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Chapter 3
Connective Tissue Proper
霍冠华
滨州医学院组织胚胎学教研室
Department of Histology & Embryology
Binzhou Medical University
September, 2013
Connective Tissue




The different types of connective tissues are
responsible for providing and maintaining form in the
body. Functioning mechanically, they provide a matrix
that connects and binds the cells and organs and
ultimately gives support to the body.
Structurally, connective tissue is formed by three
classes of components: cells, fibers, and ground
substance.
The major constituent of connective tissue is the
extracellular matrix. Extracellular matrices consist of
different combinations of protein fibers (collagen,
reticular, and elastic) and ground substance.
Connective tissue includes connective tissue proper
and specialized connective tissue.
CONNECTIVE TISSUE
PROPER
一、CHARACTERISTICS：





Fewer cells
Large amount of intercellular matrix
Cell: separated and no polarity
Intercellular matrix: fibers + ground substance + tissue
fluid
Filled with blood vessels, Lymphatic vessels and
nerves
Function: support, connect, nourish, defence, and
repair etc.
 Architectural framework of the body
 Bind together and provide mechanical support for
other tissue (metabolic, defense, transport, storage)
 Wound repair / inflammatory response
AF - Active Fibroblast IF - Inactive Fibroblast
C - Collagen
EF - Elastic Fibers
二、CLASSIFICATION
 Connective tissue proper:


Loose C.T.
Dense C.T.
Adipose tissue
Reticular tissue
Cartilage and bone
Blood and Lymph
Broadly speaking
Connective Tissue
Epithelium
Epithelium
Connective Tissue
papilla
ridge
Connective Tisue
Muscle
Connective Tissue

Extracellular Matrix
Fibers – collagen, elastic,
reticular

Ground substance
Cells
Fibers in Connective Tissue



Collagen Fibers
 most abundant protein in human body (up to 30%
dry weight)
 multiple types: fibril-forming or fibril-associated
(in skin, tendon, cartilage, bone, dentin, blood
vessels); cross-linked networks (in all basement
membranes)
Reticular Fibers – specialized type of collagen (Type
III; reticulin) associated with smooth muscle in
organs subjected to changes in volume, forms the
stroma in lymphatic and hematopoietic organs
Elastic Fibers –thin fibers or fenestrated sheets
composed of various glycoproteins, including the
protein elastin, providing elastic properties to
tissues that experience repeated deformation (in
skin, blood vessels, lung, bladder)
Major Collagen Fiber Types
Collagen Type
(out of at least 20)
Tissues
Function
I
(most abundant)
Skin, tendon, bone, dentin
Resistance to tension
II
Cartilage, vitreous of eye
Resistance to pressure
III
(reticulin)
Skin, muscle, blood vessels, liver,
etc.
Structural framework and stability
All basement membranes
Support and filtration
Fibril-forming collagens (these are visible)
Network-forming collagens
IV
Fibril-associated collagens with interrupted triple helices (FACIT)
VI, IX
Assoc. w/ type I and II fibrils
Fibril-fibril / fibril-ECM binding
Epithelia
Epidermis to basal lamina
Anchoring filament collagens
VII
Collagen fibers viewed by light microscopy
H&E
Trichrome
Collagen Fibers vs. Fibrils
H&E
fibers
fibrils
Assembly of
collagen fiber
bundles
①
Fibroblast
Synthesize:
In rER ／ GL:
→ Polypeptide
αchains ①
→ hydroxylated
αchains ②
→ Procollagen ③
In Extracellular space:
→ Tropocollagen ④
→ fibrils
⑤
→ collagenous f. ⑥
②
③
④
280x1.4nm
⑤
⑥
Clinical disorders resulting from
defects in collagen synthesis
Type
Disease
Symptoms
I
Osteogenesis
imperfecta
Spontaneous fractures,
progressive hearing loss, cardiac
insufficiency
III
Ehlers-Danlos
(type IV)
Hypermobility of digits, early
morbidity/mortality from rupture of
aorta or intestine
multiple Scurvy (lack of vit. C, Ulceration of gums, hemorrhages
a cofactor for prolyl
and lysyl
hydroxylase)
Ehlers-Danlos Syndromes
• A series of genetic diseases
with faulty assembly of
collagens (lysyl hydroxylase
deficiency).
• Hyperextensible skin and
hypermobile joints
• In some forms (e.g., type IV),
weakness in blood vessels or
intestines are life threatening.
Noncollagen Components of the
Extracellular Matrix
Elastin
 “Ground substance”

