造血系统基础知识(英文版)

Lecture 1 Hematopoiesis

1.

2.

3.

4.

5.

6.

7.

8.

Introduction

Ontogeny of hematopoiesis

Description of the Hematopoietic Stem Cell (HSC)

The concept of the stem cell niche

Anatomical description of the stem cell niche

Functional description of the stem cell niche

Role of lineage-specific Growth Factors in hematopoiesis

The formation of mature blood elements: 1. Myelopoiesis

1.

E

rythrocytes

2.

G

ranulocytes

3.

P

latelets

9.

The formation of mature blood elements: 2. Lymphopoiesis

1.

T

cells

2.

B

cells

3.

N

K cells

10. Bone Marrow Failure

1.

I

nherited disorders

2.

A

cquired disorders

1. Introduction

The formed elements of the blood, such as red cells, white cells and platelets, play a vital role in the normal

functioning of any human being. They are the end product of a highly specialized tissue called the

bone

marrow

, which resides in the cavities of all bones of the body. The process through which formed elements

of

the

blood

are

produced

is

called

Hematopoiesis.

Hematopoiesis

can

be

envisioned

as

a

hierarchical

progression

of

multipotential

hematopoietic

stem

cells

that

gradually

lose

one

or

more

developmental

options.

They

then

become

stem

or

progenitor

cells

committed

to

a

single

single

lineage

progenitor

cells

then

mature

into

the

corresponding

types

of

mature

formed

elements

of

the

blood,

also

called peripheral-blood cells.

As we will see in the following chapters, the bone marrow can be divided into two major cellular compartments:

1. One composed with hematopoietic stem cells (HSCs) which have two major physiological properties: A.

Self renewal

which is essential for the maintenance of life-long hematopoiesis, and: B

Differentiation

into

committed

progenitors.

2.

The

other

composed

of

multipotent

progenitor

cells,

which

cannot

renew

,

but

rather divide and differentiate into all mature formed elements of the blood.

At first glance, the fact that the bone marrow tissue resides in the bone cavities of all bone does not imply that

the bone itself has any role in the process of hematopoiesis. As a matter of fact, the mechanisms of bone

and blood formation have traditionally been viewed as distinct unrelated processes. Compelling evidence

now suggests that they are intertwined. It has been observed for a long time that HSCs are not randomly

distributed in the bone marrow tissue. In fact, they reside in close proximity to endosteal surfaces of the

bone. It was therefore hypothesized that

the osteoblasts

, the main bone forming cells, and therefore not

only the HSCs, play a central role in the process of hematopoiesis. The close intimacy of the HSC and the

bone endosteal surfaces is at the origin of the concept of the

Stem Cell Niche

which we will discuss in detail

later on. Therefore it seems that

normal hematopoiesis relies on the complicity of osteoblasts and HSCs.

The purpose of this lecture is to review in detail the basic physiological aspects of hematopoiesis, and discuss

briefly some bone marrow failure mechanisms.

2. Ontogeny of Hematopoiesis

Hematopoiesis begins in blood islands located in extra embryonic tissues (fetal yolk sac) in the first trimester

and

in

the

aorto-gonad-mesnenophros

(AGM)

region.

At

approximately

6

weeks

of

gestation,

hematopoiesis occurs predominantly in the fetal liver. Beginning at midterm, the medullary cavity gradually

replaces the fetal liver as the main site of hematopoiesis. In some species, such as the mouse, the spleen is

a major site of hematopoiesis in the adult.

The

yolk

sac

is

membranous

sac

attached

to

an

embryo,

providing

early

nourishment

in

the

form

of

yolk

in

primitive

mammals

and

functioning

as

the

circulatory

system

of

the

human

embryo

before

internal

circulation

begins.

The

primitive

yolksac

participates

in

nutrient

exchange

between

the

fetal

and

maternalcirculations before the formation of the placenta.

