Subclinical Hypothyroidism Rats Research Articles

Thyroid-stimulating hormone may participate in insulin resistance by activating toll-like receptor 4 in liver tissues of subclinical hypothyroid rats.

Thyroid-stimulating hormone (TSH) is an independent risk factor of and closely associated with metabolic disorders. In the present study, we explored the potential mechanism and adverse effects of TSH on insulin resistance in the liver of subclinical hypothyroidism models in vivo. The mean glucose infusion rate (GIR), free fatty acids (FFAs), the homeostatic model assessment for insulin resistance (HOMA-IR), fasting plasma insulin (FINS), the TLR4 signal pathway and its intracellular negative regulator-toll-interacting protein (Tollip), and the modulators of insulin signaling were evaluated. Compared to the normal control group (NC group), the subclinical hypothyroidism rat group (SCH group) showed decreases in GIR and increases in FFAs, FINS, and HOMA-IR. The levels of TLR4 and of its downstream molecules like p-NF-κB, p-IRAK-1, IL-6 and TNF-α were evidently higher in the SCH group than in the NC group. Conversely, the level of Tollip was significantly lower in the SCH group than in the NC group. Compared to the NC group, the levels of phosphorylated IRS-1-Tyr and GLUT2 were decreased in the SCH group. Macrophage infiltration was higher in the SCH group than in the NC group. TSH may participate in aggravating inflammation by increasing macrophage infiltration; furthermore, it may activate the TLR4-associated inflammatory signaling pathway, thus interfering with insulin signals in liver tissues. Targeting TSH may have therapeutic benefits against metabolic disorders.

Effect of Bushen Antai recipe on pyroptosis mechanism of subclinical hypothyroidism decidual cells in early pregnancy.

Subclinical hypothyroidism (SH) is common during pregnancy. It is not clear whether decidual cells in SH undergo pyroptosis during pregnancy. This study aimed to investigate the possible mechanism of Bushen Antai recipe (BAR) in the treatment of SH in early pregnancy and the relationship between SH during pregnancy and decidual cell pyroptosis through a rat model. A total of 60 female rats were divided into control group, model group, levothyroxine (L-T4) group, low-dose BAR group (6 g/kg), medium-dose BAR group (12 g/kg), and high-dose BAR group (24 g/kg). The control group underwent pseudothyroidectomy, while the remaining groups established nonpregnant SH rat models. Except for the blank control group, rats were successfully established with SH models during pregnancy. The control group and the model group were treated with saline or BAR. The animals were sacrificed 12 hours after the last administration. The levels of serum thyroid-stimulating hormone (TSH), free thyroxine (FT4), interleukin-1β (IL-1β), and interleukin-18 (IL-18) were detected by enzyme-linked immunosorbent assay (ELISA). Western blot was used to detect the expression of decidual nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) inflammasome, Caspase-1, and porin family proteins. There was no significant difference in serum FT4 among groups (P>0.05). Compared with the control group, serum TSH, IL-1β, IL-18, and NLRP3, Caspase-1, and gasdermin D (GSDMD) proteins in the decidua of the model group were significantly increased (P<0.05). Compared with the model group, the L-T4 group and the high-dose BAR group could significantly decrease the levels of serum TSH, IL-1β, IL-18, and NLRP3, Caspase-1, and GSDMD in decidual tissue (P<0.05). The medium dose of BAR could significantly decrease the levels of TSH, NLRP3, Caspase-1, and GSDMD (P<0.05), and the low dose group of BAR significantly decreased the levels of TSH, NLRP3, and GSDMD (P<0.05). Among them, the high-dose group of BAR had the best reducing effect on IL-18, NLRP3, Caspase-1, and GSDMD. The decidual cells of SH rats in early pregnancy underwent pyroptosis with a high inflammatory response. BAR could improve TSH level in SH during pregnancy, inhibit decidual cell pyroptosis, and reduce the expression of inflammatory factors.

