Assisted Reproductive Technology Flashcards
describe male reproductive system
testis, sperm duct, penis
describe female reproductive system
ovary, fallopian tubes, uterus, vagina
what needs to be functional to conceive naturally
male and female reproductive systems - all components to conceive naturally
what is ART
Specialized treatments or methods designed to increase number of oocytes and or sperm and improve likelihood of pregnancy = assisted reproductive technologies
is ART just ivf
NAHHH
many kinds of treatments
when and where to use ART - infertility
designed initially for infertiltiy
when and where to use ART - cancer
fertility preservation
adult = get cancer and have to do chemo = toxin so damages likelihood of biological kids, freeze eggs or sperm for future use
children?=complicated because how young is too young
when and where to use ART - social aspects
preserving fertility to circumvent maternal age effect = freeze eggs while young and then have children at older age
same sex couple
how do same sex couples have children
fem/fem = get sperm donor - easier
male/male = more complicated - need egg bank and surrogate
when and where to use ART - prevention
prevention of inherited diseases and genetic disorder by preforming pgt - test embryos
define term infertile
couples who have not been able to conceive after one year (12 months) of natural unprotected intercourse
what is subfertile
few couples are completely unable to have children - many are considered to be subfertile = have reduced ability to conceive and produce children = not completely infertile, can conceive with help
Clinical definition of infertility according to WHO
disease (no fault) of the reproductive system define by failure to achieve a clinical pregnancy after 12 months of more of regular unprotected sexual intercourse
what is normal fertility - parameters
50% after 3 months
70-80% within first 6 months
85% conceive by 12 months
92% after 2 years
describe likelihood of fertility - percentage
likelihood of fertility decreases to 1% after 3 years of intercourse without contraception
becomes more difficult, should approach fertility doc after 1 year
describe fertile interval
6 days prior to ovulation to day of ovulation
when is highest probability of conception
intercourse 1-2 days prior to ovulation
what is normal fertility dependent on
maternal age mostly, lesser extent = paternal age
describe life of sperm/ovum
sperm lives 3-5 days in fem repro tract
ovum survives 12-24 hrs after ovulation
cannot figure out exact implantation window but around ovulation = best
etiology of fertility
before = used to be considered only the females fault, but now know men can contribute
male factor = 30%
female factor = 30%
combined = 10%
unexplained = 25%
other =5%
must check males and females at clinic
what does detection of certain genetic causes of male infertility allow for
to be informed about potential to transmit genetic abnormalities that may affect health of offspring
describe male infertility - conditions (categories)
some conditions = identifiable (can see problem) and reversible (like surgery)
Other conditions = identifiable but not reversible
describe summary of investigations in the man - fertility
anatomical investigation = if abnormal = surgery
semen analysis = if positive antibody reaction - treat, if infection = treat, if low count = improve lifestyle and can send to art
hormone analysis = if abnormal = treat if can
if no sperm = get donation
CAN ALL BE SENT TO ART
what is male infertility due to
deficiencies in sperm
production
blockage of the sperm delivery system
antibodies against sperm
injury to testicle
disorders of hormone production
poor descent of one or both testes
the presence of a varicocele
name the most commonly identifiable female factors for infertility
ovulatory disorders - 25% -do not ovulate normally
Endometriosis –15%
Pelvic adhesions –12%
Tubal blockage –11%
Other tubal abnormalities –11%
Hyperprolactinemia (high levels of prolactin in the blood)– 7%
describe summary of investigations in the woman - fertility
swabs and cervical smear = if abnormal treat- fix before ART
fallopian tubes = if abnormal = surgery then art
ultrasound = if abnormal surgery
Hormone levels = if abnormal = surgery or medical treatment, if no eggs = egg donations
CAN ALL BE SENT TO ART
infertility in population
1 in 6 couples will face a fertility issue in their reproductive period
What are the options for Infertile couples
adopt
no children
ART
who may benefit from ivf - 5
Blocked or damaged fallopian tubes or no fallopian tubes
male factor infertility = low sperm count or poor motility
women with ovulation disorders , premature ovarian failure, uterine fibroids
Individuals with genetic disorders
sometimes unexplained fertility
describe ART - where is it now - study prediction
nearly 170 to 400 mil people in 2100 (3% of world) may be alive as result of ART
describe ART - where is it now - to date
