Post on 14-Apr-2017
Infertility, In Vitro Infertility, In Vitro Fertilization (IVF) Fertilization (IVF)
and Genetic and Genetic TestingTesting
Michele Evans, M.D.Michele Evans, M.D.
OutlineOutline InfertilityInfertility Treatment optionsTreatment options
In Vitro FertilizationIn Vitro Fertilization Genetic TestingGenetic Testing
PreconceptionPreconception PreimplantationPreimplantation Prenatal Prenatal
ControversiesControversies
Normal FertilityNormal Fertility
0
0.2
0.4
0.6
0.8
1
1 2 3 4 5 6 7 8 9 10 11 12
Months of Follow Up
Cumulative Pregnancy
Rates
Hull, et al: Br Med J 1985:291;1693
Overview of InfertilityOverview of Infertility
Definition: 1 year of well-timed, Definition: 1 year of well-timed, unprotected intercourse without a unprotected intercourse without a pregnancypregnancy
10-15% of population is infertile 10-15% of population is infertile (subfertile)(subfertile)
Causes of InfertilityCauses of Infertility
Female Reproductive Female Reproductive OrgansOrgans
FSH (+)
FSH=Follicle Stimulating Hormone
PhysiologyPhysiology
E2 (-)
E2=Estradiol
Developing follicle
Causes of Female Causes of Female InfertilityInfertility
OvaryOvary TubesTubes UterusUterus CervixCervix HormonesHormones ChromosomesChromosomes
Causes of Female Infertility Causes of Female Infertility - Ovary- Ovary
AGEAGE
Problems with ovulation Problems with ovulation
Premature ovarian failurePremature ovarian failure
Ovary - Female AgeOvary - Female Age Women are born with their lifetime Women are born with their lifetime
egg supplyegg supply 4 million at 20 weeks gestation4 million at 20 weeks gestation 400,000 at birth400,000 at birth
100,000 eggs left at time of puberty100,000 eggs left at time of puberty Fertility initially declines at age Fertility initially declines at age 2727 Significant decline at age 37-38Significant decline at age 37-38 Rare pregnancies after age 44Rare pregnancies after age 44
Percentage of Married Percentage of Married Women Who are InfertileWomen Who are InfertileFrom 3 national U.S. surveysFrom 3 national U.S. surveys
Age (years)Age (years) InfertileInfertile20-2420-24 7.07.025-2925-29 8.98.930-3430-34 14.614.635-3935-39 21.921.940-4440-44 28.728.7
Menken et al, Science 1989;23:1389
Ovarian Reserve Ovarian Reserve TestingTesting
Day 3 Day 3 FSH/EFSH/E22
FSH FSH (mIU/mL)(mIU/mL)
EE22 (pg/mL) (pg/mL)
NormalNormal <10<10 <100<100BorderlineBorderline 10-2010-20 NANAAbnormalAbnormal >20>20 >100>100
Clinical pregnancy per Clinical pregnancy per cyclecycle
as function of both FSH as function of both FSH and ageand age
Pearlstone et al, Fertil Steril 1992:58:674
Prevalence of genetically Prevalence of genetically abnormal oocytes in abnormal oocytes in
infertile womeninfertile womenAb
norm
al
(%)
Ovary - OvulationOvary - Ovulation
Ovary - OvulationOvary - Ovulation HistoryHistory
Regular menses (90%)Regular menses (90%) Premenstrual symptomsPremenstrual symptoms
Patient self-testingPatient self-testing BBT, urinary ovulation kits (LH)BBT, urinary ovulation kits (LH)
Laboratory testsLaboratory tests Mid-luteal serum progesterone > 10 Mid-luteal serum progesterone > 10
ng/mLng/mL Endometrial biopsyEndometrial biopsy
Ovary - Causes of Ovary - Causes of AnovulationAnovulation
Hormone imbalanceHormone imbalance ObesityObesity AnorexiaAnorexia Significant stressSignificant stress Patients display:Patients display:
Irregular menstrual cyclesIrregular menstrual cycles Skipped cyclesSkipped cycles Minimal or absent premenstrual symptomsMinimal or absent premenstrual symptoms
Ovary – Premature Ovarian Ovary – Premature Ovarian FailureFailure
Menopause prior to age 40Menopause prior to age 40 ↓↓EstrogenEstrogen ↑↑FSHFSH
CausesCauses AutoimmuneAutoimmune GeneticGenetic Idiopathic Idiopathic
1-2% pregnancy rate1-2% pregnancy rate
Causes of Female Infertility Causes of Female Infertility – Fallopian Tubes– Fallopian Tubes
