HORMONE DEPENDENT SYSTEMS - mechanisms Flashcards
1
Q
Name three steroid receptor co-activators
A
AIBC (amplified in breast cancer), SRC1 (steroid receptor co-activator 1) and TIF/GRIP (transcriptional intermediary factor)
2
Q
What is the difference between ppar alpha and gamma?
A
alpha - fatty acid uptake, beta oxidation and triglyceride metabolism
gamma - lipid storage, energy expenditure and brown fat beta oxidation
3
Q
What happens to the LBD after the hormone binds?
A
AF2 repositions and folds against the core of the LBD to seal the binding cavity and form a hydrophobic surface that is recognised by co-activators
4
Q
What does the hormone/NR bind to?
A
The hormone response element in the gene promoter
5
Q
What are nuclear receptors?
A
Ligand activated transcription factors
6
Q
What are the actions of co repressors
A
chromatin remodelling by hdac, inhibiting transcriptional machinery, inhibiting DNA binding, competing with co-activators and inhibit RNA processing
7
Q
How is the receptor commonly found?
A
In the cytoplasm in a complex with heat shock proteins such as the dimer hsp 90. Keeps the receptor inactive but responsive to ligand binding
8
Q
What are the three features of the AF1?
A
Ligand independent so has constitutive transcriptional activity, varies between nuclear receptors, though may rely on co-activators or co-repressors
9
Q
What is the difference between AF1 and 2?
A
AF1 is ligand independent versus AF2 is ligand dependent
10
Q
Features of the LBD?
A
Highly conserved, synergises with AF1 to promote transcription and forms a hydrophobic cavity to bind the ligand
11
Q
What are the five genes induced by ER alpha?
A
PR, VEGF, IGF, Bcl2 and cyclin D
12
Q
What are the 3 steps for ligand activated nuclear gene transcription?
A
Repression of COREPRESSORS with HISTONE DEACETYLASE ACTIVITY (HDAC), derepression to form activation complex 1 as ligand binding displaces corepressors and then TRANSACTIVATION, 2nd activator complex coactivators such as TIF and GRIP and interaction with transcriptional machinery
13
Q
General difference between ER alpha and beta
A
Alpha in endometrium, beta in ovary, prostate and brain. Highly homologous but coded on separate chromosomes on separate genes
14
Q
What is the difference between PR alpha and Beta
A
Beta is less transcriptionally active but has AF3
15
Q
What are some non-classical steroid receptor actions
A
Ligands binding to promoters/TF that lack HRE in the promoter gene. 1) ER can act as a co-activator for transactivation at other promoter sites such as c-jun and c-fos 2) PR can suppress the activities of NfKB to modulate immunosuppression in pregnancy 3)PR can repress the activity of Stat 5 and AP1 TF to mediate its actions on proliferation in the mammary gland. Non genomic actions involve interaction with GF.
16
Q
Which apoptotic pathway is more important in cancer development?
A
Intrinsic
17
Q
What is a full oestrogen receptor antagonist and how does it work?
A
FULVESTRANT. Blocks dimerisation and translocation to the nucleus. Blocks AF1 activation and promotes degradation
18
Q
How does tamoxifen work?
A
Partial antagonist. Promotes dimerisation and translocation to the nucleus but then blocks transcriptional machinery by blocking AF2 and the LBD. Additionally promotes co-reporessors such as NCOR and SMRT
19
Q
What is mifepristone?
A
Partial PR antagonist. Promotes dimerisation and translocation but then blocks AF2 and coactivators
20
Q
What is the point of a SARM?
A
To reduce prostatic hyperplasia whilst maintaining muscle strength and mass
21
Q
How may one become resistant to tamoxifen?
A
Increased co-activators AIBC1, reduced co-repressors or p53, increased GF
22
Q
What are the types of nuclear receptor?
A
Steroid, retinoic x, orphan monodimer and orphan heterodimers
23
Q
What is progesterone involved with?
A
ovary - ovulation, uterus - decidualisation, breat - proliferation and lobar development, prevention of bone loss and behaviour
24
Q
What is the difference between PR a ko and beta ko?
A
PRa KO – impaired ovulation and implantation, impaired fertility
PRb KO – reduced mammary gland ductal morphogenesis
25
What happens in decidualisation?
