ERS27 Molecular Mechanism Of Growth Control Flashcards
When to start / stop growing
- Contact inhibition
- Supply + Demand: Energy as focus
- Cell division counting
- Cell mass sensor
- Negative cellular growth regulator e.g. Tuberin (TSC2)
Hierarchy of size control
Final size of organ / organism determined by:
- Number of cells
- Size of cells
- Space between cells (~ in different animals)
3 main factors: 1. Growth factor (e.g. Insulin, IGF) —> Protein synthesis —> 控制Cell growth —> 決定Cell size
- Survival factors (IL3) / Developmental cues
—> DNA fragmentation / Protein degradation
—> 控制Cell death / Apoptosis
—> 決定Cell number - Mitogens (e.g. EGF, PDGF)
—> DNA replication / Cell-cycle progression
—> 控制Cell division
—> 決定Cell number
1+2+3 —> affect Organ size —> Organism size
Growth of organ
↑ in cell size + ↑ in cell number
GH-IGF1 axis
GH: - most important regulator of linear growth - from Somatotroph of Anterior pituitary - secreted in ***Pulsatile manner —> bind to GHR in liver —> IGF1 secretion —> GH / IGF1 —> growth, proliferation, metabolism stimulation, apoptosis inhibition
Under control of:
- GHRH (stimulatory)
- Ghrelin (stimulatory)
- Somatostatin (inhibition of Somatotroph)
Affected by:
- Nutrition
- Other hormones: **Sex steroids, **Thyroid hormone, Glucorticoid etc.
- Epigenetic
- Negative feedback by IGF1
Approach to child with short stature
History:
- Perinatal history (e.g. intrauterine growth retardation)
- Birth weight
- Growth pattern, Growth velocity (from previous growth record, know when start of stunted growth)
- Mid-parental height (MPH, from family history, indicate genetic potential)
- Symptom suggestive of systemic illness
MPH:
Boys: (sum of parents height + 13) / 2
Girls: (sum of parents height - 13) / 2
P/E:
- Dysmorphic features
- Nutritional status
- Pubertal assessment
Tests for suspected GH deficiency:
- IGF1 screening
- level in blood more stable —> can spot check
- ↓ in growth velocity / growth centile
- short stature - NOT spot GH
- Normal GH secretion is ***pulsatile (4-6 hours per 24 hours) —> random single GH measurement NOT useful - GH stimulation tests (Pharmacological tests) —> **Definitive diagnosis
- **Clonidine, L-dopa, Propranolol, **Arginine, **Glucagon and Insulin-induced hypoglycaemia
If confirmed GH deficiency
—> ***rule out Pituitary lesion (e.g. Craniopharyngioma)!!!
—> MRI pituitary
Rmb: Poor growth / Short stature = Sign / Symptoms =/ Diagnosis!!!
Rationale of GH stimulation tests
Factors that stimulate GH secretion:
- α-Adrenergic signals
- Amino acids
- Hypoglycaemia
Based on molecular mechanism of GH secretion
—> make use of factors that stimulate GH secretion
—> check blood serially every 30 mins for 2-3 hours
—> check if GH ↑
- Clonidine stimulation test
- Clonidine: α-2 agonist (anti-hypertensive) - Arginine stimulation test
- Glucagon stimulation test
- make use of sharp drop in glucose level after glucagon effect wean off (~ hypoglycaemia) - Insulin tolerance test (obsolete)
Diagnosis of GHD in children
- Clinical + Auxological assessment
- Biochemical tests
- Radiological evaluation
Confirmed GHD
—> Isolated vs With multiple pituitary hormone deficiency (MPHD)
—> Look for underlying cause (e.g. pituitary lesion?)