Infection (chlamydia)Infection (chlamydia)
EndometriosisEndometriosis
Tubal ligation (female sterilization)Tubal ligation (female sterilization)
Open TubesOpen Tubes
Blocked TubesBlocked Tubes
Female Infertility - Female Infertility - UterusUterus
UterusUterus FibroidsFibroids PolypsPolyps
Mullerian (congenital) defectsMullerian (congenital) defects AbsentAbsent Bicornuate/SeptumBicornuate/Septum
Asherman’s syndromeAsherman’s syndrome
Female Reproductive Female Reproductive OrgansOrgans
Female InfertilityFemale Infertility
Uterine muscle Uterine muscle tumortumor
Benign (>95%)Benign (>95%)
25-30% of women25-30% of women
Normal Shape of UterusNormal Shape of Uterus
Fibroid UterusFibroid Uterus
Female Infertility - Female Infertility - UterusUterus
UterusUterus TumorsTumors
FibroidsFibroids PolypsPolyps
Mullerian defects (congenital)Mullerian defects (congenital) Absent uterusAbsent uterus Bicornuate/septateBicornuate/septate
Asherman’s syndromeAsherman’s syndrome
Mullerian DefectMullerian Defect
Treatment with Treatment with HysteroscopyHysteroscopy
Treatment with Treatment with LaparoscopyLaparoscopy
Female Infertility - Female Infertility - UterusUterus
UterusUterus TumorsTumors
FibroidsFibroids PolypsPolyps
Mullerian defects (congenital)Mullerian defects (congenital) Absent uterusAbsent uterus Bicornuate/septateBicornuate/septate
Asherman’s syndromeAsherman’s syndrome
Female Infertility - Female Infertility - CervixCervix
CervixCervix Post-surgicalPost-surgical
StenosisStenosis Mucus changesMucus changes
Female Infertility - Female Infertility - HormonesHormones
Endocrine abnormality (hormones)Endocrine abnormality (hormones) ThyroidThyroid ProlactinProlactin Polycystic ovary syndrome (PCOS)Polycystic ovary syndrome (PCOS)
Estrogen, insulinEstrogen, insulin Hypothalamic hypogonadismHypothalamic hypogonadism
StressStress Exercise (ballet dancer)Exercise (ballet dancer)
Other Causes of Female Other Causes of Female InfertilityInfertility
OthersOthers Chromosome abnormalitiesChromosome abnormalities
Turner’s syndrome (XO)Turner’s syndrome (XO) Androgen Insensitivity (XY)Androgen Insensitivity (XY)
Male pseudohermaphroditeMale pseudohermaphrodite Female phenotypeFemale phenotype Blind vaginal canalBlind vaginal canal Inguinal hernia (50%)Inguinal hernia (50%)
Causes of Male InfertilityCauses of Male Infertility
Abnormality in sperm productionAbnormality in sperm production
Abnormality in sperm functionAbnormality in sperm function
Obstruction in the ductal systemObstruction in the ductal system
Male Reproductive Male Reproductive OrgansOrgans
Sperm: Semen AnalysisSperm: Semen Analysis
Volume: Volume: >> 2 mL 2 mL Concentration: Concentration: >> 20,000,000 per mL 20,000,000 per mL Motility: Motility: >> 50% 50% Normal morphology: Normal morphology: >> 40% normal 40% normal
Krueger strict criteria: Krueger strict criteria: >> 14% normal 14% normal Best predictor of fertilizing ability Best predictor of fertilizing ability
Normal Sperm Normal Sperm MorphologyMorphology
Abnormal MorphologyAbnormal Morphology
Abnormal MorphologyAbnormal Morphology
SpermSperm How many are needed for fertilization?How many are needed for fertilization? Natural conceptionNatural conception
20,000,00020,000,000 Intra-uterine inseminationIntra-uterine insemination
1,000,0001,000,000 In-vitro fertilization (IVF)In-vitro fertilization (IVF)
10,00010,000 Intra-cytoplasmic sperm injection (ICSI)Intra-cytoplasmic sperm injection (ICSI)
11
FertilizationFertilization
Binding to zona Binding to zona pellucidapellucida
Penetration of Penetration of oocyteoocyte
Decondensation Decondensation and aster and aster formationformation
Pronucleus Pronucleus formationformation
Causes of Male InfertilityCauses of Male Infertility
Abnormality in sperm productionAbnormality in sperm production
Abnormality in sperm functionAbnormality in sperm function