MORE THAN 5 mil ivf babies born
describe ART - where is it now - globally
ivf available in almost all parts of globe
doesnt mean its accessible tho - like someplaces = free while others not free = very expensive everywhere
describe normal egg development
females release one egg per month, sometimes 2 oocytes = frat twins
need one good egg (and one good sperm = pregnancy)
what is involved in ivf procedure - gen - 5 “points”
monitor and stimulate development of healthy eggs in ovaries - want as many as possible
collect eggs
secure sperm
combine eggs and sperm together in lab and provide appropriate environment for fertilization and early embryo growth (culture)
transfer embryos back into uterus
what is step 1 of IVF procedure - describe all
Fertility med prescribed to stimulate ovary - egg production
multiple eggs desired since some eggs will not develop or fertilize after retrieval
transvaginal ultrasound used to examine ovaries
blood tests = check hormone levels
describe afc - step1 ivf
count amount of follicles = inside has egg
describe standard ivf protocol - step 1
clinically prescribe to patients
oral contraceptive= administered on day 1 or 2 of menstrual cycle to prevent formation of cysts and schedule time of treatment - continued for 10-21 days - 3 weeks
designed per person tho - grow follicles and mature eggs - recruitment of follicles
how many injections - ivf step 1
10 days of injections
Subcutaneous
Financially, emotionally and physically hard
describe ultrasound image of follicular dev - step 1 ivf
look at size of follicles and measure
18-20ml = time to extract egg
follow with ultrasound
what is step 2 of IVF procedure - describe all
eggs retrieved through minor surgical procedure that uses ultrasound imaging to guide a hollow needle through pelvic cavity to remove eggs
medication provided to reduce and remove potential discomfort
describe aspects of oocyte retrieval - step 2 ivf
transvaginal ultrasound guided retrieval procedure
probe needle = bit of suction. to aspirate eggs and fluids
aspirate growing follicles one by one - hopefully good egg growing
what is step 3 of IVF procedure - describe all
male asked to produce sample of sperm - prepared for combining with eggs
what is step 4 of IVF procedure - describe all
insemination = sperm and eggs mixed together and stored in lab to encourage fertilization
some cases where there is lower probability of fertilization due to male factor (low sperm count) = icsi used = single sperm injected directly into egg
eggs are monitored to confirm fertilization and cell division are taking place
once fertilization = now considered embryos
what is step 5 of IVF procedure - describe all
embryos transferred into womans uterus 3-5 days after egg retrieval and fertilization
catheter or small tube inserted into uterus to transfer embryos
procedure painless for most women - some may experience mild cramping
if procedure successful = implantation occurs around 6-10 days following egg retrieval
step 5 of IVF procedure - describe culture gen
culture in vitro- can transfer embryo at any stage but usually wait till blastocyst, and must choose best embryo
describe lab aspects - oocyte retrieval and identification
extract eggs and pass down to embryologist in lab
look at it under microscope = find eggs and see if good
how to score oocytes - lab aspects
cumulus spread = mature
disgard morphologically abnormal oocytes = degenerated, pathenogenesis, big polar bodies
Germinal vessicle oocyte = immature
m1 oocyte = no gv, no pb
m2 oocyte = first pb- mature
weed out abnormal sizes/shapes = only take normal ones
Nuclear and cytoplasmic maturation
describe sperm analysis - gen
once have good egg then do sperm analysis
semen assessed for many factors
look and then score and decide which method to use for fertilization
describe sperm analysis - what is it assessed for - ART
count=total number of sperm
motility = % of total viable sperm
Progression = speed of forward movement
abnormality= % of abnormal sperm
describe sperm analysis - WHO parameters
sperm conc = 15mil/ml or more (usually 3-5ml)
total motility = 40% or more
progressive motility = 32% or more
morphology = 4% or more normal forms (strict criteria)
describe sperm preparation
wash sperm, swim up technique
percoll gradient
sucrose gradient
pure sperm
or just buy protocol
describe sperm morphology
All Sperm are Normal
but with Different Sizes
(the length of a healthy sperm tail can be from 40
to 250 μm)
abnoormal= big head, small head, dull head, 2 tails, pointy head
should be moving with tail forward and fast
what is globozoospermia
round headed sperm with no acrosome
cannot fertilize egg
Chromosomal abnormal = tetraploid, so embryo produced would be genetically abnormal
describe insemination
spem samples provided on day of egg collection = prepared in labelled tubes to separate out live motile sperm
each egg cumulus complex inseminated with 25,00- (100,000 sperm/ml) prepared sperm after 3-4 hr pre incubation of oocytes