GH stimulation tests: - Non-physiological (i.e. carry risks) - Carefully consider the need —> only consider when: 1. Auxological criteria - Severe short stature (height < -2.5SD / ↓ growth velocity) OR - Height percentile deviated downward for >= 2 major centiles 2. Low IGF-1 / IGF-BP3
Other important hormones for growth
- Thyroid function
2. Sex hormones (LH, FSH, Estrogen, Testosterone)
Treatment of GHD
GH injection
- daily at night
- until growth plate fused i.e. ↓ growth velocity
Laron Dwarfism / GH receptor deficiency
- GH insensitivity
- ***Mutation of GH receptor gene —> Mutated GH-R —> cannot produce IGF1
- Autosomal recessive
- **Low IGF1 —> ↓ -ve feedback —> **High GH
- ↓ growth, proliferation, metabolism, ↑ apoptosis
Molecular mechanism of GH-IGF1 axis
GH binds to GH-R —> activate ***JAK2 —> self-phosphorylate —> 2x STAT proteins bind to phosphate —> STAT become phosphorylated by JAK to form a dimer —> ***STAT dimer enter nucleus —> bind to DNA —> target gene transcription —> IGF1 production —> transported in blood as IGF1/IGFBP3/***ALS ternary complex —> bind to IGF receptor —> growth
Examples of conditions:
GH insensitivity:
1. GH-R
- Laron Dwarfism
- Intra-cellular GH signalling pathway
- e.g. STAT5b mutation
—> ***Post natal growth failure, immunodeficiency (e.g. recurrent chest infection) - Growth factors synthesis
- IGF1 gene problem
—> ***Pre + Post natal growth failure, microcephaly, deafness, dysmorphism - Transport / Bioavailability of growth factors
- ALS (acid labile subunit) deficiency
IGF1 insensitivity
- IGF-R problem
- ***Severe Pre + Post natal growth failure
IGF-1 mutation
Biochemistry:
- ***Undetectable IGF-1
- ***Elevated GH
S/S:
- severe ***intrauterine growth retardation
- ***microcephaly
- ***postnatal growth failure
- severe psychomotor retardation (affect brain development)
- sensorineural deafness
- mild dysmophic features (micrognathia, ptosis, low hairline)
Treatment:
- ***rhIGF-1 (GH replacement no use)
Signaling pathways of GH
- Extent of individuals pathways varies between cell types
- Depends on relative expression of component parts at different stages of life
IGF1 and PI3K/AKT/MTOR pathways
IGF1 bind to IGF1-R
—> activate kinase receptor
—> recruitment of PI3K to receptor complex
—> ***activated PI3K phosphorylate PIP2
—> activate PDK1, Akt
—> phosphorylation / inhibition of downstream substrate
PI3K/AKT/MTOR pathway
- most common mutation in cancer
- cause a spectrum of overgrowth syndromes
- mTOR pathway receives upstream inputs from:
—> PI3K pathway / Unknown sensors of nutrients, glucose, energy
—> PI3K signal transmitted through:
- Akt phosphorylating **TSC2 (TSC: Tuberous sclerosis protein —> Tumour suppressor)
—> **inhibition of TSC2
—> activation of mTOR - Akt phosphorylation —> direct activation of mTOR
- PDK1 phosphorylating p70-S6K (mitogen-activated Ser/Thr protein kinase)
Tuberous Sclerosis (結節硬化)
- TSC1/2 mutation —> too much mTOR activation —> abnormal cell growth/proliferation
- Rare multisystem AD genetic condition
- non-cancerous tumours in brain / other vital organ
Clinical features:
ASHLEAF
1. Ashleaf spots
2. Shagreen patches (鯊魚皮斑病變)
3. Heart rhabdomyosarcoma
4. Lung hamartoma
5. Epilepsy due to cortical tubers
6. Angiomyolipoma in kidney
7. Facial angiofibroma (acne-like)
Treatment:
Rapamycin (Sirolimus) / Everolimus (2nd gen rapamycin derivative)
—> suppress mTOR pathway
Bone growth
- Intramembranous ossification
- Flat bones - Endochondral ossification (Main determinant of final stature)
- Appendicular bones + Axial skeleton
- transformation of Cartilage (at growth plate) to Bone
Growth plate
3 distinct zones
- Resting zone (最出)
- Proliferative zone
- Hypertrophic zone (最入)
Signaling pathways regulate chondrocyte transition through growth plate:
Less differentiated cells (Resting zone)
—> enter Proliferative zone
—> divide perpendicularly (上下) to plane of growth plate (橫)
—> intercalate with each other
—> form pillars of discoid chondrocytes (Hypertrophic zone)
Known signaling pathways: 1. IHH 2. PTHrP 3. ***FGF 4. C-type natriuretic peptide (CNP) 5. TGF-β 6. Bone morphogenetic protein (BMP) 7. Notch 8. WNT (canonical, noncanonical) —> ***all aid / guide chondrocytes through growth plate —> ***regulate functions in perichondrium / periosteum —> pathways implicated in multiple skeletal dysplasia
FGF (Fibroblast growth factor) Signaling
- Important functions in developing / adult organism
- 18 FGF ligands, 4 FGF-R
FGF ligands:
- ***polypeptide growth factors
- regulate developmental processes (e.g. cellular proliferation, differentiation etc.)