Obstruction in the ductal systemObstruction in the ductal system
Abnormalities of Sperm Abnormalities of Sperm ProductionProduction
GeneticGenetic Y chromosome microdeletionsY chromosome microdeletions
Damage to testes – anatomicalDamage to testes – anatomical CryptorchidismCryptorchidism VaricoceleVaricocele
InfectionInfection Mumps orchitisMumps orchitis
GonadotoxinsGonadotoxins
Abnormalities of Sperm Abnormalities of Sperm FunctionFunction
Antisperm antibodiesAntisperm antibodies Genital tract inflammationGenital tract inflammation
prostatitisprostatitis VaricoceleVaricocele Failure of acrosome reactionFailure of acrosome reaction Problems with sperm Problems with sperm
binding/penetrationbinding/penetration
Obstructions in Ductal Obstructions in Ductal SystemSystem
VasectomyVasectomy
Congenital bilateral absence of the Congenital bilateral absence of the vas deferensvas deferens
Epididymis/ejaculatory ductsEpididymis/ejaculatory ducts Congenital or acquiredCongenital or acquired
Male Infertility - Male Infertility - LifestyleLifestyle
TobaccoTobacco MarijuanaMarijuana AlcoholAlcohol CocaineCocaine Steroids (can be permanent)Steroids (can be permanent) HeatHeat ExerciseExercise
Infertility: Initial Infertility: Initial EvaluationEvaluation
EggsEggs OvulationOvulation Egg qualityEgg quality
SpermSperm PresencePresence QualityQuality
Gamete transport/ImplantationGamete transport/Implantation HysterosalpingogramHysterosalpingogram
Unexplained InfertilityUnexplained Infertility
Work-up is negativeWork-up is negative
15-20 % of couples15-20 % of couples
TreatmentsTreatments Observation (improve timing)Observation (improve timing) Clomiphene citrate + intrauterine Clomiphene citrate + intrauterine
insemination (IUI)insemination (IUI) FSH + IUIFSH + IUI IVFIVF Egg donationEgg donation
Intrauterine Intrauterine Insemination (IUI)Insemination (IUI)
Clomiphene Citrate Clomiphene Citrate (Clomid)(Clomid)
Anti-estrogenAnti-estrogen
↑ ↑ FSHFSH
↑ ↑ egg productionegg production
$$
Estrogen AntagonistEstrogen Antagonist
FSH (+)
FSH=Follicle Stimulating Hormone
PhysiologyPhysiology
E2 (-)
E2=Estradiol
Developing follicle
Clomid
ART: Treatment ART: Treatment OptionsOptions
Clomiphene citrate + IUIClomiphene citrate + IUI Prospective randomized trialProspective randomized trial Patients with unexplained infertilityPatients with unexplained infertility
ObservatioObservationn
CC + IUICC + IUI
CyclesCycles 150150 148148PregnanciePregnanciess
55 1414
FecundityFecundity 0.0330.033 0.0950.095Deaton et al, Fertil Steril 1990;54:1083
FSH +IUIFSH +IUI Daily injections Daily injections
Goal is to make multiple follicles Goal is to make multiple follicles (eggs)(eggs)
$$$$
FSH (+)
FSH=Follicle Stimulating Hormone
PhysiologyPhysiology
E2 (-)
E2=Estradiol
Developing follicle
Infertility Treatment Infertility Treatment OptionsOptions
IUI, FSH or FSH + IUIIUI, FSH or FSH + IUI Patients with unexplained infertilityPatients with unexplained infertility
Serhall et al, Fertil Steril 1988;49:602
TreatmeTreatmentnt
CyclesCycles PregnancPregnancyy
Pregnancy Pregnancy per cycleper cycle
IUIIUI 3030 11 2.7%2.7%
FSHFSH 4949 33 6.1%6.1%
FSH+IUIFSH+IUI 3434 99 26.4%26.4%
In Vitro Fertilization - In Vitro Fertilization - HistoryHistory
1978 – First “test tube” baby was born in 1978 – First “test tube” baby was born in England England
1981 – IVF in U.S.1981 – IVF in U.S.
Started with GIFT and ZIFTStarted with GIFT and ZIFT
2004 - >98% IVF with transfer of embryo 2004 - >98% IVF with transfer of embryo to uterusto uterus
In Vitro Fertilization In Vitro Fertilization (IVF)(IVF)
Who Needs IVF?Who Needs IVF? Failed other treatmentsFailed other treatments Tubal damageTubal damage Significant male factorSignificant male factor Absent uterusAbsent uterus Carriers of genetic diseasesCarriers of genetic diseases Cancer patients?Cancer patients?
