Programming and Assisted Reproductive Technologies Modules 18 and 19

1. Programming and Assisted Reproductive TechnologiesModules 18 and 19 AnS 536 Spring 2014

2. Fetal Programming • Hypothesis • The developing fetus responds to nutritional and oxygen shortages by diverting resources from other organs to the brain • Potential adverse affects may occur later in life • Adaptations include: • Vascular response • Metabolic response • Endocrine response

3. Fetal Programming • Exogenous maternal malnutrition during pregnancy • May cause lifelong, persisting adaptation to the fetus • Low birth weight • ↑ Cardiovascular risk • Non-insulin dependent diabetes • Critical periods of vulnerability to suboptimal conditions during development • Vulnerable periods occur at different times for various tissues • Greatest risk: rapidly dividing cells

4. Fetal Programming • ‘Fetal origins’ hypothesis • Poor in utero environment induced by maternal dietary or placental insufficiency may program susceptibility later in fetal development and life • ‘Thrifty phenotype hypothesis’ • If in utero nutrition is poor, then predictive adaptive responses are made by the fetus to maximize uptake and conservation of any nutrients available, resulting in a conservative metabolism • Problems occur when postnatal diet is adequate and plentiful and exceeds the range of predicted adaptive response

5. Fetal Programming • Prevalent in developed and developing countries • Dutch famine (limited intake of 1680-3360 kJ) • During late gestation was associated with increased adult obesity and glucose intolerance • During early gestation resulted in hypertension • Disadvantageous populations in USA, South Africa, the Caribbean, India, and Australia • Shown cardiovascular risk to be greater in populations suffering from poor in utero nutrition

6. Fetal Programming • Permanent affects of programming • Modifies susceptibility to disease • Structural changes to organs • Might pass across generations • Different effects on males and females • Placental effects • Fetus will attempt to compensate for womb deficiencies

7. Metabolic Syndrome • Cluster of abnormalities occurring together, increase your risk of heart disease, stroke, and diabetes • Largely attributed to altered dietary and lifestyle factors favoring central obesity • Characterized by a group of metabolic risk factors in a person • Abdominal obesity • Atherogenic dyslipidemia • Elevated blood pressure • Insulin resistance • Proinflammatory states • Prothrombotic states

8. Metabolic Syndrome • Abdominal obesity • Strongly associated with metabolic syndrome • Atherogenic dyslipidemia • ↑ triglycerides, ↓ concentrations of HDL cholesterol, ↑ remnant lipoproteins, ↑ apolipoprotein B, small LDL particles nad small HDL particles

9. Metabolic Syndrome • Elevated blood pressure • Strongly associated with obesity • Commonly occurs in insulin-resistant individuals • Insulin resistance • Commonly associated with metabolic syndrome • Usually leads to glucose intolerance  diabetic-level hyperglycemia • Independent risk factor for cardiovascular disease

10. Metabolic Syndrome • Proinflammatory states • ↑ levels of C-reactive protein • Excess adipose tissue release inflammatory cytokines • Multiple mechanisms contribute to inflammatory state • Prothrombotic states • ↑ Plasma plasminogen activator inhibitor-1 • ↑ fibrinogen • Rises in response to a high cytokine state • Acute phase reactant • Proinglammatory and prothrombotic states are interconnected

12. Metabolic Syndrome • Underlying risk factors for this condition: • Abdominal obesity • Insulin resistance • Physical inactivity • Aging • Hormonal imbalance • Genetic predisposition • Fetal environment

13. Non-genomic Intergenerational Effects • Significant evidence that programmed phenomena can be disturbed in later generations • Offspring exposed to a poor uterine environment • Prenatal programming by nutrition or exercise (animal models) • Postnatal programming by nutrition or handling (animal models) • Effects: • Birth weight • Glucose tolerance • Hypothalamic-pituitary axis in subsequent generations

14. Non-genomic Intergenerational Effects • Effects on birth weight • Black and white hooded rats (Steward, 1975) • Continued poor maternal nutrition produced amplified effects on birth weight through a number of generations • Accidental introduction of less-palatable food in control animals resulted in a period of self-imposed calorie restriction • Evidence that poor nutrition in one generation can produce effects on birth weight in subsequent generations

15. Non-genomic Intergenerational Effects • Effects on birth weight, cont… • First generation pups (Pinto and Shetty, 1995) • Exercise during pregnancy resulted in low birth weigh first generation pups • First generation offspring were sedentary during pregnancy and second generation offspring were also found to be growth retarded • Suggesting adverse intergenerational influence of maternal exercise stress on fetal growth

16. Non-genomic Intergenerational Effects • Metabolic parameters and blood pressure • Female rabbits with surgically induced hypertension were mated with normotensive males • Female offspring had increased blood pressure as adults when compared to the offspring of sham-operated females • Blood pressure in male offspring was unaffected

