Presentations text content in Evaluation of Stillbirth
Slide1
Evaluation of Stillbirth
Katy
Kemnetz
, PGY2
Slide2objectives
Evaluation of stillbirth
Slide3Objectives
The participant will be able to:
Identify the conditions that have been best demonstrated to cause stillbirth
Evaluate a stillbirth using the most effective workup
Accurately formulate an etiology for a stillbirth, when possible
Employ the recmomended hospital policies for management of stillbirth
Slide4Outline
Definitions
Causes of stillbirth
Workup of stillbirth
Other
considerations
Conclusions
Slide5Definitions
Evaluation of Stillbirth
Slide6Definitions
Spontaneous abortion
Intrauterine fetal demise
Miscarriage<20 weeks gestation or <500g
“Stillbirth”
>20 weeks gestation or >350g—state dependent
350g is 50%ile for 20 weeks gestation
Illinois: >20 weeks gestation
“Delivery of a fetus showing no signs of life as indicated by the absence of breathing, heart beats, pulsation of the umbilical cord, or definite movements of voluntary muscles”
Does not include terminations of pregnancy or IOL for
previable
PPROM
Slide7Causes
Evaluation of stillbirth
Slide8Causes of stillbirth
>30 classification systems existImportant to distinguish betweenUnderlying cause of deathMechanism of deathRisk factors
Slide9Classification of stillbirth
The National Institute of Child Health and Human Development
Slide10Classification of Stillbirth
Eunice Kennedy Shriver workshop 2007
National Institute of Child Health and Human Development
“An optimal classification system would identify the
pathophysiological entity initiating the chain of events that irreversibly lead to death
”
Slide11Criteria for “cause”
Epidemiologic data demonstrate an
excess of stillbirth
associated with that condition
Biologic plausibility
that the condition causes stillbirth
Either
rarely seen in association with live births
or, when seen in live births, results in a
significant increase in neonatal death
A
dose-response relationship
exists
The greater the “dose” of the condition, the greater the risk of fetal death
Associated with
evidence of fetal compromise
The stillbirth
likely would not have occurred
if that condition had not been present
Slide12Causes of stillbirth
Reddy, UM et al. “Stillbirth Classification—Developing an International Consensus for Research.” Obstetrics and Gynecology,
Vol 114, No 4, October 2009.
Slide13Infections
Causes of stillbirth—NICHHD workshop consensus
Slide14Infection
Associated with
10-20%
of stillbirths in developed countries
Higher association with
preterm birth
Sometimes difficult to prove
causality
Slide15Infection
Ascending infectionamniotic fluid or fetusfetal pneumonitisHematogenous spreadvillitis
Kumar: Robbins pathologic basis of disease, 8
th
edition. 2009
Slide16Mechanism of fetal death
Severe maternal illnessPlacental infection that prevents oxygen/ nutrients from crossing to the fetusFetal infection that causes a lethal congenital deformity
Fetal infection that
damages a vital organ
Precipitation of
preterm labor
, with intrapartum fetal
death
Slide17Infections must be proven
Signs of infection in the fetusEvidence on autopsy of extensive organ involvementPositive fetal culturesPositive maternal cultures plus chorioamnionitis/ funisitis
Kumar: Robbins pathologic basis of disease, 8
th
edition. 2009
Slide18Causes of IUFD: Spirochetes
OrganismMaternal diseaseCommentTreponema palladiumSyphilisMajor cause of stillbirth when maternal prevalence is highBorrelia burgdorferiLyme diseaseTick borne; not a common cause of stillbirthBorrelia recurrentisRelapsing feverTick borne; common in the Western US; rare cause of stillbirthBorrelia duttoniiRelapsing feverTick borne; sub-Saharan Africa; important cause of stillbirthLeptospira interrogansLeptospirosisUncommon
Severe placental dysfunction
Slide19Protozoa
OrganismMaternal diseaseCommentTrypanosoma bruceiTrypanasomiasisTsetse flyTrypanosoma cruziChagas diseaseKissing bugPlasmodium falciparumMalariaCommon in endemic areasPlasmodium vivaxMalariaMosquitoesToxoplosmosis gondiiToxoplasmosisRare
Severe placental dysfunction
Slide20Viruses
OrganismMaternal diseaseCommentParvovirus B19Erythema infectiosumLikely the most common viral etiologic agentCoxsackie A and BVariousMay be importantEchovirusVariousImportance unknownEnterovirusVariousImportance unknownHepatits E virusFulminant hepatic failureEspecially in endemic areasPoliovirusPolioHistoric causeVaricella zosterChickenpoxRare causeRubellaGerman measlesRare in developed countriesMumpsParotitisRare in developed countriesRubeolaMeaslesRare in developed countriesCytomegalovirusAsymptomaticCase reportsHIVAIDSNot likely causativeInfluenzaRespiratory tract infectionSevere maternal illness
Causes lethal fetal anomalies
Slide21Bacteria
OrganismMaternal diseaseCommentE. ColiAsymptomaticProbably the most common organism assoc with stillbirthGBSAsymptomaticCommon cause of stillbirthKlebsiellaAsymptomaticCommon cause of stillbirthEnterococcusAsymptomaticUreaplasma, mycoplasmaAsymptomaticListeria monocytogenesListeriosisTransmitted transplacentallyChlamydia trachomatisPelvic infectionSuggested cause—case reportsNeisseria gonorrhoeaePelvic infectionSuggested cause—case reportsCandida albicansThrush; vaginitisConfirmed in case reports
Goldenberg RL, Thompson C. The infectious origins of stillbirth. Am J Obstet Gynecol2003;189:861–73
Severe placental dysfunction
Damage to vital organs (brain, heart)
Slide22Maternal medical conditions
Causes of stillbirth—NICHHD workshop consensus
Slide23Simpson, LL. Maternal medical disease: Risk of Antepartum Fetal Death. Semin Perinatol, 2002, 26, 47.