Subclinical hypothyroidism in pregnancy rats impaired offspring's spatial learning and memory and the cerebellar development

Subclinical hypothyroidism (SCH) is a very common preclinical condition during pregnancy. The adverse effect of maternal clinical hypothyroidism (CH) on the nervous system development of offspring is beyond doubt, but it is still controversial in SCH. The aim of this study was to investigate whether spatial learning and memory ability of offspring is inhibited in SCH rat model and its possible mechanism. 45 Wistar female rats were randomly divided into SCH, CH and control (CON) groups, which were induced by semi-thyroid electrocauterization, total thyroidectomy and sham operation, respectively. Rat pups were sacrificed at embryonic day 14 (E14), E18, postnatal day 1 (P1), P3, and P10, and pups' cerebellar tissues were collected. The proliferation, differentiation and migration of cerebellar cells were observed, and RNA level of the thyroid hormone receptor α (TRα) and TRβ in the cerebellum was detected by real-time PCR, respectively. Morris Water Maze (MWM) test was performed to detect the spatial learning and memory ability of pups at P40. Our data indicated that maternal SCH will significantly extend the offspring's escape latency time, and pups perform worse in the spatial probe test compared with the CON group. Except for E14, the proliferation of pups' cerebellar granule cells (GCs), and the migration of pups' Purkinje cells (PCs) in the SCH group was significantly inhibited compared with that in the CON group at other time points (P < 0.05 or P < 0.01), and the differentiation of cerebellar astrocytes (As) in SCH group was higher than that in CON group at P3 and P10. Except for E14, the expression of TRα mRNA in SCH group was significantly lower than that in CON group (P < 0.05 or P < 0.01). And the difference of the differentiation of As and the spatial learning and memory between SCH and CH groups was not statistically significant. Our findings suggested that SCH during pregnancy nuisances the offspring's spatial learning and memory. It may be related to the decrease of the expression of TRα in cerebellum, which may further inhibit the proliferation of GCs and the migration of PCs, and increase the differentiation of As.

Maternal Subclinical Hypothyroidism in Rats Impairs Spatial Learning and Memory in Offspring by Disrupting Balance of the TrkA/p75NTR Signal Pathway

Maternal subclinical hypothyroidism (SCH) during pregnancy can adversely affect the neurodevelopment of the offspring. The balance of nerve growth factor (NGF)-related tropomyosin receptor kinase A/p75 neurotrophin receptor (TrkA/p75NTR) signaling in the hippocampus is important in brain development, and whether it affects cognitive function in maternal SCH’s offspring is not clear. In this study, we found that compared with the control (CON) group, expression of proliferation-related proteins [NGF, p-TrkA, phospho-extracellular signal-regulated kinase 1/2 (p-ERK1/2) and phospho-cAMP response element-binding protein (p-CREB)] decreased in the hippocampus of the offspring in the SCH group, overt hypothyroidism (OHT) group, and the group with levothyroxine (L-T4) treatment for SCH from gestational day 17 (E17). In contrast, expression of apoptosis-related proteins [pro-NGF, p75NTR, phospho-C-Jun N-terminal kinase (p-JNK), p53, Bax and cleaved caspase-3] was increased. The two groups with treatment with L-T4 for SCH from E10 and E13, respectively, showed no significant difference compared with the CON group. L-T4 treatment enhanced relative expression of NGF by increasing NGF/proNGF ratio in offspring from maternal SCH rats. In conclusion, L-T4 treatment for SCH from early pregnancy dramatically ameliorated cognitive impairment via TrkA/p75NTR signaling, which involved activation of the neuronal proliferation and inhibition of neuronal apoptosis in SCH rats’ offspring.