FGF-R:
- ***Tyrosine kinase receptor
—> 1 heparin-binding sequence
—> 3 extracellular immunoglobulin-like domains (D1eD3)
—> 1 hydrophobic transmembrane domain
—> 1 split intracellular tyrosine kinase domain
FGF-FGFR signaling:
- critical to developing axial / craniofacial skeleton
- esp. **Intramembranous ossification of cranial bones + **Cranial suture homeostasis
- Adult: Tissue repair
- follow 1 of 3 transduction pathways:
1. RAS/MAP kinase
2. PI3/AKT
3. PLCg - each pathway likely regulates specific cellular behaviour
- inappropriate expression / activation of FGF / FGFR —> Unregulated cell growth, Tumorigenesis
- aberrant signaling —> implicated in many skeletal abnormalities e.g. Achondroplasia, Craniosynostosis
Process: FGF bind to FGF-R —> ligand-dependent dimerisation —> FGF binding complex (2x FGF, 2x heparin sulfate chains, 2x FGF-R) —> 1. RAS/MAP kinase - start upon FRS2 complex formation - ***Cell proliferation + differentiation
- PI3/AKT
- start upon FRS2 complex formation
- ***Cell survival + Fate determination - PLCg
- start upon binding of PLCg to activated FGF-R
—> DAG + IP3
—> PKC activation
- ***Cell morphology + Migration + Adhesion
**Overall: Regulation of Proliferation + Differentiation + Apoptosis of **Chrondrocytes
Skeletal dysplasias related to FGFR3 mutation
(1. Achondroplasia
2. Hypochondroplasia
3. Thanatophoric dysplasia)
- Autosomal dominant / Sporadic
- Short stature
- Abnormal body proportion
- Small rib cage, underdeveloped lungs
- Abnormal skull base development —> Foramen magnum stenosis / Brainstem compression
Achondroplasia (侏儒症)
- Commonest skeletal dysplasia
- AD / Sporadic
- 125cm for males, 120cm for females
Pathogenesis:
FGFR3 gain-of-function mutation (Overactivation of FGFR3 —> impair bone growth)
—> substitution of **Arginine for Glycine (G380R) within transmembrane domain
—> introduction of hydrophilic residue into hydrophobic receptor domain
—> alter signal transduction pathway due to disruption of α helical structure of transmembrane protein
—> **impair chondrocytes within growth plate
—> FGFR3 primarily expressed in proliferating chondrocytes
—> GOF mutation —> Negative regulatory functions on endochondral ossification (i.e. ↓ inhibition???)
—> **limited chondrocyte production + **limited chondrocytes maturation / ↑ size + ***limited chondrocyte converted to bone
Clinical features:
- Disproportionate short stature
- short limb esp. proximal segments (Rhizomelic shortening)
- long trunk deformed by excessive lordosis
- narrow thorax
- large head with frontal bossing
- hypoplastic midface (flat nasal ridge)
- small craniocervical junction (can cause brainstem compression) - Back, spine complications
- Bowed legs
- Dental overcrowding
- Obesity
- CVS complications
- Ear infections, sleep apnea
Treatment: 1. Limb lengthening surgery —> external fixators placed proximal and distal to osteotomy —> distraction —> extend bone length
- Attenuation of FGFR3 signaling of chondrocytes within physes
—> FGFR3 decoy receptor (sFGFR3)
—> CNP (C-type Natriuretic Peptide) Analog (Vosoritide daily SC) - GH replacement NO use (∵ abnormal skeletal development)
Attenuation of FGFR3 signaling of chondrocytes within physes
-
**FGFR3 decoy receptor (sFGFR3: lacks transmembrane domain —> secreted from cells —> unable to activate signaling cascade)
—> **avoid FGF ligand bind to actual receptor
—> ***prevent activation of mutant FGFR3
—> no inhibition of bone growth -
**CNP (C-type Natriuretic Peptide) Analog (Vosoritide daily SC)
- produced within cartilage growth plate
- works as bone growth promoter —> **opposite effect of FGFR3
- CNP bind to receptor of chondrocytes
—> **inhibit MAPK pathway at level of RAF
—> **indirectly ↓ FGFR3 pathway activity (not via STAT1 cascade) - Statin
- unknown mechanism
- accelerate FGF3 degradation on chondrocytes - Meclozine (anti-emetic)
- unknown mechanism
- in vitro effects on chondrocyte proliferation, differentiation
***Summary
Growth require interactions between
- Gene
- Metabolism
- Nutrition
- Hormones
Molecular mechanisms of growth:
1. GH-IGF1 pathway —> JAK-STAT pathway
- IGF1 —> PI3K/AKT/mTOR pathway
- inhibition of TSC: Tuberous sclerosis protein —> mTOR activation —> abnormal cell growth / proliferation —> Tuberous sclerosis - Bone growth (Chondrocytes) —> FGF/FGFR pathway
- RAS/MAP kinase
- PI3/AKT
- PLCg
—> FGFR3 gain-of-function mutation —> impair chondrocytes proliferation