ConditionEstimated stillbirth rate per 1000 births in patients with the conditionAll pregnancies6-7Chronic hypertension5-25Superimposed preeclampsia52Gestational hypertension and mild preeclampsia9Severe preeclampsia21Eclampsia18-48HELLP syndrome51
Hypertensive disorders
Slide24Hypertensive disorders
Mechanism of fetal demise:
To consider cause of death:
Placental insufficiencyIUGRAbruption
If it progresses to
eclampsia
If it is associated with placental abruption or fetal growth restriction
Slide25Simpson, LL. Maternal medical disease: Risk of Antepartum Fetal Death. Semin Perinatol, 2002, 26, 47.
ConditionEstimated stillbirth rate per 1000 births in patients with the conditionAll pregnancies6-7Gestational diabetes5-10Type 1 diabetes6-10Type 2 diabetes35
Diabetes
Slide26Diabetes
Mechanism of fetal demise:
To consider cause of death:
Congenital abnormalityPlacental dysfunctionObstructed labor and intrapartum deathMacrosomiaFetal hyperglycemiafetal insulin productionexcessive fetal growthmetabolic acidosis
Signs of intrauterine or
intrapartum
asphyxia
LGA fetus
SGA fetus
Severe malformation
Placenta demonstrates characteristic
histologic
findings
Large edematous
villi
Increased prominence of
cytotrophoblasts
Slide27Simpson, LL. Maternal medical disease: Risk of Antepartum Fetal Death. Semin Perinatol, 2002, 26, 47.
ConditionEstimated stillbirth rate per 1000 births in patients with the conditionAll pregnancies6-7Stable treated hyperthyroidism0-36Uncontrolled thyrotoxicosis100-156Subclinical hypothyroidism0-15Overt hypothyrodism15-125SLE40-150Mild chronic renal disease15Moderate and severe chronic renal disease32-200Cholestasis of pregnancy12-30
Thyroid/renal disorders
Slide28Thyroid/renal disorders
Thyroid disorders
Renal disorders
Graves disease, where thyroid-stimulating hormone receptor antibody causes fetal toxicosisUntreated thyroid disorders
Linear relationship between maternal creatinine and risk of fetal demise
Systemic Lupus Erythematosus
Stillbirth
rates are higher in the presence of HTN, nephritis, or APL
Circulating
auto-antibodies, anti-Ro, anti-La
Congenital heart block,
hydrops
Slide29Maternal medical conditions
Risk is a continuum
Reddy, UM et al, 2009
Slide30thrombophilias
Causes of stillbirth—NICHHD workshop consensus
Slide31Thrombophilias
Antiphospholipid syndrome
Inherited thrombophilias
Inflammation, thrombosis, and infarction in the placentaClear histopathological or clinical evidence of placental insufficiency
Factor V Leiden mutation, antithrombin III deficincy, prothrombin gene mutation, protein C deficiency, protein S deficiencyPlacental infarction and thrombosis
Two large prospective cohort studies found no association between factor V Leiden mutation and pregnancy loss or placental insufficiency
Thrombophilias
are common in healthy women with normal outcomes
Thrombophilias should only be considered as the cause of stillbirth with:
Evidence of placental insufficiency such as fetal growth restriction or infarction and
Recurrent fetal loss
Slide32alloimmunization
Causes of stillbirth—NICHHD workshop consensus
Slide33Alloimmunization
Red cell alloimmunization
Platelet alloimmunization
Anti-Rhesus D, anti-Rhesus C, anti-KellMust have a positive indirect Coombs testAntibody titers more than 1:16 (or 1:8 for anti-Kell)Evidence of fetal anemia with hydropsEvidence of fetal extramedullary hematopoeisis
HPA-1a, HPA-5a, HPA-4
Maternal antibodies against paternal and fetal platelet antigens
Parental platelet incompatibility for the pertinent antigen
Fetal thrombocytopenia
Massive intracranial hemorrhage
Slide34Congenital malformationschromosomal abnormalities
Causes of stillbirth—NICHHD workshop consensus
Slide35Criteria
Epidemiologic data demonstrating an
excess of intrauterine mortality
Seen
rarely in
liveborn
neonates
When seen in
liveborn
neonates, it
frequently results in neonatal death
