KERATINOCYTES AND KERATINIZATION

(1)

K

ERATINOCYTES AND

K

ERATINIZATION

The human skin is a large organ comprising a sheetlike investment of the whole body that adapts

admirably to its contours and neatly conforms to the movements of the organism within.

George Odland

Physio., Biochem., and Mol. Biol of the Skin

E

PIDERMIS

O

VERVIEW

EPIDERMAL CHARACTERISTICS

TISSUE RENEWAL

CELLULAR PROLIFERATION

C

ORNIFICATION

S

TRUCTURE/

F

UNCTION

C

ONSEQUENCES OF

F

AILURE

(2)

K

ERATINOCYTES AND

K

ERATINIZATION

E

PIDERMIS:

- the outermost layer of the skin - stratified squamous epithelia - avascular

- interactions between all the cell types of the skin are critically important for maintenance of the skin - varies in thickness depending on the anatomic site - 75-150 µm thick

(3)

C

ELLS

T

YPES IN THE

E

PIDERMIS

At least five cell types are found in the epidermis: Melanocytes Langerhans cells Merkel cells Lymphocytes Keratinocytes

(4)

E

PIDERMAL

A

PPENDAGES

- Hair

- Sebaceous glands - Eccrine glands

- The basement membrane that

separates the epidermis from the dermis is continuous throughout the appendages

(5)

F

UNCTIONS OF THE

E

PIDERMIS

- Form a protective barrier from physical insults Chemical

Biological Temperature Mechanical

- Protect body homeostasis Temperature regulation Prevent fluid loss

- Immune surveillance - Sensory organ

(6)

T

ISSUE

R

ENEWAL – Continuous self-renewal of keratinocytes

S

TRENGTH – Both intracellular and intercellular strength

C

ORNIFICATION – Process that creates a water impermeable barrier

C

HARACTERISTICS OF THE

E

PIDERMIS

(7)

C

ONSEQUENCES OF

E

PIDERMAL

F

AILURE:

DEATH

Toxic Epidermal Necrolysis – life-threatening consequences are dehydration and infection

Mutations in genes that severely compromise epidermal function are embryonic/neonatal lethal

(8)

T

ISSUE

R

ENEWAL

Fine balance between new cell replication and cell death

Characteristics of Self-renewing Epidermis: Keratinocyte Stem Cells

Keratinocyte Proliferation

Keratinocyte Terminal Differentiation

(9)

K

ERATINOCYTE

S

TEM

C

ELLS

Fuchs and Raghavan (2002), Nature Rev. 3: 199

- stem cells are slowly replicating cells

- a stem cell divides into another stem cell or

a cell committed to differentiation - stem cells are protected in a specific tissue

niche, maintained by cell-ECM and cell-cell interactions

- stem cells are located within the bulge region of the hair follicle and at the base of rete ridges of interfollicular epidermis

(10)

K

ERATINOCYTE

S

TEM

C

ELLS

Fuchs and Raghavan (2002), Nature Rev. 3: 199

- stem cells are slowly replicating cells

- a stem cell divides into another stem cell or

a cell committed to differentiation - stem cells are protected in a specific tissue

niche, maintained by cell-ECM and cell-cell interactions

- stem cells are located within the bulge region of the hair follicle and at the base of rete ridges of interfollicular epidermis

(11)

K

ERATINOCYTE

S

TEM

C

ELLS

- stem cells or the bulge region are multipotent,

they can generate hair, sebaceous gland, or epidermal keratinocytes

- stem cells allow for the maintenance of the cell population in the epidermis with little to no damage to the “tissue” genome

- at times of tissue damage, stem cells increase their rate of proliferation to repopulate the wounded area

(12)

K

ERATINOCYTE

S

TEM

C

ELLS

Control of stem cell function by Wnt family and/or Delta/Notch pathways

(13)

K

ERATINOCYTE

S

TEM

C

ELLS

arm ptch smo

shh

hh Dfz2 wg fu CI

Wnt1

β

-catenin

sonic hedgehog

patched

zw3

glycogen

kinase 3b

Wnt1

TGF

β

BMP

Gli1

hDfz2

fizzled

smoothened

Stem cell

Activated cell

(14)

S

TEM

C

ELL

C

ONSEQUENCES OF

F

AILURE

(15)

S

TEM

C

ELL

C

ONSEQUENCES OF

F

AILURE

Loss of stem cells may lead to phenotype of aged epidermis? -flattening of the epidermal/dermal junction

-keratinocyte cell size becomes variable -nuclear atypia

-loss of melanocytes

-loss of Langerhans cells -slowed injury response -slowed chemical clearance -decreased immune response

-decreased resistance to mechanical stress -increased incidence of cancer

(16)

K

ERATINOCYTE

P

ROLIFERATION

- most of proliferation done by transient

amplifying cells (TA cells)

- in normal epidermis, all TA cells remain attached to basement membrane

- transition from stem cell to TA cell is the first step in keratinocyte

differentiation

- TA cells migrate laterally along the basement membrane

(17)