17. Non-genomic Intergenerational Effects • Postnatal effects • Second generational alterations on glucose homeostasis has been seen when overfeeding takes place in the neonatal period • In rodents, naturally occurring variations in maternal behavior is associated with different hypothalamic-pituitary-adrenal stress responsiveness in offspring • Postnatal environmental manipulations to the hypothalamic-pituitary-adrenal axis stress response may produce intergenerational effects

18. Assisted Reproductive Technologies (ART) • Artificial insemination (AI) • Sexed semen and embryo sexing • Embryo transfer (ET) • In vitro fertilization (IVF) • Intracytoplasmic sperm injection (ICSI) • Gamete intrafallopian transfer (GIFT) • Zygote intrafallopian transfer (ZIFT) • Donor egg, sperm or embryo • Cloning (SCNT)

19. Artificial Insemination (AI) • Used commonly in livestock • Method of banking semen (genetics) without keeping a sire on site (cryopreservation) • Challenges • Difficulty passing AI gun through cervix • Potentially reduced pregnancy rates • Breeding when animal is in estrus • Damage to reproductive tract • Reduced fertility with cryopreserved sperm

20. Artificial Insemination (AI) • Management approaches • Goal is to increase conception rates • Implementing appropriate methods of heat detection • Skilled technician in AI • Time of year, time of day, or temperature on day of breeding can affect conception rates

21. Sexed semen and embryo sexing • Biological mechanism • Sexed semen uses flow cytometry to sort genetically male and female sperm • Female (XY) sperm have 4% more DNA than male sperm • Embryo sexing entails obtaining a biopsy of the inner cellular mass (ICM) of the embryo to determine male or female status • Used in livestock industry (dairy cattle) • Ethical considerations in humans

22. Sexed semen and embryo sexing • Challenges • Sexed semen • Reduced fertility • Higher concentration of sperm needed to ensure pregnancy • Embryo sexing • Reduced viability of embryo • Multiple pregnancies can occur

23. Sexed semen and embryo sexing Management approaches • Sexed semen is preferred method, less risk to developing embryo • Less invasive • Sexed semen is used in combination with IVF technologies or AI • Embryo sexing require embryo transfer technique

24. In vitro fertilization (IVF) • Commonly used practice in humans and livestock (cattle) • Biological mechanisms • Dam is administered a series of reproductive hormone (GnRH) to stimulate the development of Graafian follicles, a.k.a., superovulation • Oocytes are collected via aspiration and are incubated in an artificial lab environment, mimicking the environment of the uterus • Sperm is introduced to the oocytes and fertilization occurs • Embryos are developed to the blastocyst stage prior to transfer to the mother or dam

25. In vitro fertilization (IVF) • Challenges • Patients or recipients using IVF technology usually face moderate to severe infertility problems • Poor quality ovum or sperm • Uterine rejection • May be used as a ‘last ditch effort’ for pregnancy • Incidence of multiple births are high • Ectopic pregnancies

26. In vitro fertilization (IVF) • Management approaches • Age of the patient • Inversely related to the probability of multiple pregnancies and overall pregnancy success • Implantation rate • Attributed to many factors including quality of embryo • Selecting embryos with the greatest potential for survival • Matching synchrony of uterus to embryo stage of development • Number of embryos transferred • Directly related to risk of multiple pregnancies • Most controllable of the variables

27. Embryo transfer (ET) • Biological mechanisms (in livestock) • A donor animal is super ovulated, bred by a sire (AI or live cover) • Fertilization occurs in vivo and embryos are collected prior to the implantation stage • Collected embryos can be then be transferred to a recipient (surrogate) animal with the same estrus synchrony as the donor or can be cryopreserved for a later implantation date

28. Embryo transfer (ET) • Challenges • Reduced rate of pregnancy as compared to natural conception • Fresh embryos have better conception rate as compared to cryopreserved embryos • Synchronizing recipient animals with the donor animal • Retained embryos in donor animal resulting in pregnancy

29. Embryo transfer (ET) • Management approaches • Optimizing synchrony for maximum pregnancy rates • Selecting appropriate recipients for breed and birth weight of offspring • Use of prostaglandin in donor animals to eliminate pregnancy due to retained embryo

30. Intracytoplasmic sperm injection (ICSI) • Biological Mechanism • A single sperm is injected into unfertilized oocyte and is transferred to a recipient • Treatment for male factor infertility • Challenges • Potentially abnormal sperm can fertilize ova • Long term health affects, including genetic abnormalities • Lower birth weight • Abnormalities on the Y chromosome • Greater potential for developmental delays