Biologic plausibility that it
can result in death
Slide36Cause of deathStillbirth cases, % (out of total 2211 assessed)Trisomy 211.53Jugulolymphatic obstruction1.45Turner syndrome1.09Twin-twin transfusion syndrome1.09Anencephaly1.0Trisomy 180.81Amnion disruption sequence0.59Lower mesodermal defects0.50Idiopathic nonimmune fetal hydrops0.50Trisomy 130.41
Congenital malformations
Reddy, UM et al. “Stillbirth Classification—Developing an International Consensus for Research.” Obstetrics and Gynecology, Vol 114, No 4, October 2009.
Slide37Chromosomal abnormalities
Incidence
Cytogenetic abnormalities account for 6-13% of all stillbirthsThis may be higher because 40-50% attempted karyotypes fail to grow23% monosomy X, 23% trisomy 21, 21% trisomy 18, 8% trisomy 13
Kumar: Robbins pathologic basis of disease, 8
th
edition. 2009
Slide38Chromosomal abnormalities
Autosomal recessive disorders
Autosomal dominant disorders
Alpha thalessemia, Smith Lemli OpitzPhenotype in lethal cases may differ from live cases
Skeletal dysplasiasMore often spontaneous mutations
Firestein: Kelley’s Textbook of Rheumatology, 9
th
ed. Saunders 2012
Slide39Fetomaternal hemorrhage
Causes of stillbirth—NICHHD workshop consensus
Slide40Fetomaternal hemorrhage
The cause
4% of all stillbirths
Risk factors:
Placental abruption
Abdominal trauma
Multiple gestation
Abnormal fetal testing
Slide41Fetomaternal hemorrhage
Risk of stillbirth depends on
Amount of hemorrhage
Acute/chronic
Gestational age
A threshold of
20
mL
/kg of fetal bleeding
is associated with increased risk of stillbirth
Autopsy confirmation of
fetal anemia and hypoxia
Slide42Placental causes
Causes of stillbirth—NICHHD workshop consensus
Slide43Placental causes
Placenta previa, vasa previa, neoplasmsPlacental abruption has 8.9 relative risk of stillbirthMay be considered the cause of death if >30% of the placenta shows signs of abruption
Reddy, UM et al, 2009
Slide44Placental causes
Any disease that causes an SGA placenta may result in stillbirth
<5% expected weight for gestational age
Preeclampsia, DM, HTN, renal, chronic infections
Any disease that causes an LGA placenta may result in stillbirth
>95% expected weight for gestational age
Hydrops
fetalis
, DM, syphilis
Slide45Umbilical cord pathology
Stillbirth classification—NICHHD workshop consensus
Slide46Umbilical cord pathology
Account for
3.4-15% of stillbirths
Velamentous
insertion
If it leads to a
vasa
previa
or bleeding
during labor
Umbilical cord
prolapse
Associated with prematurity,
malpresentation
,
mutiparity
, obstetric manipulation
Umbilical cord occlusion
Cord
prolapse
, entanglement (mono-mono twins)
Torsion
Rupture, strictures, hematomas
Slide47Velamentous insertion
Slide48Umbilical cord pathology
Nuchal cordOccurs in up to 30% of normal pregnanciesNot associated with an increased risk of stillbirth in study of 14,000 deliveriesTrue knotAlso common in live birthsGrooving of the cord, constriction of the umbilical vessels, edema, congestion, thrombosisrequired to claim it is the etiology
Isolated finding of a nuchal cord or a true knot at the time of delivery is insufficient evidence that cord accident is the cause of stillbirth
Exclude other relevant causes of stillbirth
Find evidence of
hypoxia and cord occlusion
on postmortem examination
Slide49Multifetal gestation
Stillbirth classification—NICHHD workshop consensus
Slide50Complications of multifetal gestation
Monochorionic placentationTwin-twin transfusion syndrome occurs in 9% of mono-di twinsMortality can be 90% in untreated cases
Slide51Complications of multifetal gestation
Mono-mono twinsCord entanglement, preterm birth, growth impairment, malformations, genetic abnormalities, vascular anastomoses
Gabbe: Obstetrics: normal and problem pregnancies, 6
th
ed. Saunders 2012
Slide52Uterine complications
Causes of stillbirth—NICHHD workshop consensus