Early L-T4 intervention improves fetal heart development in pregnant rats with subclinical hypothyroidism rats by activating BMP4/Smad4 signaling pathway

BackgroundIt is unclear whether the offspring of subclinical hypothyroidism (SCH) pregnant rats still have abnormal cardiac development, and whether early intervention with L-T4 can improve the abnormality of these offspring. Therefore, the aim of this study was to investigate the effect of early L-T4 intervention on the heart development of offspring of SCH pregnant rats and its possible molecular mechanism.MethodsEighty female Wistar rats were randomly divided into Sham group (placebo control), SCH group, LT4-E10 group (L-T4 treatment started on the 10th day of gestation), and LT4-E13 group (L-T4 treatment started on the 13th day of gestation). Each group was further divided into E16 (16th day of gestation), E18 (18th day of gestation), P5 (5th day postnatal day), and P10 (10th day postnatal day) subgroups. The levels of serum TT4 and TSH, the ratio of heart weight to body weight of offspring rats, the expression of metabolic enzymes, and the histopathology of cardiomyocytes were determined. To elucidate the effects of L-T4 on cardiac development of offspring of SCH pregnant rats, the expression levels of GATA4, Nkx2–5 and proteins involved in BMP4/Smad4 signaling pathway were detected by immunohistochemistry, real time quantitative polymerase chain reaction and Western blotting to elucidate the molecular mechanism of L-T4 regulating the heart development of the offspring of SCH pregnant rats.ResultsCompared with Sham group, serum TSH was significantly increased in SCH pregnant rats. Moreover, early L-T4 intervention significantly reduced the levels of serum TSH. Compared with the offspring in the SCH group, early L-T4 intervention significantly increased the heart weight, heart weight to body weight ratio, the activities of succinate dehydrogenase (SDH), Na+/K+-ATPase and Ca2+-ATPase, but reduced myocardial cell shrinkage and nuclear staining, hyperemia/congestion and vacuolar degeneration. In addition, early L-T4 intervention not only significantly increased the mRNA and protein expression of Gata4 and Nkx2–5, but also increased the protein expression involved in BMP4/Smad4 signal pathway in myocardium of the offspring of SCH pregnant rats.ConclusionsEarly L-T4 intervention can regulate the cardiac development of the offspring of SCH pregnant rats by activating BMP4/Smad4 signaling pathway and increasing the expression of Gata4 and Nkx2–5 proteins.

Thyrotropin Regulates eNOS Expression in the Endothelium by PGRN Through Akt Pathway.

To investigate the expression of endothelial nitric oxide synthase (eNOS) and nitric oxide (NO) in the aorta of subclinical hypothyroidism (SCH) rat model. The mechanisms underlying thyrotropin (TSH) affecting eNOS and PGRN expression in human umbilical vein endothelial cells (HUVECs) cultured in vitro were investigated. In the current study, SCH rat models were established by the administration of L-T4 injection after thyroidectomy in Wistar rats, as opposed to that in the normal and clinical hypothyroidism (CH) groups. The concentrations of NO (pmol/μL) in the SCH and CH groups were significantly lower than that in the normal group (40.8 ± 7.6 and 32.9 ± 10.8 vs. 51.2 ± 12.1, P < 0.05). However, the expression level of eNOS is increased significantly (P < 0.05) in both SCH and CH groups; a similar result was observed for the PGRN protein. In cultured HUVECs, TSH can also up-regulate the expression of eNOS; however, it is accompanied by a reduced concentration of NO and increased level of superoxide anion, thereby indicating uncoupled eNOS. As eNOS is increased, we found that Akt in HUVECs were upregulated by TSH, as well as PGRN expression. While inhibiting the expression of PGRN in HUVECs using siRNA, the expression of eNOS, as well as Akt were also inhibited. In conclusion, SCH can induce vascular endothelial dysfunction in rats, and PGRN participated in the process of TSH-induced expression of Akt/eNOS in the endothelium.

Adipokine expression and endothelial function in subclinical hypothyroidism rats.

