Epithelia and Glands

Jean-Pierre Louboutin
7/9/12

Jean-Pierre Louboutin
- Neurologist
- MD: dissertation on correlations between clinical,
neurophysiological and neuroradiological data in Multiple
Sclerosis
- PhD: dissertation on morphological and physiological
properties
during skeletal muscle regeneration
- Research in USA (Philadelphia) focused on gene/cell
therapy/immunology (University of Pennsylvania, UPENN, 6
years) and also on physiopathology and molecular therapy of
HIV-1-associated dementia (Thomas Jefferson University, 7
years)

Cell

Basic structural and functional unit of all living organism but

Tissue

Humans are not unicellular organisms and cells are organized in human
tissues, ensemble of similar cells of the same origin, that together carr
t a specific function- Study of tissues: histology

+ Epithelia
- Definitions
- Functions
- Different types
- Special cytological features
+ Glands
- Definitions and functions
- Different types
+ Different staining methods in histology
+ Examples of experimental therapies targeting epithelia
- Gene transfer
- Regenerative medicine
+ Objectives
+ Few words of advice

+ Epithelia
- Definitions
- Functions
- Different types
- Special cytological features
+ Glands
- Definitions and functions
- Different types
+ Different staining methods in histology
+ Examples of experimental therapies targeting epithelia
- Gene transfer
- Regenerative medicine
+ Objectives
+ Few words of advice

Definitions
Four basic types of animal tissues:
. Epithelia
. Connective tissue
. Muscle tissue
. Nervous tissue

. Epithelia: tissues consisting of closely apposed cells,

without intervening intercellular substances

Epithelia are avascular but innervated; all epithelia

grow" on an underlying layer of vascular connective tissue

The connective tissue and the epithelium are separated

by abasement membrane
Epithelium covers all free surfaces of the body (i.e., skin)

Epithelium also lines the large internal body cavities,

where it is termedmesothelium

The internal surfaces of blood and lymph vessels are

ined by epithelium, here calledendothelium

Functions of epithelia

. Protection from toxins, physical trauma, radiation (e.g., skin)

. Absorption: cells of the gastrointestinal tract can absorb

nutrients from the digestive tract lumen (lumen: inside space of
a tubular structure)

. Excretion: epithelial tissue lines the excretory lumina like the

urinary tracts from the kidneys through the urethra

. Secretion of sweat, mucus, enzymes delivered by ducts

(channels/canals) of the glandular epithelium

Types of epithelia

. Epithelia are classified on the basis of the number

of cell layers and the shape of the cellsin the surface layer

. If there is only one layer of cells in the epithelium,

t is designatedsimple

. If there are two or more layers of cells, it is calledstratifi

. Cells in the surface layer are described according

to their height assquamous(scale- or plate-like),cuboidal
orcolumnar

Simple squamous epithelium

Cells: single layer, flattened

Nuclei: flattened, centrally located
Functions: diffusion, lubrication
Examples: blood vessels, heart lined by simple squam. epithelium

Simple cuboidal epithelium

Cells: single square-like layer

Nuclei: centrally located and spherical
Functions: absorption, secretion, protection
Examples: tubules of kidney, follicles of thyroid, surfaces of
varies, excretory ducts of many glands

Simple columnar epithelium

Cells: tall, single-layered

Nuclei: basally located and elongated
Functions: absorption, protection, secretion (goblet cells with
microvilli, cilia)
Examples: gastrointestinal tract (cardia of stomach to rectum )

Stratified squamous epithelium (1)

. Cells: squamous cells apically (at the top), but basal layers vary
from cuboidal to columnar
. Nuclei: centrally located
. Function: protection
. Examples: skin, oesophagus
Stratified squamous keratinized
epithelium: skin

Stratified squamous epithelium (2)

. Cells: squamous cells apically (at the top), but basal layers vary
from cuboidal to columnar
. Nuclei: centrally located
. Function: protection
. Examples: skin, oesophagus
Stratified squamous nonkeratinized epithelium: oesophagus

Stratified cuboidal epithelium

.
.
.
.

Cells: two layers

Nuclei: centrally located and spherical
Functions: absorption, secretion
Examples: duct of the sweat glands

Stratified columnar epithelium

Cells: single layer of columnar cells on one or several

ayers of cuboidal cells
Nuclei: basal and oval
Functions: protection, secretion
Examples: duct of the mammary gland and large salivary gland

Transitional epithelium
.
.
.
.

