Thyrotropin-releasing hormone overexpression induces structural changes of the left ventricle in the normal rat heart.

Thyrotropin-releasing hormone (TRH) hyperactivity has been observed in the left ventricle of spontaneously hypertensive rats. Its long-term inhibition suppresses the development of hypertrophy, specifically preventing fibrosis. The presence of diverse systemic abnormalities in spontaneously hypertensive rat hearts has raised the question of whether specific TRH overexpression might be capable of inducing structural changes in favor of the hypertrophic phenotype in normal rat hearts. We produced TRH overexpression in normal rats by injecting into their left ventricular wall a plasmid driving expression of the preproTRH gene (PCMV-TRH). TRH content and expression of preproTRH, collagen type III, brain natriuretic peptide, ß-myosin heavy chain, Bax-to-Bcl-2 ratio, and caspase-3 were measured. The overexpression maneuver was a success, as we found a significant increase in both tripeptide and preproTRH mRNA levels in the PCMV-TRH group compared with the control group. Immunohistochemical staining against TRH showed markedly positive brown signals only in the PCMV-TRH group. TRH overexpression induced a significant increase in fibrosis, evident in the increase of collagen type III expression accompanied by a significant increase in extracellular matrix expansion. We found a significant increase in brain natriuretic peptide and ß-myosin heavy chain expression (recognized markers of hypertrophy). Moreover, TRH overexpression induced a slight but significant increase in myocyte diameter, indicating the onset of cell hypertrophy. We confirmed the data "in vitro" using primary cardiac cell cultures (fibroblasts and myocytes). In conclusion, these results show that a specific TRH increase in the left ventricle induced structural changes in the normal heart, thus making the cardiac TRH system a promising therapeutic target.

Cardiac thyrotropin-releasing hormone mediates left ventricular hypertrophy in spontaneously hypertensive rats.

Local thyrotropin-releasing hormone (TRH) may be involved in cardiac pathophysiology, but its role in left ventricular hypertrophy (LVH) is still unknown. We studied whether local TRH is involved in LVH of spontaneously hypertensive rats (SHR) by investigating TRH expression and its long-term inhibition by interference RNA (TRH-iRNA) during LVH development at 2 stages (prehypertrophy and hypertrophy). SHR and their control rats (WKY) were compared. Cardiac hypertrophy was expressed as heart/total body weight (HW/BW) ratio. TRH content (radioimmuno assay), preproTRH, TRH receptor type I, brain natriuretic peptide (BNP), and collagen mRNA expressions (real-time polymerase chain reaction) were measured. For long-term inhibition of TRH, TRH-iRNA was injected into the left ventricle (LV) wall for 8 weeks. Hearts were processed for morphometric studies and immunohistochemical analysis using antibodies against α-smooth muscle actin and collagen type III. LV preproTRH-mRNA abundance was similar in both strains at 7 weeks of age. At the hypertrophic stage (18 weeks old), however, there was a 15-fold increase in SHR versus WKY, consistent with a significant increase in tripeptide levels and the expression of its receptor. Specific LV-TRH inhibition at the prehypertensive stage with TRH-iRNA, which decreased >50% preproTRH expression and tripeptide levels, prevented LVH development as shown by the normal HW/BW ratio observed in TRH-iRNA-treated SHR. In addition, TRH-iRNA impeded the increase in BNP and type III collagen expressions and prevented the increase in cardiomyocyte diameter evident in mismatch iRNA-treated adult SHR. These results show for the first time that the cardiac TRH system is involved in the development of LVH in SHR.

Association between diencephalic thyroliberin and arterial blood pressure in agouti-yellow and ob/ob mice may be mediated by leptin.