Convergence of the PR classical nuclear receptor pathway and g protein coupled PR
26
What is the key TSG in type 1 endometrial cancer vs t2?
Type 1 - PTEN, 2 - p53. Oncogene ras
27
What do caspases do?
exist as procaspases and lead to proteolysis of key structural elements and activates DNAses
28
What binds to the extrinsic pathway.
TNFR family of which the ligands are TNF and Fas (Fas expressed by cytotoxic t lymphocytes. The intracellular DD of TNFR recruits FADD (adaptor molecule) for caspase 8 cleavage and activation of the caspase cascade. Binding of Fas induced trimeristion of CD 95 forming the DISC. FLIP stops procaspase 8> capsize. Apoptosis occuring is a balance between FADD and FLIP
29
What initiates the intrinsic pathway? What happens in it?
Loss of mitochondrial membrane potential and the main initiator apoptotic protein is cytochrome c. Cytochrome c binds to APAF 1 card (caspase activation and recruitment domain). APAF-1 undergoes an ATP dependent conformational change allowing it to bind to procaspase 9 in the CARD domain and activate apoptosis. Balance between pro-apoptotic Bid, Bad, Bax and antiapoptotic Bcl and BCLx
30
Controlling one area of the cell cycle, what is an initiator of apoptosis?
ATR-ATM detect double stranded DNA breaks that activated P53 that initiates apoptosis
31
What can be used to treat endometrial carcinoma?
Progestins to reverse hyperplasia and induce decidualisation and endometrial thinning
32
What are glycoprotein hormone receptors? Which hormones and how is specificity achieve?
G-protein linked including LH, FSH and TSH. Through the leucine rich repeats in the ectodomain
33
What happens when a ligand binds to the glycoprotein receptor?
Recruitment of alpha, beta and gamma subunits. Alpha subunit releases GDP for GTP, then there is dissociation of the alpha and beta/gamma subunits. Then regulate respective effector proteins. The serpentine transmembrane domain transmits the signal the g protein receptor.
The ectodomain acts as an inverse agonist
34
Discuss the TSHR. Where is it found? What does it act as and do? What do germline, both activating and inactivation mutations cause? other factors
It is a glycoprotein g protein linked receptor that is found in the thyroid, thymus, brain, pituitary and lymphocytes. Acting as a dimer, it stimulates thyroid cell proliferation and differentiation through transport of sodium-iodide transporter, thyroperoxidase and thyroglobulin.
Germline activating mutations lead to autosomal dominant non autoimmune hyperthyroidism whereas inactivating lead to TSH resistance (congenital hypothyroidism).
TSHR stimulating antibodies - graves, blocking is hashimotos hypothyroidism. Can by activated promiscuously by bhCG in pregnancy for gestational hyperthyroidism
35
What do glycoprotein hormones consist of?
They are heterodimeric and consist of a common alpha subunit and varying beta
36
What can thyroid hormones do?
Heterodimerise with the retinoic x receptor
37
Describe the DBD?
2 zinc fingers each with 4 conserved cysteine residues, The nuclear receptors can bind as homodimers, heterodimers or monomers. The DNA helix makes base specific contacts in the major groove.
38
Describe the LBD
Highly conserved amongst receptors. WEDGE SHAPED HYDROPHOBIC CAVITY which contains the ligand binding site and is recognized by co-activators. Synergises with AF1 to promote transcriptional activation. It displaces co-repressors and activators. Allows heterodimerization
39
What do steroid hormones mainly bind as?
Homodimers
40
Where are steroid receptors found
In the nucleus or the cytoplasm
41
What is the androgen receptor like?
Androgen receptor: a single isoform that is responsive to testosterone and 5DHT
42
What are three classic non-classical activities of the steroid receptors?
Ligands binding to promoters/TF that lack HRE in the promoter gene. 1) ER can act as a co-activator for transactivation at other promoter sites such as c-jun and c-fos 2) PR can suppress the activities of NfKB to modulate immunosuppression in pregnancy 3)PR can repress the activity of Stat 5 and AP1 TF to mediate its actions on proliferation in the mammary gland. Non genomic actions involve interaction with GF
43
What can steroid receptors do, non genomically?
STEROID RECEPTORS AND GROWTH FACTORS CROSS TALK.
