Lecture 20 Flashcards
List the reasons that infertility is a growing problem in the US.
- Reduced morbidity
- Societal changes (delayed childbearing, marriage and first pregnancy)
- More therapies available
- Reduced expense and less taboo
List the forms of Assisted Reproductive Technologies (ART) in women.
- Sperm donation
- Egg donation
- Embryo donation/adoption
- In vitro fertilization (IVF)
- Intrauterine insemination (IUI)
- Traditional surrogacy
- Gestational surrogacy
- Sperm cryopreservation
- Egg cryopreservation
- Embryo cryopreservation
- Post-mortem gamete harvesting
- Post-death conception
List the stages and times of a typical IVF cycle.
- TVOR, Sperm Prep and Insemination (Day 0)
- Fertilization check (Day 1)
- Embryo transfer (Day 3-5)
- Embryo freeze (Day 5-6)
In IVF, when does the embryo culture occur?
Days 1 through 6
What does TVOR stand for?
Transvaginal oocyte retrieval
In IVF, what hormones are injected and when?
-FSH (-14 to -10 days till day 0) to produce eggs
- hCG (day 0) triggers egg maturation and ovulation
- Leuprolide acetate (day 0), a GnRH agonist that can control the stimulation process or be used as a trigger shot
- Progesterone supports endometrial development and early pregnancy
- Estrogen stimulates egg growth and thickens uterine lining
- Corticosteroids improves outcome of IVF with conventional insemination when sperm has autoantibodies
What stage of the menstrual cycle is progesterone associated with?
Luteal phase
What occurs during the luteal phase in terms of follicular development and in the endometrial cycle?
Corpus luteum forms and the uterine wall thickens
What hormones spike to trigger ovulation?
- LH
- Estrogen
- FSH
What is the process of IVF egg maturation?
- Ovarian stimulation/ovulation induction: adminster FSH, LH, and GnRH for 8-14 days
- Development of eggs is monitered using trans-vaginal ultrasound
- Blood is drawn frequently to assess ovarian estrogen production
- When eggs are mature hCG injection initiates ovulation process
What is the process of IVF egg retrieval?
- 34-36 hours after hCG injection, a mld sedative and painkiller is given to the patient.
- An ultrasound probe is inserted into the vagina to visualize the ovaries and follicles
- Needle is inserted through wall of vagina to ovaries to extract the eggs
What is the process of IVF fertilization?
- If sperm are healthy, placed in a dish with the egg in an incubator over use
- If they can’t fertilize the egg, a single sperm is injected into an egg using a neeedle in a process called Intracytoplasmic Sperm Injection (ICSI)
- About 60% of IVF is performed with ICSI
- Couples should consider genetic testing if sperm cannot fertilize the egg on their own
What is the process of IVF embryo transfer?
- Embryos that develop from IVF are placed into the uterus 1-6 days after retrieval
- Procedure is normally painless but patient many experience cramping
- A long thin tube is inserted through the vagina and into the uterus to inject the embryo
- Embryo should implant into the lining 6-10 days after retrieval
Third Party Assisted ARTs
- Sperm Donation
- Egg Donation
- Surrogates and gestational carriers
When is egg donation a good option for women?
- Have primary ovary insufficiency (POI)
- Have had chemotherapy or radiation therapy
- Have had surgical removal of the ovaries
- Were born without ovaries
- Carriers of known genetic diseases
- Are infertile because of poor egg quality
What is a surrogate carrier?
A woman inseminated with sperm from the male partner of the couple
What is a gestational carrier?
A woman implanted with an embryo that is not biologically related to her.
What are the risks of traditional vs gestational surrogacy?
Surrogate: poses legal issues due to being genetically related to surrogate, surrogate undergoes IUI, comes with risks during development
Gestational: Costs more money, IVF is required
What is GIFT?
Laparoscopically placing mature eggs into the healthy fallopian tube along with washed sperm (Gamete intrafallopian transfer)
What is ZIFT?
Zygotes are placed directely into the fallopian tubes via lapaosocpy or trasncervical falloopian tube caterization (Zygote intrafallopian transfer)
How can a scientist tell if fertilization was a success?
Presence of a second polar body (meaning meiosis II occured) and male and female pronuclei both present
What is PGS?
Preimplantation genetic screening screens embryos to ensure 23 pairs of chromosomes
What is PGD?
