2014 Group Project 8

2014 Student Projects
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The Group assessment for 2014 will be an online project on Fetal Development of a specific System. This page is an undergraduate science embryology student and may contain inaccuracies in either description or acknowledgements.

Musculoskeletal

This webpage will be focusing on fetal muscular development.

Muscle development General Timeline

Primary myofibers develop in first trimester, Secondary myofibers develop during Second and Third Trimester We should do a brief overview of primary myofiber formation, and extensive overview of secondary myofiber formation)

Background Embryonic development

Mesenchymal progenitor cells from somites(occiptal, cervical, thoracic, lumbar, sacral), undergo multiple differentiation stages to create muscle fibers.

http://www.mdconsult.com/books/figure.do?figure=true&eid=4-u1.0-B978-1-4377-2002-0..00015-1--f0025&sectionEid=4-u1.0-B978-1-4377-2002-0..00015-1&isbn=978-1-4377-2002-0&uniqId=464007141-2 (demonstration of myotomes in week 6 and 8)

Skeletal muscle Muscle develops from a process known as myogenesis. Mesenchymal cells differentiates into embryonic muscle cells, myoblasts. Myoblasts fuse and elongate to form myotubes which are multinucleated and cylindrical.

The neural tube and notochord release signaling molecules like Shh, Wnts and [BMP]-4. These signaling molecules act on transcription factors of the MyoD family and Pax .The MyoD or MrF family includes MyoD, Myf-5 , myogenin and Myrf4 . The MyoD family of transcription factors, like MyoD are myogenic bHLH (basic helix loop helix) transcription factors. Pax-3 and the MyoD induce myogenesis, formation of myoblasts. Pax-3 also acts on c-met which is a migratory peptide. Skeletal muscle is derived from the somites. Paraxial mesoderm segments into somite structures on both sides of the notochord and neural tube. Somites are mesodermal structures where the dorsal most end of the somite, which is known as the dermomyotome, becomes skeletal muscle and dermis . A small anterior portion of the paraxial mesoderm remains un-segmented and eventually forms some muscles of the head .

The myotome lies in-between the scleratome which is ventrally located. The scleratome forms the cartilage and bone of the axial skeleton of the embryo. The dermomyotome is located dorsally and forms the first skeletal muscle in the embryo. The medial part of the dermomyotome forms the dorsal and intecostal muscles whilst the lateral part of the dermomyotome forms the limb and ventral muscles .

Myogenesis occurs in two phases; primary and secondary which occur in embryonic and fetal periods Primary myotubes express MHC slow myosin heavy chains . The primary myotubes form the structure and scaffold upon which secondary myotubes form during secondary myogenesis which occurs in the fetal period .

Molecular and Cellular regulation of fetal myogenesis

Skeletal myofiber number is set at birth . This was found using experiments on mice and pigs. Similar trends have been observed in humans as found by Widdowson et al (1972) where a huge increase and then levelling of gastrocnemius myofibers were found in the gestational period .

Skeletal muscle development Multipotent mesenchymal cells (MSC) form myoblasts as well as adipocytes and fibroblasts . Therefore specific the nutrients and growth are important in directing the growth of these MSC into either adipocytes or myoblasts . Myoblasts differentiate form the embryonic, fetal and adult skeletal muscle and in fact myoblasts differentiate to form either three early on. Myoblasts fuse and form primary myoblast or myotubes. As discussed previously the primary myotubes form the scaffolding for which the fetal myoblasts will differentiate into secondary myotubes and add too . There are a greater deal of secondary myofibers than primary and are receptive to nutrients and growth factors. Muscle regulatory factors (MRFs) control the proliferation of secondary myoblasts. MRF’s are helix-loop-helix transcription factors. Myoblasts early on express PAX7 and MYF5. PAX7 and MYF5 act on regulatory proteins of the cell cycle like CDK4 and Cyclin D1. CDK4 and Cyclin D1 dephosphorylate RB. RB when phosphorylated is active and inhibits the cell-cell progression in the cell cycle and hence inhibits proliferation. Therefore Cyclin and CDK4 in this case induces cell proliferation. ; , These creates more fetal myoblasts committed which are committed or programmed to form fetal or secondary myofibers later on. These committed myoblasts express MRF, MYOD. MYOD acts on myostatin to take the myoblast out of the cell cycle and stop it from proliferating ; .Now the myoblast is ready for differentiation. The phosphorylation of RB at this period helps to slow the activity of the cell cycle. The myoblasts differentiate and fuse to become multinucleated myotubes as discussed previously. In the formed myotubes MRFs like Myogenin and MRF4 act to inhibit cell cycle and proliferation by acting regulatory proteins like P21 to stop the cell cycle ; This allows the conversion of the myotube into a myofiber. The myofiber can express desmin an intermediate filament that is increasingly seen as gestation progresses. Growth factors, amino acids and stretch/load activity act on myofibers to affect subsequent hypertrophy maturation of myofibers