Glycosaminoglycans (GAG’s)
Proteoglycans
Multiadhesive matrix proteins
 laminin
 fibronectin
2、Elastic fibers
Structure:
LM:
EM：
elastin +
microfibrils (no
bandings)
F.B.
Elastic Fibers
LM: Visualized by selectively staining with Weigert’s,
resorcin-fuchsin, or aldehyde-fuchsin
EM: Consist of amorphous core of elastin surrounded
by microfibrillar glycoprotein, fibrillin (8-10nm).
Elastin: is rich in glycine and proline, but it contains little
or no hydroxyproline and hydroxylysine . uniquely
contains desmosine and isodesmosine, which are
thought to cross-link the molecules into a network of
randomly coiled chains. This cross-linking is
responsible for its rubber-like properties.
Confers elasticity: present in large amounts in ligaments,
lung, skin, bladder, and walls of blood vessels.
Marfan Syndrome: defect in elastic fiber synthesis;
reduced elasticity in skin and lungs, skeletal defects
(bones are longer and thinner than usual),
cardiovascular complications (aneurism, valve
prolapse)
Elastin appears amorphous (not
fibrillar) in the electron microscope
E=elastin; C, collagen fibrils
M/L=microfibrils of fibrillin, a scaffolding glycoprotein involved
elastin deposition
Marfan Syndrome: defect in fibrillin gene, results in weakened
elastic fibers
Elastin molecules are joined by covalent bonds to
generate an extensive cross-linked network. Because
each elastin molecule in the network can expand and
contract like a random coil, the entire network can stretch
and recoil like a rubber band.
3、Reticular fibers
Sructure:
LM: argyrophilia,
PAS(+)
EM: fibrils w/ periodic
cross-bandings at
intervals of 64nm,
typeIII collagen
covered carbohydrate
Reticular fibers
in liver
Reticular fibers
in lymph node

Electron micrograph of cross sections of reticular (left) and collagen (right)
fibers. Note that each fiber type is composed of numerous smaller collagen
fibrils. Reticular fibrils (R) are significantly narrower in diameter than
collagen fibrils of collagen fibers (C; see histogram inset); in addition, the
constituent fibrils of the reticular fibers reveal an abundant surfaceassociated granularity not present on regular collagen fibrils (right).
x70,000.
Ground
Substance
Ground Substance of the
Extracellular Matrix (ECM)
1.
2.
3.
Glycosaminoglycans (GAG)
• linear (unbranched) polysaccharides, e.g. heparan sulfate,
condroitin sulfate, keratan sulfate, hyaluronic acid
• very hydrophilic due to abundant negative charges (e.g. SO4groups).
• except for hyaluronic acid, are usually bound covalently to
protein core as part of a proteoglycan
Proteoglycans
• core protein + GAG side chains (like a bottle brush)
• bind cells, other proteins, and/or ECM components
Multiadhesive glycoproteins
• small glycosylated proteins containing NUMEROUS binding
sites to cells, signaling molecules, and other ECM components
• e.g. fibronectin and laminin: important for adhesion of
epithelial cells to the basal lamina via transmembrane integrin
receptors.
Ground substance

Jelly-like & amorphous substance;
 Proteoglycan: in molecular sieve
hyaluronic acid
GAG(glycosaminoglycan) chondroitin sulfate
keratin sulfate etc.
protein: core protein & link protein
glycoprotein： fibronectin (FN)
laminin (LN)
chondronectin (ChN)；
Side chain
subnit
hyaluronic
acid
Collagenous
fiber
Ground substance
Intercellular material
hyaluronic
acid
molecular sieve:
hyaluronic acid -- link
pr.-- side chain subunit
subunit: core protein
+ chondroitin sulfate &
keratin sulfate
tissue fluid: flowing
through the sieve pores
side
chain
subunit
Core pr.
link pr.
chondroitin
sulfate
Link pr.
keratin sulfate
core pr.
glycoprotein：
fibronectin (FN):
produced by epithelial cells and fibroblasts
play a role in events of identification,
adhesion, migration and proliferation
 laminin (LN) :
located in B.M., produced by epi. cells,
function: adhesion the epi.cells and B.M.
 chondronectin (ChN):
in cartilage tissue,
fuction: a component of ground substances;
adhesion chondrocyteS and colagen typeII