EGF signaling can either occur through Ras/MAPK or through PI3/AKT. It is mitogenic in breast or uterus and its signaling is enhanced by oestradiol or progesterone. Membrane associated ERalpha can activate EGF family of receptors.
ER acts synergistically with IGF1 and insulin to promote tumour cell division in utero. Serines in ERalpha can be phosphorylated by activated MAPK which makes it hypersensitive to oestradiol
44
What evidence is there that oestrogen and egf cross talk?
EVIDENCE: UTERINE AND BREAST TISSUE PROLIFERATIVE RESPONSE TO EGF IS BLOCKED BY OESTROGEN SIGNALLING BLOCKADE, and the inverse that response to oestradiol is blocked by blocking egf signalling.
Er alpha directly communicates with p85 subunit of PI3K
45
What factors can oestrogen amplify>
Mapkinase signalling through EGF and PI3 kinase signalling through IGF1
46
What makes oestrogen receptors hypersensitive to E2? What is this similar to?
Serines in ERalpha can be phosphorylated by activated MAPK which makes it hypersensitive to oestradiol. What happens in progesterone
47
What is one mechanisms by which oestrogen receptor cross talk occurs?
Er alpha directly communicates with p85 subunit of PI3K
48
What is c-Src essential for?
The non genomic proliferative effects or ER, AR and PR
49
What are the 2 main co-repressors?
NCoR: nuclear receptor corepressor
| SMRT: silencing mediator for retinoid and thyroid receptor
50
Overall how do steroid receptors work?
* lipophilic steroid hormone passes across cell membrane
* steroid hormone is bound by receptor
* ligand-activated receptor localises in the nucleus
* ligand-activated receptor binds to SRE
* ligand-activated receptor initiates DNA transcription
51
What is the role of ER alpha vs Beta?
ER alpha and beta are highly homologous but are coded on separate genes, can form heterodimers and have different AF1 binding domains. Alpha more in endometrium and beta more in ovary, prostate and brain. Very similar DBD but only 56% in LBD
52
What genes does ER alpha induce?
Genes induced by ER alpha. PR, VEGF, IGF, Bcl2 and cyclin D
53
AF2 role?
AF2 activity requires ligand binding which enduces a conformational change that activates the LBD. AF2 repositions and folds against the core of the LBD to seal the binding cavity and form a hydrophobic surface that is recognised by co-activators
54
How do direct oestrogen antagonists vs agonists work?
OESTROGEN DIRECT ANTAGONISTS -bind to the LBD and prevent h12 from adopting the agonist confirmation, h12 is positioned over the site where co-activators would normally contact the LBD
INDIRECT – lack a side chain that displaces H12
55
Describe the progesterone receptor?
2 isoforms (alpha and beta) with similar binding affinities. They are derived from the same gene with alternate promoters. Beta generally less transcriptionally active but has AF3 and a proleine rich sequence. PR is required for side branching In Puberty and Alveologenesis in Pregnancy.
56
What happens in PR KO models?
PRa KO – impaired ovulation and implantation, impaired fertility
PRb KO – reduced mammary gland ductal morphogenesis
57
How can progesterone derivatives be used in cancer?
PROGESTINS CAN BE USED TO TREAT ENDOMETRIAL HYPERPLASIA BY INDUCING DECIDUALISATION AND ENDOMETRIAL THINNING.
58
How does egf contribute to steroid receptors?
Ras/MAPK or through PI3/AKT. It is mitogenic in breast or uterus and its signaling is enhanced by oestradiol or progesterone. Membrane associated ERalpha can activate EGF family of receptors
59
What is EGF?
It is a receptor tyrosine kinase. It is mitogenic in breast or uterus and its signaling is enhanced by oestradiol or progesterone. Membrane associated ERalpha can activate EGF family of receptors.
60
How can EGF be activated by steroids? What else can be activated at this point
EGF receptors can be activated by intracellular tyrosine kinase SRC which in turn can be activated by ER, AR and PR. IGF-1 pathway
61
How does steroidogenesis occur?
Only the dominant follicle gets LH receptors in granulosa cells.
LH stimulates the theca cells to produce androgens.
FSH (receptors in granulosa cells) converts androgens to oestrogens by aromatase
62
What is the precursor for steroidogenesis?
cholesterol available for steroidogenesis: cholesterol is stored in the cell as lipid droplets which can be made into free cholesterol. It can also come into the cell via LDL receptors through endosomes or made de novo via endoplasmic reticulum.