Myofiber hypertrophy is occurs when protein synthesis is greater than protein degradation. The resulting accumulation of protein results in hypertrophy. Therefore maintaining protein levels is important in the hypertrophy of myofibers. This is regulated by nutrients and growth factors (GFs). Growth factors There are a large number of growth factors that affect fetal myogenesis. Some of the main GFs as Brown (2014) describes in her article “IGF1, insulin, basic fibroblast growth factor (bFGF), transforming growth factor-β (TGF- β)” ; , . Experiments have shown that if IGF1 is removed reduced muscle mass and hypoplasia has resulted. And increased IFG1 expression has resulted in hyperplasia and increased skeletal muscle. IFG1 has also been found to enhance protein synthesis. ; , . Insulin similarly has been found to promote fetal muscle growth and protein synthesis. Basic fibroblast growth factor (bFGF) and transforming growth factor-β (TGF- β) induce proliferation and myogenesis by upregulating cyclin D.

Nutrients Being the building block of proteins, amino acids are important in muscle protein synthesis. But they are more important in adult muscle protein synthesis than fetal muscle protein synthesis. Experiments have shown for example an amino acid infusion did not always result in a fetal muscle growth, only when there was a rise in the insulin levels. And little is known of the interaction of amino acids and growth factors in the context of fetal myogenesis. Research has shown that fetal or secondary myofibers are more prone to suffer from nutrient deficiency than primary myofibers in pigs and sheep. , , .

Fetal myofiber number have been observed to decrease with detal nutrient deficiency. This is concerning as myofiber numbers are set at birth. Nutrients are also important for fetal myofiber hypertrophy. Stretch and loading Stretch and loading also affect hypertrophy. .

Tendon Development

Tendons are connective tissue which join muscle and bone allowing the transmission of force. Tendons primary embryonic structure originate from mesenchymal progenitor somite cells with further contributions of Neural Crest and Lateral plate mesoderm.

Appearance of tendons begins in the 20th Carnegie stage and marks the beginning of fibrillogenesis. This process is initiated by fibroblasts in series of extracellular compartments; they enlarge the cells domain into extracellular space. Channels deep in the cytoplasm drive the process of elongation, these channels location is associated with Golgi bodies.

First type of compartments are formed by collagen containing secretory vacuoles which fuse with surrounding cell membranes. Initial fibrillogenesis is mediated my macromolecular interactions based on vacuole content, with a lesser input from receptor membrane interactions. Fibril groups as fibres close to the cell surface and Secondary extracellular compartments form; at this stage fibroblast are arranged adjacently. Third level of compartmentalization forms later when fibroblast are adjacent with 2 or more other fibroblasts. As the tendon matures fibres coalesce invading each other with interdigitating processes. Secretory vacuole persist, laterally aggregating with further growth preserving sites for fibril deposit. Collagen fiber assembly branch creating fibre networks along fascicles. Fibres branch within tendon fascicle

The main regulatory factors of tendon fibrillogenesis are Leucine-rich repeat proteoglycans.

Non-fibrillar components	Molecular Characteristics	Function
Fibromodulin	Keratin sulphate, proteoglycan	Fibromodulin modulates the site-specific cross-linking ultrastructure of collagen, ensuring mechanical strength. Control the pattern of lysyl oxidase-mediated collagen cross-linking by reducing access of the enzyme to telopeptides, by binding to the collagen.
Decorin	Chondoirin sulphate, proteoglycan	Regulates expression of multiple leucine-rich proteoglycansins(SLRP) during tendon fibrillogenesis, via Class I and II Small(SLRP). Competes with Biglycan for binding sights on collagen types I-VI . Concentration increases as fetal development continues.
Biglycan	Chrondroitin sulphate and Dermatan sulphate, proteoglycan	Process of regulation closely mimic Decorin though concentration is maximal expression at day 16-18 during embryonic development, reducing after this point. Competes with Decorin for bonding sights on collagen I-VI.

Historical findings

Gluteus Maximus muscle Morphogenesis

A 1985 study investigated the development of Gluteus Maximus during the embryonic and fetal periods. Pelvic micro-dissection of human embryos and foetuses with crown length varying from 22-215mm were compared with newborns and adults. The presence of a muscle not present in post-natal adults was discovered called the coccygeofemoralis or pars coccygea; this muscle originates from sides of coccyx and inserts onto the gluteal tuberosity. In the adult human this muscle fuses with the larger pars sacroiliac or fetal gluteus maximus to create the adult gluteus maximus. Pars sacroiliac originates from ilium and sacrum and inserts onto the gluteal tuberosity.

