Transcript Biochemistry of skin - Univerzita Karlova. Prague

Biochemistry of skin
Jana Novotná
Department of Med. Biochemistry
2nd Faculty of Medicine
Charles University
Skin
• it provides barrier against a range of noxious stress (UV irradiation,
mechanical, chemical and biological insults);
• acts as the periphery „sensing“ system;
• system which maintaining body homeostasis.
• 2 m2 in area
• ~ 2.5 mm thick on average
• constitutes 6% of our total body weight (5 – 6 kg)
• barrier to prevent a desiccation and temperature balance
• protection to the UV radiation – absorbing pigmentation system
• complex immuno-regulatory network protection
• normal skin pH is somewhat acidic - the range of 4.2. to 5.6.
Human skin layers
• Mammalian skin is composed of two primary layers:
– the epidermis, which provides waterproofing and serves as a
barrier to infection;
– the dermis is responsible for the tensile strength of skin. Its main
functions are to regulate temperature and to supply the
epidermis with nutrient-saturated blood. Much of the body's
water supply is stored within the dermis.
Epidermis
• An external stratified, non-vasularized
epithelium (75 – 150 mm thick),
continually keratinizing
– Stratum corneum – 15 – 30 sheet
of non-viable, but biochemicaly
active corneocytes
– Stratum granulosum – 3 – 5 sheet
of non-dividing keratinocytes,
producing keratino-hyalin
– Stratum spinosum – 8 – 10 sheet
of keratinocytes with limited
dividing capacity, Langerhan´s
cells
– Stratum basale – maturing/aging
keratinocytes, melanocytes, Merkel
cells (receptor cells)
Keratins
• keratinocytes contain filaments of the
keratin intermediate filament (KIF)
family (cyto-skeleton)
• hair, nails, horny layers of the skin – are
formed from keratin cytoskeleton of
dead cells.
• two primary groups of keratins, the akeratins and the b-keratins
 a-keratins occur in mammals, b-keratins
in birds, reptiles
• both form are right handed helical
structure
• 2 types
– type I – acidic keratins
– type II – basic keratins
– heterodimer – type I forming a coild coil with
type II
Composition and Structure of
Keratin
• Human skin contains ~ 20 genetically
different keratins
• Long stretches a-helix is interrupted
by short non-helical segments
• The most abundant amino acid are
glycine and alanine, cysteine can
account for up to 24%
• Contact between 2 a-helices are
formed by hydrophobic amino acid
side chain on 1 edge of each helix
– two polypeptides form a dimeric colid
coil
– protofilaments are formed from two
staggered rows of head-to-tail
associated coils
– protofilaments dimerize to form a
protofibril and four of which form a
microfibril
http://biochem118.stanford.edu/Papers/Protein%20Papers/Voet%26Voet%20chapter6.pdf
Composition and Structure of
Keratin
• Intra- and intermolecular hydrogen bonds, disufid
bridges occure at all keratins.
• In cells, keratin type I forms pair with keratine type II
• Different keratin types are expressed in different cell
types and different layers of epidermis:
– cytoskeleton of epithelial cells - K14 (type I) & K5
(type II), K18 (type I) & K8 (type II)
– Basal layer – K13 (type I) & K4 (type II)
– Spinus and granular layer – K10 (type I) & K1 (type II)
– Stratum corneum – K3 (type I) & K12 (type II)
– Hairs and nails – various other keratin pairs
The Epidermal Permeability
Barrier
• Barrier function in human epidermis depends on
transglutaminase-mediated cross-linking of structural
proteins and lipids („biological glues“)
– post-translation modification of proteins – formation of covalent
bond between a e-amino group of lysine and the free amine
group of glutamine.
• Bonds formed by transglutaminase exhibit high
resistance to proteolytic degradation.
• Proteins are than highly resistent to mechanical
perturbation and proteolysis.
• The quality of the S. corneum barrier depends on the
presence of equimolar concentration of ceramides,
cholesterol and fatty acids.
• Changes in the concentration of any of these can affect
barrier quality.
Fatty Acids in Epidermis
• Arachidonic acid and 20-carbon PUFA can be
metabolized by either cyclooxygenase or lipoxygenase
pathways → prostaglandins, hydroxyeicosatetraenoic
acids.