Slide53Uterine complications
Uterine rupture
Evidence of obstructed circulation
Uterine abnormalities
There is an increased risk of uterine abnormalities in women with recurrent pregnancy loss/stillbirth
Possibly due to
poorly
vascularized
uterine tissue
or
space constraints
Increased risk of PPROM, cervical insufficiency, preterm labor
Septate
uterus
has highest risk of stillbirth and placental abruption
Slide54Workup
Evaluation of stillbirth
Slide55importance
Workup of stillbirth
Slide56Importance of a stillbirth evaluation
Counseling for
risk of recurrence
Possible
intervention
to reduce recurrence risk
Facilitate
emotional closure and healing
Slide57Most stillbirths remain unexplained
Incomplete evaluation
Lack of clinician awareness
Concerns of the family
Lack of single universally accepted
classification scheme
Difficult to assign a definitive cause
Unknown cause
Sometimes despite thorough evaluation
Slide58overview
Workup of stillbirth
Slide59Overview
Recommended studiesSometimes helpfulNot generally usefulAutopsyPlacental pathologyKaryotypeKliehauer-BetkeIndirect CoombsAcquired thrombophilia panelAnti B2-glycoprotein abToxicology screenSyphilis serologyInherited thrombophilia panelGlucose screeningTSHCMV, toxoplasmosis, other infectiousBile acidsSonohysterogramRoutine TORCH titersANA testingCultures of placental membranes
Silver et al, 2010
Slide60How do we know what works?
“
Evaluation of 1025 fetal deaths: proposed diagnostic workup
.”
Korteweg
FJ,
Erwich
JJHM,
Timmer
A, et al. AJOG 2012.
Prospective cohort study in 50 Dutch hospitals
Singleton IUFDs at >=20wks gestation, excluding pregnancy terminations and
intrapartum
deaths
Cause of death determined by a
multidisciplinary panel
(2 obstetricians, an OB resident,
perinatal
pathologist, neonatologist, genetics, microbiologist)
Initial
demonstratable
pathophysiological
entity
initiating the chain of events that irreversibly led to death
Contribution of each diagnostic test for determination of cause of death was determined by the panel
Valuable for “
establishing cause of death
” or “
excluding cause of death
”
Slide61Medical history
Workup of stillbirth
Slide62Thorough medical/obstetrical history
Pregnancy complications
Maternal medical history
Genetic abnormalities
Substance use
Pregnancy history
Family history
Slide63Examinationof the fetusand placenta
Workup of stillbirth
Slide64Examination of the fetus
Workup of stillbirth
Slide65Physician’s exam
Weight, head circumference, lengthPhotographsFrontal and profieWhole body, face, extremeties, palms, abnormalities
Kumar: Robbins pathologic basis of disease, 8th edition. 2009
Slide66FindingTime of demiseBrown or red discoloration of the cord stumpDesquamation >1cm>6 hours agoDesquamation of face, back, abdomen>12 hours agoDesquamation >5% of the body or >2 body zones>18 hours agoSkin color brown or tan>24 hours agoMummification(reduced soft tissue, leathery skin, dark brown)>2 weeks ago
Time of demise
“Estimating the time of death in stillborn fetuses: III. External fetal examination; a study of 86 stillborns.” Genest DR, Singer DB. Obstet and Gynecol, 1992; 80(4), 593
Slide67Autopsy
New information that influences counseling in
26-51%
of cases
Dutch study: valuable in
72% of cases
However, it is performed in
<50% of cases
Clinician hesitation to recommend autopsy
Patient reservations
Slide68Autopsy
Ask about patient reservations
Emphasize potential benefits to family
Allow the family ample time to hold the baby and perform religious/cultural activities first
Slide69Autopsy
Medical autopsies require consent of next-of-kinConsent should be explicit enough that parents can limit the autopsy procedure
Slide70Illinois Masonic Policy
“An autopsy is an option for fetuses 20 weeks or greater, but if clinically indicated or requested by the family, an autopsy can be offered before 20 weeks.”