Ovarian Ovarian HyperstimulationHyperstimulation
Egg RetrievalEgg Retrieval
Good EggGood Egg
Bad EggBad Egg
FertilizationFertilization
2 Pronuclei (2PN)2 Pronuclei (2PN)
1 day after egg 1 day after egg retrievalretrieval
Day 3 EmbryoDay 3 Embryo
Day 3 EmbryoDay 3 Embryo
Day 3 EmbryoDay 3 Embryo
Day 3 EmbryoDay 3 Embryo
Blastocyst – Day 5Blastocyst – Day 5
Hatching BlastocystHatching Blastocyst
Embryo TransferEmbryo Transfer
Embryo TransferEmbryo Transfer
Special IVF ProceduresSpecial IVF Procedures Assisted hatchingAssisted hatching Intracytoplasmic sperm injection Intracytoplasmic sperm injection
(ICSI)(ICSI) Preimplantation genetic diagnosis Preimplantation genetic diagnosis
(PGD)(PGD) FreezingFreezing Egg donationEgg donation SurrogacySurrogacy
Assisted HatchingAssisted Hatching
Empty ZonaEmpty Zona
ICSIICSI
ICSIICSI
ICSIICSI
How Many Embryos are How Many Embryos are Transferred?Transferred?
Related to age and embryo qualityRelated to age and embryo quality <35 = 2<35 = 2 35-37 = 2-335-37 = 2-3 38-40 = 3-438-40 = 3-4 >40 = up to 5>40 = up to 5
For patients with 2 or more failed IVF cycles, For patients with 2 or more failed IVF cycles, or a poor prognosis, can add more based on or a poor prognosis, can add more based on clinical judgementclinical judgement
IVF Success Rates - 2000IVF Success Rates - 2000
U.S. Fertility Centers (SART/CDC)U.S. Fertility Centers (SART/CDC) Female ageFemale age
<35 – 33%<35 – 33% 35-37 – 27%35-37 – 27% 38-40 – 18.6%38-40 – 18.6% >40 – 7.8%>40 – 7.8%
IVF Statistics - 2000IVF Statistics - 2000
65.1% singletons (16,533)65.1% singletons (16,533)
30.8% twins (7,817)30.8% twins (7,817)
3.9% triplets (1,000)3.9% triplets (1,000)
0.2% higher order multiples (44)0.2% higher order multiples (44)
Twin PregnancyTwin Pregnancy
IVF and Multiple IVF and Multiple PregnancyPregnancy
Maternal complicationsMaternal complications Fetal complicationsFetal complications CostCost ““Selective reduction”Selective reduction” Single embryo transfer vs. success Single embryo transfer vs. success
ratesrates
IVF Statistics - 2000IVF Statistics - 2000 383 U.S. programs offer IVF383 U.S. programs offer IVF
99,989 cycles of ART treatment99,989 cycles of ART treatment
7,581 donor oocyte cycles7,581 donor oocyte cycles
25,394 deliveries (birth of 35,345 25,394 deliveries (birth of 35,345 neonates)neonates)
Current status of ART in Current status of ART in the USAthe USA
SART registry, 1998
Pregnancy and live birth ratesfor ART cyclesusing fresh,non-donor eggs,by age of woman
Cost of IVFCost of IVF IVF cycle + medications = $10,000-IVF cycle + medications = $10,000-
15,00015,000 Assisted hatching = $500Assisted hatching = $500 ICSI = $1,500ICSI = $1,500 Freezing = $650Freezing = $650 Storage = $360 Storage = $360 Egg Donor = $5,000Egg Donor = $5,000 Surrogate = $10,000-15,000Surrogate = $10,000-15,000
Egg DonationEgg Donation
Egg Egg donationdonation
IVF for twoIVF for two
DonorDonor Standard controlled Standard controlled
ovarian ovarian hyperstimulationhyperstimulation
Egg retrievalEgg retrieval
RecipientRecipient Embryo transferEmbryo transfer
Egg DonationEgg Donation
Known/anonymous donorKnown/anonymous donor
<35 years old<35 years old
Current status of ART Current status of ART in the USAin the USA
SART registry, 1998
Live birth rates per transferfor fresh embryos from own anddonor eggs, by age of recipient
Cumulative pregnancy ratesafter oocyte donation
Human Reprod 1997;12:835
Egg DonationEgg Donation Grade A: The Market for a Yale Grade A: The Market for a Yale
Woman’s EggsWoman’s Eggs When a Yale undergraduate explored When a Yale undergraduate explored becoming an egg donor for a wealthy becoming an egg donor for a wealthy couple willing to pay top dollar to the couple willing to pay top dollar to the right candidate, she didn't realize how right candidate, she didn't realize how unsettling the process of candidacy would unsettling the process of candidacy would prove to beprove to be
by Jessica Cohen by Jessica Cohen
Egg DonationEgg Donation October 23, 1999October 23, 1999 Selling Fashion Models' Eggs Online Raises Ethics Selling Fashion Models' Eggs Online Raises Ethics
IssuesIssues By CAREY GOLDBERG By CAREY GOLDBERG
CAMBRIDGE, Mass. -- To the horror and disgust of CAMBRIDGE, Mass. -- To the horror and disgust of mainstream infertility groups, a longtime fashion mainstream infertility groups, a longtime fashion photographer has begun offering up models as egg photographer has begun offering up models as egg donors to the highest bidders, auctioning their ova donors to the highest bidders, auctioning their ova via the Internet to would-be parents willing to pay via the Internet to would-be parents willing to pay up to $150,000 in hopes of having a beautiful child. up to $150,000 in hopes of having a beautiful child.