dishes containing egg and sperm are incubated overnight in incubator - see if fertilized
what is icsi - describe
insertion of a single sperm into the oocyte
bypassing all the oocyte coat penetration and gamete fusion steps (characteristic of natural fertilization)
sperms must be immobilized = nick tail
polar body must be at 6 o clock - so won’t disturb chroms
which step is bypassed in icsi
sperm fusion step
male pronuclear development generally required oocyte activation but in humans
in icsi - how is human oocyte activated
vigorous suction of ooplasm and sperm nuclear insertion - injection process itself activates egg
when is icsi primarily used
Treatment of sever cases of male factor infertility with an abnormal semen analysis
do not need many good sperm to fertilize egg
describe normal and abnormal fertilization -ivf
all morphological
presence of 2 pronuclei = normal and 2 polar bodies
zygotes with triple or more pronuclei = discarded
zygotes with normal fertilization at time of scoring transferred into new dishes containing pre incubated embryo culture medium for further developmental culture
describe embryo culture - culture media
media and components crucial parts of what is needed at each developmental stages of embryos
was hard to determine bc started with mouse - but humans more complicated
describe embryo culture - environment
many factors
homeostatic environment with as little stress as possible
very delicate
need to get environment right = temp and stress free
what are incubators
device used to grow and maintain cell and embryo cultures
maintains optimal etmp, humidity and other conditions such as co2 and o2 content of atmosphere inside
must get it right bc ph of media is important
describe regular air vs incubator settings
regular air = 78% nitrogen gas (N2) and 22% oxygen (O2), CO2 content of air is less than one tenth of 1%
incubator = with 6% CO2 and 5% O2 control - interacts with culture media used to determine the ph within system
describe ivf incubators - gen
early days = glass cylinder, can open and put embryo in media then close it, stick in 37 degree ovens
now = incubators connected to computer system - if something goes wrong like concentration of gases computer will know
diff types = tri gas incubators - can open only one - so then not affect others, exposing to room temp = bad all the time
describe scoring day 1 zygote
2 pronuclei
2 polar bodies
describe scoring day 2 cleavage
Should get 2-4 cells
describe scoring cleavage stage grading - gen
grading based on how looks under microscope
worse grade embryos probably will not develop same as a better grade embryo
describe scoring cleavage stage grading - GRADE 1
Evenly sized blastomeres
spherical
Moderate refractility (not very dark)
describe scoring cleavage stage grading - GRADE 2
Uneven or irregularly
shaped blastomeres
Variation in refractility
< 10% fragmentation
describe scoring cleavage stage grading - GRADE 3
< 50% fragmentation
Remaining blastomeres should be in Grade 2 condition
describe scoring cleavage stage grading - GRADE 4
> 50% fragmentation
Remaining blastomeres
should appear viable
describe scoring - day 3 cleavage stage embryo
Transition from maternal to zygotic gene activation at around 4-8 cells
many more cells - seems to compact
describe scoring day 4 - compaction
morula
tight junctions (outer)
gap junctions (inner)
start of differentiation
tries to form blastocyst cavity, nicely compacted
describe scoring day 5/6
should see icm (becomes embryo) and te (placenta)
do not develop at same rate
Choosing morphologically good quality blastocyst for transfer
describe grading blastocyst - all
ICM = A- numerous and tightly packed B- several and loosely packed cells C- few cells
TE = A- many tightly packed cells organized into epithelium B- several cells organized into loose epithelium C- few cells
GRADE A - MORE LIKELY TO IMPLANT AND SUCCEED
do all morphologically good looking embryos work well and produce good babies
nooooo
do not have equal potential for implantation
since just looking at morphology
do not know genetics - chromosome abnormalities
describe real time monitoring device for embryo selection - gen
idea to apply time-lapse techniques in mammalian embryology is to observe some changes in structure elements or to analyze mechanism of dynamic processes including
expansion and hatching
describe real time monitoring device for embryo selection - purpose
measure the length of cell cycle, compaction, blastocoel formation, and compare the effects of various culture conditions
if does a,b,c,d = best - at certain times, how long it took untill 1st cell division, how many cell divides at what time, at what rate, when blastocele cavity forms
describe real time monitoring device for embryo selection - mechanism
may help to understand the mechanism of polar body
extrusion and pronuclear formation and provide a direct
evidence about the reversible nature of fragmentation
