Dietary salt restriction in hyperthyroid rats. Differential influence on left and right ventricular mass

This study assessed the impact of salt restriction on cardiac morphology and biochemistry and its effects on hemodynamic and renal variables in experimental hyperthyroidism. Four groups of male Wistar rats were used: control, hyperthyroid, and the same groups under low salt intake. Body weight, blood pressure (BP), and heart rate (HR) were recorded weekly for 4 weeks. Morphologic, metabolic, plasma, cardiac, and renal variables were also measured. Low salt intake decreased BP in T₄-treated rats but not in controls. Low salt intake reduced relative left ventricular mass but increased absolute right ventricular weight and right ventricular weight/BW ratio in both control and hyperthyroid groups. Low salt intake increased Na⁺/H⁺ exchanger-1 (NHE-1) protein abundance in both ventricles in normal rats but not in hyperthyroid rats, independently of its effect on ventricular mass. Mammalian target of rapamycin (mTOR) protein abundance was not related to left or right ventricular mass in hyperthyroid or controls rats under normal or low salt conditions. Proteinuria was increased in hyperthyroid rats and attenuated by low salt intake. In this study, low salt intake produced an increase in right ventricular mass in normal and hyperthyroid rats. Changes in the left or right ventricular mass of control and hyperthyroid rats under low salt intake were not explained by the NHE-1 or mTOR protein abundance values observed. In hyperthyroid rats, low salt intake also slightly reduced BP and decreased HR, proteinuria, and water and sodium balances.     

Effect of thyroid state on susceptibility to oxidants and swelling of mitochondria from rat tissues

The effects of the thyroid state on oxidative damage, antioxidant capacity, susceptibility to in vitro oxidative stress and Ca(2+)-induced permeabilization of mitochondria from rat tissues (liver, heart, and gastrocnemious muscle) were examined. Hypothyroidism was induced by administering methimazole in drinking water for 15 d. Hyperthyroidism was elicited by a 10 d treatment of hypothyroid rats with triiodothyronine (10 micro g/100 g body weight). Mitochondrial levels of hydroperoxides and protein-bound carbonyls significantly decreased in hypothyroid tissues and were reported above euthroid values in hypothyroid rats after T(3) treatment. Mitochondrial vitamin E levels were not affected by changes of animal thyroid state. Mitochondrial Coenzyme Q9 levels decreased in liver and heart from hypothyroid rats and increased in all hyperthyroid tissues, while Coenzyme Q10 levels decreased in hypothyroid liver and increased in all hyperthyroid tissues. The antioxidant capacity of mitochondria was not significantly different in hypothyroid and euthyroid tissues, whereas it decreased in the hyperthyroid ones. Susceptibility to in vitro oxidative challenge decreased in mitochondria from hypothyroid tissues and increased in mitochondria from hyperthyroid tissues, while susceptibility to Ca(2+)-induced swelling decreased only in hypothyroid liver mitochondria and increased in mitochondria from all hyperthyroid tissues. The tissue-dependence of the mitochondrial susceptibility to stressful conditions in altered thyroid states can be explained by different thyroid hormone-induced changes in mitochondrial ROS production and relative amounts of mitochondrial hemoproteins and antioxidants. We suggest that susceptibilities to oxidants and Ca(2+)-induced swelling may have important implications for the thyroid hormone regulation of the turnover of proteins and whole mitochondria, respectively.     

Influence of thyroid disorders on kidney angiotensinase activity.

Thyroid disorders affect renal function, which involves changes in local renin angiotensin system (RAS). Angiotensin peptide levels in the tissue are regulated by the activity of several aminopeptidases (AP) known as angiotensinases. The nature and consequences of the thyroid-induced RAS changes are not completely understood. We investigated the relationship between thyroid status (hyper- and hypothyroidism) and several kidney AP actions involved in RAS control. We have determined fluorometrically soluble (SOL) and membrane-bound (M-B) alanylaminopeptidase (AlaAP), glutamylaminopeptidase (GluAP) and aspartylaminopeptidase (AspAP) activity using naphthylamide derivatives as substrates. Sprague-Dawley rats were divided into three groups--control, hyperthyroid, and hypothyroid. Hyperthyroidism was induced by daily subcutaneous injection of L-thyroxin (300 microg/kg/day). Hypothyroidism was induced by continuous administration of methimazole (0.03%) in drinking water. Hypothyroid animals demonstrated a significant increase in SOL and M-B GluAP activity in renal cortex and a decrease in M-B AlaAP compared to euthyroid rats. This result may suggest higher Ang III availability. In hyperthyroid animals, M-B AlaAP and M-B AspAP activity increased significantly, which may suggest increased Ang III to Ang IV metabolism and greater formation of Ang 2-10, respectively. In contrast, no differences were observed between euthyroid and hypothyroid animals for SOL and M-B AP activity in renal medulla. However, hyperthyroid animals demonstrated a significant decrease in SOL and M-B GluAP activity compared to euthyroid rats, which may suggest a greater availability of Ang II in renal medulla. Alterations in angiotensin metabolism may, in part, account for some changes in renal function during thyroid disorders.     