31. Intracytoplasmic sperm injection (ICSI) Management approaches • Men and women should have genetic screening for potential chromosomal abnormalities prior to fertility treatment • Men lacking a vas deferens can carry mutations increasing the risk of offspring with cystic fibrosis

32. Gamete Intrafallopian transfer (GIFT) • Biological mechanisms • An unfertilized oocyte and sperm are combined outside of the uterus and are surgically transferred to the site of normal fertilization in the fallopian tube via laparoscopic technique • Fertilization occurs in vivo • Implantation occurs naturally

33. Gamete Intrafallopian transfer (GIFT) • Challenges • Surgical intervention causes trauma and scarring • More invasive technique • Multiple pregnancies • Management approaches • Other techniques are more widely used (IVF) due to higher success rates

34. Zygote intrafallopian transfer (ZIFT) • Biological mechanisms • Similar to GIFT however, oocyte and sperm are combined outside of the uterus and are not transferred until an embryo is produced • Management approaches • Other techniques are more widely used (IVF) due to higher success rates

35. Donor ova, sperm or embryo • Donor oocyte, sperm or embryos can be used to generate offspring if poor quality ova or sperm exist or if there is a lack of a female or male counterpart • Challenges • Social implications • Lack of genetic history • Predisposition to risk of disease • Children may never know their parents

36. Cloning (SCNT) • Producing genetically identical copies of a biological entity • Different types of methods: • Reproductive • Natural identical twinning • Somatic cell nuclear transfer (SCNT) • Non-reproductive • Recombinant DNA Technology • Therapeutic cloning

37. Cloning (SCNT) • Challenges • Low conception rates • Increased birth weights • Increased incidence of genetic abnormalities • Decreased neonatal survival • Increased placentation abnormalities • Decreased life span of animal?? • Increased dystocia and prolonged gestation • Decreased genetic variation

38. Cloning (SCNT) • Biological mechanisms • Low conception rates • Research is being done to explore this reality • Current methods of cloning are very artificial and vastly differ from normal in vivo embryo development • Methods to promote a more similar environment to what the embryo experiences in vivo • Increased birth weights • Possible link to media used in incubating cloned embryos • Fetal calf serum (FCS) promotes excessive growth of embryo

39. Cloning (SCNT) • Biological mechanisms, cont… • Increased incidence of genetic abnormalities • Possible link to problems in cell reprogramming with SCNT • Electric charge fuses cells to promote cell proliferation • Decreased neonatal survival • Offspring can be less vigorous initially after birth • Anemia, enlarged organs, metabolic disturbances, problems thermoregulating, hypoxia can all contribute

40. Cloning (SCNT) • Biological mechanisms, cont… • Increased placentation abnormalities • Mechanisms unknown • Hydrops amnion is a condition that is seen during gestation in cattle and sheep • Less frequent attachment sites but increased size of codyledons as compared to normal pregnancies in cattle • Intrauterine Growth Restriction (IUGR) • Decreased life span of animal ?? • “Dolly” the sheep only lived to 6 years of age • Controversial studies that cloning affects life span of offspring • Decreased telomere length has been associated with a decreased life span • Age of animal being cloned may affect life span of offspring (increased age shortens telomere length)

41. Cloning (SCNT) • Biological mechanisms, cont… • Increased dystocia and prolonged gestation • Recipient animals carrying cloned animals fail to recognize the onset of parturition near term or the cloned fetus fails to induce parturition • Increased birth weights contribute to dystocia • Decreased genetic variation • Selection of cloned animal can potentially promote a genetically inferior or superior animal • Breeding pool can be narrowed • Long term effects?

42. Cloning (SCNT) • Management approaches • Low conception rates • Matching synchrony of recipient animal with stage of embryo • Increased birth weights • Selecting larger framed, multi-parous recipient animals • Awareness of breed of embryo and potential birth weight • Caesarian section deliveries

43. Cloning (SCNT) • Management approaches, cont… • Increased incidence of genetic abnormalities • Humane euthanasia or abortion in severe cases • Preventing the perpetuation of genetically inferior animals through selection • Decreased neonatal survival • Intensive care and monitoring of animal first week of life • Ensuring colostrum uptake • Temperature regulation

44. Cloning (SCNT) • Management approaches, cont… • Increased placentation abnormalities • Close monitoring of recipient animals for hydrops amnion • Abort early in gestation if necessary • Pregnancy palpations/ultrasound to determine fetal well being • Decreased life span of animal ?? • Age of animal being cloned may affect life span of offspring (increased age shortens telomere length)

45. Cloning (SCNT) • Management approaches, cont… • Increased dystocia and prolonged gestation • Know expected parturition dates • Induce parturition if necessary • Caesarian sections • Decreased genetic variation • Criteria for animal selection • Promoting healthy animals – not just based on phenotype