Thyroid Hormone Analogues: Promising Therapeutic Avenues to Improve Neurodevelopmental Outcomes of Intrauterine Growth Restriction
Creators: name: Chincarini, Ginevra
Publisher: name: RMIT University
Year: 2025
Subjects: classificationCode: 321501
subject: Foetal development and medicine
classificationCode: 320901
subject: Autonomic nervous system

description: Intrauterine growth restriction (IUGR) is a pathological restriction of fetal weight commonly caused by placental insufficiency.1 Currently, there are no in utero interventions to prevent or correct placental dysfunction, which can severely compromise brain development.2-9 Thyroid hormone (TH) is crucial for brain maturation, and decreased expression of monocarboxylate transporter 8 (MCT8), a specific TH transporter, has been reported in the brains of IUGR human fetuses10 and rodents (Azhan, A., unpublished thesis, 2019). 
The thyromimetic 3,5-diiodothyropropanoic acid (DITPA) does not require MCT8 for cellular uptake,11 making it a promising therapeutic candidate. Our laboratory has previously shown that short-term DITPA administration to IUGR rats from postnatal day (P) 1 to P6 improves myelination without detrimental effects on body composition when assessed immediately after treatment at P7 (Azhan, A., unpublished thesis, 2019). However, when DITPA was administered for a prolonged period, from P1 to P13, at P14 myelination was improved in the IUGR offspring only in some of the brain regions assessed (Kondos-Devcic, D., unpublished thesis, 2021). In fact, this prolonged treatment duration showed negative effects in IUGR rats, including reduced overall myelination in the cerebral cortex, as well as in the corpus callosum (CC) and external capsule (EC). These data suggest that a shorter treatment duration might be safer and sufficient to protect the developing IUGR brain, leading to question whether the beneficial effects of DITPA are sustained in the long-term. 
The present thesis first conducted a systematic review (Chapter 3) to assess the long-term neurobehavioural effects of IUGR in humans and animal models caused by placental insufficiency. More specifically, the translatability of neurodevelopmental outcomes reported in preclinical models of IUGR to the human condition was systematically assessed. Findings in animal models were consistent with human outcomes, showing that individuals born with IUGR are at higher risk of motor and cognitive problems during adolescence and young adulthood. IUGR may also correlate with anxiety, although further clinical studies are needed to reach firm conclusions. This systematic review therefore served as a starting point for the present thesis, confirming the hypothesis that preclinical models of IUGR, including the rat model of IUGR, are valuable tools for investigating the underlying neurodevelopmental deficits and testing novel therapies. 
Chapters 4 and 5 examined whether DITPA reduced long-term adverse neurostructural and functional outcomes following IUGR using a rat model induced by bilateral uterine vessel ligation (BUVL), a well-established technique to mimic the effects of placental insufficiency.12,13 In chapter 4, the optimal age for DITPA treatment was initially studied using immunohistochemical analysis (Chapter 4). The results confirmed decreased MCT8+ cell density in the CC, EC, and primary motor cortex (M1) in the P7 IUGR rat brain, which resolved only in the M1 by P10 and P14. Combined with the positive outcomes of DITPA treatment in the previous short-term administration study (P1-P6; Azhan, A., unpublished thesis, 2019) and the negative effects following a longer DITPA treatment duration (P1-P13; Kondos-Devcic, D., unpublished thesis, 2021), a 7-day administration regime was chosen for the present study. DITPA (0.5 mg/100 g) or saline (equivalent volume) were therefore injected daily to newborn IUGR and control rats from P1 to P7. 
Behavioural tests were performed on all rats between P28 and P33, equivalent to early adolescence in humans. At P34, tissues were collected for wellbeing assessments (results reported in Chapter 6), and brains were processed for immunohistochemical analyses of oligodendrocyte (OL) density and myelination in major white and grey matter regions. Chapter 4 assessed motor function and anxiety-like behaviours, as well as OL density and myelination in the CC, EC, and M1. Chapter 5 examined memory function as well as OL density and myelination in the hippocampal cornu Ammonis (CA) 1 and CA3 regions and the fimbria. 
In Chapter 4, IUGR rats exhibited no motor impairments compared to controls, but anxietylike behaviours in IUGR offspring increased and were alleviated by DITPA treatment. Immunohistochemical analysis in IUGR compared to control rats showed reduced density of the entire population of OLs (oligodendrocyte transcription factor 2; Olig2), and a reduced density of mature OLs (adenomatous polyposis coli; APC) in the CC, EC, and M1, with increased myelination (assessed using myelin basic protein, MBP) in all three regions. DITPA treatment in IUGR rats increased Olig2+ cell density in all three regions, however only improved mature APC+ cell density in the CC and EC. Furthermore, DITPA normalised myelination in IUGR rats compared to saline treated rats in all three regions. In control rats, DITPA decreased mature APC+ cell density in the EC, suggesting possible adverse effects in healthy animals. Chapter 4 highlights the region-specific impacts of IUGR and the restorative potential of DITPA, while cautioning against its use in healthy neonates. 
In Chapter 5, male IUGR rats exhibited long-term non-spatial memory impairments, which were resolved with DITPA treatment. Spatial memory was not affected by IUGR or treatment. In the female offsprings, no differences in memory function were found between any of the groups. Both male and female IUGR rats showed reduced Olig2+ and APC+ OL densities in the hippocampal CA1 and CA3 regions, and in the fimbria, all of which were corrected by DITPA. Myelination was increased in the fimbria of IUGR + Saline rats compared to Control + Saline counterparts, but trended towards normalisation in IUGR + DITPA animals, while myelination in the CA1 and CA3 regions remained unaffected in IUGR rats. These results suggest that early postnatal TH-based therapy can restore neurodevelopmental processes in male and female IUGR rats and improve cognitive function in IUGR males. 
Chapter 6 examined the effects of early-life DITPA administration on growth and wellbeing in IUGR and control rats during early adolescence. IUGR + Saline animals had reduced body and organ (liver and kidney) weights, total brain and cerebral hemisphere weights, as well as body morphometry (head and hip circumference, crown-rump length) and body composition (DEXA; bone area, bone mineral density, bone mineral content, lean mass, fat mass) parameters. Furthermore, male IUGR + Saline rats had reduced circulating free thyroxine (FT4) concentrations, along with elevated liver alkaline phosphatase (ALP) levels compared to Control + Saline males. In IUGR + Saline females, alkaline transaminase (ALT) and ALP concentrations were increased compared to Control + Saline females, however all values remained within normal ranges. Free triiodothyronine (FT3) and thyroid stimulating hormone (TSH) serum levels were also within normal ranges, although IUGR rats had higher FT3 values than controls. DITPA treatment normalised cerebral hemisphere weights and FT4 concentrations in IUGR rats but had no effect on the other parameters. In control rats, DITPA reduced fat mass and FT4 levels, and increased ALP enzyme levels in males, though all remained within safe limits. Overall, DITPA had no adverse effects on postnatal growth, supporting it as a suitable therapy in the context of IUGR. 
In summary, this thesis provides evidence that early postnatal short-term administration of DITPA has sustained benefits on behavioural outcomes, OL density, and myelination in early adolescent-equivalent IUGR rats. DITPA did not negatively impact postnatal growth and metabolic parameters in IUGR animals, though it led to some undesirable effects in control animals. This work suggests that DITPA has the potential to improve in the long-term neurodevelopmental outcomes of IUGR in rats without harmful off-target effects. However further preclinical research using a large animal model is necessary before considering it as a therapeutic option.