Figure 1: Primitive yolk sac

The yolk sac is an extra embryonic structure responsible for the initial and transient production of red cells in the

embryo, mainly during the first two weeks of gestation. Before placental circulation is established, the yolk

sac constitutes the primary source of exchange between the mother and the embryo (between 7-11 weeks

= 7mm in diameter) and constitutes the very first site of hematopoiesis. The first blood cells observed in the

embryo are large nucleated erythroblasts generated in blood islands of the extra embryonic yolk sac. These

unique

red

cells

have

been

termed

primitive

because

of

their

resemblance

to

nucleated

erythroblasts

of

non-mammalian species. It is now widely assumed that hematopoiesis in the yolk sac is

primitive

and that

definitive

hematopoiesis has its origins in the

aorta/gonad/mesonephros

(AGM) region. The first maturing

blood

cells

and

committed progenitors

are provided

by

the

yolk sac,

allowing

survival

until

AGM-derived

hematopoietic

stem

cells

can

emerge,

seed

the

liver

and

differentiate

into

mature

blood

cells.

Stem-cell

activity in the human yolk sac has not been reported. The AGM is a region of embryonic mesoderm that

develops

during

embryonic

development

and

is

the

site

of

origin

of

the

definitive

HSC.

It

has

an

intra

embryonic

location

(as

opposed

to

extra

embryonic

for

the

yolk

sac)

and

is

the

site

of

residence

and

amplification of the definitive hematopoietic stem cells that eventually seed the fetal liver and adult bone

marrow (see figure 2).

Figure 2: The embryonic Aorto-Gonad- Mesonephros (AGM) region

Initiation of hematopoietic stem cells (HSC) in the aorta-gonad-mesonephros (AGM) region in 10- day embryos

is

observed

with

additional

expansion

and

migration

to

the

fetal

liver

(FL).

In

the

adult

mouse,

both

the

spleen (SP) and bone marrow (BM) have hematopoietic activity.

At

approximately

6

weeks

of

gestation,

hematopoiesis

occurs

predominantly

in

the

fetal

liver.

Yolk

sac

hematopoietic cells are largely

a transient

embryonic population and the definitive stem cell, in fact, derives

from AGM region. Beginning at midterm, the

medullary cavity

gradually replaces the fetal liver as the main

site of hematopoiesis.

Figure 3: Summary of the ontogeny of hematopoiesis

Figure 3 shows the chronological description and respective sites of hematopoiesis during the development of a

human being.

3. Description of the Hematopoietic Stem Cell (HSC).

Hematopoietic stem cells (HSCs) are a subset of bone marrow cells that are capable of

self-renewal

and of

terminal differentiation into all types of mature formed elements of the blood (see figure 4). The HSC “

niche

”,

the

in vivo

regulatory microenvironment where HSCs reside, and the mechanisms involved in controlling the

number of adult HSCs will be discussed later

Each

day

an

adult

produces

approximately

200

billionerythrocytes,

100

billion

leukocytes,

and

100

billion

platelets. Moreover, these rates can increase by a factor or 10 or morewhen the demand for blood cells

increases. All this production relies on the presence of an adequate number of HSCs.

In the haematopoietic system, HSCs are heterogeneous with respect to their ability to self-renew. Multipotent

progenitors

constitute

0.05%

of

bone- marrow

cells,

and

can

be

divided

into

three

different

populations:

long-term

self- renewing

HSCs

(LT-HSC)

short-term

self-renewing

HSCs

(ST-HSC)

and

multipotent

progenitors

(MPP) without detectable self-renewal potential (see figure 4). These populations form a lineage

in which the long-term HSCs give rise to short-term HSCs, which in turn give rise to multipotent progenitors.

As HSCs mature from the long-term self-renewing pool to multipotent progenitors,

they progressively lose

their

potential

to

self-renew

but

become

more

mitotically

active.

Whereas

long-term

HSCs

give

rise

to

mature

hematopoietic

cells

for

the

lifetime

of

the

mouse,

short-term

HSCs

and

multipotent

progenitors

reconstitute lethally irradiated mice for less than eight weeks.