63
What are the key regulators of steroidogenesis
Gonadotrophins. • IR/IGFR stimulation is part of the MAPK and PI3K pathway which stimulates steroidogenesis.
• There is also interaction of PDK by IGFR/IR which can go and activate PKA and this stimulates steroigogensis as well. It has been sown that insulin/igfs stimulate steroiogenesis (progesterone
• It has also been seen that insulin amplifies LH induced progesterone production by granulosa cells.
64
What biochemical abnormalities are there in PCOS?
Raised LH and androgens (also insulin increases LH)
65
What does cyclin D do?
cell cycle progression. The synthesis of cyclin D is initiated during G1 and drives the G1/S phase transition. ER alpha codes for it
66
What does oestrogen do to the endometrial lining?
Estrogen promotes proliferation and growth of the endometrial lining
67
How does progesterone signal?
Through the classical steroid receptor pathway (activates CREB and AP1), but can also activate MAPK and PI3K through SRC or Non-genomic progesterone signaling: mPR Membrane-bound PR
G-protein coupled receptor
7 transmembrane domain, that binds progesterone on its extracellular domain
Explains quick non-genomic actions of progesterone
It activates the MAPK cascade
KO of the membrane receptor has no effect
CAMP seems to be needed for progesterone's effects
68
What are the main glycoprotein hormones?
FSH, LH and TSH. common alpha subunit and varying beta
69
What is key in glycoprotein receptors?
G protein without ectodomain is active. The ectodomain shuts this off acting as an inverse agonist. When ligand binds, it becomes an agonist again
70
How does radioimmuno assay work? Key definitions
Amount of bound labelled antigen is inversely proportional to the amount of unlabelled antigen.
Once equilibrium attained, the antibody-bound and free antigen are separated and the radioactivity in the bound fraction is measured.
Efficient separation of the bound and free radioactivity is essential.
Total Counts= total amount of radioactive peptide added to each tube
Non-specific binding= binding of label to tube
(cpm in absence of Ab and standard/unlabelled peptide)
Maximum binding
= binding of radioactive peptide to Ab in the absence of standard / unlabelled peptide
Standard curve
= series of reference standards producing a displacement curve from which the displacement of samples with unknown concentration can be quantified
71
How many ligands are required to activate the glycoprotein family?
1 or 2. TSHR functional dimerization: if you get dimerization with an ectodomain you get no signal (found in the cytoplasm – inverse agonist) If there is no ectodomain you get low grade signalling, Having one ligand activates the receptor a bit (partial agonist), two ligands results in maximal activation (full agonist)- there is interaction between the serpentine domains (TM5 and TM6)
72
Why is the TSHR susceptible to mutations compared to other glycoprotein receptors?
• Unlike other glycoprotein receptors, the TSHR is constitutively active and susceptible to enhanced constitutive activation by mutation and deletions
73
How can the glycoprotein TSH receptor be described
A mutation or ligand binding, an “open” conformation ensues. This 2-state model predicts that the open format of the receptor, when stabilized, would lead to full activation. Support: constitutive activation developed when the TSHR ectodomain was truncated, which suggested that the presence of the ectodomain dampened a constitutively active β subunit
74
What role in embryogenesis does the TSHR have?
• It is expressed from 12 weeks gestation and is essential for terminal thyroid maturation and growth
75
What can bind to the TSHR? What is this termed?
LH and hCG can interact with the TSHR —termed specificity crossover
76
What can be said about HRT, breast and endometrial caner?
Use of tamoxifen in treatment of breast cancer outweighs the risk of developing endometrial cancer (120 vs 6 /1000).
77
What does AKT allow?
Progression to the S phase of the cell cycle
78
What is the difference between the IGF1 and insulin pathways?
THE IGF1 PATHWAY EXACTLY THE SAME BUT NO GLUCONEOGENESIS OR HEPATIC TRIGLYCERIDE PRODUCTION INVOLVED
79
What happens in farnesylation?