Egg Donation - Ethical Egg Donation - Ethical IssuesIssues
Egg Donor Egg Donor Known or anonymousKnown or anonymous How many times to donate?How many times to donate?
RecipientRecipient How old is too old?How old is too old?
Pregnancy in the Sixth Pregnancy in the Sixth Decade of LifeDecade of Life
USC experience: 1991-2000USC experience: 1991-2000
77 recipients of egg donation77 recipients of egg donation
Mean age 52.8 + 2.9 yearsMean age 52.8 + 2.9 years
Paulson, Tourgeman, Boostanfar et al, JAMA 2002:228;2320.
Pregnancy in the Sixth Pregnancy in the Sixth Decade of LifeDecade of Life
Medical screeningMedical screening EKG and treadmillEKG and treadmill Mammogram, PapMammogram, Pap Chest x-rayChest x-ray Glucose tolerance testGlucose tolerance test Complete blood countComplete blood count Blood chemistry with cholesterolBlood chemistry with cholesterol
Pregnancy in the Sixth Pregnancy in the Sixth Decade of Life: Pregnancy Decade of Life: Pregnancy
OutcomeOutcome Of the 77 women, 42 (54.5%) had Of the 77 women, 42 (54.5%) had
live birthslive births
45 deliveries in 42 women45 deliveries in 42 women
Pregnancy in the Sixth Pregnancy in the Sixth Decade of Life: Pregnancy Decade of Life: Pregnancy
OutcomeOutcome 45 Live births45 Live births
78% Cesarean delivery78% Cesarean delivery 31 Singletons31 Singletons
68% Cesarean delivery68% Cesarean delivery 6% Vacuum assisted6% Vacuum assisted 26% NSVD26% NSVD
14 Multiple gestations14 Multiple gestations 100% Cesarean delivery100% Cesarean delivery
Pregnancy in the Sixth Pregnancy in the Sixth Decade of Life: Obstetric Decade of Life: Obstetric
ComplicationsComplications 35% Pre-eclampsia35% Pre-eclampsia
25% mild25% mild 10% severe10% severe
Background incidenceBackground incidence 3-5% in young women3-5% in young women 10% in women over age 4010% in women over age 40
Pregnancy in the Sixth Pregnancy in the Sixth Decade of Life: Decade of Life:
Obstetric ComplicationsObstetric Complications 20% gestational diabetes20% gestational diabetes
17.5% diet controlled17.5% diet controlled 2.5% insulin2.5% insulin
Background incidenceBackground incidence 5% overall, increasing with age5% overall, increasing with age
<20 years of age: 3.7%<20 years of age: 3.7% 20-30 years of age: 7.5%20-30 years of age: 7.5% >30 years of age: 13.8%>30 years of age: 13.8%
Is 55 a “physiological limit”?Is 55 a “physiological limit”?
Marked increase in pre-eclampsiaMarked increase in pre-eclampsia
Increase in diabetesIncrease in diabetes
How old is too old?How old is too old?
