Steroid Hormones and Vit D

Question 1

Basics of steroids

Answer 1

-cholesterol is the precursor of all classes of steroid hormones: glucocorticoids (cortisol), mineralocorticoids (aldoesterone), sex hormones (androgens, estrogens, progestins)
-synthesis and secretion occurs in several organs:
adrenal cortex- cortisol, aldosterone, and androgens
ovaries and placenta- estrogens and progestins
testes- testosterone
-steroid hormones travel in the blood from their point of synthesis to their target using nonspecific (albumin) vs specific (corticosteroid-binding globulin,aka transcortin, transports cortisol) carrier proteins
-once reaching the target the steroid hormone enters cells through the plasma membrane and once inside binds a receptor in the cytoplasm or nucleus. The receptor can bind both the steroid hormone and specific DNA hormone response elements, once homo-dimerized, resulting in altered transcription

Question 2

Synthesis of steriod hormones

Answer 2

• involves shortening the hydrocarbon chain of cholesterol and hydroxylating the steroid nucleus
• rate limiting step is conversion of cholesterol to the 21 carbon pregnenolone
• this reaction is catalyzed by the cholesterol side-chain cleavage enzyme- which is a cytochrome P450 mixed function oxidase located on the inner mitochondrial membrane- requires NADPH and O2
• cholesterol within the cell moves to the mitochondrial outer membrane and then moves to the inner mitochondrial membrane. THe latter step is mediated by StAR

Question 3

Congenital Adrenal Hyperplasias

Answer 3

• pregnenolone is then the precursor of all steroid hormones
• it is oxidized and isomerized to progesterone which is further modified by hydroxylaton reactions occuring in the mitochondria and ER. The enzymes are primarily cytochrome P450 (CYP) proteins
• a defect at any step in these pathways can cause several diminished products at later steps and at the same time a build up of substrates at prior steps
• serious metabolic imbalance occurs with these enzyme deficiencies which cause disorders known as congenital adrenal hyperplasias

Question 4

3 Beta- hydroxysteroid dehydrogenase deficiency

Answer 4

• problem with 3-beta hydroxysteroid dehydrogenase
• virtually no glucocorticoids, mineralocorticoids, active androgens, or estroges
• salt excretion in urine
• female like genitalia
• autosomal recessive with incidence of 1:10,000
• build up of pregnenolone

Question 5

17-alpha-hydroxylase deficiency

Answer 5

• virtually no sex hormones or cortisol
• increased production of mineralocorticoids causing sodium and fluid retention and therefore hypertension
• female like genitalial
• build up of progesterone

Question 6

21-alpha hydroxylase deficency

Answer 6

• most common form of Congenital Adrenal Hyperplasias
• partial and virtually complete deficiencies known
• mineralocorticoids and glucocorticoids virtually absent (salt wasting classic form) or deficient (nonclassic form)
• overproduction of androgens leading to masculinzation of external genitalia in females and early virilization in males-

Question 7

11- Beta1 hydroxylase deficiency

Answer 7

• decrease in serum cortisol, aldoesterone, and corticosterone
• increased production of deoxycorticosterone causes fluid retention (because this hormone suppresses the renin-angiotensin system, it causes low renin hypertension
• overproduction of androgens causing masculinzation and virilization

Question 8

Secretion of Adrenal Cortical Steroid Hormones

Answer 8

• secreted from tissue of origin in response to hormonal signals- corticosteriods and androgens are produced in different regions of the adrenal cortex and are secreted into the blood
• cortisol is produced in the middle layer (zona fasciculata) of the adrenal cortex. It’s production and secretion is controlled by the hypothalamus to which the pituitary gland is attached
• in response to stress (infection) corticotropin releasing hormone (CRH), produced by hypothalamus travels through the capillaries to the anterior lobe of the pituitary. There it induces the production and secretion of adrenocorticotropic hormone (ACTH) - polypeptide stress hormone
• causes the adrenal cortex to synthesize and secrete the glucocorticoid cortisol. Cortisol helps the body to respond to stress through effects on metabolism (ex stimulating gluconeogensis) and the inflammatory and immune responses
• as corticol concentrations rise the release of CRH and ACTH is inhibited

Question 9

Role of ACTH

Answer 9

• binds to G protein coupled receptor which leads to increased cAMP which in turn activates protein kinase A
• PKA phosphorylates and activates the lipase which converts cholesterol ester to cholesterol and StAR protein which allows the cholesterol to move to the inner mitochondrial membrane where it is converted to pregnenolone
• pregnenolone is returned to the cytosol where it is converted to progesterone
• in two ER membrane located hydroxylation steps catalyzed by CYP17 and 21 progesterone is converted to 11- Deoxycortisol
• 11-deoxycortisol is returned to the inner mitochondrial membrane where CYP11B1 catalyzes the Beta- hydroxylation at C21 yielding cortisol which can exit the cell

