HEMATOLOGY LECTURE NOTES

Dr. Brady-West

White Blood Cell Disorders
Learning Objectives:
At the end of these lectures, the student should understand:
1. The normal process of white cell production, differentiation and
maturation.
2. The etiolog and patholog of reactive changes in the number and
morpholog of granuloctes
!. The etiolog and patholog of reactive changes in the number and
morpholog of lmphoctes and monoctes
". The difference between a leu#emia and a leu#emoid reaction
$. The indication, procedure and interpretation of the leu#octe
al#aline phosphatase test %LA&'
(. The morphological definition and the implication of a leuco)
erthroblastic blood picture
*. The epidemiolog clinical features, laborator diagnosis, and
complications of +nfectious ,ononucleosis
-. The clinical, morphological, ctochemical and immunological
basis for the diagnosis and classification of leu#emia
.. The general scheme of treatment of acute leu#emia, and the
prognostic factors which affect the outcome of such therap
1/.The definition, classification, differential diagnosis and
management of the ,eloproliferative diseases
11.The epidemiolog, ctogenetics, clinical features, laborator
diagnosis, natural evolution and therapeutic options of 0hronic
,eloid Leu#emia
WHTE CELL DSORDERS
1e2uirements for leu#opoesis %white cell production':
1 a. Ade2uate numbers of normal stem cells
b. 3uitable microenvironment provided b a stromal matri4 on which adherent
stem cells can proliferate and differentiate
c. Ade2uate levels of growth factors %0olon 3timulating 5actors'
6ranulocte maturation
The earliest identifiable granulocte precursor is the !yelo"last, usuall found in small
numbers in the bone marrow but absent from the peripheral blood in health individuals.
There are three pools of marrow granuloctes
a. The mitotic pool which comprises all cells from the meloblast to the
melocte. These are all capable of self 7renewal b mitosis. 8ifferentiation
into neutrophil basophil and eosinophil is evident at the melocte stage.
b. The maturation pool which e4tends from the metamelocte to the mature
granulocte
c. The storage pool of mature granuloctes
There are two components of the peripheral blood granulocte pool
a. circulating
b. marginating % adherent to endothelium of small venules and capillaries'
6ranuloctosis ma occur b several mechanisms
a. ,obili9ation of marginating cells
b. +ncreased rate of maturation
c. +ncreased rate of mitosis
6ranules
&rimar % a9urophilic ' seen at the meloblast and promelocte stage ,and
contain the en9me ,eopero4idase
3econdar : these appear at the melocte stage. The are neutral staining in the
neutrophil, red) orange in the eosinophil and blue in the basophil.
:eutrophil
:umber 2.$ 7 *.$41/
.
;L
5unction %see illustration'
a. ,igration to the site of infection or inflammation
b. &hagoctosis
c. <illing microorganisms b o#y$e% de&e%de%t mechanisms. This
involves the production of hdrogen pero4ide and the supero4ide
anion b the en9me :A8= o4idase
d. <illing microorganisms b o#y$e% i%de&e%de%t mechanisms 7
intracellular acid ph, or en9mes lso9me and lactoferrin that are
contents of the secondar granules.
Lifespan of neutrophils in the marrow is 11 das. >hen neutrophils enter the
peripheral pool, the onl survive for hours. %=alf) life of ()- hours'. 3urvival in
tissues for 1)2 das
Ne'tro&hilia( Ca'ses
A. &hsiological
. ?igorous e4ercise
. &regnanc
. :ewborn
@. &athological
. @acterial infections
. +nflammation or necrosis
. ,etabolic disorders e.g. diabetic #etoacidosis, uremia, and eclampsia
. 3teroid therap
. Acute hemorrhage or hemolsis
0hanges in neutrophil morpholog in disease states include:
Left shift ) this is the appearance in the peripheral blood of more immature
components of the maturation pool
8ohle bodies and ctoplasmic vacuolation
To4ic granulation 7 increase in the number and intensit of secondar granules
Le')e!oid rea*tio%
8efinition: A4tremel high leu#octe counts seen in a non) leu#emic state and
ma be lmphoid or granuloctic in nature
0auses:
3evere infections
A4tensive burns
,alignancies with bone marrow infiltration
3evere hemorrhage
Lmphoid reactions seen usuall in children in response to viral infections
8ifferentiation from leu#emia b the following features:
1. &resence of an appropriate underling condition
2. ,orpholog of white blood cells: reactive e.g. to4ic changes vs. neoplastic
!. :o evidence of bone marrow failure %anemia or thromboctopenia'
". =igh LA& score in granuloctic reactions
LA+ test
This is a semi 2uantitative assessment of the level of functional al#aline
phosphatase in the ctoplasm of neutrophils.