If an autopsy is not requested, a Surgical Pathology examination can be performed
Slide71Components of an autopsy
Review of the clinical history, laboratories, antenatal imaging
External examination:
Weight, crown-rump length, foot length, crown-heel length, occipital-frontal circumference
Joint mobility, facial features, hands, feet, genitalia
Intraoral examination:
Pharynx, soft palate, uvula
Take into consideration changes from maceration
Photographs
Slide72External examination
Lentz: Comprehensive Gynecology, 6
th ed
Kumar: Robbins pathologic basis of disease, 8
th
edition. 2009
Slide73Components of an autopsy
Internal examination
Y or T shaped incision on anterior trunk
Organs examined in situ
Heart sometimes opened in situ, sometimes removed and fixed
Vascular connections, septal defects
Organs removed in bloc for detailed dissection
Spinal cord removed
Slide74Components of an autopsy
Brain
Coronal scalp incision extending to behind the ears
Skull is opened along cranial sutures
Can extend into posterior neck for posterior
fossa
(Dandy-Walker,
Chiari
) by removing occipital bone and cervical vertebrae
Brain is placed in formalin for 10 days before examination
Slide75Brain
Chiari
malformation—downward displacement of medulla
Fligner
2011
Slide76Components of an autopsy
Samples of tissue for
Microscopic evaluation
Viral cultures
Bacterial cultures
Metabolic studies (Guthrie card)
Slide77Histologic exam
Kumar: Robbins pathologic basis of disease, 8
th edition. 2009
Extramedullary
hematopoeisis
in fetal liver in fetus with non-immune
hydrops
Slide78What autopsy can identify
Markers of IUGR
Head circumference percentile higher than weight percentile
Brain weight appropriate for gestation while other organs are appropriate for body weight
Altered brain
weight:liver
weight
Kidneys
Polycystic kidney disease
Certain brain pathologies
Congenital anomalies
Slide79Kidneys
A: 28 wks, suspicious for
autosomal
recessive polycystic kidney disease
B: confirmed on autopsy
C: 22 wks, suspicious for ARPKD
D: Autopsy showed diffuse renal cystic dysplasia
Slide80Autopsy can provide evidence
InfectionAnemiaHypoxia
Kumar: Robbins pathologic basis of disease, 8
th
edition. 2009
Slide81Alternatives to autopsy
Partial autopsy
Head-sparing autopsy (may miss CNS pathology)
External examination by a trained pathologist
Can identify syndromes, congenital anomalies, timing of death, growth anomalies
Will likely miss fetal infections and internal anomalies
External examination with selected biopsies
More likely to identify fetal infections
Slide82Alternatives to autopsy: MRI
Very good for CNS pathologySometimes better than autopsy, because fetal brain has high water content and liquefiesFluid collections and effusions in the body
May miss cardiac anomalies, bowel anomaliesCannot diagnose infections or metabolic disease
Advantages
Disadvantages
Slide83MRI
24 wks, massive bilateral
intraventricular hemorrhage
Fligner
2011
Slide84MRI
Image-guided percutaneous biopsies<50% success rate at obtaining tissue in one studySomewhat inferior to autopsy:12 cases where “minimally invasive autopsy” provided equivalent information to autopsy12 cases where “minimally invansive autopsy” provided inferior information than autopsy2 cases where MRI and biopsy provided superior informationHowever, 73% of minimally invasive autopsies provided Information of equivalent clinical significance to autopsy
Fligner
2011
Slide85Alternatives to autopsy
X ray
Useful for skeletal
dysplasias
Ribs/vertebrae as part of diagnosis of VACTERL or diabetic
fetopathy
Slide86X ray
Achondrogenesis
1b (absence of vertebral bodies)
Fligner
2011
Slide87X ray
Hemivertebrae
(VACTERL)
Fligner
2011
Slide88Examination of the placenta
Suggested workup of stillbirth
Slide89Examination of the placenta
The most valuable diagnostic test in most studies
Dutch study showed it to be valuable in 95% of
cases
Provides additional information in 30% of cases
Slide90Examination of the placenta
Weight
in relation to norms for gestational age
Evidence of abruption, infarction,
thrombophilias
Hemosiderin
deposits
chronic
abruption
Perivillious
and marginal fibrin deposition
Decidual
necrosis
Evidence of infarction
Slide91Examination of the placenta
Multiples:
c
horionicity
, vascular
anastomoses
in
multifetal
gestations
Cord: thrombosis
,
velamentous
cord insertion,
vasa
previa
Evidence of infections
More common in preterm stillbirth
Viral
nuclic
acid
amplification
Bacterial cultures
Slide92Examination of the placenta