Upregulation of Slc39a10 gene expression in response to thyroid hormones in intestine and kidney

A novel zinc transporter has been purified and cloned from rat renal brush border membrane. This transporter was designated as Zip10 encoded by Slc39a10 gene and characterized as zinc importer. Present study documents the impact of thyroid hormones on the expression of Zip10 encoded by Slc39a10 gene in rat model of hypo and hyperthyroidism. Serum T(3) and T(4) levels were reduced significantly in hypothyroid rats whereas these levels were significantly elevated in hyperthyroid rats as compared to euthyroid rats thereby confirming the validity of the model. Kinetic studies revealed a significant increase in the initial and equilibrium uptake of Zn(++) in both intestinal and renal BBMV of hyperthyroid rats in comparison to hypothyroid and euthyroid rats. By RT-PCR, Slc39a10 mRNA expression was found to be significantly decreased in hypothyroid and increased in hyperthyroid as compared to euthyroid rats. These findings are in conformity with the immunofluorescence studies that revealed markedly higher fluorescence intensity at periphery of both intestinal and renal cells isolated from hyperthyroid rats as compared to hypothyroid and euthyroid rats. Higher expression of Zip10 protein in hyperthyroid group was also confirmed by western blot. These findings suggest that expression of zinc transporter protein Zip10 (Slc39a10) in intestine and kidney is positively regulated by thyroid hormones.     

Changes in lipid peroxidation and free radical scavengers in kidney of hypothyroid and hyperthyroid rats

Kidney weight was significantly decreased in hypothyroidism (induced by Na131I administration) and increased in hyperthyroidism (induced by thyroxine treatment) as compared to control in female Wistar rats. The tissue lipid peroxidation level remained unchanged in hyperthyroid rats but significantly increased in hypothyroid rats. Superoxide dismutase was decreased in both experimental groups but more so in hyperthyroid rats. Catalase was reduced significantly in hyperthyroid rats but remained unaffected in hypothyroid rats. Tissue glutathione peroxidase (GPx) activity was increased while reduced glutathione levels remained unaltered in both hypothyroid and hyperthyroid rats. Plasma GPx activity was significantly low in both the hypothyroid and hyperthyroid rats. The results suggest alterations in the oxidative stress in hypothyroid and hyperthyroid rat kidneys with concomitant changes of free radical scavengers.     

Mineralocorticoid receptor activation: a major contributor to salt-induced renal injury and hypertension in young rats

Excessive salt intake is known to preferentially increase blood pressure (BP) and promote kidney damage in young, salt-sensitive hypertensive human and animal models. We have suggested that mineralocorticoid receptor (MR) activation plays a major role in kidney injury in young rats. BP and urinary protein were compared in young (3-wk-old) and adult (10-wk-old) uninephrectomized (UNx) Sprague-Dawley rats fed a high (8.0%)-salt diet for 4 wk. The effects of the MR blocker eplerenone on BP and renal injury were examined in the high-salt diet-fed young UNx rats. Renal expression of renin-angiotensin-aldosterone (RAA) system components and of inflammatory and oxidative stress markers was also measured. The effects of the angiotensin receptor blocker olmesartan with or without low-dose aldosterone infusion, the aldosterone synthase inhibitor FAD286, and the antioxidant tempol were also studied. Excessive salt intake induced greater hypertension and proteinuria in young rats than in adult rats. The kidneys of young salt-loaded rats showed marked histological injury, overexpression of RAA system components, and an increase in inflammatory and oxidative stress markers. These changes were markedly ameliorated by eplerenone treatment. Olmesartan also ameliorated salt-induced renal injury but failed to do so when combined with low-dose aldosterone infusion. FAD286 and tempol also markedly reduced urinary protein. UNx rats exposed to excessive salt at a young age showed severe hypertension and renal injury, likely primarily due to MR activation and secondarily due to angiotensin receptor activation, which may be mediated by inflammation and oxidative stress.     

