title: Investigating the effect of low dietary iodine on cardiovascular disease parameters, gene expression and thyroid function in stroke-prone spontaneously hypertensive rats
Creators: name: Moseki, Phillip
Publisher: name: University of Glasgow
Year: 2024

description: Despite progress in strategies for preventing and controlling iodine deficiency, iodine deficiency disorders (IDD) continue to be a major public health problem throughout the world. At the same time, cardiovascular disease (CVD) remains a global health problem. Hypertension is the leading risk factor for CVD. The number of people with hypertension is expected to rise to 1.56 billion in 2025 while it is also estimated that &gt;2000 million people worldwide are still at risk of insufficient iodine intake. Iodine is an essential microelement in human physiology. Its role in thyroid function is well known and documented in the literature. However, recent evidence links iodine with direct cardiovascular functional changes independent of thyroid hormones (TH). The stroke-prone spontaneously hypertensive (SHRSP) rat is a well-established model of primary hypertension and endothelial dysfunction which can be used for investigating CVD mechanisms and novel interventions. It has been established that Wistar Kyoto (WKY) and SHRSP rats have similar thyroid function under baseline conditions. 

In this thesis, I aimed to understand the direct effect of iodine on cardiovascular function and disease development in young Wistar Kyoto (WKY) and stroke-prone spontaneously hypertensive (SHRSP) male and female rats. I also employed RNA sequencing (RNA-Seq) to determine the effect of low iodine diet (LID) on protein coding genes in thoracic aorta from WKY and SHRSP male rats. Moreover, I determined the effect of iodide on contractile signalling in mesenteric artery vascular smooth muscle cells (VSMCs) isolated from WKY and SHRSP males. 

In chapter 3, 5-week-old WKY and SHRSP males and females (n =7-8 per group) were assigned a normal (NID) or LID for 4 weeks. Tail cuff blood pressure (BP) and body weight (BW) were measured weekly while water consumption, urine production and echocardiography measurements were determined fortnightly. Plasma was collected for assessment of thyroid-stimulating hormone (TSH), free triiodothyronine (fT3), free thyroxine (fT4) and thyroglobulin (Tg). At sacrifice, thyroid gland and heart tissues were snap-frozen for assessing sodium iodide symporter (SLC5a5) and pendrin (SLC26a4) gene expression. Mesenteric arteries were assessed for vascular function using wire myography. At baseline there were no significant differences in urinary iodine concentration (UIC) and TH profiles between WKY and SHRSP males and females, however following the 4- week dietary period, UIC significantly reduced in LID fed rats irrespective of strain and sex. Contrary to the stable levels of fT3 and fT4, levels of TSH significantly increased in SHRSP but remained unchanged in WKY males and females. Strain differences in homeostatic mechanisms in response to LID were further revealed when SLC5a5 gene expression was significantly increased in the thyroid gland from WKY males but not SHRSP males fed LID. There were no significant effects of LID on BW, BP and cardiac function parameters. Contractile responses to U46619 in mesenteric arteries from SHRSP and WKY males fed LID increased in a dose dependent manner concomitant with significantly reduced UIC which were independent of fT3 and fT4. 

In chapter 4, thoracic aorta was snap frozen for transcriptomic profiling using RNA sequencing (RNA-Seq). Moreover, ingenuity pathway analysis (IPA) software was used to understand the biology linking to the differential gene expressions (DEGs). In support of the vascular function data (chapter 3), RNA-Seq analysis demonstrated DEGs in SHRSP fed LID. 438 DEGs were significantly increased in thoracic aorta from SHRSP fed LID when compared to those fed NID. The DEGs in IPA identified dilated cardiomyopathy as the most significant canonical pathway and within this pathway contractile protein coding genes such as Mhy7, Myh6, Actc1, Tnnc1, Tnni3, Tnnt2 and Ttn were significantly increased. Moreover, expression of these genes was also significantly increased in Ca2+, integrinlinked kinase (ILK) and actin cytoskeleton signalling pathways. The DEGs in ingenuity pathway analysis (IPA) software identified growth failure or short stature as one of the diseases and functions associated with LID. This suggest that SHRSP fed LID are likely to experience growth restriction independent of thyroid dysfunction. 

In chapter 5, mesenteric artery VSMCs were isolated from WKY and SHRSP males and used for assessing the effect of iodide on contractile signalling. This allowed investigation of the molecular mechanisms underlying enhanced vascular contractility secondary to iodine modification. I demonstrated that phosphomyosin light chain (phospho-MLC) was significantly reduced in SHRSP VSMCs incubated in cell culture media supplemented with iodide while there was no significant difference in VSMCs isolated from WKY male rats. In general, I demonstrated an anticontractile effect of iodide on VSMC isolated from SHRSP male rats. Furthermore, I demonstrated the presence of SLC5a5 protein in VSMCs from WKY and SHRSP male rats. 

To conclude, the results in this thesis demonstrate that short-term LID increases vascular reactivity in SHRSP and WKY male rats through increased expression of genes encoding contractile proteins in SHRSP but not WKY rats. Moreover, enhanced contractile response occurred independent of changes in TH profiles suggesting a direct effect of iodine on vascular function. This PhD thesis work therefore suggests that short-term LID is likely to have a greater detrimental effect on the hypertensive than the normotensive rat model and could potentially effect individuals with hypertension. Our work supports the hypothesis that dietary iodine has a direct effect on cardiovascular function and disease outcomes independent of TH levels and might be a new modifiable risk factor for CVD.