Genetic TestingGenetic Testing
PreconceptionPreconception
PreimplantationPreimplantation
PrenatalPrenatal
PostnatalPostnatal
Preconception Preconception CounselingCounseling
Offered to all womenOffered to all women Prenatal vitamins – 400 micrograms folic Prenatal vitamins – 400 micrograms folic
acid/dayacid/day Rubella immunityRubella immunity Varicella immunityVaricella immunity Rh statusRh status HIVHIV Hepatitis B screenHepatitis B screen Cystic Fibrosis screeningCystic Fibrosis screening
Preconception Preconception CounselingCounseling
Offered to certain ethnic groupsOffered to certain ethnic groups Mediterranean – thalassemiaMediterranean – thalassemia African-American – sickle-cell anemiaAfrican-American – sickle-cell anemia Caucasian/Hispanic – cystic fibrosisCaucasian/Hispanic – cystic fibrosis Ashkenazi Jews – 7 autosomal recessive Ashkenazi Jews – 7 autosomal recessive
disordersdisorders Gaucher disease (1/13), Tay-Sachs (1/30), Gaucher disease (1/13), Tay-Sachs (1/30),
Familial dysautonomia (1/30), Canavan disease Familial dysautonomia (1/30), Canavan disease (1/40), Fanconi anemia (1/89), Niemann-Pick (1/40), Fanconi anemia (1/89), Niemann-Pick disease (1/90), Bloom syndrome (1/100)disease (1/90), Bloom syndrome (1/100)
Prenatal TestingPrenatal Testing UltrasoundsUltrasounds
Serum screensSerum screens
Chorionic villus sampling (CVS)Chorionic villus sampling (CVS)
AmniocentesisAmniocentesis
Prenatal Tests - Prenatal Tests - UltrasoundUltrasound
Nuchal translucency screeningNuchal translucency screening
Performed between 10-13 weeks Performed between 10-13 weeks gestationgestation
Screen for Down SyndromeScreen for Down Syndrome
Nuchal TranslucencyNuchal Translucency
Prenatal Tests - Prenatal Tests - UltrasoundUltrasound
Second trimester ultrasoundSecond trimester ultrasound Detailed examDetailed exam Down Syndrome, other trisomiesDown Syndrome, other trisomies Cardiac, renal, spinal, limb, brain Cardiac, renal, spinal, limb, brain
deformitiesdeformities Cleft lip/palateCleft lip/palate
Ultrasound Ultrasound AbnormalitiesAbnormalities
Ultrasound Ultrasound AbnormalitiesAbnormalities
Prenatal Tests - SerumPrenatal Tests - Serum 15-20 weeks gestation15-20 weeks gestation
Quad ScreenQuad Screen Tests for AFP, hCG, uE3 and inhibin ATests for AFP, hCG, uE3 and inhibin A Neural tube defects, Down syndrome Neural tube defects, Down syndrome
and trisomy 18and trisomy 18 Readjusts age-related risksReadjusts age-related risks
Prenatal TestsPrenatal Tests Who is offered further testing?Who is offered further testing?
Advanced maternal ageAdvanced maternal age Previous child or pregnancy with birth Previous child or pregnancy with birth
defectdefect Suggestive screening test resultsSuggestive screening test results Family historyFamily history
Prenatal Tests – CVSPrenatal Tests – CVS Chorionic Villus SamplingChorionic Villus Sampling
11-13 weeks gestation11-13 weeks gestation Catheter/needle biopsy of placental Catheter/needle biopsy of placental
cellscells Performed through cervix or abdomenPerformed through cervix or abdomen Can tests for aneuploidy and enzyme Can tests for aneuploidy and enzyme
defectsdefects 1-2% miscarriage rate1-2% miscarriage rate Digit/limb deficiencies (10 weeks)Digit/limb deficiencies (10 weeks)
Prenatal Tests - Prenatal Tests - AmniocentesisAmniocentesis
Performed at 15-18 weeks gestationPerformed at 15-18 weeks gestation 10 cc amniotic fluid10 cc amniotic fluid Living cells from fetus in amniotic Living cells from fetus in amniotic
fluidfluid Cells grown in lab for 1-2 weeksCells grown in lab for 1-2 weeks Results in 3 weeksResults in 3 weeks
Postnatal TestingPostnatal Testing Most done during first day of lifeMost done during first day of life Heel stickHeel stick CaliforniaCalifornia
GalactosemiaGalactosemia Hypothyroidism (congenital) Hypothyroidism (congenital) Phenylketonuria (PKU) Phenylketonuria (PKU) Sickle Cell Disease (SCD) and Sickle Cell Disease (SCD) and
Hemoglobinopathies Hemoglobinopathies
Postnatal Screening – Postnatal Screening – Tandem Mass SpectrometyTandem Mass Spectromety
Fatty Acid Oxidation Disorders Carnitine/Acylcarnitine Translocase Deficiency (Translocase)
Carnitine Palmitoyl Transferase Deficiency Type I (CPT-I)2
3-Hydroxy Long Chain Acyl-CoA Dehydrogenase Deficiency (LCHAD)2,4-Dienoyl-CoA Reductase Deficiency2
Medium Chain Acyl-CoA Dehydrogenase Deficiency (MCAD)Multiple Acyl-CoA Dehydrogenase Deficiency (MADD or Glutaric Acidemia-Type II)Neonatal Carnitine Palmitoyl Transferase Deficiency-Type II(CPT-II)Short Chain Acyl-CoA Dehydrogenase Deficiency (SCAD)Short Chain Hydroxy Acyl-CoA Dehydrogenase Deficiency (SCHAD)Trifunctional Protein Deficiency (TFP Deficiency)Very Long Chain Acyl-CoA Dehydrogenase Deficiency (VLCAD)
