural tube.

• Rhombomeres: Eight bulges (Rhombomeres 1-8) in the hindbrain that define rostral-caudal patterning and determine the destination of NCC populations.

• Migration Streams:

  • NCCs from midbrain and Rhombomeres 1 and 2: Move to the first branchial arch and the face (contributing to craniofacial connective tissue).

  • First Stream: Face.

  • Second Stream: First branchial arch.

  • Rhombomeres $\\ge 3$: Contribute to other structures.

Transcription Factors in Craniofacial Development

• Hox Genes: Ancient body-patterning genes used for the trunk and rhombomeres $\\ge 3$.

• Hox-free Status: Because craniofacial structures are newer evolutionary developments, the face and first branchial arch (populated by NCCs from midbrain and rhombomeres 1 and 2) are "Hox-free."

• Craniofacial Organizers: Instead of Hox genes, these transcription factors direct development:

  • $Otx2$ (Orthodenticle homeobox 2).

  • $Msx$ (Muscle segment homeobox 1-3).

  • $Dlx$ (Distal-less homeobox 1-7).

  • $Barx$ (BarH-like homeobox 1-2).

• Homeobox TFs: Contain a specific ~180 base pair sequence called a "homeobox."

  • Homeobox amino acid sequence: RRRKRTA-YTRYQLLE-LEKEFLF-NRYLTRRRRIELAHSL-NLTERHIKIWFQNRRMKWKKEN.

Anatomy and Fates of Branchial Arches

• General Anatomy of an Arch:

  • Arch: External bumps (Ectoderm), internal bumps (Endoderm), internal core (mix of NCC and mesoderm).

  • Groove (Cleft): External depression separating arches, lined by ectoderm.

  • Pouch: Internal depression separating arches, lined by endoderm.

• Branchial Arch 1 (Mandibular Arch):

  • Splits into Right/Left Maxillary processes and Right/Left Mandibular processes.

  • Derivatives: Mandible, Maxilla, teeth, and middle ear.

  • Meckel’s Cartilage: The cartilage of the first arch.

• Branchial Arch 2 (Hyoid Arch):

  • Reichert’s Cartilage: The cartilage of the second arch.

  • Operculum: An embryonic covering fold formed by Arch 2 that grows over Arches 3 and 4 (along with Clefts II, III, and IV), eventually forming the sides of the neck.

• Branchial Cleft 1: Remains as the External Auditory Meatus.

• Commissure Site: The point where the maxillary and mandibular processes meet (commissura labiorum oris; corner of the mouth).

Research and Clinical Minutes

• Research Minute: DLX Family:

  • In vivo function is studied via "knockout" mice.

  • $Dlx1/2$ knockout mice lacks maxillary molars and has altered craniofacial morphology.

• Clinic Minute: Treacher-Collins Syndrome (Mandibulofacial Dysostosis):

  • Underdevelopment of the mandible and craniofacial region.

  • Caused by impairment/failure of NCC migration to the face.

  • Mutations: $TCOF1, POLR1C$ (TCS3, Autosomal Recessive), or $POLR1D$.

  • Symptoms: Downward slanting palpebral fissures, colobomas of lower eyelids, facial bone hypoplasia, cleft palate, ear malformation/atresia, hearing loss.

• Clinic Minute: OMIM (Online Mendelian Inheritance in Man):

  • Database (omim.org) of human genes, mutations, and diseases; useful for tracking genetic heterogeneity in syndromes like Treacher-Collins.

Questions & Discussion

• Quick Quiz 1: (Reference to Socrative content throughout the first half of the lecture).

• Quick Quiz 2: (Reference to Socrative content involving Branchial arches and NCCs).

• Identifying Images: Students must identify structures such as Rathke's pouch, the stomodeum, and the cardiac plate on sagittal sections of 3-4 week embryos.

• Distinguishing Structures: Students should differentiate between the maxillary process, mandibular process, and various branchial clefts in frontal or lateral views of the 4th-week embryo.

• Operculum Role: Confirming that the operculum (from Arch II) causes the disappearance of Clefts II, III, and IV.

• Derivatives: Identify that Branchial Cleft I becomes the external auditory meatus.