ART ISSUES

1. ART ISSUES Tyler Dohlman

2. Poor Success • SCNT • High rates of embryonic and fetal mortality • Rates have been reported to be 50-100% embryonic loss • Success rates have been consistently around 4-12% to live birth • Many postnatal issues (discussed later)

3. Large Offspring Syndrome(Large Placenta Syndrome?) • 50% of SCNT cloned calves • 53.1 kg (117#) average weight compared to 44.5kg (98#) from IVF calves (Cloning and Gestation Loss in Cattle Heyman et al 2002) • At day 150 of gestation, NT calves 17% heavier than AI calf controls (Rita Lee et al 2003) • Reduced number of placentomes, heavier placentomes, increased birth weight, enlarged organs (heart, liver, kidneys, umbilical cords), postnatal morbidity/mortality • Along with other abnormalities (hydroallantosis, fetal ascites, other fetal anomalies)

4. LOS study • Perinatology Cloned Calves (Batchelderet al 2007) • 8 cloned calves (Herford and Holstein) vs 6 ET (Hereford and Holstein) birth to 48hrs • Decrease in placentomes, 2 fold increase in surface area and mass, large and flat • Found other defects that were significant • Decrease in glucose and increase in fructose fetal blood = measurements of carbohydate metabolism • Fructose – produced by placenta and probably accelerates umbilical and cardiac anomalies

6. LOS

7. LOS study • Neonatal Study (Palmer 2005 ACVIM) • 41 Holstein clones • 27% death loss • 45% Persistent Pulmonary Hypertension of the Neonate (PPHN) • Reversed fetal circulation • Decrease oxygen flow • Due to patent ductusarteriosis (PDA) • Increase partial pressure oxygen and prostaglandins • 27% Musculoskeletal Issues • 18% congenital heart defects • 77% sepsis (umbilical infections) • 15/41 (37%) Neonatal Encephalopathy • 8/41 (20%) Neonatal Nephropathy

8. Delivery of cloned calves (study) • 26 cloned calves delivered by induction and c-section • 100% enlarged umbilicus with excessive bleeding • 56% umbilical infection • 35% cardiorespiratory maladaptation (respiratory failure) • 50% hypoxemic with hypercapnia • 35% apnea • 27% hypoventilation • 27% musculoskeletal abnormalities (bulldog or arthrogryposis) • 17/26 (65%) alive and healthy at 3 months of age • (Fectean et al 2009)

9. Bulldog Calf

10. Arthrogryposis

11. Fetal Anasarca

12. Factors of LOS • Epigenetic and aberrant expression of genes • Insulin Growth Factor – 1 • Concentration in fetal plasma significantly higher in NT pregnancies (Ravelich et al 2003) • Allantoic fluid levels of IGFBP-1 significantly higher in NT pregnancies • These findings suggest a disturbance in endocrine and paracrine function and equilibrium • Altered expression of some genes may play a huge role in imprinting genes that regulate fetal growth and development • XIST mRNA in the chorions of SCNT • CDKN1 and TP73 are key roles of expression of VEGF (important key player in angiogenic factor of the placenta (Farin et al 2006) • Methylation factors DNMT 3a and 3b may play a role in expression of altering preimplantation embryos

13. LOS • Monitoring for LOS (Heyman et al 2002) • Ultrasound and maternal pregnancy serum protein B (PSP60) • Ultrasound has been used in the human and on equine side to monitor fetal health and placental attachments • PSP60 is a common pregnancy diagnostic indicator procedure done with maternal blood • PSP60 by the binucleated cells of the placenta • Pregnancy loss for SCNT had significantly higher PSP60 levels by Day 50 than maintained pregnancies • PSP60 was also higher between 120 Day – Parturition in SCNT calves that had pathological issues in which placentome sizes were significantly larger and severe hydroallanotosis developed

17. IVF Human Issues • Multiple pregnancies (twins and more) • Preterm labor • Maternal hemorrhage • Cesarean delivery • Pregnancy induced high blood pressure • Gestational diabetes • Ectopic (tubal) pregnancies – 2-4%

18. IVF Human Issues • Preterm issues • Breathing and feeding difficulties • Cerebral palsy • Developmental delay • Vision and hearing problems/impairments • Birth defects (normal conception = 2-3%, IVF = 2.6-3.9%) • Sex chromosome abnormalities • Cardiovascular and musculoskeletal abnormalities • Hypospadia • Imprinting/epigenetic disorders especially if ICSI • Angelman and Beckwith-Wiedemann syndromes

21. IVF ISSUES • What is the biggest factor affecting outcome of ART success (beyond the scope of multiple pregnancies)? (Lambert, 2003) • Infertility itself was the highest factor for health problems with ART • Untreated infertility is 3.3X more likely to have perinatal death compared to fertile woman

22. Possible Solutions • Fresh or vitrification IVF embryos produce bigger/healthier babies?? (Wenhao Shi, 2012) • 494 vitrified embryo babies compared to 808 fresh cleaved embryo babies • Statistical differences: • Mean birth weight higher in vitrified babies (p<.05) for single and multiple gestations • Vitrified babies had lower neonatal ICU admission rate on multiple gestations (30% compared to 44% fresh ET)

23. Possible solutions • Longer incubation of IVF embryos produce bigger babies? (Makinen et al, 2012) • 1079 infant data (2000-2010) comparing Day 2, 3 and 5-6 culturing differences • Statistical analysis: • Proportion of large of gestation age (LGA) increased with longer incubation • D2 = 9.4%, D3 = 11.5%, D5-6 = 18.8% • Pvalue=<.004 (D2:D5-6) • Pvalue = <.05 (D3:D5-6) • Is LGA good? Maybe correlated to early childhood obesity

24. Possible solutions • Does the type of media to incubate and mature embryos control neonatal birthweight? (Vergouw et al, 2012) • 358 fresh D3 singletons vs 159 frozen-thawed Day5 singletons • No difference in type of media used • However, frozen-thawed embryo babies had higher birthweight (pvalue=<.0001)