Question 10

Physiological Functions of Aldoesterone

Answer 10

• produced in the outer layer of the adrenal cortex (zona glomerulosa) Its production is stimulated by a decrease in plasma Na+/K+ ratio and by angiotensin II
• angiotensin II is a peptide hormone (octapeptide) produced by cleavage of the decapeptide angiotensin I by angiotensin converting enzyme (ACE)
• angiotensin I is produced in the blood by specific cleavage of angiotensinogen, an inactive precursor secreted by the liver, by renin a proteolytic enzyme synthesized and secreted by the kidneys
• angiotensin II binds to a specific G coupled cell surface receptor and acts through the phosphatidylinositol 4,5 bisphosphate pathway
• aldosterones primary effect is upon kidney tubules where it enhances Na+ and water uptake and K+ efflux
• aldosterone increases BP and ACE inhibitors are used to treat renin dependent hypertension

Question 11

Androgens

Answer 11

• produced by inner (zona reticularis) and middle layers of the adrenal cortex
• the adrenal androgens (primarily androsterone and androstenedione) are converted to testerone and estrogen in peripheral tissues

Question 12

Testes and ovaries and hormones

Answer 12

• they synthesize hormones that are required for sexual differentiation and reproduction
• a hypothalmic releasing factor, Gonadotropin-releasing hormone (GRH) stimulates the anterior pituitary to release luteinizing hormone and follicle stimulating formone both glycoproteins
• LH and FSH bind to G-protein linked surface receptors and their functions are mediated by increase in cAMP and stimulation of protein kinase A

Question 13

Luteinizing hormone (LH)

Answer 13

-stimulates the testes to produce testosterone and the ovaries to produce estrogens and progesterone

Question 14

Follicle stimulating hormone (FSH)

Answer 14

-regulates the growth of ovarian follicles and stimulates spermatogenesis within the testes

Question 15

Estrogens

Answer 15

• produced from androstenedione and then testosterone
• the enzyme involved in this latter conversion is Aromatase
• Aromatase inhibitors are currently used as a treatment for hormone positive (estrogen responsive) breast cancer in post menopausal women

Question 16

Steroid Hormone at Cellular Level

Answer 16

• sufficiently hydrophobic to diffuse through the plasma membrane of their target cells
• inside cell they bind to cytoplasmic or nuclear receptor
• the ligand receptor complex enters the nucleus
• once in the nucleus the ligand-receptor complex dimerizes and in association with coactivator proteins, binds to a specific regulatory sequence called the hormone response element
• HRE is promoter or an enhancer element for genes to a specific steroid hormone- in presence of coactivator protein mRNA transcription is increased for these specific genes
• binding of ligand receptor causes conformational change in the latter thus exposing a DNA binding domain- complex can now associated with zinc finger motif
• receptors that bind steroid hormones, thyroid hormone, retinoic acid and Vit D and all function in a similar fashion

Question 17

Further Metabolsim and Excretion of Steriod Hormones

Answer 17

• typically converted into inactive excretionproducts in the liver
• reactions involved include reduction of unsaturated bonds and introduction of additional hydroxyl groups
• conjugation with glucuronic acid or sulfate (from 3–phosphoadenosyl-5–phosphosulfate) makes the excretion products more water soluble
• 20-30% of these metabolites are secreted into the bile and excreted in feces. The remainder are released in the blood and filtered in the kidney passing into the urine for excretion
• as these excretion products are water soluble they do not require protein carriers

Question 18

Vitamin D Basics

Answer 18

• D Vitamins are a group of sterols that function like hormones
• the active molecule: 1,25-dihydroxylcholecalciferol (aka 1,25-diOH-D3 or calcitriol) binds to receptor proteins within the cell. The ligand receptor complex interacts with DNA in a manner similar to the steroid hormnes and enhanced or represses transcription of a coordinated set of genes
• the most important functions of 1,25-diOH-D3 is to regulate plasma levels of calcium and phosphorous

Question 19

Endogenous Vitamin

Answer 19

• 7-Dehydrocholestero, an intermediate in cholesterol biosynthesis, is converted to cholecalciferol in the dermis and epidermis when we are exposed to sunlight
• cholecalciferol is then transported to the liver while bound to a vitamin D-binding protein

Question 20

Exogenous Source/Diet

Answer 20

• Ergocalciferol (vitamin D2), which is found in plants, and cholecalciferol (vitamin D3) found in animal tissues are sources of preformed vitamin D.
• Dietary vitamin D is packaged into chylomicrons
• supplementation with preformed vitamin D is a dietary requirement in individuals with limited exposure to sunlight