GTP-bound ras must become physically associated with the cell membrane for downstream effects on growth and cell division
this requires farnesylation of ras by farnesyl transferase
range of different compounds currently being studied as farnesyl transferase inhibitors
80
What happens in histone modification?
is the mechanism by which DNA methylation modulates gene transcription
proteins that bind to methylated CpG recruit histone methyl transferases and histone deacetylases:
-histone methylation packs the nucleosome to prevent gene transcription which can be retained through cell division
-histone deacetylation packs the nucleosome to prevent gene transcription and aids histone methylation
81
What actions does TSH have on the thyroid?
Stimulates production, release of thyroid hormone and proliferation, growth and terminal differentiation of follicular cells oStimulates thyroid peroxidase (uptake of iodine into the thyroid gland, being incorporated into thyroglobulin) oTSH secreted into the portal system and then peripheral system
82
What does thyroxine travel in the blood bound as, and then what does it bind to?
Bound to TBG or albumin. The TR is A NUCLEAR RECEPTOR
83
How does the T3/T4 enter cell?
Through specific transporters such as MCT8.
84
What determines how much active T3 there is in the cell?
T4 converted to T3 by type 2 diodinase to activate. Type 3 deiodinase (D3) inactivates T4 to produce the inactive metabolite reverse T3 . rT3 cannot bind to the thyroid hormone receptor. ALL THESE DETERMINE HOW MUCH T3 ENTERS THE NUCLEUS THROUGH ACTIVATION AND DEACTIVATION
85
What happens in hyperthyroidism in terms of the thyroid hormones?
Type 3 deiodinase (D3) inactivates T4 to produce the inactive metabolite reverse T3 (removal of 5 I atom) . rT3 cannot bind to the thyroid hormone receptor.
86
What does T3 bind to? What is the receptor like?
•T3 acts through receptors TRAlpha and TRB that are encoded by THRA and THRB. TRalpha is more ubiquitous. Don't forget it is a nuclear receptor and so only binds T3, T4 is a pro hormone. Unlike the steroid receptors, the TR is not bound to heat shock protein
87
Within the nucleus, what does TR recognise?
Thyroid R recognizes TREs located within the target gene
88
What do thyroid receptors bind with?
Thyroid receptors bind with retinoid X receptors as heterodimers to promoter region
89
What happens in the absence of a ligand in terms of thyroid receptors?
In the absence of hormone, the thyroid receptor still enters nucleus (unlike GR) and forms heterodimer with RXR and binds to direct repeat sequence in promoter region
In absence of hormone, it acts as a repressor: recruits NCoR, which recruits other proteins with histone deacetylase activity. This causes deacetylation of histones- tight packaging of chromatin in target genes, making it inaccessible to transcription machinery.
90
What happens to the TR when a ligand is present?
In the presence of thyroid hormone, binding of the hormone to the receptor causes conformational change in the structure of thyroid receptor. This releases the NCOR corepressor and exposes another binding site for the coactivator SRC-1. Histone acetylation occurs and opens up chromatin and allows transcription to begin.Cofactor exchange to recruit TRAP complex and this fully activates transcription
91
What do T3 and T4 act as?
TR as a repressor and T3 as an inducible TF.
92
What isoforms are there or thyroid receptor?
TRa1 and TRb1 bind to T3 and are true receptors. TRalpha 2 function unknown
93
What are thyroid hormone cell membrane transporters?
MCT8, MCT10 Organic anion transporting polypeptides (OATP), L-type amino acid transporters (LAT1, LAT2) oTransport T4 and T3 oWidely expressed
94
What is MCT8 like? What specific cell is it involved in?
High affinity for T4, T3, rT3, T2. Widely expressed. MCT8 mutations cause severe X-linked psychomotor retardation with elevated serum T3, slightly low T4 and normal TSH
Availability of T3 from astrocytes -> neurons require MCT8 for uptake of T3 from astrocytes
95
What is MCT10 Like?
Transports T3 in preference to T4 oWidely expressed, high levels in cartilage
96
What is OATP like?
OATPC1 has high affinity for T4 and rT3 and facilitates influx and efflux, Restricted expression pattern, Regulates T4 transport across blood-brain barrier
97
Where is D3 particularly important?
D3 inactivates thyroid hormones during pregnancy. Increased level of T3 in placenta, uterus and foetal liver; flow of maternal blood to foetus enters through uterine, placental & liver arteries; thyroid hormones critical for angiogenesis and development; thyroid hormones stop cell proliferation and triggers differentiation; in early pregnancy need proliferation of cells before they differentiate. Then D3 switched off and D2 switched on- molecular switch which triggers increased thyroid hormone levels which stimulates differentiation
98
What is resistance to thyroid hormone?