Organic Acid Disorders 3-Hydroxy-3-Methylglutaryl-CoA Lyase Deficiency (HMG)
Glutaric Acidemia-Type I (GA I)Isobutyryl-CoA Dehydrogenase DeficiencyIsovaleric Acidemia (IVA) Acute onset Chronic2-Methylbutryl-CoA Dehydrogenase Deficiency3-Methylcrotonyl-CoA Carboxylase Deficiency (3MCC Deficiency)3-Methylglutaconyl-CoA Hydratase DeficiencyMethylmalonic Acidemias Methylmalonyl-CoA Mutase Deficiency 0 Methylmalonyl-CoA Mutase Deficiency + Some Adenosylcobalamin Synthesis Defects Maternal Vitamin B12 DeficiencyMitochondrial Acetoacetyl-CoA Thiolase Deficiency (3-Ketothiolase Def.)Propionic Acidemia (PA) Acute onset Late onsetMultiple-CoA Carboxylase DeficiencyMalonic Aciduria
Amino Acid Disorders Argininemia
Argininosuccinic Aciduria (ASA Lyase Deficiency) Acute onset Late onsetCarbamoylphosphate Synthetase Deficiency (CPS Def.)2
Citrullinemia (ASA Synthetase Deficiency) Acute onset Late onsetHomocystinuriaHypermethioninemiaHyperammonemia, Hyperornithinemia, Homocitrullinemia Syndrome (HHH)2
Hyperornithinemia with Gyral Atrophy2
Maple Syrup Urine Disease (MSUD) Classical MSUD Intermediate MSUD5-Oxoprolinuria (pyroglutamic Aciduria)2
Phenylketonuria (PKU) Classical PKU Hyperphenylalaninemia Biopterin Cofactor Deficiencies (4)TyrosinemiaTransient Neonatal Tyrosinemia Tyrosinemia Type I (Tyr I)2
Tyrosinemia Type II (Tyr II) Tyrosimenia Type III (Tyr III)
Other Abnormal Profiles Hyperalimentation
Liver DiseaseMedium Chain Triglyceride (MCT) Oil AdministrationPresence of EDTA Antigoagulants in blood specimenTreatment with Benzoate, Pyvalic Acid, or Valproic AcidCarnitine Uptake Deficiency2
Preimplantation Genetic Preimplantation Genetic Diagnosis (PGD)Diagnosis (PGD)
Can test embryos for genetic Can test embryos for genetic abnormalities prior to implantationabnormalities prior to implantation
Uses single cell (blastomere) at 8-Uses single cell (blastomere) at 8-cell stagecell stage
Which Embryo is Which Embryo is Disease-Free?Disease-Free?
PGD – Clinical PGD – Clinical IndicationsIndications
Single gene defectsSingle gene defects Balanced translocationsBalanced translocations Advanced maternal age (aneuploidy)Advanced maternal age (aneuploidy) Repetitive IVF failureRepetitive IVF failure Recurrent pregnancy lossRecurrent pregnancy loss Embryo selectionEmbryo selection
Fluorescence in situ hybridization Fluorescence in situ hybridization (FISH)(FISH) Aneuploidy/translocations and Aneuploidy/translocations and
sexing (5-9 chromosomes)sexing (5-9 chromosomes) PCRPCR
specific single gene disordersspecific single gene disorders
PGD
•Achondroplasia •ADPKD1 •ADPKD2 •Adrenoleukodystroph •Age-related aneuploidies •Alpha-thalassemia •Alpha-1-antitrypsin •Alport disease •Amyloid precursor protein (APP) mutation •ARPKD •Becker muscular dystrophy •Beta-thalassemia •Charcot Marie Tooth disease •Chromosomal translocations •Congenital adrenal hyperplasia •Cystic fibrosis •Down syndrome •Duchenne muscular dystophy •Dystonia •Epidermolysis bullosa •Familial dysautonomia •Fanconi anemia •FAP •Fragile X syndrome •Gaucher disease •Hemophilia A and B •HLA genotyping •HSNF5 mutation
•Huntington disease •Hypophosphatasia •Incontinentia pigmenti •Kell disease •Klinefelter syndrome •LCHAD •Lesch Nyhan syndrome •Marfan syndrome •Multiple epiphysial dysplasia •Myotonic dystophy •Myotubular myopathy •NF1 and NF2 •Norrie disease •Osteogenesis imperfecta •OTC deficiency •P53 mutations •PKU •Retinitis pigmentosa •SCA6 •Sickle cell anemia •Sonic hedgehog mutations •Spinal muscular atrophy (SMA) •Tay-Sachs disease •Tuberous sclerosis •Turner syndrome •Von Hippel Lindau •X-linked hydrocephaly •X-linked hyper IgM syndrome
PGD for Single Gene PGD for Single Gene Disorders - AdvantagesDisorders - Advantages
Safer than elective terminationSafer than elective termination More psychologically acceptable for More psychologically acceptable for
couplescouples Provides couples with another optionProvides couples with another option
AdoptionAdoption SterilizationSterilization Donor gametesDonor gametes
PGDPGD Pre-implantation Pre-implantation
genetic diagnosis genetic diagnosis (PGD)(PGD) has been has been successfully used in successfully used in diagnosing and diagnosing and preventing inherited preventing inherited genetic diseases like genetic diseases like Cystic Fibrosis, Tay Cystic Fibrosis, Tay Sach’s, Thalassemia, Sach’s, Thalassemia, Sickle Cell Anemia and Sickle Cell Anemia and may be potentially may be potentially used to screen for used to screen for cancer mutations.cancer mutations.