Question 21

Converting Inactive Vit D to Active Vit D

Answer 21

• vitamin D2 and D3 are not biologically active on their own. They are converted in vivo to the active form (1,25-diOH-D3) by two sequential hydroxylation reactions
• the first occurs in the liver and the enzyme is 24 hydroxylase yielding 25-hydroxycholecalciferol (25-OH-D3 or calcidiol). Calcidiol is the major form of vitamin D in the plasma and the major storage form
• 25-OH-D3 is further hydroxylated at the 1-position by 25-hydroxylcholecalciferol 1-hydroxylase, in the kidney to form 1,25-diOH-D3 (calcitriol)
• both hydrolylases are cytochrome P450 proteins

Question 22

Regulation of the 25-hydroxylcholecalciferol 1-hydroxylase

Answer 22

• 25-hydroxycholecalciferol 1-hydroxylase activity is increased directly by low plasma phosphate and indirectly by low plasma calcium
• low calcium triggers secretion of PTH. PTH upregulates the 25-hydroxycholecalciferol 1-hydroxylase
• therefore hypocalcemia from insufficient dietary calcium results in elevated plasma 1,25-diOH-D3
• elevated 1,25-diOH-D3 inhibits activity of 25-hydroxycholecalciferol 1 hydroxylase and the expression of PTH forming a negative feedback loop

Question 23

Regulation of Plasma Calcium Levels by 1,25-diOH-D3

Answer 23

• low plasma calcium ->
• increase parathyroid hormone (also effects increased calcium mobilization from bone) ->
• increase 1,25-diOH-D3 ->
• increase calcium mobilization from bone ->
• increase renal reabsorption of calcium ->
• decrease renal excretion of calcium ->
• increase calcium absorption from intestine ->
• increase plasma calcium

Question 24

Vit D Receptor Interaction

Answer 24

• 1,25-DiOH-D3 enters cell and binds to ligand domain within vitamin D receptor (VDR) within cytoplasm of intestinal cell
• ligand-VDR complex enters nucleus, forms heterodimer with retinoid-X-receptor (RXR) and binds coactivator proteins
• recognize Vit D response element- VDRE
• can enhance or diminish cell type specific transcripts influencing the expression of specific proteins

Question 25

Calbindin-D9K

Answer 25

• within human intestinal epithelial cells (enterocytes)
• member of the S100 family of calcium binding proteins
• mediates the transport of calcium across the enterocytes from the apical side
• another protein which is increased in expression is the calcium transport protein TRPV5 which allows entry of calcium into the epithelial cell
• the transport of calcium across the enterocyte cytoplasm appears to be rate-limiting for calcium absorption in the intestine
• the presence of calbindin increases the amount of calcium crossing the cell without raising the free concentration

Question 26

Vit D and Calcium Reabsorption from Bones

Answer 26

• low plasma calcium causes elevated of 1,25-DiOH-D3 and PTH both act to increase calcium absorption and bone resorption (demineralization) and inhibit calcium excretion
• high plasma calcium blocks production of PTH, low PTH results in conversion of 25-OH-D3 to 24, 25-DiOH-D3 instead of 1,25-DiOH-D3. Lowered levels of PTH and 1,25-DiOH-D3 comes elevated expression of calcitonin resulting in inhibition of bone resorption and enhanced calcium exretion. High levels of plasma calcium and phosphate also increase bone mineralization. With dietary vitamin D and calcium are adequate no net loss of bone calcium occurs
• when vitamin D is deficient there is demineralization of bone due to increased PTH

Question 27

Vitamin D Requirements

Answer 27

• in fatty fish, liver and egg yolks, milk must be fortified
• 25-OH-D3 is the major form of Vit D found in serum and is measured to determine Vit levels in patients
• 50 nmol/L, >20 ng/mL- generally considered adequate for bone and overall health in healthy individals
• > 125 nmol/L, >50 ng/mL- emerging evidence links potential adverse effects to such high levels, particularly >150 nmol/L

Question 28

Vit D Deficency

Answer 28

• results in net diminerization of bone resulting in rickets in children and osteomalacia in adults
• Rickets is characterized by formation of the colalgen matrix but insufficient mineralization resulting in soft and pliable bones
• in the case of osteomalacia demineralization of existing bones makes them more susceptible to fracture

Question 29

Renal Osteodystrophy

Answer 29

• chronic kidney disease causes decreased synthesis of active Vit D and increased retention of phosphate
• resulting in hypocalcemia and hyperphosphatemia
• low blood calcium leads to increased PTH and the associated bone demineralization
• treatment includes supplementation with Calcitriol while reducing phosphate

Question 30

Hyperthyroidism

Answer 30

• lack of PTH causes hypocalcemia and hyperphosphatemia

- these patients can be treated with calcium and calcitriol

Question 31

Toxicity of Vitamin D

Answer 31

• a fat soluble vitamin and therefore can be stored in the body and is slowly metabolized
• in those taking excessive supplements there can be loss of appetite, nausea, thirst and stupor
• the enhanced calcium absorption and bone resorption results in hypercalcemia which in turn leads to calcium deposits in many organs but particularly in the arteries and kidneys
• the UL is 4000 IU/day for 9 years and older with lower levels for younger children