Figure 4

Description

of

the

HSC

compartment

with

LT-HSC,

ST-HSC

and

the

multipotential

progenitor

compartment

(MPP). Blood- cell development progresses from a hematopoietic stem cell (HSC), which can undergo either

self-renewal or differentiation into ST-HSC and the multipotential progenitor cells (MPP). MPP give rise to

two

major

multilineage

committed

progenitor

cells:

a

common

lymphoid

progenitor

(CLP)

or

a

common

myeloid progenitor (CMP). These cells then give rise to more- differentiated progenitors ultimately giving rise

to unilineage committed progenitors for B cells NK cells T cells granulocytes monocytes erythrocytes and

platelets.

HSCs cannot undergo normal somatic mitotic activity as any other kind of cells in the body as illustrated in figure

5.

Figure 5

Symmetrical division during normal mitotic activity.

If it would, then, by definition, by giving rise to two identical daughter cells trough symmetrical division would

exhaust

the HSC pool. In order to maintain a constant the pool of long-term HSCs an individual stem cell

has

to

give

rise

to

two

non- identical

daughter

cells,

one

maintaining

stem-cell

identity

and

the

other

becoming a

differentiated cell

by divisional asymetry. There are two mechanisms by which this asymmetry

can

be

achieved,

depending

on

whether

it

occurs

pre-

(divisional

asymmetry),

or

post-

(environmental

asymmetry) cell division.

Figure 6

Asymmetrical division in HSCs in the bone marrow

As shown in figure 6, in a, cell-fate determinants are asymmetrically localized to only one of the two daughter

cells, which

retains stem- cell fate

, while the second daughter cell

differentiates

. In b, during environmental

asymmetry,

after

division,

one

of

two

identical

daughter

cells

remains

in

the

self-renewing

niche

microenvironment

while

the

other

relocates

outside

the

niche

to

a

different,

differentiation-promoting

microenvironment.

Maintenance

of

long-term

HSCs

and

regulation

of

their

self-renewal

and

differentiation

is

thus

maintained

through asymetrical division.

The

main

characteristis

of

HSC are

their

quiescent

state

.

They

divide

infrequently

and can

be

quiescent

for

weeks or even months. They also are very few in numbers, representing 0.05% of all bone marrow cells.

Finally, they are the only cells able so self-renew for ever, for life-time, and able at the same time to give

birth to cells that will divide and mature into formed elements of the blood.

4. Concept of the Hematopoietic Stem Cell niche.

HSCs

reside

in

the

cavity

of

long

and

axial

bones.

As

we

will

see,

they

are

surrounded

by

a

special

microenvironment defined as

the stroma

. The stroma is composed of cells derived from mesenchymal stem

cells (MSCs),

which give rise to a mixture of cells including

fibroblasts, adipocytes, endothelial cells and

osteoblasts

.

Each

of

these

stromal

cells

is

essential

for

supporting

the

HSCs

in

their

physiological

role.

Hematopoiesis cannot occur alone and needs all these cell partners forming the microenvironment or

niche

.

This is where the concept of a HSC niche comes into play. As we will see, in the niche, the

osteoblasts

play

the most important role in supporting hematopoiesis.

A stem-cell niche can be defined as a spatial structure in which HSCs are housed and maintained by allowing

self- renewal

in

the

absence of

differentiation.

The stem-cell

niche

functions

include

storage

of quiescent

stem cells,self-renewal and inhibition of differentiation.

The main function of a self-renewing niche would be

to guarantee that by environmental and/or divisional asymmetry, one of the two daughters of a dividing stem

cell maintains the stem-cell fate while the other produces differentiating progenitors.

In

that niche,

HSCs are

in

intimate

contact

with

bone,

more

specifically,

in

close proximity

to

bone surfaces

(endosteal surfaces), supporting the concept of an endosteal niche (see figure 7)

Bone

marrow

niche

organization

showing

that

the

HSCs

are

not

randomly

distributed

in the

bone

cavity

but

rather concentrated and attached to the endosteal surfaces of the bone, more specifically to osteoblasts.

The more mature progenitor cells tend to localize in the middle of the cavity.

The mechanisms of bone and blood formation have traditionally been viewed as distinct, unrelated processes,

but compelling evidence suggests that

they are intertwined

. Based on observations

that HSCs reside close

to endosteal surfaces of the bones

as

shown in figure 7, it was hypothesized that

osteoblasts

play a central

role inhematopoiesis. We will see that osteoblasts are critical in the regulation of hematopoiesis and are one

of

the

most

important

regulatory

cells

in

the

stem

cell

niche.