Leptin, a hormone secreted by the adipose tissue, stimulates anorexigenic peptides and also inhibits orexigenic peptides in hypothalamic arcuate nuclei-located neurons. It also counteracts the starvation-induced suppression of thyroid hormones by up-regulating the expression of preproTRH gene. On the other hand, in addition to its role as a modulator of the thyroid-hypothalamic-hypophysial axis, thyrotropin-releasing hormone (TRH) acts as a modulator of the cardiovascular system. In fact, we reported that overexpression of diencephalic TRH (dTRH) induces hypertension. We have recently shown that, in rats with obesity-induced hypertension, hyperleptinemia may produce an increase of dTRH together with an elevation of arterial blood pressure (ABP) through an increase of sympathetic activity and that these alterations were reversed by antisense oligonucleotide and small interfering RNA against preproTRH treatments. Here we explore the possible role of dTRH as a mediator involved in leptin-induced hypertension in 2 obesity mouse models: agouti-yellow mice, which are hyperleptinemic and hypertensive, and ob/ob mice, which lack functional circulating leptin. These 2 models share some characteristics, but ob/ob mice show lower ABP and plasma catecholamines levels. Then, for the first time, we report that there is a clear association between ABP and dTRH levels in both mouse models, as we have found that dTRH content was elevated in agouti-yellow mice and diminished in ob/ob mice compared with their controls. We also show that, after 3 days of subcutaneous leptin injections (10 microg/12 hours), ABP and dTRH increased significantly in ob/ob mice with no alterations of thyroid hormone levels. These results add evidence to the putative molecular mechanisms for the strong association between obesity and hypertension.

Knocking down the diencephalic thyrotropin-releasing hormone precursor gene normalizes obesity-induced hypertension in the rat.

We recently showed that diencephalic TRH may mediate the central leptin-induced pressor effect. Here, to study the role of TRH in obesity-induced hypertension (OIH), we used a model of OIH produced by a high-fat diet (HFD, 45 days) in male Wistar rats. After 4 wk, body weight and systolic arterial blood pressure (SABP) increased in HFD animals. Plasma leptin was correlated with peritoneal adipose tissue. Then, we treated OIH animals with an antisense oligodeoxynucleotide and small interfering (si)RNA against the prepro-TRH. Antisense significantly decreased diencephalic TRH content and SABP at 24 and 48 h posttreatment. Similar effects were observed with siRNA against prepro-TRH but for up to 4 wk. Conversely, vehicle, an inverted antisense sequence and siRNA against green fluorescence protein, produced no changes. SABP decrease seems to be owing to an inhibition of the obesity-enhanced sympathetic outflow but not to an alteration in thyroid status. Using a simple OIH model we demonstrated, for the first time, that central TRH participates in the hypertension induced by body weight gain probably through its well-known action on sympathetic activity. Thus the TRH-leptin interaction may contribute to the strong association between hypertension and obesity.

Thyrotropin-releasing hormone decreases leptin and mediates the leptin-induced pressor effect.

Leptin, an adipocyte-released hormone, modifies food intake and energy expenditure regulating hypothalamic-pituitary-thyroid axis function. We previously reported that thyrotropin-releasing hormone (TRH) precursor gene overexpression induces hypertension in the normal rat and that spontaneously hypertensive rats have central TRH hyperactivity with increased TRH synthesis and release and an elevated TRH receptor number. In both models, intracerebroventricular antisense (AS) treatment against the TRH precursor produced a dose-dependent reduction of the increased diencephalic TRH content while normalizing high arterial blood pressure. In this article, we report that male Wistar rats that were made hypertensive by intracerebroventricular injection of a eucaryotic expression plasmid containing the pre-TRH cDNA showed decreased leptin plasma levels and that pre-TRH AS treatment reversed this phenomenon. In addition, male and female spontaneously hypertensive rats showed lower levels of circulating leptin than did sex-matched Wistar-Kyoto control rats. This difference also was abated by the pre-TRH AS treatment. Conversely, 20 microg ICV leptin induced a long-lasting pressor effect (18 +/- 5 mm Hg, n=6, P<0.01, >60 minutes) that was not observed in pre-TRH AS pretreated rats (2 +/- 3 mm Hg, n=6) but persisted in rats used as controls that were treated with inverted oligonucleotide (20 +/- 6 mm Hg, n=4, P<0.01). These data suggest that in rats with TRH-induced hypertension, leptin is decreased, inducing compensatory adiposity. We propose that because leptin produces central TRH synthesis and release, obesity may induce hypertension through TRH system activation and that the TRH-leptin interaction may thus contribute to the strong association between hypertension and obesity.