Effects of chronic inhibition of inducible nitric oxide synthase in hyperthyroid rats

We hypothesized that nitric oxide generated by inducible nitric oxide synthase (iNOS) may contribute to the homeostatic role of this agent in hyperthyroidism and may, therefore, participate in long-term control of blood pressure (BP). The effects of chronic iNOS inhibition by oral aminoguanidine (AG) administration on BP and morphological and renal variables in hyperthyroid rats were analyzed. The following four groups (n = 8 each) of male Wistar rats were used: control group and groups treated with AG (50 mg.kg(-1).day(-1), via drinking water), thyroxine (T4, 50 microg.rat(-1).day(-1)), or AG + T4. All treatments were maintained for 3 wk. Tail systolic BP and heart rate (HR) were recorded weekly. Finally, we measured BP (mmHg) and HR in conscious rats and morphological, plasma, and renal variables. T(4) administration produced a small BP (125 +/- 2, P < 0.05) increase vs. control (115 +/- 2) rats. AG administration to normal rats did not modify BP (109 +/- 3) or any other hemodynamic variable. However, coadministration of T4 and AG produced a marked increase in BP (140 +/- 3, P < 0.01 vs. T4). Pulse pressure and HR were increased in both T4- and T4 + AG -treated groups without differences between them. Plasma NOx (micromol/l) were increased in the T4 group (10.02 +/- 0.15, P < 0.05 vs. controls 6.1 +/- 0.10), and AG reduced this variable in T4-treated rats (6.81 +/- 0.14, P < 0.05 vs. T4) but not in normal rats (5.78 +/- 0.20). Renal and ventricular hypertrophy and proteinuria of hyperthyroid rats were unaffected by AG treatment. In conclusion, the results of the present paper indicate that iNOS activity may counterbalance the prohypertensive effects of T4.     

Upregulation of Slc39a10 gene expression in response to thyroid hormones in intestine and kidney

A novel zinc transporter has been purified and cloned from rat renal brush border membrane. This transporter was designated as Zip10 encoded by Slc39a10 gene and characterized as zinc importer. Present study documents the impact of thyroid hormones on the expression of Zip10 encoded by Slc39a10 gene in rat model of hypo and hyperthyroidism. Serum T₃ and T₄ levels were reduced significantly in hypothyroid rats whereas these levels were significantly elevated in hyperthyroid rats as compared to euthyroid rats thereby confirming the validity of the model. Kinetic studies revealed a significant increase in the initial and equilibrium uptake of Zn⁺⁺ in both intestinal and renal BBMV of hyperthyroid rats in comparison to hypothyroid and euthyroid rats. By RT-PCR, Slc39a10 mRNA expression was found to be significantly decreased in hypothyroid and increased in hyperthyroid as compared to euthyroid rats. These findings are in conformity with the immunofluorescence studies that revealed markedly higher fluorescence intensity at periphery of both intestinal and renal cells isolated from hyperthyroid rats as compared to hypothyroid and euthyroid rats. Higher expression of Zip10 protein in hyperthroid group was also confirmed by western blot. These findings suggest that expression of zinc transporter protein Zip10 (Slc39a10) in intestine and kidney is positively regulated by thyroid hormones.     

Effects of chronic treatment with 7-nitroindazole in hyperthyroid rats

This study analyzed the contribution of neuronal nitric oxide synthase (nNOS) to the hemodynamic manifestations of hyperthyroidism. The effects on hyperthyroid rats of the chronic administration of 7-nitroindazole (7-NI), an inhibitor of nNOS, were studied. Six groups of male Wistar rats were used: control, 7-NI (30 mg.kg-1.day-1 by gavage), T(4)50, T(4)75 (50 or 75 microg thyroxine.rat-1.day-1, respectively), T(4)50+7-NI, and T(4)75+7-NI. All treatments were maintained for 4 wk. Body weight, tail systolic blood pressure (SBP), and heart rate (HR) were recorded weekly. Finally, SBP, pulse pressure (PP), and HR were measured in conscious rats, and morphological, metabolic, plasma, and renal variables were determined. Expression of nNOS in the hypothalamus of T(4)75 and control rats was analyzed by Western blot analysis. The response of mean arterial pressure (MAP) to pentolinium (10 mg/kg iv) was used to evaluate the sympathetic contribution to BP in T(4)75 and T(4)75+7-NI rats. T(4) produced an increased hypothalamic nNOS expression and dose-related increases in blood pressure (BP), HR, and PP vs. control rats. 7-NI did not modify BP or any other hemodynamic variable in normal rats. However, 7-NI produced a marked reduction in BP, HR, PP, and food and water intake in both hyperthyroid groups and improved creatinine clearance in the T(4)75 group. Pentolinium produced a greater MAP decrease in the T(4)75+7-NI than in the T(4)75 group. In conclusion, administration of 7-NI attenuates the hemodynamic and metabolic manifestations of hyperthyroidism, suggesting that nNOS contributes to the hyperdynamic circulation of this endocrine disease by modulating sympathetic activity.     

