<b>Ontological changes in the swine fetus and placenta from mid- to late-gestation</b>
Creators: name: Rudy, Kaylyn G
Publisher: name: Purdue University Graduate School
Year: 2024
Subjects: classificationCode: 300301
subject: Animal growth and development
classificationCode: 300305
subject: Animal reproduction and breeding

description: Porcine reproductive and respiratory syndrome virus (PRRSV) is a devastating virus that is endemic to the swine industry. This virus has little direct effect on the dam but results in abortions, stillborn, and delivery of viremic piglets. PRRSV is unable to cross the swine placenta in early gestation but as gestation progresses, the placenta becomes permissible during late gestation. The mechanisms that allow the virus to cross the late gestation placenta are not well understood, but several theories have been presented regarding changes in placental morphology or enzymatic changes. Additionally, piglets who experience IUGR due to uterine crowding have been found to be more resistant to PRRSV infection, having lower viral levels than their normal litter mates. When vertical transmission from the dam to fetus occurs not only can the effects previously mentioned occur, but PRRSV is also known to cause suppression of maternal and fetal thyroid hormone. Thyroid hormone plays numerous roles in fetal development such as accretion of fetal mass, appetite regulation, and coincidently follows a similar increase trajectory to that of fetal growth during mid- to late-gestation. Consequently, any dysregulation of thyroid hormone has the potential to cause severe side-effects and may alter fetal growth. The relationship between thyroid hormone and fetal growth and development is not well understood. Chapter 2 investigates the potential cause-and-effect relationship between fetal growth and thyroid hormone through the induction of fetal hypothyroidism. Pregnant gilts (n=24) were given a sham treatment (CON; n=12) or treated with methimazole (MMI; n=12), a goitrogen capable of crossing the placenta. These gilts were then further subdivided across four gestational timepoints spanning mid- to late-gestation, these being days 55, 66, 76, and 86. Treatment started 21 days prior to these desired dates. Upon completion of treatment the gilts were humanely euthanized, and fetuses were extracted (resulting populations of n=174 MMI and n=166 CON) and fetal body and organ weights were recorded. Collected tissues included heart, liver, lung, kidneys, spleen, brain and thyroid. Fetuses were imaged in the left and right lateral recumbency for phenotypic analysis, including novel head measurements. Placental sample were also taken. Additionally fore- and hind limbs were taken from the centermost male and female from each litter so that radiographs could be taken to analyze bone growth. Statistical analysis of all phenotypic differences was carried out using a linear mixed effect model including gestational age and treatment as fixed effects and gilt as a random effect. The data revealed that the left and right phenotypic parameters are highly correlated (R2&gt;0.9). Upon extraction, goiters were present in the MMI fetuses and there was a significant increase in both absolute and relative thyroid weights. Thus, the use of MMI during this period of gestation was successful in inducing hypothyroidism. Additionally, the MMI treated thyroids had a significant treatment by time interaction with 0.014g and 0.21g increase at day 55 and 66 respectively indicating reduced compensatory action within the fetal hypothalamic-pituitary-thyroid axis during this earliest period. Liver weight as a percentage of body weight decreased from 6.06% to 2.56% between days 55 and 86 in the CON group but, was significantly increased at all time points in response to MMI induced hypothyroidism (P&lt;0.01). Thus, the in brain to liver weight ratio decreases over time, in MMI fetuses (P&lt;0.05). While all other phenotypic parameters were significantly altered by gestation age, there was no significant impact of fetal hypothyroidism. This indicates that fetal thyroid hormone is not the driving factor for the exponential fetal growth seen in mid- to late-gestation. PRRSV virus is a complex and devastating virus to the swine industry, especially when it infects pregnant gilts and sows. PRRSV is unable to cross the swine placenta during mid-gestation but as gestation progresses the virus readily crosses the placenta and is able to infect piglets during this late gestation period. The mechanisms by which PRRSV crosses the highly restrictive porcine placenta are not clear. Additionally, piglets who experience intrauterine growth retardation experience lower virus levels than their normal counterparts. Chapter 3 investigates the changes in three genes of interest that we hypothesized, had the potential to fluctuate throughout gestation and facilitate PRRSV transfer, as well as the morphological changes that occur in the maternal-fetal interface through mid- to late-gestation and how these aspects may vary between IUGR and normal piglets. Placental samples were collected from pregnant gilts (n=12) equally divided across days 55, 66, 76, and 86 of gestation. Samples were taken of each fetus’s placenta adjacent to the umbilical cord. A portion of the sample was cut into 1 cm2 and placed into a mold with optimal cutting temperature media (OCT) for later cryo-sectioning and histology. The remaining portion had the fetal placenta peeled from the endometrium and flash frozen in liquid nitrogen for RNA extraction. A subset of samples was chosen based on fetus’s brain to liver weight ratios (n=96). From each litter two males and two females with the most extreme case of IUGR, based on z-scores, were chosen and the same was done for the two males and females with lowest brain to liver weight ratios, the later were classified as large for gestational age (LGA). 56 of the original 64 had acceptable levels of placental RNA for analysis. A total of 3 genes were chosen for analysis based on their function and previous literature. These included CD163, SIGLEC1 and IL-10. No significant up or down regulation was seen in any of the selected genes and there was no variation between IUGR and LGA fetuses. Additionally, placenta histology was conducted to evaluate populations of CD163 positive macrophages throughout the maternal fetal interface across mid- to late-gestation. Populations of CD163 positive macrophages were found on both the maternal and fetal sides of the maternal fetal interface at all timepoints. Collectively these results show there is no fluctuation in CD163, SIGLEC1, or IL-10 among timepoints or between IUGR and LGA fetuses. Additionally, the histology samples confirm the presence of resident populations of CD163 positive macrophages on maternal and fetal sides of the MFI. Collectively these results indicate that more research needs to be done to determine the underlying mechanisms of PRRSV transmission during late gestation.