Cells in Connective Tissue
1.
Fixed
2.
(permanent
residents)
3.
4.
5.
Free
(transient
residents)
•
•
•
Fibroblasts
Adipose (fat) cells
Macrophages**
Mast cells**
undifferentiated mesenchymal cells
Lymphocytes
Plasma Cells (differentiated B-cells)
**
Leukocytes**
(specifically, neutrophils,
eosinophils, & basophils)
** derived from hematopoietic stem cells and involved in
immune function and inflammation
Connective Cell Lineages
Functions of Connective Tissue Cells.
Cell Type
Representative Product or Activity
Representative
Function
Fibroblast,
chondroblast,
osteoblast
Production of fibers and ground
substance
Structural
Plasma cell
Production of antibodies
Immunological
(defense)
Lymphocyte
(several types)
Production of immunocompetent cells
Immunological
(defense)
Eosinophilic
leukocyte
Participation in allergic and vasoactive
reactions, modulation of mast cell
activities and the inflammatory process
Immunological
(defense)
Neutrophilic
leukocyte
Phagocytosis of foreign substances,
bacteria
Defense
Macrophage
Secretion of cytokines and other
molecules, phagocytosis of foreign
substances and bacteria, antigen
processing and presentation to other
cells
Defense
Mast cell and
basophilic
leukocyte
Liberation of pharmacologically active
molecules (eg, histamine)
Defense
(participate in
allergic reactions)
Adipose (fat) cell Storage of neutral fats
Energy reservoir,
heat production
1. Fibroblasts are the most common cells
in connective tissue

Synthesize & secrete collagenous pro. & elastic pro.
（to form collagenous f., elastic f. & reticular f. ）;
and ground substance (proteoglycan & glycoprotein).

Active and quiescent stages (when quiescent
sometimes called fibrocytes or mature fibroblasts).

Synthesize growth factors.

Rarely undergo cell division unless tissue is injured,
which activates the quiescent cells.

Play a major role in the process of wound healing
and respond to an injury by proliferating and
enhanced fiber formation.
A connective tissue
layer (dermis) shows
several fibroblasts
(F), which are
elongated cells. H&E
stain.

Structurally, fibroblasts are of 2 types, one of which resembles mesenchymal
cells. This type is stellate, with long cytoplasmic processes and a large, ovoid,
pale-staining nucleus. The cytoplasm contains abundant RER and Golgi
complex, and this type is important in the production of collagen and other
matrix components. Also, lysosomes and vacuoles of secretion product are
prominent features of the cells. Fibroblast secrets collagen, fibronectin,
glycoproteins, and proteoglycans. The cells have many actin-containing
microfilaments because they are highly motile cells. They also contain
numerous microtubules, which probably help maintain the cell morphology.

Electron micrograph
revealing portions of
several flattened
fibroblasts in dense
connective tissue.
Abundant
mitochondria, rough
endoplasmic reticulum,
and vesicles
distinguish these cells
from the less active
fibrocytes. Multiple
strata of collagen
fibrils (C) lie among
the fibroblasts.
x30,000.

Cells of the second type are less active
and are sometimes termed fibrocytes,
because they are believed to be more
mature. They are smaller and spindleshaped, with a dark, elongate nucleus
and fewer cytoplasmic organelles. They
can revert to the fibroblast state and
participate in tissue repair.
External morphologic
characteristics and
ultrastructure of
each cell are shown.
Fibroblasts that are
actively engaged in
synthesis are richer
in mitochondria,
lipid droplets, Golgi
complex, and rough
endoplasmic
reticulum than are
quiescent
fibroblasts
(fibrocytes).
Active
quiescent
Quiescent fibroblasts are elongated cells with thin cytoplasmic
extensions and condensed chromatin. Pararosaniline-toluidine blue
(PT) stain.
2. Macrophages
LM:
Large, stellate, cytoplasm is
acidophilic
Cytoplasmic granules
Vacuoles
Macrophages