K

ERATINOCYTE

P

ROLIFERATION

- TA cells only divide 2-3 times before

they withdraw from the cell cycle - 2 mm punch biopsy contains a

replication unit

- TA cells carry-out cell proliferation for tissue without risk for long-term genetic damage to tissue

- increased expression of c-myc

associated with transition from stem cell to TA cell

- NFκB, retinoids, vitamin D, AP1 regulate proliferation and

(18)

K

ERATINOCYTE

P

ROLIFERATION

C

ONSEQUENCES OF

F

AILURE

Dysregulation of proliferation can lead to hypo- and hyperproliferative diseases

(19)

K

ERATINOCYTE

P

ROLIFERATION

C

ONSEQUENCES OF

F

AILURE

Dysregulation of proliferation can lead to hypo- and hyperproliferative diseases

(20)

K

ERATINOCYTE

T

ERMINAL

D

IFFERENTIATION

- when a keratinocyte releases from the basement membrane, it undergoes changes in morphology and gene expression

- gradual change in cell strength and water impermeability

- terminally differentiated keratinocytes synthesize a cornified cell envelope and undergo programmed cell death

(21)

K

ERATINOCYTE

T

ERMINAL

D

IFFERENTIATION

Keratinocyte morphology

and function

Differentiation-specific proteins expressed

stratum corneum – keratinocytes contain thickened cell envelopes, contain no nucleus, imbedded in lipid matrix

stratum granulosum – cells become elongated, usually 1-2 cell layers thick, accumulate amorphous keratohyaline granules

stratum spinosum – cells increase in size, increased cytoplasm:nucleus ratio, cell layer 4-6 cells thick, no further cell division

stratum basale – cuboidal cells, cells within this layer proliferate, all cells attached to the basement membrane, one cell layer thick

no new proteins expressed

keratins K1 and K10 loricrin, filaggrin transglutaminase 3 keratins K1 and K10 involucrin, envoplakin periplakin, 14-3-3σ keratins K5 and K14 integrins, p63

(22)

K

ERATINOCYTE

T

ERMINAL

D

IFFERENTIATION

and function

stratum corneum– keratinocytes contain thickened cell envelopes, contain no nucleus, imbedded in lipid matrix

stratum granulosum– cells become elongated, usually 1-2 cell layers thick, accumulate amorphous keratohyaline granules

stratum spinosum– cells increase in size, increased cytoplasm:nucleus ratio, cell layer 4-6 cells thick, no further cell division

(23)

K

ERATINOCYTE

T

ERMINAL

D

IFFERENTIATION

and function

(24)

K

ERATINOCYTE

T

ERMINAL

D

IFFERENTIATION

and function

(25)

K

ERATINOCYTE

T

ERMINAL

D

IFFERENTIATION

C

ONSEQUENCES OF

F

AILURE

Stained with an α-filaggrin antibody NORMAL SKIN ICHTHYOSIS VULGARIS LAMELLAR ICHTHYOSIS

(26)

K

ERATINOCYTE

I

NTRACELLULAR

S

TRENGTH

- Keratins are members of the intermediate

filament (IF) gene family

- there are over 50 members of the IF gene family that are expressed in a tissue- and differentiation-specific manner

- IF proteins have a conserved central rod domain of helical coiled-coil segments

- the amino- and carboxy-terminal sequences of IF proteins are variable

(27)

K

ERATINOCYTE

I

NTRACELLULAR

S

TRENGTH

- keratins heterodimerize with specific pairing partners: one Type I family

one Type II family

- the heterodimers then oligomerize into longer fibrils

(28)

K

ERATINOCYTE

I

NTRACELLULAR

S

TRENGTH

Fuchs and Cleveland, Science 279: 514

- keratins heterodimerize with specific pairing partners: one Type I family

one Type II family

- the heterodimers then oligomerize into longer fibrils

- fibrils continue to assemble until IF is 10-12 nm in diameter

(29)

K

ERATINOCYTE

I

NTRACELLULAR

S

TRENGTH

Tissue Type I keratin expressed Type II keratin expressed Basal cells Suprabasal cells Hyperproliferative skin Simple epithelia (also Merkel cells) K14 (50 kD) K10 (56.5 kD) K11 (56 kD) K16 (48 kD) K18 (46 kD) K5 (58 kD) K1 (67 kD) K2 (65 kD) K6 (56 kD) K8 (52 kD)

(30)

K

ERATINOCYTE

I

NTRACELLULAR

S

TRENGTH

- keratin filaments extend from the nuclear membrane to desmosomal plaques at the cell membrane

- keratins enable keratinocytes to sustain mechanical and non-mechanical stress

(31)

K

ERATINOCYTE

I

NTRACELLULAR

S

TRENGTH

C

ONSEQUENCES OF

F

AILURE

-Major defects in keratin molecules are incompatible with life

-Minor defects in keratin molecules lead to major debilitating skin disease

Genodermatoses Keratin mutation identified

Epidermolysis Bullosa Simplex K5, K14

Epidermolytic hyperkeratosis K1, K10

Palmoplantar keratoderma, epidermolytic K1, K9

Palmoplantar keratoderma, diffuse non-epidermolytic K1 Palmoplantar keratoderma, focal non-epidermolytic K16