,ethod: 5ilm is made from freshl collected blood, and immediatel fi4ed.
+ncubate in a phosphate solution, then rinse and counterstain.
+nterpretation: assess the number and intensit of blue ctoplasmic granules in
1// cells.
5or each cell score /)". ,a4imum score is "//. :ormal !$ )1//
/: :o stained granules
1: few granules
2: moderate staining
!: :umerous granules, strongl positive
": :umerous intensel stained granules

:eutropenia
8efined as a neutrophil count of less than 2.$ 4 1/
.
;L. Bsuall smptomatic at
C1./ 4 1/
.
;L., with recurrent infections, oral ulcers. 3erious or life)threatening reactions
occur at C /.2 1/
.
;L.
Classi,i*atio%
@enign familial
0clic: neutrophil counts fall at 21 da intervals and remain low for $)*
das. 8ue to failure of normal humoral feedbac# mechanism
3econdar: due to viral infections, autoimmune disease or
drug induced 7 most common adult cause of isolated neutropenia, associated with
anti)inflammator, antithroid, antihpertensive and oral hpoglcemic agents
Aosinophilia
8efined as an absolute eosinophil count D /.* 4 1/
.
;l
0auses
&arasitic infestation, especiall b organisms which invade tissues
Allergic disorders : bronchial asthma, urticariaE ha fever
8rug reactions
=ematologic diseases: 0hronic meloid leu#emia. &ernicious anemia,
=odg#in disease
@asophils
3imilar to mast cells found in tissues
+nvolved in +gA mediated hpersensitivit reactions. 3ubse2uent to reaction
between allergen and +gA the release of basophil granule contents e.g. histamine, lead to
the recogni9ed clinical features of allerg or hpersensitivit.

0auses of @asophilia
=pothroidism
,eloproliferative diseases
0hic#en po4
,ononuclear 0ells
Lmphoctes : &roduced in the bone marrow from pluri) potent stem cells.
T lmphoctes account for ($)-/F of peripheral blood lmphoctes and
are functionall divided into T helper cells %predominate in blood' and T suppressor cells
%predominate in marrow'
@ lmphoctes : these have endogenousl produced +g molecules on the
cell surface , which act as receptors for specific antigens.
Lmphoctosis : absolute lmphocte count D "./4 1/
.
;l. Levels are higher in
infanc and graduall decrease toward adult levels.
Ca'ses o, ly!&ho*ytosis
1. Acute infections : pertussis, hepatitis, infectious mononucleosis
2. 0hronic infections : tuberculosis , congenital sphilis
!. Lmphoma or leu#emia
,orphologic variations in lmphoctes in reactive states:
1. increased si9e
2. increase in ctoplasm cf to the nucleus
Mo%o*ytes
@one marrow monoctes arise from the same precursor cell as granuloctes. @one
marrow monoctes give rise to peripheral blood monoctes and tissue macrophages.
Tissue macrophages constitute part of the !o%o%'*lear &ha$o*yte syste!.
Mor&holo$y o, !o%o*ytes
?ariable si9e
Abundant gra ctoplasm, often vacuolated
Larger than lmphoctes
+ndented nuclei
,a combine to form giant cells
Mo%o*ytosis: 0auses
1. @acterial infections % most cause neutrophilia' sphilis, bacterial endocarditis
2. 1ecover from acute infections
!. &roto9oan infections
". 0ollagen vascular diseases
$. chronic steroid therap
(. 6ranulomatous diseases: sarcoidosis, ulcerative colitis.