Cells: apical cells often large, round and bi-nucleated

Nuclei: centrally located
Functions: distention
Examples: bladder, ureter and urethra

Pseudostratified columnar epithelium

ells: differ in height, not all cells reach apical surface

Nuclei: at various positions
unctions: absorption, secretion, transportation (goblet cells, cili
xamples: trachea, excretory ducts (canals) of many glands

Different epithelia can be seen in the same organ

Trachea
Pseudostratified columnar
epithelium

Bronchus
Pseudostratified columnar
epithelium

Terminal bronchiole
Simple columnar epithelium

Alveoli
Squamous epithelial
cells (type I pneumocytes)

Respiratory bronchiole
Simple cuboidal epithelium

Special cytological features of epithelia

Basement membrane (BM):
. Epithelia separated from underlying connective tissue by
extracellular layer: BM
. BM acts as a scaffolding on which epithelium can grow and
regenerate after injury
. BM composed of 2 sublayers:
1) Basal lamina (80 nm thick) consists of protein filaments
embedded in amorphous matrix. Major components of basal
lamina are two glycoproteins: laminin and collagen
Basal lamina: selectively permeable filter between epithelium
and connective tissue
2) Reticular lamina consists of reticular fibres embedded in
ground substance. Fibres of the reticular lamina connect
basal lamina with underlying connective tissue. Contains
fibronectin

Epithelium
Basement Membrane
Connective Tissue

Basal Lamina
Reticular Lamina
Connective Tissue

Miicrovilli

Special features of the apical surface:

. Microvilli and stereocilia are fingershaped extensions of epithelial cells
. Function: to increase the surface area of
epithelial cells
. Found in epithelia active in absorption
. Microvilli contain actin filaments
. Microvilli are much shorter than
stereocilia
. Stereocilia are not moving structures
. Goblet cells: mucus-secreting cells found in pseudostratified and columnar epithelia (e.g., gastrointestinal tract and trachea)
. Cilia can help move molecules within the lumen.
Found in pseudostratified and columnar
epithelia (e.g., trachea)

Isolated cell

Tissue

Nature of the junction between 2 cells ?

Special features of the basal and lateral surfaces: cell

junctions

. Multicellular organisms
require adhesion for cells to
adhere to each other or to the
extracellular matrix

. Cell adhesion occurs through

specific cellular specializations
and
molecules and has both static
and dynamic functions
. Cell junctions are especially
abundant in epithelia
. 3 major types of cell
junctions:
+ Gap junctions:
communicating junctions
+ Adherens junctions,
desmosome, hemidesmosome:
anchoring junctions
+ Tight junctions: occluding

Communicating junctions
. Gap junctions connect cytoplasm of
2 cells and are made up of proteins
called connexins

Anchoring junctions

Gap junctions

. Desmosomes: Group of
glycoproteins (cadherins) inserted
into the opposing plasma membrane
(like a zipper linking both cells)Mediate cell-to-cell adhesion
. Hemidesmosomes resemble
desmosomes but are made up of
integrins (transmembrane protein)
instead of cadherin

Desmosomes

. Zonula adherens (adherens

junctions): cytoplasmic face is
linked to the actin cytoskeleton.
They can appear as bands
encircling the cell or as spots of
attachment to the extracellular
matrix (alpha and beta catenins
links to actin filaments and
transmembrane anchors are
cadherins)

Occluding junctions
. Tight junctions (zonula
occludens): isolate the interior of
the body from the outside world.
Proteins inserted into the cell
membranes of adjacent cells
'stitch' the membranes of the
cells together. Provide an
effective barrier to the diffusion
of substances from the outside of

Tight junction

+ Epithelia
- Definitions
- Functions
- Different types
- Special cytological features
+ Glands
- Definitions and functions
- Different types
+ Different staining methods in histology
+ Examples of experimental therapies targeting epithelia
- Gene transfer
- Regenerative medicine
+ Objectives
+ Few words of advice

Glands: definitions and functions

. Glands are aggregations of cells whose function is
secretion
. Exocrine glandsrelease the secretory product via
ducts (channels) that open upon one of the
surfaces of the body in contact with the external
world (gastrointestinal tract, etc.)

. Endocrine glandsrelease their secretory product

(typically hormones) into the spaces between the
secretory cells (extracellular space) from which it
enters the bloodstream
. Both endocrine and exocrine glands are
developmentally derived from epithelia. Exocrine
glands maintain the connection with the surface
epithelium, whereas the connection is lost by
endocrine glands

Different types of exocrine glands

. Exocrine glands may be classified according to cell
number, and/or the shape and branching pattern of their
secretory portions and ducts
. Unicellular or multicellular describes a single-celled
gland or a gland made of many cells, respectively . A
multicellular gland consists of a group of secretory cells
and a duct through which the secretions pass as they exit
the gland
. Branched glands refers to the branching arrangement of
secretory cells
. Simple or compound refers to whether the duct of the
gland (not the secretory cells) does or does not branch,
respectively
. Tubular describes a gland whose secretory cells form a
tube, while alveolar (or acinar) describes secretory cells