Autosomal dominant. THRB mutations cause RTH, in which negative feedback regulation of TSH is disrupted
oMutations interfere with T3 binding, co-repressor release or co- activator recruitment
Mutant TR acts as dominant-negative antagonist
Phenotype is variable
THRB mutation absent in 15% of cases
99
How has the thyroid hormone been implicated in cancer?
Cancer
Somatic mutation or aberrant expression of
THRB identified in thyroid, liver and renal cell tumours
100
What about mutations in THRA?
Only 3 patients have been identified since 2012, further work is needed.
101
What happens in hyper/hypothyroidism in children/adults?
In children hypothyroidism causes growth arrest and epiphyseal dysgenesis.
Hyperthyroidism still short stature – accelerated growth but also accelerated epiphyseal fusion. In adults – increased risk of osteoporosis and fracture.
102
Which is worse, thyroid hormone deficiency or a lack of receptors?
ApoTR has shown that absolute hormone deficiency is worse
103
Where are TRa and b found?
* TRalpha- brain, heart, spleen, gut, bone and cartilage
| * TRbeta – liver, hypothalamus, pituitary and cochlea
104
What does deletion of TRalpha cause?
Deletion of TRa Growth retardation Delayed ossification Impaired bone resorption High bone mass (due to impaired bone resorption and turnover). Phenotype like HYPOTHYROIDISM
105
What happens in TRb KO?
impaired auditory evoked brainstem response, absent m-opsin and redistribution of s-opsin cones in retina less formation but increased resorption – PHENOTYPE LIKE HYPERTHYROIDISM. This is because TRbeta is in the pituitary gland so through negative feedback you get increase t3/4. TRB expressed in cochlea and retina and responsible for actions of TH during development of hearing and colour vision
106
Whats the difference between stature in TRalpha and beta KO?
TR alpha knock out causes transiently short stature whereas TRB knock out causes persistent short stature.
107
In the thyroid receptor, what is the ectodomain?
extracellular (ectodomain) part looks like a base ball glove (TSHR has 9 leucine rich repeats and alpha hel and beta pleats) and then there is a trans membrane domain. The leucine rich repeats are the A subunit, then the serpentine domain is the beta subunit that transmits the signal to the g protein
108
What is TSHR required for?
TSH is not needed for early organogenesis and migration of the thyroid to the correct position or for follicle formation. TSHR is expressed from 12 weeks gestation and is needed for terminal thyroid maturation (ability for thyroid follicular cells to make hormone) and growth.
109
What does hypo/hyperthyroidism in adults do to bone?
Adults
Hypothyroidism Increased fracture risk in population studies (increased mineralization of skeleton)
Thyrotoxicosis: Accelerated bone loss, osteoporosis with increased susceptibility to fracture
110
What does hypo/hyperthyroidism in children do to bone?
Hypothyroidism: Growth arrest, delayed bone age, epiphyseal dysgenesis (abnormal formation of growth plates), immature body proportion
Thyrotoxicosis: Accelerated growth (overproduction of TH), advanced bone age, short stature (because growth plates fuse too early), craniosynostosis (due to
abnormally advanced prematuration of skeleton)
111
How do you experimentally measure the effect of TRa and b knock out? What happens?
KO mice and wild type and measure TSH and T3/T4 levels. n TRa KO mouse: T4, T3 and TSH levels are normal. Negative feedback normal and no effect on thyroid status. In TRb KO mouse: disrupt negative feedback of TSH in pituitary. 12 times amount of TSH, drives up T3 and T4- increase approximately 5 fold
112
In TR a and b KO, what happens to osteoclasts?
TRa KO Decrease in osteoclasts TRb KO Increased osteoclasts- bone resorption increases
113
What does TSH do to osteoclasts?
TSH inhibits: osteoclast formation and osteoblast differentiation.
114
What happens in TSHR--mice?
TSHR -/- mice: Thyroid hypoplasia, undetectable T3/T4 but TSH 500x.They have a severe growth delay and die by ten weeks unless given thyroxine. Young ages have high bone turnover osteoporosis.