– phospholipids are starting point for the arachidonic
acid pathway during inflammation (allergic reaction)
• Some of these metabolites can interact with signaling
system in proliferating and differentiating epidermal cells
→ modulation of protein kinase C, nuclear MAP-kinase
Epidermal Cell Differentiation
and Turnover
• Basal keratinocytes → transformation ~ 30 days to
corneocytes.
• Damage cells are removed by normal squamation.
• Genetic damage - (UV-R) → trigger apoptosis (within
hours) – „sunburn“ cells.
• Skin protection against UV-R – concentrating transferred
melanin over vulnerable keratocyte nucleus.
• Other insults can induce keratnocyte apoptosis –
chemical, mechanical, immunological.
• The principal marker for keratinocyte/epidermal
differentiation is expression of particular keratin pairs.
Epidermal Cell Differentiation
and Turnover
• Majority of over 30 keratins currently known.
• Proliferative basal keratinocytes express K5 and K14;
• keratinocytes in the early stages of
maturation/differentiation switch to K1 and K10.
• The „pluri-potent“ stem cells for keratinocytes sebaceous
gland and epidermis rised from hair folicules.
• Ca2+ plays pivotal role in epidermal differentiation - 4-fold
increase of extracellular Ca2+ in S. corneum.
• Keratinocyte differentiation is regulated by hormones
and vitamins - D3 and retinol from diet, thyroid hormones
and steroid hormones.
• The skin has nucleas receptors for glucocorticoids,
estrogen, androgen and progesterone.
Epidermal Cell Differentiation
and Turnover
• Importan factors for keratocyte differentiation are Ca2+-dependent
integrins – the receptors for the extracellular matrix fibronectin
binding.
• Laminin and collagen IV and VII (basemen membrane components)
– regulation of keratinocytes migration on basement membrane
(very important during wound healing).
• Migrating keratinocytes produce many matrix metalloproteinases.
• Mature keratinocytes (in S. graulosum) contain protein-rich,
keratohyalin granules and lipid-rich, lamellar granules.
• Lipids from lamelar granules form the sheets of the lipid permeability
of the lipid permeability barrier of the epidermis.
Melanocytes
•
•
•
•
Melanocytes are melanin-producing cells located in S. basale
Precursor - melanoblast
Melanin is stored in the melanosomes.
„Epidermal melanin unit“ - the anatomical relationship between
keratinocytes and melanocytes.
• 1 melanocyte is in contact with ~ 40 keratinocytes
• Melanocytes extend arms to transfer melanosomes into the
keratinocytes
Melanocytes
• Cover picture:The Rab27a
GTPase associates with
melanosomes and regulates
their transport to, and retention
in, the peripheral cytoplasm in
skin melanocytes.
Melanosome transport also
requires the microtubule and
actin cytoskeleton. Staining of
a cultured murine melanocyte
for filamentous actin (red) and
microtubules (blue) reveals a
close relationship between
Rab27a-labelled melanosomes
(green) and these cytoskeletal
elements.
Hume et al. JCB 2001;152 (4): 795
Formation of melanosomes
• Melanosomes - elliptic membranebound organelles (melanin
synthesis).
• Synthesis of matrix proteins and
tyrosinase (TYR) on the rough
endoplasmic reticulum.
• TYR undergoes post translational
modification in the form of
glycosylation in the Golgi apparatus.
• Fusion of premelanosomes with
coated vesicles containing tyrosinase
- formation of the melanosome.
• Melanosome migrates into one of the
dendrites of the melanocyte →
transfer to a neighboring
keratinocyte.
Production of Melanin
• Three enzymes in melanosomes whih absolutely required for
different melanin type synthesis
– tyrosinase (TYR) – responsible for critical step of melanogenesis (tyrosine
hydroxylation)
– tyrosinase-related protein 1 (TYR1) and DOPAchrome tautomerase
(DHI = 5,6-dihydroxyindole; DHICA = 5,6-dihydroxyindole-2carboxylic acid)
Melanins
• Melanins are polymorphous and multifunctional polymers of
eumelanin, pheomelanin, mixed melanins (a combination of the
two); and neuromelanin
• Mammalian cells produce black-brown eumelanin and yellow-redish
pheomelanin
• Eumelanin - highly heterogenous polymer consisting of DHI and
DHICA units in reduced or oxidized states.