The purpose of our study was to observe adipokine expression and endothelial function in subclinical hypothyroidism (sHT) rats and to determine whether levothyroxine (LT4) treatment affects these changes. Sixty-five male Wistar rats were randomly divided into five groups: the control group; sHT A, B and C groups and the sHT + T4 group. The sHT rats were induced by methimazole (MMI) and the sHT + T4 rats were administered LT4 treatment after 8 weeks of MMI administration. Thyroid function and lipid levels were measured using radioimmunoassays and enzymatic colorimetric methods, respectively. Serum adiponectin (APN), chemerin, TNF-α, endothelin (ET-1) and nitric oxide (NO) levels were measured using ELISA kits and a nitric-reductive assay. The expression of APN, chemerin and TNF-α in visceral adipose tissue (VAT) was measured in experimental rats using RT-PCR and Western blotting. Hematoxylin–eosin (HE) staining was used to observe changes in adipose tissue. The sHT rats had significantly higher levels of thyroid-stimulating hormone (TSH), TNF-α, chemerin, ET-1, total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) and lower levels of APN and NO than those in control and sHT + T4 rats. Based on Pearson correlation analysis, the levels of chemerin, TNF-α, ET-1, LDL-C, TC and triglyceride (TG) were positively correlated with TSH, but APN and NO levels were negatively correlated with TSH. These findings demonstrated that high TSH levels contribute to the changes of adipokines and endothelial dysfunction in sHT, but LT4 treatment ameliorates those changes.

Effect of Thyrotropin on Osteopontin, Integrin αvβ3, and VCAM-1 in the Endothelium via Activation of Akt.

Numerous epidemiological studies have shown that subclinical hypothyroidism (SCH) can impair endothelial function and cause dyslipidemia. Studies have evaluated the effects of thyroid stimulating hormone (TSH) on endothelial cells, but the mechanism underlying the proatherosclerotic effect of increased TSH levels remains unclear. In the present study, SCH rat models were established in thyroidectomized Wistar rats that were given l-T4 daily. The results showed that in vivo, the expression of osteopontin (OPN) vascular cell adhesion molecule (VCAM-1), and levels of integrin αvβ3 in the aortic tissue in SCH and Hypothyroidism (CH) groups was higher than in the control group. However, the effect in the SCH group was higher than in the CH group. In vitro, results showed that different concentration and time gradients of TSH stimulation could increase the expression of OPN, VCAM-1, and integrin αvβ3, and this was accompanied by extracellular signal regulated kinase 1/2 (Erk1/2) and Akt activation in human umbilical vein endothelial cells (HUVECs). TSH induced elevation of these proatherosclerotic factors was partially suppressed by a specific Akt inhibitor but not by a specific Erk inhibitor. Findings suggested that the endothelial dysfunction caused by SCH was related to increased proatherosclerotic factors induced by TSH via Akt activation.

Resveratrol Ameliorates the Anxiety- and Depression-Like Behavior of Subclinical Hypothyroidism Rat: Possible Involvement of the HPT Axis, HPA Axis, and Wnt/β-Catenin Pathway.