describe real time monitoring device for embryo selection - correlations
early disappearance of pronuclei and onset of the first cleavage were found to be correlated with a higher number of blastomeres on day 2 after oocyte retrieval providing the predictive potential of early-stage development of human
embryos
describe embryoscope
Chamber and put dish and monitor embryos = video
time lapse incubator
embryo development not uniform = different rates
describe human blastocyst hatching
proteases digest proteins and zona, once blastocyst expanded= egg must hatch out for implantation
if no hatching = embyro dies
describe laser assisted hatching
makes hole and wait for it to hatch out
before = used to be solution used to make hole
name the indications of assisted hatching - 9
2 previous implantation failures
37 years old and over
Day 3 FSH ≥ 15 mIU/ml
Thickened ZP (≥ 15 μm thick)
< 5 cells on Day 3
> 20% of the perivitelline space occupied by extracellular fragments
IVM embryos
Thawed oocytes/embryos
Patient request
what is ivm
in vitro maturation of oocytes
why ivm - all
some women v sensitive to stimulation with exogenous gonadotropins
increased risk of developing ovarian hyperstimulation syndrome (ohss) = nausea, vomiting, liquid retention, shortness of breath- during ivf process
long term side effects of repeated ovarian stimulation may increase risk of ovarian, endometrial and breast cancers - some patients worry, studies have not shown this tho
describe ivm
do no give hormones
immature oocytes can be matured in vitro - after releasing from antral follicles in vitro - extract cells then mature in vitro - hard
culture conditions affect oocyte maturation rate and quality during ivm - must mimic natural envir
cumulus cells associated with oocyte quality during ivm
oocyte maturation rates in vitro are different from diff reports (50-75%, immature eggs that mature, culture for 24-36 hrs, some keep till 48, but after 36 hrs probably wont mature)
describe ivm - procedure
immature oocytes –> ivm –> fertilization –> culture in vitro –> cryopreseve or insert to uterus
what is cumulative morphological assessment
optimal evaluation of an embryo’s potential would necessitate multiple assessments of the embryo at each of the developmental checkpoints;
Embryo development makes its transition from a single-celled zygote after fertilization of the oocyte to a blastocyst and from maternal to embryonic genome activation, initiation of protein synthesis, and cell differentiation;
what is most predictive information regarding embryo quality today
conducting noninvasive evaluation of the preimplantation period in a systematic, cumulative fashion
name all noninvasive and invasive methods to select best quality embryo - 6
Metabolic Parameters for assessing Embryo Viability(all non invasive)
Oxygen Consumption as an indicator of Embryo Viability
Amino Acid Turnover as a Biomarker of Embryo Viability
Gene Expression Changes during Embryo Development: New Application for Embryo Selection (will tell you which embryo best for implantation)
Real-Time Embryo Monitoring Device for Embryo Selection
Embryos biopsy and Preimplantation genetic testing for chromosome compliments
describe embryo transfer
load embryos on catheter under microscope - the best looking ones
ultrasound guided embryo transfer - transvaginal = outer sheath of catheter and deposit into endometrial cavity
what to do with rest of embryos - 2 methods
1 = slow freezing = slowly reduce temp - takes around 3 hrs, then stick into liquid nitro
2 = vitrification or fast freezing = more effective and better results
describe embryo vitrification
shrink at blastocyst stage - liquid can cause ice crystals - when shrink = remove liquid and only have cells, put on media - buffer or washing media, equilibration solution (3-5mins, 7.5%dmso + 7.5% eg) then vitrification solution (30 sec, 0.5M sucrose, 15%dmso +15%eg) then put on freezing straw then put into liquid nitrogen
describe storage of spare of supernumerary embryos
cryo
be careful when labelling - for each patient
descrive embryo thawing
run through process =
tm (1 min, 1.0M sucrose) then d1 (2min, 0.5M sucrose) then d2 (3 mins, 0.25M sucrose) then wash (5 mins, buffer or washing media) then repeat wash
expands again - takes a couple of hours for embryo to be back to normal
describe pregnancy test
beta hcg test at around 14 days after embryo test
if at certain number = positive for pregnancy
descrive viability test
see if fetal heart beat - embryo viable then send to obstetrician
describe historical view of human ivf
took a while for scientists to culture embryo in vitro
have been taken from the mother before ovulation, fertilized in vitro and grow in vitro to the eight or sixteen-celled stage in various media
discover = 1970
describe worlds first ivf baby
1978
in UK - cambridge
louise brown
needed a clinician and a scientist = Patrick steptoe and robert edwards
all IVF BABIES ARE NOT INFERTILE dude