TSHR +/-: euthyroid, normal growth and intermediate skeletal phenotype
115
What happens in TRa/b KO and FGF levels?
In TR alpha knock out mice you get decreased FGFR1 expression in osteoblasts, unchanged FGFR2 and decreased FGFR3 in chondrocytes.
In TR beta knock out mice you get increased FGFR1 in osteoblasts and in chondrocytes, and increased FGFR3 in chondrocytes.
EXPERIMENTALLY SHOWN by decreased receptor mRNA in bone.
116
Have any TRalpha or beta mutations been identified in humans?
Patient documented in 2012- 9 yrs old Mutation in TRa- identical region to TRa mutant mice Phenotype: oVery short- 1m tall oMarked shortening of distal bones than proximal bones (subischial leg length much lower than sitting height- legs shorter than torso) oProfound constipation- marked transit time in gut. TRa important in gut
117
What is the thyroid/iodine status in pregnancy? What happens if there is insufficient?
What evidence is there in the UK?
increased production of thyroid hormone daily requirement: 150 mcg > 250mcg per day in pregnant and lactating women.
Being iodine deficient will cause inc TSH to be made and goiter formation, worsened brain development (look at neurocognitive scores of infants), increased misscarriage rate and post partum thyroiditis
Looked at urinary iodine excretion in the UK and found that majority of girls in reproductive age were iodine deficient.
118
What naturally happens in pregnancy concerning tsh levels?
In pregnancy thyroid is a vascular organ and blood supply to it increases to allow iodine in blood to be transferred better. This can cause enlargement of the thyroid gland .
hCG promotes maintenance of the corpus luteum during beninning of pregnancy allowing corpus luteum to secrete progesterone.
It has a very small effect on Tsh receptors normally but in pregnancy there is an enormous amount of hCG so this can act on TSH receptors more causing a relative hyperthyroidism which also causes the thyroid gland to grow due to hcg stimulation. the excess stimulation making more t3/t4 causes dec TSH being made.
So in normal pregnancy the TSH levels fall at 10 weeks then come back to normal after pregnancy. 10 weeks is when HCG is most elevated so the relative hyperthyroidism and goiter formation in early pregnancy is due to
this
119
What happens to T3/T4 levels and TBG?
* Normally thryroixine carried in blood by TBG, albumin and transthyretin (unbound 0.04% T4 and 0.5% T3). In pregnancy even more is carried by TBG due to increased oestrogen
* The increase in TBG leads to increased T3/T4 (plateau at 20 weeks and then maintained till full term).
120
What epidemiological data is there of iodine deficiency?
Countries of severe iodine deficiency- Peru, Ecuador, severe deficiency leads to increased proportion of cretinism. Can bring down by giving iodine.
121
What is the placenta permeable to?
* Placenta is permeable to TRH but not TSH
* T4 and T3 do not cross easily except early pregnancy
* Elevated deiodinase activity in the placenta probably plays an important role in metabolism of maternal thyroid hormones. There are 6 thyroid transported detected in the placenta and the plasma membrane allowing differences between maternal and foetal circulation
122
What is iodine normally in an equilibrium between?
• Iodine pool in dynamic equilibrium between kidney and thyroid. In pregnancy, increased renal clearance and thus lower circulating levels of iodine. Thus increase amount of thyroxine entering gland → indicates that elevated thyroid levels in pregnancy
123
What is further evidence of oestrogen and thyroxine?
In oestrogen theray of postmenopausal women, total thryoxine increases
124
If mother is hypothyroid in pregnancy what does this predispose her to?
Post partum thyroiditis
125
What are causes of high and low uptake thyroiditis?
Causes of thyrotoxicosis High uptake Graves’ disease 40-60% Toxic multinodular goitre 30-50% Single toxic adenoma 5% TSH induced Thyroid cancer induced Trophoblastic tumour and Struma ovarii Low uptake Subacute thyroiditis Postpartum thyroiditis Amiodarone Silent thyroiditis (immune and amiodarone)
126
What is the prevalence of hypothyroidism?
2.5%
127
What study for women with TPO antibody in pregnancy?
• Levothyroixine treatment in euthyroid preg women with autoimmune thyroid disease: women with TPO antiboidy with normal TSH due to being given small amounts of levothyroixine and women with TPO antibody with no treatment and women with no TPO antibody. This showed that women with TPO antibody who don't get thyroixine had inc miscarriages and inc preterm deliveries.