Case History
A 2/)ear)old student presents with a *)da histor of fever sore throat, letharg and
tender enlarged glands in the nec#.
&hsical e4amination reveals fever, mild jaundice, inflamed pharngeal mucosa and
cervical adenopath
@lood results
=bE 12.$ g;dl, wbc 1-./41/
.
;l , differential !/F neutrophils "/F lmphoctes !/F
abnormal lmphoctes. &latelets 1// 41/
.
;l
Throat swab: :o bacterial growth
=+? test negative
,onospot test: positive
%,e*tio's Mo%o%'*leosis
0aused b infection with Apstein)@arr virus %A@?' and characteri9ed b:
5ever and pharngitis
Lmphadenopath and mild splenomegal
+ncreased circulating atpical mononuclear cells
=igh titers of heterophile antibodies
&ea# incidence at ages 1$ 72$ rs.
Cli%i*al ,eat'res
. +ncubation period of $)- wee#s
. &hangitis with edema and adenoidal hpertroph
. Lmphadenopath 7 tender, bilateral and smmetrical
. ,ild to moderate splenomegal in $/)*$ F
. Atpical features include s#in rash, hepatitis and encephalitis
Di,,ere%tial dia$%osis
1. Acute viral pharngitis caused b other organisms ) serological tests are
negative
2. Acute leu#emia 7 usuall significant anemia and ;or thromboctopeniaE also
peripheral blood lmphoid cells are blasts %with nucleoli'. &eripheral blood
picture will be the same or worse after 1/)1" das %will show improvement in
+.,.'
He!atolo$i*al ,eat'res
1. Leucoctosis of 12)1- 41/;l with atpical mononuclear cells. The majorit of
these are activated T lmphoctes.
2. Anemia and thromboctopenia are uncommon, and usuall autoimmune in
nature
Serolo$i*al -eat'res
1. A@?) specific antibodies
a. Antibodies to ?iral 0apsid Antigen %?0A' : +g, antibodies produced
during incubation period and pea# after 2)! wee#s then decline. +g6
antibodies subse2uentl appear and persist for life
b. Antibodies to :uclear antigen %A@:A' begin wee#s after onset of
illness and persist indefinitel
2. Autoantibodies: uncommon, ma cause autoimmune anemia or
thromboctopenia
!. =eterophil antibodies
These are non)specific serum agglutinins that will agglutinate sheep or
horse red cells. +, heterophile antibodies are differentiated b the
failure to be absorbed b guinea pig #idne cells. This is the basis
of the G,onospotH test.
Therap
1. Treatment is smptomatic and antibiotics do not positivel alter the course of
the illness
2. 3teroids are indicated for severe and complicated cases eg autoimmune
ctopenias or encephalitis
Acute Leu#emia
8efinition
A leu#emia is a clonal neoplastic proliferation of white cells in blood and;or bone
marrow .
0lassification of leu#emia
Acute ,eloid %A,L' or Acute Lmphoblastic %ALL'
0hronic ,eloid %0,L' or 0hronic Lmphoctic %0LL'
Case History
A ( ear old female presents with a ! wee# histor of fever, being less active than normal
and becoming easil tired.
3he also has bleeding gums and eas bruising for 1 wee#
&hsical A4amination:
&ale and febrile
Tender over ribs and sternum
,ultiple cutaneous hemorrhagic lesions
Anlarged cervical lmph nodes
Anlarged spleen
Laborator results
=b. (./g;dlE plats. 124 1/
.
;lE >@0 -$41/
.