Kumar: Robbins pathologic basis of disease, 8
th
edition. 2009
Slide93genetics
Workup of stillbirth
Slide94Karyotype
Abnormal fetal
karyotype
noted in
8-13% of all stillbirths
and in >20% of those with morphologic abnormalities or IUGR
Dutch study:
11.9% prevalence of a chromosomal abnormality
in the 362
IUFDs
who underwent
karyotyping
37%
trisomy
21, 16%
monosomy
X, 4%
trisomy
13
Karyotype
was
valuable in 29%
of cases
Slide95Cell culture
Successful in
only
~50
%
of
cases
Cells must be cultured
in
vitro—need metaphase chromosomes
Only
1% of cells
are dividing at any time
Slide96Kumar: Robbins pathologic basis of disease, 8
th edition. 2009
Slide97Cell culture
Acceptable
cytologic
specimens:
Amniotic fluid
(yield is 84%) obtained by amniocentesis at the time of prenatal diagnosis of demise, 20-30mL
1x1cm placental block
from below the cord insertion site on an unfixed placenta
1.5cm
segement
of
umbilical cord
Internal fetal tissue specimen
(
costochondral
junction, patella, lung, kidney, gonad,
not skin
)
Slide98Cell culture
Collect
the specimen in a
sterile
manner
Prokaryotic cells outcompete and overgrow any human cells
Transport to the laboratory
as soon as possible
Blood, amniotic fluid, chorionic
villi
at
room temperature
Solid tissue transported on
wet ice
Lysosomal
enzyme activity
is inhibited at 4 C and viability is maintained
Slide99Cell culture
Monolayer in situ
U
p
to
2 weeks
for solid tissue culture
Cells are swelled
hypotonically
, ruptured (blowing on the
coverslip
) and metaphase chromosomes are spread
Automatic robotic harvesters
Stained with
Giemsa
or Wright
Results in banding pattern unique to each chromosome
Slide100Karyotype
Count the chromosomes in each cellTechnologist at a microscopeMust analyze 15-20 cells to get karyotypePhotograph of karyogram or computer-assisted imagingManipulate/move chromosomesPattern-recognition software to generate karyogramComputer must be double-checked by a technologist
Stein 2011
Slide101If culture is unsuccessful
Flourescent
in-situ
hybridization (FISH)
Can test for the most common
aneuploidies
Can test for specific mutations
Slide102FISH
Molecular
probe
Fluorescent
dye
V
isualized
using fluorescence microscopy
Molecular probes can be
Specific to certain mutations
A cocktail that coats the whole chromosome
FISH can be performed on metaphase or
interphase
chromosomes—
cell culture not required for FISH
Slide103FISH
DNA is denatured and allowed to hybridize with the probe
Nonhybridized
DNA is then
counterstained
Slide104FISH
X, X
Trisomy
21
Normal chromosome 21
Stein 2011
Slide105FISH
Microdeletion
in chromosome 22 associated with DiGeorge syndrome
Stein 2011
Slide106FISH
Probes must be known genes/mutationsLimited number of colors can be seen with fluorescent microscope
Does not require metaphase chromosomesCan obtain karyotype and information about mutations
Advantages
Disadvantages
Slide107FISH with computer-assisted system
Trisomy
18 and XXY
Fluorochromes can be mixedToo subtle for human eye, but computer assisted systems allow detection of all 24 chromosomesKaryotype in interphase
Stein 2011
Slide108If live cells are not available: Microarray
Screens the genome for
copy number variations (CNPs)
BAC arrays provide
overview of genome
SNP arrays provide
more detailed coverage
with probes on every 100-1000 base pairs
Detects
deletions, duplications,
aneuploidies
, unbalanced translocations
with a gain/loss of sequences
Good for
small deletions
or cryptic changes
Cytogenetics
resolution is only 5-10Mb
Slide109Microarray
Stein 2011
Slide110Limitations
Many copy number changes are of
uncertain clinical significance
More CNVs are being identified every day
It will not detect
balanced translocations or
inversions
Slide111Microarray versus karyotyping
Reddy UM et al,
2012:
Prospective population-based study of
532 stillbirths
over
2 years
Patients with IUFD
underwent:
Interview
, chart abstraction, postpartum examination, placental pathology,
karyotype
analysis, and specimen collection
DNA analyzed with an
SNP microarray
with data aligned to Human Genome release 18
Slide112Microarray versus karyotyping
Benign CNV
: full length listed in any of three databases of unaffected persons
Pathogenic CNV
:
E
vidence
of
pathogenicity
in literature
A
gene listed in the OMIM database that is known to cause developmental/lethal
disease
In
a pathogenic database
Probably benign CNV
: no evidence of
pathogenicity
in literature
Unknown CNV
Slide113Microarray versus karyotyping