Unicellular glands
. Consist of a single secretory cell
. In mammals, the only example of unicellular exocrine
glands are goblet cells, which are found in the
epithelium of many mucous membranes
. Goblet cells secrete the glycoprotein mucin, which by
the uptake of water, is converted into a slimy substance,
mucus
Rectum
Jejunum

Multicellular glands
. The simplest form of a multicellular gland is
asecretory epithelial sheath

Acidic content

. It is a surface epithelium consisting entirely of

secretory cells(e.g., the epithelium lining the inner
surface of the stomach, where the mucous secretion
protects the stomach wall from the acidic contents of
the stomach)

Other multicellular glands have their secretory portion

mbedded in the connective tissue underlying the epithelium

The secretion is either discharged directly from the secretory

ortion onto the epithelium or reaches the epithelium via a duct
ystem that consists of non-secretory cells

The secretory portion may have a variety of shapes. Secretory

ells may form:
+ tubesintubular glands,
+ aciniinacinar glandsor
+ alveoliinalveolarglands

. Branched glands refers to the branching arrangement of cells

. Simple or compound refers to whether the duct of the gland

(not the secretory portion) does or does not branch, respectivel

Exocrine glands are also classified according to their function:

In merocrine glands, secretions pass through the cell membranes of th
secretory cells (exocytosis). For example, goblet cells of the trachea
elease mucus via exocytosis

In apocrine glands, a portion of the cell containing secretions is release

as it separates from the rest of the cell. For example, the apical portion
of lactiferous glands release milk in this manner

In holocrine glands, entire secretory cells disintegrate and are released

along with their contents. For example, sebaceous glands release sebum
o lubricate the skin

Serous and mucous acini

. Two morphologically
distinct forms:
serousormucous
. Serous secretions have a low
viscosity, i.e. they are
rather "watery". The apical
cytoplasm of the cells
formingserous aciniis usually
well-stained

. Mucous secretions have a

high viscosity, i.e. they are
rather "slimy".
The contents of the secretory
vesicles in the apical cytoplasm
of cells ofmucous aciniare
only weakly stained, giving
a foamy appearance

Endocrine glands
. Endocrine glandssecretehormonesdirectly into
the bloodstream.
. For example, the thyroid gland secretes the
hormone thyroxin into the bloodstream, where it is
distributed throughout the body, stimulating an
increase in the metabolic rate of body cells.

+ Epithelia
- Definitions
- Functions
- Different types
- Special cytological features
+ Glands
- Definitions and functions
- Different types
+ Different staining methods in histology
+ Examples of experimental therapies targeting epithelia
- Gene transfer
- Regenerative medicine
+ Objectives
+ Few words of advice

Different staining methods

. Hematoxylin and eosin (H&E)
staining
. H&E stains nuclei in dark
blue and cytoplasm and other
structures in pink/red
. Example: trachea
. Other methods can be used
to see molecular and structural
components of the trachea

Immunohistochemistry: technique using antibodies to target

pecific proteins or peptides antigens in the cell

Bound antibodies can be detected using several methods

e.g., fluorophores)

High Magn.

Low Magn.

Example: detection of a molecule involved in innate immune

ystem, Toll-like receptor 2 (TLR2) in the apex of the trachea
TLR2

TLR2

TLR2

Control

TLR2

Control

Double Immunohistochemistry: technique allowing to detect 2

ifferent antigens in a cell/tissue
Ex: detection of another molecule, Toll-like receptor 5 (TLR5) and
mucin-1 (present in goblet cells) in the trachea
TLR5

TLR5

Control

TLR5

Mucin-1

TLR5 + Mucin-1 +
DAPI (nuclei)

Histochemistry: identification and localization of molecular

components, metabolic and enzymatic activities in tissues
Example: gene transfer of the lac Z gene into the trachea using
viral vector
Lac Z gene encodes for beta-galactosidase enzyme
Staining for beta-galactosidase enzymatic activity: proof of
ene transfer
Virus
Lac Z gene

Control

Beta-Galactosidase

Beta-Galactosidase

lectron Microscopy (EM): provides high-powered resolution imag

Transmission EM (TEM)

Scanning EM (TEM)

+ Epithelia
- Definitions
- Functions
- Different types
- Special cytological features
+ Glands
- Definitions and functions
- Different types
+ Different staining methods in histology
+ Examples of experimental therapies targeting epithelia
- Gene transfer
- Regenerative medicine
+ Objectives
+ Few words of advice