• Pheomelanin - mainly sulfur-containing benzothiazine derivatives.
• Neuromelanin is produces in dopaminergic neurons of substantia
nigra.
• Melanin absorbs UV light at a wavelength of 280 - 320 nm
• Both eumelanin and pheomelanin play important protective role in
binding to cations, anions, drugs, chemicals, etc.
Factors Involve in Melanin
Production
• The melanin granules accumulate above the nuclei of keratinocytes
and absorb harmful UV-R before it can reach the nucleus and
damage the DNA.
• Quick responds of the melanocyte-keratinocyte complex to a wide
range of environmental stimuli (paracrine and/or autocrine) - to UVR, melanocyte-stimulating hormone (MSH), endothelins, growth
factors, cytokines, etc.
• UV-R exposure → melanocytes increase their expression of
proopiomelanocortin (POMC, the precursor of MSH) and its receptor
melanocortin 1 receptor (MC1-R), TYR and TYRP1, protein kinase
C (PKC), and other signaling factors.
Factors Involve in Melanin
Production
• Fibroblasts (possibly other cells in skin) - produce
cytokines, growth factors, and inflammatory mediators
that can increase melanin production and/or stimulate
melanin transfer to keratinocytes by melanocytes.
• Other factors derived from keratinocytes which can
regulate proliferation and/or differentiation of
melanocytes:
– α-MSH, ACTH, basic fibroblast growth factor (b-FGF),
nerve growth factor (NGF), endothelins, granulocytemacrophage colony-stimulating factor (GM-CSF),
leukemia inhibitory factor (LIF), and hepatocyte
growth factor (HGF).
Other Epidermal Cells
• Langerhans cells - dendritic cells - arise from bone marrow early in
embryonic development, occupie 2 - 8% of epidermis
• important element of the immune system, interacting with T-cells
• resided in suprabasal layer - attracted to keranocytes by Ecadherin receptor
• their motion is regulated by specific integrin receptor and by α –
TNF
• in the stratum germinativum interacts with the allergen and migrates
to the lymphoid gland - then “teache” the T cells about the allergen
• interact specifically with T-lymphocytes and keratinocytes to initiate
host response to antigens (allergens)
• UV B stimulates synthesis and release of TNF-a by keratinocytes
which in turn modifies the behavior and morphology of Langerhans
cells, decreases their total number.
Langehans
cell
Allergen
cytokine
T cell
Activated
T cell
Other Epidermal Cells
• Merkel cells – location in S. germinativum
• have synaptic contacts with sensory nerve endings
• associated with the sense of „light touch“ discrimination
of shapes and textures.
Dermis
• responsible for the tensile strength of skin
• main functions – regulation of temperature and to supply the
epidermis with nutrient
• much of the body's water supply is stored within the dermis
• components:
–
–
–
–
–
–
–
connective tissue
hair follicles
sweat glands
sebaceous or oil glands
apocrine glands
lymph vessels
blood vessels
• The main cell type - fibroblast
Dermal Proteins and Extracellular
Matrix
• Collagen – about 90% of total dermal proteins
– predominatly type I (85 – 90%),
– type III (8 -11%),
– minor type V (2 – 4%), (papillary dermis, matrix around vessels,
nerves),
– type VI – associated with fibrils and interfibrillar spaces
(responsible for fine structure in early prenatal development of
skin).
• Elastin, proteoglycans, glycoproteins, water and
hyaluronic acid
Collagen structure - refer to lecture on Collagens
Elastin, proteoglycans, glycoproteins – refer to lecture on
Extracellular matrix
Skin Appendages
• Skin plays in the body homeostasis, therefore is
well-equiped with secretory (release of chemicals
from cells for physiological function) and excretory
(elimination of weste products of metabolism) capacity.
– sweat glands [can be sweat secreted with strong
odour (apocrine) or with a faint odour (eccrine)].
– sebaceous glands (secrete sebum onto hair follicle to
oil the hair).
– hair follicle
Sweat Glands
• 3 – 4 million eccrine sweat glands are
in our skin – each producing water
perspiration (serves mainly to cool us)
and maintain core temperature at
37.5oC.
• At maximum output the eccrine sweat
glands can excrete as much as 3 l/hour,
and heat loss is more than 18 kcal min1.
• Humans utilize eccrine sweat glands as
primary form of cooling.