128
How do you treat hyperthyroidism in pregnancy?
treat with carbimazole, propylthiuracil, small dose possible cant give RAI, if allergic to drugs may need surgery
129
What other action does oestrogen have on TBG?
Estrogen mediated prolongation of TBG half-life from 15 minutes to 3 days
130
What happens to T3/T4 levels?
They increase following the oestrogen driven increase in TBG
131
What is hyperthyroidism associated with?
Severe maternal hyperthyroidism is associated with increased risk of stillbirth, preterm delivery, intrauterine growth restriction, preeclampsia, and heart failure. Also, thyrotoxicosis at conception increases the risk for spontaneous abortion
132
What does breast ERa and b KO cause?
a - immature ductal system
| b - nothing
133
What happens to fertility in Era and ERb KO?
ERa both males and females infertile, with ErB males are fertile
134
What happens in the pituitary in ERa and b KO?
ERa LH elevated and Prolactin low whilst in Erb nothing
135
What happens in ovary Era and ERb KO?
Elevated oestrogen, follices do not mature, haemorrhagic cystic follicles begin developing at puberty due to LH elevation.
ERB there is a reduced number of corpa lutea, ineffiecient ovulation and normal steroid and gonadotrophin levels
136
What happens in uterus ERa KO?
Immature, insensitive to oestrogen and no epithelial proliferation. ER beta nothing
137
What happens in aromatase KO?
Immature mammary glands,
Immature uteri,
haemorrhagic cysts,
males infertile due to loss of spermatids
Deficiencies is sexual behaviour in males (mounting) - also seen in ßERKO
138
What does aromatase convert?
In post-menopausal women androstenedione conversion occurs at peripheral sites such as fat, liver, muscle
139
What two different types of aromatase inhibitors do you have?
Suicide inhibitors initially compete with the natural substrate (i.e., androstenedione and testosterone) for binding to the active site of the enzyme. The enzyme, then form covalent bonds at or near the active site of the enzyme. Through this mechanism, the enzyme is irreversibly inactivated.
Competitive inhibitors bind reversibly to the active site of the enzyme and prevent product formation only as long as the inhibitor occupies the catalytic site
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What two benefits does tamoxifen have? Two potential disadvantages.
It is oestrogenic in bone and in the cardiovascular system. (Anecdotal reports) associating the administration of tamoxifen for advanced breast cancer with subsequent thromboembolic episodes.
Tamoxifen is known to produce endometrial thickening, hyperplasia, and fibroids following several years of therapy, and may increase eye cataracts, hot flushes
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What other potential use is there for tamoxifen?
Prevention of breast cancer, e.g. in high risk ladies with atypical ductal hyperplasia
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Why do 15% of breast cancers become resistant to tamoxifen?
Although 15% of patients who develop resistance to tamoxifen lose ERα expression
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Why might age be a risk factor for breast cancer?
ER alpha expression increases with age
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What indirect actions do the oestrogen receptors have?
ERs can regulate gene expression by indirect recruitment to gene promoters through interaction with other transcription factors, in particular AP1, Sp1, and NF-κB
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What does tamoxifen binding do?
• Binding of the LBD by tamoxifen and raloxifene causes a conformational change that positions helix 12 over the coactivator binding groove to prevent coactivator recruitment. Recruit co repressors
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What do co-activators for example oestrogen do?
they function as platforms for the recruitment of histone acetyltransferases
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What do co-repressors do?
NCoR and SMRT act as molecular scaffolds for the recruitment of histone deacetylases
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What does ligand binding do to the LBD?
Ligand binding results in a remarkable conformational change in the LBD, such that a surface for the recruitment of co-activators is revealed
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Why is tamoxifen an agonist in some tissues and an antagonist in others?
Anti-oestrogens fall into two broad categories. The first class, exemplified by tamoxifen and raloxifene, inhibits AF2 activation and thereby ER activity, but does not prevent activation of AF1. Because the ER activity in breast epithelium is due mainly to AF2, tamoxifen acts largely as an antagonist in breast cells, whereas in other tissues, such as the uterus, AF1 activ- ity is more significant, resulting in greater agonistic activity