To

put

it

very

simply,

“no

osteoblasts,

no

hematopoiesis”. Several animal modelsstrongly implicate osteoblasts in

hematopoiesis by virtue ofcreating

a niche. In mice with a maturational arrest of osteoblasts, there is a total lack of bone marrow throughout the

entire skeleton and therefore total absence of hematopoiesis.

5. Anatomical description of the Hematopoietic Stem Cell niche.

Where do we find the HSCs niches that are so important for maintaining hematopoiesis for life by virtue of their

protective effect on LT-HSCs? If one looks carefully at the anatomical aspects bones in general, including

long bones, there is a special area called spongy bone or cancellous bone (see figure 8). For long bones,

cancellous bone is at the

epiphysis.

Figure 8

Anatomical schema of a long bone with spongy or “cancellous bone” at the epiphysis this is the anatomical area

where most LT- HSCs reside in contact with osteoblasts in a close network of trabecula (see text). This type

of bone (spongy or cancellous) is mostly found in the axial skeleton. As shown in figure 3, the axial skeleton

is the major site of hematopoiesis in the adult.

Cancellous bone is a spongy type of bone

with a very high surface area, found at the ends of long bones and

axial bones (vertebrae). The very high surface area is the result of a complex network of trabecula,

which

are

fine

bone

spicules

.

The

spicules

form

a

latticework,

with

interstices

filled

with

bone

marrow

where

LT-HSC

are

in

intimate

contact

with

osteoblasts

and

other

mesenchymal

cells

such

as

adipocytes,

endothelial cells and fibroblasts (see figure 9).

Figure 9

Anatomy

of

the

HSC

niche

in

the

cancellous

or

spongy

bone

with

the

presence

of

a

network

of

trabeculae

creating multiple spaces thereby increasing the surface area where HSCs can come in intimate contact with

osteoblasts and provide life-long hematopoiesis

In the HSC niche, although the survival of HSCs requires intimate cell-cell contact with osteoblasts, one must

remember

that

bone

marrow

stromal

cells

derived

from

mesenchymal

stem

cells,

including

fibroblasts,

adipocytes and endothelial cells are also important in

supporting HSCs by secretion of important survival

proteins. In summary, the main anatomical site of the HSC niche is in the spongy or cancellous bone where

one

finds

a

significant

increase

in

bone

surface

area

ensuring

and

increase

and

adequate

number

of

LT-HSCs. As we will see,

the control of the HSC numbers is directly related to the number of osteoblasts

.

By increasing the bone surface area (by the same token the number of osteoblasts) through the network of

bone

trabecula,

there

is

a

parallel

increase

in

the

number

of

LT-HSCs.

In

the

adult,

the

spongy

or

cancellous

bone

is

found

mainly

at

the

proximal

ends

of

the

humerus

and

femur,

in

the

vertebrae,

ribs,

sternum and pelvis

.

6. Functional description of the Hematopoietic Stem Cell niche.

The functions of the niche include adhesive interaction between HSCs and the niche.

Although,

as

stated

earlier,

four

different

stromal

cells

interplay

within

that

niche,

such

as

adipocytes,

endothelial cells, fibroblasts and osteoblasts, the major function of the HSC niche is to regulate the number

of HSCs through a complex interaction with osteoblasts. This is why HSCs localize close to the endosteal

lining

of

bone- marrow

cavities

in

trabecular

regions

of

long

bones,

whereas

more

differentiated

hematopoietic

progenitors

are

found

mainly

in

the

central

bone- marrow

region

(see

figure

7).

Quiescent

HSCs detach from the endosteal niche and migrate towards the centre of the bone marrow to the vascular

zone

from

where

they

establish

hematopoiesis.

This

specific

site

(center

of

the

bone

marrow)

is

mostly

populated with endothelial cells, fibroblasts and adipocytes. It is called the vascular niche, as opposed to

the

endosteal

niche.

Collectively,

the

vascular

and

endosteal

niches

strongly

cooperate

to

control

HSC