Metabolic disease subclinical hypothyroidism (SCH) is closely associated with depression-like behavior both in human and animal studies, and our previous studies have identified the antidepressant effect of resveratrol (RES) in stressed rat model. The aim of this study was to investigate whether RES would manifest an antidepressant effect in SCH rat model and explore the possible mechanism. A SCH rat model was induced by hemi-thyroid electrocauterization, after which the model rats in the RES and LT4 groups received a daily intragastric injection of RES at the dose of 15 mg/kg or LT4 at the dose of 60 μg/kg for 16 days. The rats’ plasma concentrations of thyroid hormones were measured. Behavioral performance and hypothalamic–pituitary–adrenal (HPA) activity were evaluated. The protein expression levels of the Wnt/β-catenin in the hippocampus were detected by western blot. The results showed that RES treatment downregulated the elevated plasma thyroid-stimulating hormone concentration and the hypothalamic mRNA expression of thyrotropin-releasing hormone in the SCH rats. RES-treated rats showed increased rearing frequency and distance in the open-field test, increased sucrose preference in the sucrose preference test, and decreased immobility in the forced swimming test compared with SCH rats. The ratio of the adrenal gland weight to body weight, the plasma corticosterone levels, and the hypothalamic corticotrophin-releasing hormone mRNA expression were reduced in the RES-treated rats. Moreover, RES treatment upregulated the relative ratio of phosphorylated-GSK3β (p-GSK3β)/GSK3β and protein levels of p-GSK3β, cyclin D1, and c-myc, while downregulating the relative ratio of phosphorylated-β-catenin (p-β-catenin)/β-catenin and expression of GSK3β in the hippocampus. These findings suggest that RES exerts anxiolytic- and antidepressant-like effect in SCH rats by downregulating hyperactivity of the HPA axis and regulating both the HPT axis and the Wnt/β-catenin pathway.

SERUM LIPOCALIN-2 PREDICTS CARDIOVASCULAR MORBIDITY RISKS IN SUBCLINICAL HYPOTHYROID RAT MODEL

Background: Subclinical hypothyroidism (SCH) and its associations with cardiovascular disease (CVD) remain controversial. Previous studies revealed that lipocalin -2 (LCN2) associated with increased cardiovascular mortality risk. Objective: Evaluation of the serum level of LCN2 and its association with some cardiovascular risk factors in SCH rat model. Material and methods: Twenty four adult male albino rats weighing 150-170 g were used and divided into two equal groups: control euthyroid group and SCH-induced group where SCH was induced by administration of methimazole (MMI) (10 mg/kg body weight) by gavage once daily for 3 months. Groups were compared for body mass index (BMI), serum thyroid-stimulating hormone (TSH), triiodothyronine (T3) thyroxin (T4), glucose, insulin, lipid profiles, C reactive protein (CRP), LCN2 ,Homeostasis model assessment of insulin resistance (HOMA-IR), atherogenic index ,systolic (SBP) and diastolic blood pressure (DBP). Results: SCH induced group showed significantly higher mean serum levels of TSH, LCN2, insulin, total cholesterol, low density lipoprotein - cholesterol (LDL- C), triglycerides (TG), CRP than that of control. Also, BMI, calculated HOMA, atherogenic index, SBP and DBP were significantly higher than that of control and all these parameters were positively correlated to LCN2. However, high density lipoproteins- cholesterol (HDL- c) levels were significantly lower than that of control group and negatively correlated with serum LCN2 levels. Conclusion: Serum lipocalin-2 may be associated and can predict cardiovascular risk factors in SCH.

Endothelial Functioning and Hemodynamic Parameters in Rats with Subclinical Hypothyroid and the Effects of Thyroxine Replacement.

ObjectiveSubclinical hypothyroidism (SCH) and its associations with atherosclerosis (AS) and cardiovascular disease remain controversial. The purpose of our study was to observe changes in endothelial functioning and hemodynamics in rats with SCH and to determine whether L-thyroxine (L-T4) administration affects these changes.MethodsIn total, sixty male Wistar rats were randomly divided into the following three groups with 20 rats each: control euthyroid rats, SCH rats and SCH rats that had been treated with thyroxine (SCH+T4). The SCH rats were induced by administration of 10 mg.kg-1.d-1 methimazole (MMI) once daily by gavage for 3 months. The SCH+T4 rats were administered the same dose of MMI for three months in addition to 2 μg.kg-1.d-1 L-T4 once daily by gavage after 45 days of MMI administration. The control rats received physiological saline via gavage.ResultsThe SCH group had significantly higher thyroid-stimulating hormone (TSH), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), and endothelin (ET) levels and a lower nitric oxide (NO) level than the control and SCH+T4 groups. The tail and carotid artery blood pressures, left ventricular systolic pressure, heart rate and aorta ventralis blood flow were significantly lower in the SCH group than in the control and SCH+T4 groups. ACH treatment caused concentration-dependent relaxation, which was reduced in the SCH arteries compared with the control and SCH+T4 arteries. Histopathological examination revealed the absence of pathological changes in the SCH rat arteries.ConclusionsThese findings demonstrate that L-T4 treatment ameliorates endothelial dysfunction and hemodynamic changes in SCH rats.