(32)

K

ERATINOCYTE

I

NTRACELLULAR

S

TRENGTH

C

ONSEQUENCES OF

F

AILURE

EPIDERMOLYSIS BULLOSUS SIMPLEX

-mutation of K5 or K14

-fragility in basal layer keratinocytes

(33)

K

ERATINOCYTE

I

NTRACELLULAR

S

TRENGTH

C

ONSEQUENCES OF

F

AILURE

E

PIDERMOLYSIS

B

ULLOSUS

S

IMPLEX

(34)

K

ERATINOCYTE

I

NTRACELLULAR

S

TRENGTH

C

ONSEQUENCES OF

F

AILURE

E

PIDERMOLYTIC

H

YPERKERATOSIS

-mutation in either K1 or K10

(35)

K

ERATINOCYTE

I

NTERCELLULAR

S

TRENGTH

Three types of interactions hold keratinocytes together in epidermal sheets: Hemidesmosomes

Desmosomes

Adherens Junctions

(36)

K

ERATINOCYTE

I

NTERCELLULAR

S

TRENGTH

HEMIDESMOSOMES

- adhesion site that links the keratin cytoskeletal components of a cell to extracellular matrix (basement membrane) - Transmembrane components: integrins α6β4 BPAG2 (BP180) -Plaque components: BPAG1 (BP230) plectin -Cytoskeletal component: keratin

(37)

K

ERATINOCYTE

I

NTERCELLULAR

S

TRENGTH

D

ESMOSOMES

- adhesion site that links the keratin cytoskeletal

components of two cells - Transmembrane components: desmogleins desmocollins -Plaque components: desmoplakins plakoglobin plakophilin keratoclamin -Cytoskeletal component: keratin

(38)

K

ERATINOCYTE

I

NTERCELLULAR

S

TRENGTH

A

DHERENS

J

UNCTIONS

- adhesion site that links the actin cytoskeletal components of two cells - Transmembrane components: E-cadherin -Plaque components: β-catenin α-catenin vinculin VASP p120ctn -Cytoskeletal component: actin

(39)

K

ERATINOCYTE

I

NTERCELLULAR

S

TRENGTH

C

ONSEQUENCES OF

F

AILURE

Autoimmune Bullous Diseases

Genodermatoses Bullous Diseases

Bullous pemphigoid (α-BPAG1) Generalized atrophic benign EB (BPAG2) EB simplex-MD (plectin)

Junctional EB-pyloric atresia (α6-integrin or β4-integrin) Pemphigus foliaceus (α-desmoglein 1) Pemphigus vulgaris (α-desmoglein 3) Bullous pemphigoid (α-BPAG2)

(40)

K

ERATINOCYTE

I

NTERCELLULAR

S

TRENGTH

C

ONSEQUENCES OF

F

AILURE

(41)

K

ERATINOCYTE

I

NTERCELLULAR

S

TRENGTH

C

ONSEQUENCES OF

F

AILURE

PEMPHIGUS

F

OLIACEUS

PEMPHIGUS

V

ULGARIS

(42)

K

ERATINOCYTE

C

ORNIFICATION

-process that begins in cells of the upper spinous layers

-the induction of proteins that comprise the cornified cell envelope (CCE) are expressed as intracellular [Ca2+] rise in differentiating keratinocytes

-chromosome 1q21 contains cluster of genes called the Epidermal

(43)

K

ERATINOCYTE

C

ORNIFICATION

ENVOPLAKIN PERIPLAKIN INVOLUCRIN TG 1 TG 3 K5/K14 K1/K10

- newly synthesized envoplakin, periplakin, and involucrin form heterotrimers that associate with the cell membrane - transglutaminase 1 crosslinks these proteins via ε-(γ-lysine)

isopeptide linkages

- this scaffold forms along the entire inner surface of the cell membrane

(44)

ENVOPLAKIN PERIPLAKIN INVOLUCRIN TG 1 TG 3 K5/K14 K1/K10 PHOSPHOLIPID ω-OH-CERAMIDE FATTY ACIDS CHOLESTEROL

K

ERATINOCYTE

C

ORNIFICATION

- lamellar bodies (containing free-fatty acids, cholesterols, and ceramides) bud off from the Golgi complex

- at the transition from the granular layer to the cornified

layer, the lamellar bodies fuse with the cell membrane and extrude their contents into extracellular space

- also at transition, filaggrin becomes post-translationally modified and directs the aggregation of keratin proteins

(45)

- loricrin that was produced in the granular cell is crosslinked to SPR proteins via transglutaminase 3

- the loricrin-SPR complexes become fixed to the previously assembled scaffold via transglutaminase 1

- organelles, microtubules, microfilaments, and the nucleus are degraded via programmed cell death

- the mature corneocyte contains bundled keratins contained within CCE imbedded in lipid lamellae

ENVOPLAKIN PERIPLAKIN INVOLUCRIN TG 1 TG 3 K5/K14 K1/K10 PHOSPHOLIPID ω-OH-CERAMIDE FATTY ACIDS CHOLESTEROL LORICRIN SMALL PROLINE-RICH PROTEIN OTHER PROTEINS