;l
./F blasts
0I1: enlarged hilar lmph nodes
@, aspirate D ./F blasts
Cli%i*al ,eat'res o, a*'te le')e!ia
a. 8ue to organ infiltration
@one pain
Lmphadenopath or hepatosplenomegal
b. @one marrow failure
Anemia 7 dspnea, fatigue, palpitations
:eutropenia 7 fever, infections
Thromboctopenia 7 bleeding from s#in or mucosa
c. =permetabolic state
5ever
8renching night sweats
Apidemiolog
,ost common childhood malignanc is ALL
-/:2/ ALL: A,L in childhood, the ratio is reversed in adults
Anvironmental ris# factors
@en9ene
+oni9ing radiation
0hemotherap
6enetic disorders
8owns sndrome
<linefelter sndrome
:eurofibromatosis
0lassification of A,L is based on the degree of differentiation or maturation of the
neoplastic cells
,/: 7 A,L with minimal differentiation not recogni9able b morpholog
,1: A,L with no maturation ofD ./F of meloid blasts
,2: A,L with maturation : Auer rods and primar granules visible
,!: A&L % promeloctic maturation'
,": Acute ,elomonoctic leu#emia
,$: Acute ,onoctic leu#emia
,(: Arthroleu#emia
,*: Acute ,ega#aroctic leu#emia
0lassification of ALL ma be based on morpholog or immunolog,
,orphologic classification %5A@'
L1: @lasts are homogenous, small, with scant nucleoli and high :;0 ratio
L2 @lasts are larger, heterogeneous with prominent nucleoli
L!: @lasts are large with basophilic ctoplasm and ctoplasmic vacuoles
+mmunologic classification
T)ALL shows earl T cell antigensE ma be of L1 or L2 morpholog
0)ALL shows earl @ cell antigens
J @) ALL shows mature @ cell antigensE this is alwas L! in morpholog
8iagnosis of acute leu#emia
At least !/F blasts in bone marrow aspirate
A,L and ALL are differentiated b morphological appearance and
0tochemical stains 7 melopero4idase Kve for A,L and &A3Kve for ALL
&rognostic factors in AML
Age less than 2 or greater than (/ ears
&receding hematological disorder
>@0 greater than 1//
Tpes ,/ , ,( and ,*
&rognostic factors in ALL
5avorable Bnfavorable
Age : 2)1/ ears C 2 or D 1/ ears
>@0: 1/ or less D $/
6ender: female male
Tpe: L1 ; 0)ALL L! ; @)ALL
1emission: earl late
A,8: absent present
Treatment and outcome of A,L
+nduction of remission with intensive chemotherap with 0tosine, 8aunorubicin
0onsolidation therap with repeated courses of similar agents
($)-/F achieve complete remission
1/)!/F cure rate
Treatment and outcome of ALL : treatment is stratified according to ris#
1emission induction
0onsolidation; intensification
0:3 prophla4is with intrathecal methotre4ate and ;or radiation
,aintenance %prevention of bone marrow relapse' for 2) ! ears
@one marrow transplantation
1/ ear survival "/ 7-/ F
,elodsplasia
,elodsplastic 3ndromes %,83'
8efinition : =eterogenous group of clonal disorders characteri9ed b :
1. peripheral blood ctopenias with normal or increased marrow cellularit
2. morphological and functional abnormalities
!. pea# incidence in the elderl
0auses
,ost are idiopathic
A4posure to al#lating agents
+oni9ing radiation
0linical features
Anemia
1ecurrent infections
Abnormal bleeding
8splastic changes in peripheral blood or marrow
,acroctic anemia
,egaloblastoid erthropoesis
Agranular neutrophils
1inged sideroblasts
monoctosis
Therap and outcome
Treatment depends on age, tpe of ,83, general condition of the patient
3upportive therap with transfusion of red cells or platelets
0hemotherap for advanced disease similar to treatment for A,L
@one marrow transplant 7onl in relativel oung patients.
Outcome is variableE 2$)"$F transform to A,L
MYELO+ROL-ERAT.E DSEASES
De,i%itio%: These are a group of related chronic marrow diseases that have in common
the hperplasia of cellular and ;or stromal bone marrow components. The are classified
based on the nature of the &redo!i%a%t proliferating cell line:
1. &rimar polcthemia % erthroid'
2. Assential thrombocthemia %mega#aroctic'
!. 0hronic meloid leu#emia % granuloctes'
". &rimar melofibrosis % fibrous tissue'
Cli%i*al ,eat'res
1. :on)specific features common to all, due to a hpermetabolic state
a. 5ever , weight loss and drenching night sweats
b. 3plenomegal : most prominent in ,5 and 0,L
2. 3pecific features such as bleeding or thrombosis in &1? and AT
!. All ma be incidentall discovered on routine phsical or laborator tests
Dia$%osis
1. A4clude a secondar or reactive state that can mimic the primar disorder.
There are four such reactive conditions
a. 3econdar polcthemia %vs. &1?'
b. 1eactive thromboctosis %vs. AT'
c. Leu#emoid reaction %vs. 0,L'
d. 3econdar melofibrosis %vs. ,5'
2. +dentif the specific ,&8 b the presence of diagnostic criteria eg.
a. +ncreased red cell mass or pac#ed cell volume
b. &latelet count above (//4 1/
.