Microarray analysis yielded a result in
87.4%
stillbirths compared to
70.5%
for
karyotype
85.2% of these were
benign, too small, or probably benign
2.6% were pathogenic, 6.9% were
aneuploid
Microarray detected CNV consistent
with
DiGeorge
syndrome
not detected by
karyotype
in 3 cases
Slide114Conclusions: microarray
Can detect mutations too small for FISHHigher chance of obtaining a result
More expensive
Advantages
Disadvantages
Genetic workup should be guided by family history and presence of congenital
anomalies
Consider microarray when
karyotype
fails to grow in culture
Slide115Maternal workup
Workup of stillbirth
Slide116Immediate maternal workup
Workup of
stillbirth
Slide117Laboratories (Recommended)
CBC
Kliehauer-Betke
Human
parvovirus B-19
IgG
and
IgM
Lupus anticoagulant,
anticardiolipin
antibodies
Indirect
Coombs
If not already done
antepartum
Toxicology
screen
Slide118Kliehauer Betke
Recommended
to do
before induction
of labor
However, given that only massive hemorrhage is likely to cause fetal death,
can also be done up
to
2-3 weeks after delivery
In
Dutch study, FMH was a
contributing factor in 10.6%
of the total cohort
Slide119Kliehauer Betke
Reddy et al 2009
Slide120Antiphospholipid antibodies
One fetal death satisfies criteria for testingConfirm with repeat testing in 6-12 weeksMore likely positive if stillbirth was accompanied by IUGR or severe preeclampsiaTwo dutch studies (750 fetal deaths in Korteweg et al 2010, 1025 fetal deaths in Korteweg et al 2012) showed that neither testing for acquired nor inherited thrombophilia is valuableUnless the patient has a family or personal history of thrombophilia
Given significant implications for future pregnancies and future health of the mother, testing is generally recommended
Slide121Laboratories (Sometimes useful)
SyphilisTSHInherited thrombophilia workupFactor V Leiden, prothrombin gene mutation, antithrombin III, fasting homocysteineGlucose screeningSonohysterogramEspecially if loss associated with preterm labor, PPROM, cervical insufficiency, previable gestations, fetal malpresentation
Guided by maternal history and risk factors
Slide122Inherited thrombophilia
Korteweg et al 2010. Multicenter, prospective study. 750 singleton fetal deaths >=20 wks, excluding terminationsTested for vWF, antithrombin, protein C, total and free protein S, prothrombin gene mutation, factor V LeidenCause of death classified by a panel“Except for vWF and paternal free protein S, acquired and thrombophilic defects were not more prevalent after fetal death.”However, many case-control studies show an association
Consider it
in cases of
severe placental pathology, IUGR, or a history of thrombosis
However, remember that most women with these conditions will have uncomplicated pregnancies
Slide123Laboratories (unproven benefit)
Toxoplasmosis, rubella, CMV, HSV, other infections
Viruses for which vaccines are prevalent are uncommon in developed countries
However, if
autopsy, pathology, or history is suggestive
, take maternal/neonatal serology, special tissue
staines
, testing for nucleic acids
ANA
Slide124Routine TORCH
Dutch study: of the women tested for TORCH, 17.0% had positive
IgM
for TORCH,
parvo
, syphilis, or hepatitis
B
However only
1.8% of the total cohort
had infection as the cause of death
Slide125Postpartum maternal workup
Suggested workup of stillbirth
Slide126In selected cases
Paternal karyotypeProtein S and C activityKliehauer-Betke
Slide127Considerations
Evaluation of stillbirth
Slide128AIMMC protocol
All
intact fetal remains are sent to the morgue within 4 hours of expiration,
UNLESS
a Surgical Pathology examination is
requested
If
a Surgical Pathology examination is
requested
Remains with a Surgical
Pathology requisition to the Histology
Laboratory
Central
Transport
and
Nursing
are
responsible for the initial transport of the body
to
the
morgue
and filling out the morgue census sheet
Slide129AIMMC protocol
If autopsy is requested, the fetus is obtained from the morgueAn autopsy is an option for fetuses >=20 weeksBut if clinically indicated or requested by the family, an autopsy can be offered before 20 weeks
Slide130AIMMC protocol
Autopsies are performed by AIMMC pathologists
Trained in fetal/neonatal autopsies
Consult specialists when needed
Preliminary result in 24 hours; final result in 30 days
Genetic and molecular testing at ACL
A fetus that undergoes an autopsy
can
still receive private burial
Slide131Fetal death disposition notification