Renal endothelin in chronic angiotensin II hypertension

To determine the influence of chronic ANG II infusion on urinary, plasma, and renal tissue levels of immunoreactive endothelin (ET), ANG II (65 ng/min) or saline vehicle was delivered via osmotic minipump in male Sprague-Dawley rats given either a high-salt diet (10% NaCl) or normal-salt diet (0.8% NaCl). High-salt diet alone caused a slight but not statistically significant increase (7 +/- 1%) in mean arterial pressure (MAP). MAP was significantly increased in ANG II-infused rats (41 +/- 10%), and the increase in MAP was significantly greater in ANG II rats given a high-salt diet (59 +/- 1%) compared with the increase observed in rats given a high-salt diet alone or ANG II infusion and normal-salt diet. After a 2-wk treatment, urinary excretion of immunoreactive ET was significantly increased by approximately 50% in ANG II-infused animals and by over 250% in rats on high-salt diet, with or without ANG II infusion. ANG II infusion combined with high-salt diet significantly increased immunoreactive ET content in the cortex and outer medulla, but this effect was not observed in other groups. In contrast, high-salt diet, with or without ANG II infusion, significantly decreased immunoreactive ET content within the inner medulla. These data indicate that chronic elevations in ANG II levels and sodium intake differentially affect ET levels within the kidney and provide further support for the hypothesis that the hypertensive effects of ANG II may be due to interaction with the renal ET system.     

High dietary salt intake increases carotid blood pressure and wave reflection in normotensive healthy young men

Dietary salt intake is associated with high brachial blood pressure (BP) and increased risk of cardiovascular disease. We investigated whether changes in dietary salt intake are associated with changes in central BP and wave reflection in healthy volunteers. Ten healthy normotensive male volunteers (22-40 yr) participated in a 6-wk double-blind randomized crossover study to compare a low-dietary salt intake (60-80 mmol sodium/day) with a high-salt intake (low salt intake supplemented with 128 mmol sodium/day) on central BP and wave reflection. Brachial and carotid BP, carotid blood flow velocity, forward (P(f)) and backward (P(b)) pressure, wave intensity, body weight, and urinary electrolyte excretion were measured at the end of each crossover period. High salt intake significantly increased carotid systolic BP [98 (SD 11) vs. 91 mmHg (SD 13), P < 0.01] and increased wave reflection [ratio of backward to forward pressure (P(b)/P(f)) 0.13 (SD 0.02) vs. 0.11 (SD 0.03), P = 0.04] despite only small effects on brachial BP [114 (SD 9) vs. 112 mmHg (SD 6), P = 0.1]. Urinary sodium excretion and body weight were also increased following high salt intake. High salt intake disproportionately increases central BP compared with brachial BP as a result of enhanced wave reflection. These effects may contribute to the adverse effect of high dietary salt intake on the risk of cardiovascular disease.     

Thyroid hormone-induced haemoglobin changes and antioxidant enzymes response in erythrocytes

Thyroid hormones modulate haemoglobin and reactive oxygen species (ROS) production, leading to antioxidant changes. This study evaluated the antioxidant response to ROS in erythrocytes in hypothyroid and hyperthyroid rats. Wistar rats were divided into four groups: control; hyperthyroid (T4-12 mg 1(-1) in drinking water); sham operated (simulation of thyroidectomy); and hypothyroid (thyroidectomized). Four weeks after, blood was collected and haemoglobin and T(4) levels, lipid peroxidation (LPO), protein oxidation, superoxide dismutase (SOD), catalase (CAT) , glutathione S-transferase (GST) and glutathione peroxidase (GPx) activities, and total radical antioxidant potential (TRAP) were measured. SOD, CAT and GST immunocontent was evaluated. Haemoglobin levels were increased in hyperthyroid erythrocytes. LPO and carbonyls were augmented (65% and 55%, respectively) in hyperthyroid and reduced (31% and 56%, respectively) in hypothyroid group. SOD and CAT activities have not changed, as well as CAT immunocontent. TRAP was diminished in both hyperthyroid and hypothyroid groups (36% and 37%, respectively). GST activity and immunocontent, as well as GPx activity, were increased in hyper and hypothyroid rats. The data suggest that thyroid hormone changes determine ROS concentration changes and decrease of some antioxidant defences that would lead to a compensatory answer of the GST and GPx enzymes, which could be consider as credible biomarkers.     