;l
c. &hiladelphia chromosome
d. @one marrow fibrosis D 1;!
0ase =istor
A $- ear old 0aucasian man is admitted for elective repair of an inguinal hernia.
1outine 0@0 : =b. 21.$ g;dl, &0? /.(1E >@0 1( 4 1/
.
;LE platelets $2/4 1/
.
;L
&hsical e4amination: enlarged spleen
=e admits to having recurrent headache and blurred vision for the past ( months
+ri!ary +oly*ythe!ia /+R.0
&olcthemia is defined as an elevation of the pac#ed cell volumeE and ma be:
1. Absolute &olcthemia: the red cell mass is actuall increasedE this increase
ma be :
a. +diopathic : this is primar proliferative polcthemia %&1?'
b. 3econdar to underling diseases which produce increased A&O
i' =po4ic states eg. 0anotic heart disease, chronic lung
disease
ii' +nappropriate A&O production eg. 1enal csts, renal cancer,
phaeochromoctoma
2. 1elative polcthemia: there is no increase of red cell mass, but a relative
decrease in plasma volume causes an increased &0?
+ri!ary +oly*ythe!ia
&ea# incidence in the (
th
decade, but ma be seen in oung adults
0ommon signs and smptoms
&lethoric s#in
3plenomegal
=eadache and di99iness
?enous or arterial thrombosis
0riteria for diagnosis
+ncreased &0? D /.$$
Arterial o4gen saturation D .2 F
3plenomegal
Leucoctosis ; thromboctosis
,anagement
1. reduce blood volume b phlebotom 1)2 per wee# until &0? is
C"$
2. allopurinol to prevent urate nephropath
!. ,elosuppression with hdro4urea
". 1adioactive phosphorus in older patients
+ri!ary !yelo,i"rosis
&resents with smptoms of anemia or of massive splenomegal .
Laborator features
1. leucoerthroblastic blood picture which comprises:
a. tear drop shaped red cells
b. nucleated red cells in peripheral blood
c. immature granuloctes
2. :ormochromic anemia
!. ?ariable white cell and platelet counts
". Alevated LA& score
$. &rogressive bone marrow fibrosis
0auses of secondar fibrosis must be e4cludedE
1.,arrow infiltration b lmphoma, leu#emia or solid tumors
2. 6ranulomatous diseases eg. Tuberculosis or sarcoidosis
.,anagement
3mptomatic with transfusion of blood products
3plenic si9e ma be reduced b chemotherap or splenic
irradiation
,edian survival is !)* ears. 2/F transform to acute meloid
leu#emia
Chro%i* Myeloid Le')e!ia
Case History
A !2)ear)old man presents with a left upper abdominal mass for 1 month, but has no
other smptoms.
=b. 1$./ g;dlE platelets normalE >@0 "!4 1/
.
;lE
>@0 differential: "$F :E 2F LE -F AoE (F @asoE !$F meloctesE "F
metameloctes
LA& score: 2-
De,i%itio%: A clonal stem cell disorder characteri9ed b increased granuloctic
proliferation at all stages of maturation.
=ematological features:
1. Leucoctosis
2. 5ull spectrum differential with pea#s at melocte and mature
neutrophil
!. Aosinophilia and basophilia
". LA& score low
8istinguish from leu#emoid reaction b:
1. :o clinical histor of infection or inflammation etc.