In fetus with crown length of less than 45 mm the two muscle primordia are separated by small amount of loose connective tissue. This point onwards the muscles become fused by a small furrow which persists until 215mm crown length. By the time of birth the furrow is absence and the muscles are entirely fused . The coccygeofemoralis in long tailed mammals remains separate from the gluteus maximus, known as the caudofemeralis muscle . In these animals its function is lateral flexion of the tail . This is the sole morphogenesis study on any of the large muscles; it is likely other muscles with multiple origins and insertion have separate fetal muscle primordia.

Morphogenesis of human sphincter urethrae muscle

This study was completed in 1989 at the same Czechoslovakian institute of that of Gluteus maximus morphogenesis. . Study displayed three developmental phasess differentiating them by morphogenesis, histology and sexual dimorphism. External urethral sphincter in embryos and fetuses with crown length varying 18-320mm, neonates, children and adults was fixed in formaldehyde and embedded in paraplast. Then cut in series and stained with hematoxylin and eosin for histological analysis. Morphogenesis and Sexual dimorphism specimens were micro-dissected using a stereomicroscope.

Phase	Time Period	Characteristics
Indifferent Phase	Before 10th week	Muscle primordia discernible from neighbouring muscle by week 8. Grow to forms a shallow arch; connecting the urethra and urogenital diaphragm. Consists of condensation of myoblasts up to 9.5 weeks, past this point myotubes and muscle fibres appear.
Sexual dimorphic Phase	10th week to Birth	Associated with the development of prostate and vagina. Primordia spread along urethra wall posteriorly. In Males: Spreads to create the infraprostatic part of external urethral sphincter. Arches anteriorly to join prostate and urethra In Female: Spreads to create upper part of external urethral sphincter. Lower sixth of sphincter connects anterior and lateral urethral walls, additionally projects to lateral vaginal walls.
Definite Structuring Phase	After birth	Position of urethral sphincter does not alter in relation with prostate and inferior part of vagina. Infraprostatic region in males and upper part in females grow to form a complete ring.

Abnormalities

Duchenne Muscular Dystrophy(DMD)

Caused by a mutation of the dystrophin protein on locus Xp21; this protein complex connects the cytoskeleton of muscle fibres to the extracellular matrix. Abnormal dystrophin results in a degradation of cellular integrity, excessive penetration of sarcolemma by calcium and water entering mitochondria increasing pressure and bursting. The lack of significant load during fetal development results in minimal wasting, it is detected in postnatal babies at 3-5 years old with muscles resisting gravity being the first to waste. Patients are restricted to wheelchairs by their early teens and have a life expectancy of 25 years. The incidence in male infants is 1 in 36,000. Typically males are affected while females are carriers. In the offspring of a carrier mother and unaffected father; sons have 50% chance of affected and daughter’s 50% chance of becoming carriers. Since the disease is a terminal illness killing in mid-twenties; affected fathers are not considered in the situation. Absence of affected fathers means it is very unlikely for daughters to be affected.

Duchenne Muscular Dystrophy has no present cure, treatment is aimed at altering onset of symptoms and maximising quality of life, these include:

• Corticosteroid based medication significantly improves muscle strength and function over the short term. Patients using the drugs long term gradually have reduced doses to avoid severe side effects which can include; weight gain, behavioural disorders and osteoporosis. Most effective corticosteroids are prednisolone, and deflazacort. Theories on how the steroids work include; include activation of T-Cell pathways, directly reducing muscle regeneration, modulating cell inflammation and enhancement of myogenic precursors.

Amyoplasia

Amyoplasia is characterised by replacement of newborn muscle tissue with fat and dense fibrous tissue. Is the most common of Arthrogryposis multiplex congetia disorders; these result in multiple joint contractures. Affected limbs have significantly altered positioning, typically clubfoot is present and elbows are extended. Primary cause is limited fetal movement, muscle is replaced by dense fibrous tissue. Conditions which limit fetal movement include abnormal uterus morphology and reduced amniotic fluid. Presently no genes have been linked to the deformity. 85-90% of newborns undergo surgery within days of birth primarily on the legs and hips, releasing tendons from contractures. Motion can be improved by casting and splinting, lower limbs are typically cast and upper limbs are typically splinted.

Involvement subtype	Percentage of Case
Four-limb symmetrical	55%
Three limb	5%
Upper limb only	17%
Lower limb only	16%

Types of Muscular Dystrophy; Duchenne Muscular Dystrophy, Becker type muscular Dystrophy, Occulophayngeal muscular Dystrophy,

Absence of specific muscles;

References

Anatomy and variations of palmaris longus in fetuses.

Development of the rectus abdominis and its sheath in the human fetus.

Sonic hedgehog acts cell-autonomously on muscle precursor cells to generate limb muscle diversity.

The normal growth of the biceps brachii muscle in human fetuses.

References to format in later

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