Derived from monocytes
EM：
 a large number of Ly.,
 pinosome & phagosomes
 Well-developed Golgi complex
 bundles of microtubules and microfilaments
Mononuclear phagocyte system: monocytes, macrophages,
Kupffer cells (liver), osteoclasts (bone), dust cells (lung),
Langerhans cells (skin), microglial cells (central nervous
system)
Ultrastructural features of a
Macrophage
secondary lysosomes
phagocytic
vacuoles
Function:
Mobility
chemotaxis
chemotactic factor:
complement C5a,
bacterial products etc.
 Phagocytize
specific:depend on
identify factors: Ab, C,
Fibronectin etc （their
receptors are on surface
of the macrophge）.
non-specific:
independently

indigestible remnant
adsorb
phagosome
fuse





Functions:
Their role is to phagocytose, or engulf and then digest,
cellular debris and pathogens, either as stationary or as
mobile cells. They are specialized phagocytic cells that attack
foreign substances, infectious microbes and cancer cells
through destruction and ingestion.
They participate in the immune response by presenting
phagocytosed antigens to lymphocytes, and also stimulate
lymphocytes and other immune cells to respond to pathogens.
Secret bioactivators (bioactive substances ):
 lysozymes kill bacteria, complement participating in
process of inflammation
 Cytokines: IL-1, Interferon, hematopoietic cell colony
stimulating factor, tumour necrosis factor (TNF) etc.
Identified by specific expression of a number of proteins
including CD14, CD40, CD11b, CD64, F4/80 (mice)/EMR1
(human), lysozyme M, MAC-1/MAC-3 and CD68
antigens
ingest
phagocytic vacuole
Primary lysosomes
secondary lysosome
MHC-II moleculesantigen complex
MHC: major Histo-compatibility complex molecules
type II sets specially at surface of some immune cells, type I occurs in all
types of cells.
Macrophage functions as a antigen presenting cell
3. Plasma Cells, the mature B lymphocytes
that constitutively secrete antibodies
chromatin
Two more plasma cells (P) are shown in this field. One shows the
cartwheel nucleus well. The cytoplasm is basophilic. Also, note
three macrophages which are
detected by their content of ingested black deposits. Perhaps this
came from a lymph node near the lung (of a smoker). The cell
labeled is an Eosinophil.

The plasma cells are characterized by their size and abundant
basophilic cytoplasm (rough endoplasmic reticulum) and are
involved in the synthesis of antibodies. A large Golgi complex
(arrows) is where the terminal glycosylation of the antibodies
(glycoproteins) occurs. Plasma cells produce antibodies of
importance in immune reacions.
Plasma cells
 Structure:
Nu.:eccentrically，heterochromatin in
wheel shape
Cytoplasm: strong basophilic w/ lightlystained area near Nu.
Plasma Cells
EM：rich in rER &
ribosomes,
developed Golgi
complex
 Source:
B lmphocytes
 Function:
 synthesize
antibody &
cytokines
 participate in
humoral immunity
Golgi complex
RER
4. Mast cell




The thin strands: Elastic fibers
The thick strands: Collagen fibers
The small dark purple structures: Nuclei of fibroblasts
The large granular-looking cell: Mast cell
Basophilic
granules
Metachromasia – when stained with toluidine blue, the
granules bind the dye and change its color to red.
EM：
a great number of granules w/ crystals,
containing:
heparin,
histamine,
leukotriene,
slow-reacting
substance,
eosinophil
chemotactic factors
(ECF-A)
degranulation
Functions

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
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
Granules: degranulation
Heparin: is an anticoagulant, anti coagulation
Histamine: causes vasoconstriction and increased
permeability of small venules
eosinophil chemotactic factor: attract eosinophils
migration to the allergic reaction site
Cells secret Luekotriene: cytoplasm, likes histamine
Principal function is storage in secretory granules
and REGULATED release (degranulation) of
histamine and other vasoactive mediators of
inflammation.
Responsible for the immediate hypersensitivity
response characteristic of allergies, asthma[‘æzmə]
and anaphylactic shock.

Mast cells are filled with conspicuous large
granules containing Heparin and Histamine,
mast cells can "degranulate" - often in
response to injury - and release Heparin, which
slows blood clotting, and Histamine, which
increases capillary permeability. In this manner,
mast cells participate in the inflammatory
response; they promote the flow of blood out
of the bloodstream and into tissue spaces,
where blood cells and antibodies can fight
infection.
5. Adipocytes, predominate in adipose tissue
Very active cells with many functions:
• Triglyceride storage and glucose metabolism (insulin and glucagon
receptors)
• Secretion of many bioactive molecules: leptin (regulates satiety);
angiotensinogen (blood pressure); steroids (glucocorticoids & sex
hormones); growth factors (e.g. insulin-like growth factor, tumor necrosis
factor ); cytokines (e.g. interleukin-6)
6. Leucocytes
Leucocytes