2. The presence of significant splenomegal
!. The low LA& score
". The wbc differential
$. The presence of &hi chromosome
Cyto$e%eti* ,eat're
&hiladelphia chromosome: mutual translocation with e4change of genetic
material between chr . and chr 22. This results in he formation of an abnormal hbrid
gene %@cr)Abl' that results in increased cell proliferation
:ote: The &hiladelphia chromosome is %ot specific for 0,L. &resent in .$F of patients
with 0,L, also found in some cases of ALL
Role o, +hi i% &atho$e%esis o, CML
6enetic se2uence on 0hr 22 % bcr ' are fused with se2uences translocated from chr .
abl' . This fusion gene codes for an abnormal protein with Tyrosi%e 1i%ase activit. This
Trosine <inase is involved in signal transduction and activates pathwas within the
affected cells leading to malignant tra%s,or!atio%. Tyrosi%e )i%ases 2or) "y
tra%s,erri%$ a &hos&hate $ro'& ,ro! AT+ to i%tra*ell'lar &rotei%s that re$'late *ell
di3isio%.
Nat'ral E3ol'tio% o, CML
1. 0hronic phase: usuall presents in this phase, with progressive leucoctosis
and splenomegal. The disease is responsive to ctoto4ic therap.
2. Accelerated phase: 3mptoms are worse, with night sweats, bone pain,
splenomegal less responsive to therap. &eripheral blood shows increasing
numbers of basophilic, blasts and promeloctes. 8isease is difficult to
control.
!. @lastic phase: smptoms progress, e4tramedullar deposits %chloromas'
appear, D !/F blasts in blood or bone marrow.
@last transformation ma be lmphoid %1$F' or meloid %-$F'
Treat!e%t o, CML
A. 3upportive treatment
1. =dration and allopurinol prior to starting ctoto4ic therap for prevention of urate
nephropath %nucleic acid brea#down'
2. Analgesics for bone pain or splenic pain 7 these are more commonl seen in the
accelerated and blast phases
!.3plenic irradiation ma be used for palliation of massive splenomegal
@. 3pecific treatment
1. 0hoice of therap depends on the age of the patient, phase of disease and the
availabilit of a matched bone marrow donor.
2. 0hronic phase
a. Treatment of choice used to be bone marrow transplant if patient is
less than "$ ears old, and a matched donor is available for allogeneic
bone marrow transplant. Onl 2$F of patients are eligible.
b. +f bone marrow transplant is not possible 7 ) interferon will
effectivel lower the cell count. +nterferon is administered
subcutaneousl. ,ain side effect is fever ; malgia. E#&e%si3e
+nterferon is not used if @,T is planned 7 worsens outcome
c. 6livec % imatinib' inhibits the trosine #inase activit of the protein
produced b the fused gene on the &hi chr. :ow approved for first
line treatment. Ad3a%ta$e ( oral ad!i%istratio%4 also a*ts at as
tar$eted thera&y to &re3e%t &rod'*tio% o, the !ali$%a%t *lo%e.
This has %o2 re&la*ed "o%e !arro2 tra%s&la%t as treat!e%t o,
,irst *hoi*e. +rod'*es *li%i*al5 hae!atolo$i*al a%d *yto$e%eti*
re!issio%s i% a hi$h &er*e%ta$e o, &atie%ts i% the *hro%i* &hase.
d. =dro4urea: 6iven orall $oomg 7 ! 6m dail. 5re2uent monitoring
of >@0 is necessar because the rate of fall is unpredictable. 0ounts
recover 2uic#l after cessation or lowering of dose. ,a be used in
pregnanc. Not *ar*i%o$e%i*
e. @usulfan: 6iven orall as dail low dose or pulse high dose.
&redictable rates of fall of >@0 so fre2uent monitoring of counts not
necessar. Low counts do not recover promptl after cessation. ,a
cause prolonged marrow aplasia. 0ontraindicated in pregnanc. Bsed
in older patients when fre2uent clinic visits are inconvenient. 3ide
effects include pulmonar fibrosis
!. Accelerated phase
a. 0toto4ic therap used in higher doses, or switch agents.
b. 0ombination of agents 7 add ctosar to oral agent
". @last phase
a. Lmphoid : vincristine and prednisone will induce remissions in some
patients % bac# to chronic phase' E remissions are of short duration
b. ,eloid : ctosar and adriamcin , remissions are harder to induce
than in de)novo A,L.