Preimplatation genetic diagnosis diagnoses embryos for known genetic disorders that both the patient and partners are carriers of
What is CCS?
Comprehensive chromosomal screening is a next generation sequencing that detects aneuploidy with a more in-depth scan of the genoe to find abnormalities
Who should use biopsy/genetic screening?
- Women over 35 years
- Women who have had repeated miscarriages
- Couples with multiple failed IVF cycles
- Previous pregnancy with chromosomal abnormality
What is a mosaic embryo?
When two or more cell populations with different genotypes are present within the same embryo
Which mosaicism types are potentially diagnosable?
- Total mosaic
- TE mosaic
Which mosaicism types are never diagnosable?
- ICM mosaic
- ICM/TE mosaic type I
- ICM/TE mosaic type II
Consequences for embryo survival in terms of mosaic embryos are unknown
TRUE
What are the features of slow-rate cryopreservation?
- Low concentration of cryoprotectant
- Programmable freezer
- Slow cooling rate
- Closed devices
- Inexpensive devices
What are the features of vitrification cryopreservation?
- High concentration of cryoprotectant
- No mechanical equipment
- Rapid cooling
- Open or closed devices
- Expensive devices
When is totipotency lost?
When blastomeres reach the 8 cell stage after which differentiation starts
Are embryonic stem cells completely totipotent?
No, as they are not interchangeable as differentiation starts.
How do embryonic stem cells (ES) differ from all other cells?
- They can be induced to change or differentiate into virtually any cell before differentiation starts
- Somatic cells which have finite lifespans, ES cells can grow indefinitely in culture
What are factors of somatic cells compared to induced pluripotent stem cells?
- Limited proliferation
- Pluripotency genes methylated
- Tissue-specific cell morphology
- X-chromosome inactivated
- Active G1 celll cycle checkpoint
- Expression of somatic cell specific markers
What are factors of induced pluripotent stem cells compared to somatic cells?
- Self-renewal
- Pluripotency
- ES cell morphology
- Demethylation of pluripotency genes
- Reactivation of pluripotency genes
- X-chromosome reactivation (female cells)
- Telomerase activity
- Loss of G1 checkpoint
What reprogramming factors can turn somatic cells into induced pluripotent stem cells?
- Sox2, Oct4, Klf4, c-Myc
- Sox2, Oct4, Nanog, Lin28
- Sox2, Oct4, Klf4
- Etc.
How does differentiation capability change throughout development?
- Pre-blastocyst: totipotent
- Blastocyst: pluripotent
- Fetus: multipotent
- Adult: unipotent
How exactly did ovarian function regenerate using placental-derived (PD) mesenchymal stem cells?
- PD-MSC transplatation increased estradiol level in ovarietomized (Ovx) rats compared with the non-transplantation group (NTx)
- Spheroid PD-MSCs exhibited a significantly higher efficiency of engraftment onto ovarian tissues at 2 weeks.
- Nanos3, Nobox and LHx8 were also significantly increased in Spheroid group
- PD-MSC transplantation restored ovarian function in Ovx rats by increasing esttrogen production & enhancing folliculogenesis-related gene expression levels
- Spheroid-cultured PD-MSCs have enhanced therapeutic potential via increased engraftment efficiency.
How can oocytes be developed from stem cells?
- ES cells and induced pluripotent stem cells have been differentiated into oocytes that have grown to full follicular maturity in mice
- In humans, follicle maturation through oocyte IVM to produce heathy offspring has been achieved
- No offspring production as of yet
- Further research needs to be done which is difficult as ethics still needs to be considered.
What is a Free Martin?
Sterile heifer sharing placenta with twin bull fetus.
How was Dolly the sheep cloned?
Nucleus from a ewe’s somatic cell was taken and implanted in an enucleated egg cell from another ewe, which was then fused with electrical pulses to induce fertilization.
Four methods to alter gene expression in livestock in vivo
- RNA interference (RNAi)
- Zinc-finger nucleases (ZFN)
- Transcription activatory-like effector nucleases (TALENS)
- Clustered regularly interspersed palindromic repeats (CRISPR) and CRISPR associated protein 9 (Cas 9)
Advantages and disadvantages for RNAi
- Advantages: diminishes gene expression
- Disadvantages: diminishes genes expression and variability between animals over time
Advantages and disadvantages for CRISPR-Cas 9
- Advantages: Ablates gene expression
- Disadvantages: May be lethal