Thyroid hormone may induce changes in the concentration of the mitochondrial calcium uniporter

We explored the possibility that the hormone 3,3',5-tri-iodothyronine can regulate the biosynthesis of the mitochondrial calcium uniporter. To meet this objective experiments on Ca(2+) transport, and binding of the specific inhibitor Ru(360) were carried out in mitochondria isolated from euthyroid, hyperthyroid and hypothyroid rats. It was found that V(max) for Ca(2+) transport increased from 11.67+/-0.8 in euthyroid to 14.36+/-0.44 in hyperthyroid, and decreased in hypothyroid mitochondria to 8.62+/-0.63 nmol Ca(2+)/mg/s. Furthermore, the K(i) for the specific inhibitor Ru(360), depends on the thyroid status, i.e. 18, 19 and 13 nM for control, hyper- and hypothyroid mitochondria, respectively. In addition, the binding of 103Ru(360) was increased in hyperthyroid and decreased in hypothyroid mitochondria. Scatchard analysis for the binding of 103Ru(360) showed the following values: 28, 40 and 23 pmol/mg for control, hyper- and hypothyroid mitochondria, respectively. The K(d) for 103Ru(360) was found to be 30.39, 37.03 and 35.71 nM for controls, hyper- and hypothyroid groups, respectively. When hypothyroid rats were treated with thyroid hormone, mitochondrial Ca(2+) transport, as well as 103Ru(360) binding, reached similar values to those found for euthyroid mitochondria.     

Atrial natriuretic polypeptide in atria and plasma in experimental hyperthyroidism and hypothyroidism

To investigate the involvement of thyroid hormone on the release of atrial natriuretic polypeptide (ANP), we have measured immunoreactive ANP in the atria and plasma of experimental hyperthyroid and hypothyroid rats. Plasma ANP was higher (p less than 0.05) in hyperthyroid rats and was lower (p less than 0.05) in hypothyroid rats than in euthyroid rats. ANP content and concentration in the atria were lower (p less than 0.01) in hyperthyroid rats than in hypothyroid rats. An inverse correlation was found between the plasma ANP and ANP concentration in the atria (n = 15, r = 0.60, p less than 0.01). The results indicate an increased systemic release of ANP from the atria in hyperthyroidism and a decreased systemic release in hypothyroidism.     

Influence of hyper‐ and hypothyroidism on lipid peroxidation,unsaturation of phospholipids,glutathione system and oxidative damage to nuclear and mitochondrial DNA in mice skeletal muscle

While the biochemical literature on free radical metabolism is extensive, there is little information on the endocrine control of tissue oxidative stress, and in the case of thyroid hormones it is mainly limited to liver tissue and to short-term effects on a few selected biochemical parameters. In this investigation, chronic hypothyroidism and hyperthyroidism were successfully induced in mice, and various oxidative-stress-related parameters were studied in skeletal muscle. In vivo and in vitro lipid peroxidation significantly increased in hyperthyroidism and did not change in the hypothyroid state. The fatty acid composition of the major phospholipid classes was affected by thyroid hormones, leading to a significant decrease in total fatty acid unsaturation both in hypothyroid and hyperthyroid muscle in phosphatidylcholine and phosphatidylethanolamine fractions. In cardiolipin, however, the double bond content significantly increased as a function of thyroid status, leading to a 2.7 fold increase in the peroxidizability index from euthyroid to hyperthyroid muscle. Cardiolipin content was also directly and significantly related to thyroid state across the three groups. Glutathione system was not modified by thyroid state. The oxidative damage marker 8-oxo-7,8-dihydro-2'-deoxyguanosine did not change in mitochondrial DNA, and decreased in genomic DNA both in hypothyroid and hyperthyroid muscle. The results indicate that chronic alterations in thyroid status specially affect oxidative damage to lipids in skeletal muscle, with a probably stronger effect on mitochondrial membranes, whereas the cytosolic redox potential and DNA are better protected possibly due to homeostatic compensatory reactions on the long-term.     

Effects of thyroid state and fasting on the concentrations of CoA and malonyl-CoA in rat liver

Liver CoA is markedly higher in hyperthyroid rats as compared to hypothyroid rats, and in fasted rats as compared to fed rats. In hyperthyroid rats the CoA is increased mainly in the cytosol, while in fasted rats the increase is mainly in the mitochondria. Malonyl-CoA is markedly higher in hypothyroid rats than in euthyroid and hyperthyroid rats. With fasting, malonyl-CoA drops 80-85% in all thyroid states. These findings are discussed in relation to the regulation of fatty acid oxidation in the liver.