Examples of experimental therapies targeting epitheli

Gene transfer/Gene therapy

Gene therapy based on the use of DNA as a pharmaceutical agent to

eat a disease

Most common form of gene therapy involves using DNA that encodes
therapeutic gene in order to replace amutated gene (i.e., sickle cell
nemia, cystic fibrosis)

Other forms involve using DNA that encodes a therapeutic protein drug
ather than a natural human gene) to provide treatment

In gene therapy, DNA that encodes a therapeutic protein is packaged

ithin a "vector", usually a virus, which is used to get the DNA inside cell
the body. Once inside the cell, the DNA becomes expressed by
e cell machinery, resulting in the production of therapeutic protein,
hich in turn treats the patient's disease

. Gene delivery to tissues/organs

. Virus can target tissues, leading to gene delivery and
transgene expression

Virus
AU1
transgene

Alveoli

DAPI: staining of
nuclei

Staining of
transgene AU1 in
alveolar cells

Merged DAPI + AU1

Regenerative medicine

Regenerative medicineis the process of replacing or regenerating

uman cells, tissues or organs to restore or establish normal function

This field holds the promise of regenerating damaged tissues and orga
the body by replacing damaged tissue and/or by stimulating the body's
wn repair mechanisms to heal previously irreparable tissues or organs

Potential to solve the problem of the shortage of organs available for

onation

Refers to a group of biomedical approaches to clinical therapies that

ay involve the use ofstem cells

Examples include the injection ofstem cells or progenitor cells

ell therapies); the induction ofregeneration by biologically active
olecules administered alone or as a secretion by infused cells
mmunomodulation therapy); andtransplantation ofin vitrogrown
gans and tissues (tissue engineering)

Targeting of bone marrow (BM) stem cells in vivo by direct injection

into the femoral cavity
- Using a viral vector carrying a transgene
- Following the distribution of BM stem cells in the body by studying
transgene expression

Virus
Transgene

BM stem cells

Study of the distribution of

the transgene in the body

Migration of BM stem cells ?

Transgene expression in the rat lung at the alveolar level

- DAPI staining of nuclei; FLAG: transgene;
- TTF-1: marker of precursor cells in the lungs

Louboutin et al., J Gene Med, 2007

Transgene expression in the rabbit lung at the alveolar level

- DAPI staining of nuclei; AU1: transgene;
- TTF-1: marker of precursor cells in the lungs

Conclusions
- Bone marrow stem cells can be
transduced in situ in the BM
- They migrate to the lungs
- They differentiate into
precursor cells in the lungs

Study of capillaries of the rat brain after BM injection of a viral vector

carrying the gene for the transgene FLAG

- DAPI: marker of nuclei; FLAG: transgene; CD31: marker of endothelia

cells and capillaries

Study of capillaries of the rabbit brain after BM injection of

a viral vector carrying the gene for the transgene AU1
- DAPI: marker of nuclei; AU1: transgene; CD31: marker of
endothelial cells and capillaries

Louboutin et al., Neuroscience, 201

Conclusions
- BM stem cells can migrate to the brain
vessels
- They can differentiate into endothelial cells

+ Epithelia
- Definitions
- Functions
- Different types
- Special cytological features
+ Glands
- Definitions and functions
- Different types
+ Different staining methods in histology
+ Examples of experimental therapies targeting epithelia
- Gene transfer
- Regenerative medicine
+ Objectives
+ Few words of advice

Objectives

- To know the major features of epithelial tissue ++

- To know the different types of epithelial tissues
(including the corresponding organs) +++
- To know the special cytological features of epithelia ++
- To know the different types of glandular tissues
(including the corresponding organs) +++

+ Epithelia
- Definitions
- Functions
- Different types
- Special cytological features
+ Glands
- Definitions and functions
- Different types
+ Different staining methods in histology
+ Examples of experimental therapies targeting epithelia
- Gene transfer
- Regenerative medicine
+ Objectives
+ Few words of advice

Few words of advice

- Attend the lectures:
+ It helps for memorizing the topics and for putting things in order
+ Take notes
+ Better for getting hints about the exams
- Stick to the lectures rather than to the books:
+ Do not get overwhelmed by the books
+ Use the books as references to check the lectures or the points not
understood during the lectures
+ Stick to the essential at first; enrich your knowledge with details
progressively
- Read and reread the lectures while taking notes or making drawings:
+ Make sure to work on the lectures regularly and to memorize the
lectures at the end of each week
+ 4 hours each day in average til mid November (for all fields)
+ 6-8 hours a day in average from mid November til the exams
- Spend time in the Laboratory bringing lectures and/or books
- Do not be afraid to ask questions or ask for help +++