PGDPGD After a cycle of After a cycle of
in- vitro in- vitro fertilization, fertilization, biopsy of a single biopsy of a single cell can be cell can be performed from performed from an 8 cell embryo an 8 cell embryo obtained after 3 obtained after 3 days of culture in days of culture in the laboratory.the laboratory.
PGD – Timing of BiopsyPGD – Timing of Biopsy <67 hours post-<67 hours post-
retrievalretrieval
Implantation rates Implantation rates significantly lower if significantly lower if >70 hours>70 hours
Probably represents Probably represents technical issues with technical issues with compacting embryocompacting embryo
PGDPGD The genetic The genetic
material of this material of this single cell can be single cell can be amplified by PCR amplified by PCR and the and the chromosomal chromosomal mutation or an mutation or an aneuploidy can be aneuploidy can be identified in the identified in the embryo that embryo that underwent a biopsy.underwent a biopsy.
PGDPGD The embryos would The embryos would
continue to grow for continue to grow for 2 more days in the 2 more days in the laboratory, awaiting laboratory, awaiting genetic analysis, genetic analysis, and confirmation of and confirmation of which embryos were which embryos were unaffected with the unaffected with the mutation or mutation or aneuploidy.aneuploidy.
PGDPGD
The unaffected The unaffected embryos are then embryos are then transferred to the transferred to the uterus at the uterus at the blastocyst stage on day blastocyst stage on day 5 of embryo culture 5 of embryo culture and subsequently a and subsequently a child would be born child would be born unaffected from the unaffected from the screened genetic screened genetic disease.disease.
Balanced TranslocationsBalanced Translocations
PGD – Female EmbryoPGD – Female Embryo
Uses fluorescence Uses fluorescence in-situ hybridization in-situ hybridization (FISH) technique to (FISH) technique to identify XXidentify XX
Sex-linked diseasesSex-linked diseases
““Family balancing”Family balancing”
PGD – Male EmbryoPGD – Male Embryo
Uses FISH to Uses FISH to identify XY embryoidentify XY embryo
Prenatal vs. Prenatal vs. Preimplantation DiagnosisPreimplantation Diagnosis
PNDPND PGD PGDCellsCells >100,000 >100,000 11Time Time 2 weeks2 weeks6-10 hrs6-10 hrsAccuracyAccuracy 99%99% 90%90%
Who Would Benefit Who Would Benefit From PGD?From PGD?
Couples with a history of --Couples with a history of -- Abnormal numbers of Abnormal numbers of
chromosomeschromosomes Single gene disordersSingle gene disorders Balanced translocationsBalanced translocations
Couples who --Couples who -- Desire an offspring of a certain sex Desire an offspring of a certain sex
Future considerationsFuture considerations Cytoplasmic transferCytoplasmic transfer
Donation of enucleated oocytesDonation of enucleated oocytes Oocyte cryopreservationOocyte cryopreservation
““Pausing the biological clock”Pausing the biological clock” Reproduction without gametesReproduction without gametes
Use of nuclear material from somatic Use of nuclear material from somatic cellscells
Donated or synthetic cytoplasmDonated or synthetic cytoplasm Reconstituted oocytesReconstituted oocytes
CasesCases Sperm donorSperm donor Female coupleFemale couple Huntington’s diseaseHuntington’s disease Surrogacy – Baby “M”Surrogacy – Baby “M” Single womanSingle woman Sex-linked diseaseSex-linked disease Family balancingFamily balancing ““Wrongful death” of discarded embryoWrongful death” of discarded embryo
Stem Cell Stem Cell Research:Research:
PromiseandControversy
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