Within 24 hours after an abortion/miscarriage the mother must be informed of her right to determine the final disposition of the remainsPrivate burialCremationHospital disposition
Slide132Fetal death disposition notification
http://www.idph.state.il.us/vitalrecords/pdf/Fetal%20Death%20Disposition%20Notification.pdf
Slide133Fetal death disposition notification
If patient is unclear,
sign the form
without making a
selection
RN notes “
undecided at
discharge”
Fax a copy to
Mission and Spiritual Care
at 773-296-5010 and place a copy in the chart
Mission
and Spiritual Care or the
Perinatal
Loss Coordinator will follow up with the family
If the family can not be reached
within
2
weeks
, Pathology
will initiate the procedure for hospital disposition
Slide134Private burial
Perinatal
loss coordinator has a list of funeral homes
Cremation or burial
Only a
licensed funeral director
can pick up the fetus
Parents provide the name to Health Information Management
Cost: some funeral home directors
reduce the rate
Medicaid pays for modest funeral and burial
For
very young fetuses
, parents advised that there might not be many remains after autopsy
Slide135Hospital disposition
St Luke’s Cemetery5300 N. Pulaski Road, Chicago, IL 60630“Babyland,” exact location not identifiedNo charge
Slide136Perinatal loss coordinator
Always notifiy Anna Zieba (pager 61-7656, phone 61-7127)Always notify Mission and Spiritual Care 61-5005Can provide religious services from most religions
Slide137Considerations
Parents benefit from
seeing/holding
the infant
Warn them about how the baby will appear
Use the term “
baby
”
Encourage parents to
name
the infant
Knowing the
sex
is important
Fetal loss can be devastation at
any gestational age
Different
cultures
grieve in different ways
Slide138Considerations
Keepsakes and memory boxes
Child
Life specialist
can aid with
siblings
For a loss in the ER, consider consulting
crisis
if parents show signs of depression
Bereavement care track
after discharge
Slide139Conclusions
Evaluation of stillbirth
Slide140Conclusions
The cause of a stillbirth is the
initial
pathophysiologic
entity
that irreversibly led to fetal death
Cause must be proven
with evidence of fetal harm
There are
many benefits
to finding a cause
Encourage
patients
to allow an evaluation within the boundaries of their personal and cultural values
Slide141Conclusions
Recommended laboratories are
CBC
,
Kliehauer-Betke
,
parvovirus
B-19
IgG
and
IgM
,
lupus
anticoagulant
,
anticardiolipin
antibodies
, and
toxicology screen
Only perform other labs as indicated by
maternal history
Encourage patients to receive an
autopsy
Partial autopsy
and
MRI
are alternatives
Always send the
placenta
to pathology
Slide142Conclusions
Encourage
karyotyping
Base evaluation for other chromosomal abnormalities on
family history
and results of
other studies
Consider
FISH or microarray
if low likelihood of getting a result from cell culture
Utilize
hospital resources
for
perinatal
loss
Slide143Thank you
Slide144Sources
Evaluation of stillbirth
Slide145ACOG Practice Bulletin Number 102. “Management of Stillbirth.” American College of Obstetricians and Gynecologists. March 2009. acog.org
Fligner
CL,
Dighe
M. “Fetal and
Perinatal
Death Investigation: Redefining the Autopsy and the Role of Radiologic Imaging.” Ultrasound
Clin
6, 2011 (105-117
).
Illinois Masonic Medical Center
Perinatal
Loss Policy. Policy 20.118.044
Korteweg
FJ,
Erwich
JJHM,
Timmer
A, et al. “Evaluation of 1025 fetal deaths: proposed diagnostic workup.” Am J
Obstet
Gynecol
2012; 206:53.e1-12
.
Korteweg
FJ,
Erwich
JJHM, et al. “Prevalence of Parental
Thrombophilic
Defects after Fetal Death and Relation to Cause.”
Obstet
and Gynecol. August 2010,
Vol
116, No 2, part 1.
Kumar: Robbins and
Coltran
Pathologic Basis of Disease, Professional Edition, 8
th
ed
. 2009 Saunders, an imprint of Elsevier.
Reddy UM,
Goldenburg
R, et al. “Stillbirth Classification—Developing an International Consensus for Research.”
Obstet
and Gynecol
. Oct 2009,
Vol
114, No 4.
Reddy UM, Page GP,
Saade
GR. “
Karyotype
versus Microarray testing for Genetic Abnormalities after Stillbirth.” N
Engl
J med 367; 23, December 2012
Silver R,
Heuser
C. “Stillbirth Workup and Delivery Management.”
Clin
Obstet
and
Gynecol
2010; 53, 3.
Stein, CK. “Applications of
cytogenetics
in Modern Pathology.”
McPherson: Henry’s Clinical Diagnosis and Management by Laboratory Methods, 22
nd
ed
. 2011 Saunders, an imprint of Elsevier.
Evaluation of Stillbirth
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