• Apocrine sweat glands are larger, have
different mechanism of secretion, and
are limited to axila and perianal area.
Sweat Secretion
• Eccrine gland activity is regulated via neural
stimulation using sympathetic nerve fibers
distributed around the gland.
• Neurotransmitter is acelylcholine
• Sweating is controlled from hypothalamus (a
center in the preoptic and anterior regions),
where thermosensitive neurons are located.
• The stimulus for perspiration:
– direct heating alone (39 to 46oC)
– physiological sweating due to nerve reflex arise from
sweat centers in brain cortex (emotional),
hypothalamus (thermoregulation)
Eccrine Sweat
• contains mainly water (99.0 – 99.5%). It also
contains electrolytes NaCl, K+ and HCO3-, and
other simpl molecules - lactate, urea, ammonia,
amino acids (serin ornithin, citrulin, aspartic
acid) and minerals.
• Mineral composition varies with the individual:
–
–
–
–
their acclimatisation to heat,
exercise and sweating,
the particular stress source (exercise, sauna, etc.),
the duration of sweating, and the composition of
minerals in the body
Apocrine Sweat
• In lower mammals – secretion of pheromones (trigger
sexual and territorial response)
• In humans – the significance of apocrine secretion of
pheromones is not completely understood.
• Apocrine gland begin secreting at puberty
• Apocrine duct exit to the surface via he hair follicle.
• Apocrine sweat – more viscous, with milky consistency
due to high content of fatty acids, cholesterol, squalene,
triglycerides, androgens, ammonia, sugars.
Mineral Composition of Sweat
sodium
0.9 g/l
potassium
0.2 g/l
calcium
15 mg/l
magnesium
1.3 mg/l
zinc
Microelements
0.4 mg/l
copper
0.3 – 0.8 mg/l
iron
1 mg/l
chromium
0.1 mg/l
nickel
0.05 mg/l
lead
0.05 mg/l
Sebaceous Glands
• Glands secrete an oily/waxy
matter, called sebum, to
lubricate the skin and hair
• Composition – 25% wax
monoesters, 41% triglycerides,
16% free fatty acids, 1%
squalene, small amount
cholesterol esters and
cholesterol.
Skin Metabolism
• Primary source for energy production in epidermis is glucose
from circulation – diffuses into keratinocytes without effect of
insulin. Large proportion of glucose is catabolized up lactate
(even in presence of oxygen)
• citric acid cycle does operate in epidermis – explanation why this
cycle is inefficient is due to wide fluctuation of temperature and
blood flow in skin.
• 20% of glucose is metabolized by pentose-phosphate pathway
(PPP) – production of NADPH and pentose for both FA
synthesis and nucleic acids.
• Secondary source of energy - fatty acids derived from both
epidermal stores and exogenous sources (when glucose flow is
limited, then FA are metabolized).
Skin Metabolism
• Glycogen – small amount under physiological
conditions, however, elevation in all manner of injury of
epidermis or during hair growth in follicle – explanation –
energy when skin needs to be repaired or to use glucose
immediately, most probably – disequilibrilium in
metabolic processes.
• Furthermore, glucose is substrate also for synthesis of
lipids, polysaccharides, glycoproteins and nucleic acids.
• GAG and proteoglycans – highly charged and attract
water – forming gels (see also lecture about ECM).
Skin Metabolism
Lipid metabolism - components:
a) membranes,
b) major constituents of permeability barrier,
c) energy supply
• Synthesis from both glucose catabolism, from AA and
circulating FA - lipogenesis is going on in all layers of
epidermis - sebum synthesis → in sebaceous glands
(higher synthesis of sebum is after sexual maturation).
• Degradation - generally with lipases (yields in FA for
neutral lipids – TG, sterol esters) – in outermost layers of
epidermis (e.g. formation of prostaglandins)
Skin Immune System
• Skin not only provides immune protection for itself, but
also for the whole body.
• Cell types containing battery of mediators of immune
response
– Langerhanse cells, monocytes, macrophages, mast cells
(cooperation with T-cells)
• Cell types producing free radicals, anti-bacterial
peptides, cytokines chemokines, pro- and antiinflammatory mediators
– Neutrophils, eosinophils, basophils.
• B-cells secrete immunoglobulins (antibodies)