Depression-like behavior in subclinical hypothyroidism rat induced by hemi-thyroid electrocauterization

The purpose of this study was to investigate the depression-like behavior performances of subclinical hypothyroidism (SCH) rat. SCH rat model was induced by hemi-thyroid electrocauterization, and the behavior performances were measured by sucrose preference test, force swimming test (FST), and tail suspension test (TST). SCH rat model was established successfully by hemi-thyroid electrocauterization. In the behavior tasks, SCH rats displayed depression-like behavior were indicated as a significant elevation of immobility time in both the TST and FST, though the sucrose preference was not significantly decreased. The index of left adrenal cortex in both SCH and clinical hypothyroidism (CH) group significantly increased, and many large lipid vacuoles were observed in the zona fasciculata cells. The serum corticosterone concentration and hypothalamic corticotropin-releasing hormone mRNA expression 2 h after behavior test was markedly up-regulated in CH rats, but not SCH rats, indicated that SCH induced a less impairment of HPA axis than CH did. The important finding of this study was that the concentration of hippocampal T3 was lower in SCH group than that of the sham group. Furthermore, the results of Pearson correlation test showed that the immobility behaviors in TST and FST were both negatively correlated with hippocampal T3 concentration. Taking together, our results indicated that SCH could result in depression-like behavior, accompanied with subtle hyperactivity of HPA axis. The reduced hippocampal T3 prior to the reduction of thyroid hormone in serum might be taken as an early sign of hippocampus impairment in the progression from SCH to CH.

Impaired Learning and Memory Performance in a Subclinical Hypothyroidism Rat Model Induced by Hemi‐Thyroid Electrocauterisation

It is well known that clinical hypothyroidism (CH) can induce cognitive deficits, and the decision to start treatment for CH with thyroxine is usually straightforward. However, the relationship of cognition dysfunction with subclinical hypothyroidism (SCH) is inconsistent, and the decision concerning the need to treat SCH is controversial. In the present study, we induced a SCH rat model by hemi-thyroid electrocauterisation; then employed a serial of behavioural tests, including a beam balance, open field task and Morris water maze (MWM), to investigate the behaviour performance of SCH rats; and finally explored the protein expression of phosphorylated extracellular signal-regulated kinase (ERK)1/2 in the hippocampus by western blotting. The results demonstrated that hemi-thyroid electrocauterised rats had an elevated plasma thyrotrophin-stimulating hormone (TSH) level, with normal free thyroxine (fT4) and triiodothyronine (T3) concentrations, which defines SCH in humans. If rat SCH is diagnosed according to measurements of both plasma TSH higher than 97.5 percentile for the sham group and fT4 in the range 2.5-97.5 percentile for the sham group, the success rate of SCH modelling was 66.6%. SCH decreased exploratory behaviour but did not affect motor function in rats, showing a negative correlation of exploratory behaviour with plasma TSH concentration. Moreover, SCH rats displayed an impairment of learning and memory ability in the MWM task, with a longer escape latency in the acquisition phase and a shorter duration in the target quadrant in the test phase compared to that of sham rats. The mechanism for this might be related to the increased plasma TSH concentration, the decreased hippocampal T3 level and the enhanced expression of phosphorylated ERK1/2 in the hippocampus. The results of the present study, together with the results obtained in other studies, suggest that treatment is necessary for SCH.