Neutrophils: Small cells with multi-lobed, heterochromatic
nuclei (or “polymorphonuclear neutrophils”).
L
N
7. Undifferentiated mesenchyme cells
Tissue fluid
dynamic balance
Function:
Tissue fluid nourishes the cells & tissues；
 Molecular sieve acts as a barrier: to prevent the
spread of bacteria & other microorganisms
*haemolytic streptococci produce hyaluronidase
& promote the invasion
 Glycoproteins (FN/LN/chondronectin):
identification/ adherence/ migration/ proliferation
etc.


Edema: swelling from excessive accumulation of serous fluid
in tissue. Tends to appear first where tissue turgor is normally
lowest. E.g. chronic nephritis, nephrotic syndrome, hepatitis,
heart failure, cancer. Eyelid edema: common in
nephritis, chronic liver disease, malnutrition, anemia,
blood vessels, such as edema. Edema of lower limb.



Dehydration: to lose too much water from body; to make a
person's body lose too much water. E.g. got
diarrhoea(diarrhea ) and is suffering from dehydration;
excessive sweating; burn
The eyes are often retracted from dehydration.
The major factors causing death are electrolyte imbalance and
dehydration, leading to cardiac arrest.
Types of Connective Tissue Proper
Loose (areolar) connective tissue – delicate, vascularized,
cellular; supports the epithelia of the major organs and glands and
fills the space between muscle tissue. - not very resistant to stress
Dense connective tissue (many more fibers than cells)
Dense irregular: meshwork of coarse fibers; dermis of skin,
organ capsules, fascia - resists multi-directional forces
Dense regular:
collagenous: fibers aligned in defined pattern; tendons,
ligaments, etc. - resists linear mechanical stresses
elastic: elastin and microfibrils (fibrillin) - elasticity
Adipose - fat storage, glucose regulation, satiety
Reticular - argyrophilic fibers of type III collagen - forms
stroma of highly cellular organs (e.g. liver, lymph nodes,
spleen)
LOOSE CONNECTIVE TISSUE
I. Characteristics：






A small amount of cells & many
kinds of cells
small number of fibers & great
amount of ground substance
The cells have no polarity
Sponge-like structure (areolar
tissue)
Distributed widely between cells,
tissues and organs.
Function: support, connect,
defence, and repair etc.
Loose connective tissue:
delicate, vascularized, flexible; facilitates transport of cells
and materials (secretion, absorption, immunity)
small intestine lamina propria
mammary gland intralobular connective tissue
Dense Irregular CT
Densely packed collagen fibers, often in perpendicular bundles; resists
tension in many directions and provides mechanical support.
Collagen
Fibroblast nucleus
Skin dermis, H &E
Dense Irregular CT from human dermis contains thick bundles of collagen
fibers, fibroblast nuclei (arrow) , and a few small blood vessels (bv). H&E
stain.
Dermis of Skin has both Loose and
Dense Irregular CT
Loose CT
sweat
gland
H&E
Dense Regular CT
(collagenous)
consists of a small
population of
fibroblasts
and lots of
collagenous fibers
Dense Regular CT (elastic)
Wall of large artery: ①Elastic membrane
Adipose Tissue
Structure:
Loose C.T.+ fat
cells (in large
aggregations)
HE: / Osmic acid:
Function:
energy storage,
shock absorber,
insulating layer
Unilocular adipose tissue.
Brown (Multilocular) Adipose Tissue
Present in newborns, e.g. hips, back, (and hibernating冬眠 mammals)
and involved in thermoregulation
Mitochondria of brown fat cells
express uncoupling protein which
“short circuits” the electron
transport chain producing HEAT
rather than ATP.
RETICULAR TISSUE
Structure:
 Reticular cells + reticular
f. + ground substance
 Reticular cell: stellate,
pale nucleus, obvious
nucleoli, processes (rich
in RER)
Function:
architectural framework
of lymphatic & hematopoietic tissues
cells
Lymph node
fibers
macrophage
reticular fiber
reticular cell
lymphocyte
reticular cell: cytoplasmic processes, cross-linked
networks; in rich in cytoplasm, more RER.
Function: produces reticular fibers and ground
substances.