Genes Influencing Circadian Differences in Blood Pressure in Hypertensive Mice

Essential hypertension is a common multifactorial heritable condition in which increased sympathetic outflow from the central nervous system is involved in the elevation in blood pressure (BP), as well as the exaggerated morning surge in BP that is a risk factor for myocardial infarction and stroke in hypertensive patients. The Schlager BPH/2J mouse is a genetic model of hypertension in which increased sympathetic outflow from the hypothalamus has an important etiological role in the elevation of BP. Schlager hypertensive mice exhibit a large variation in BP between the active and inactive periods of the day, and also show a morning surge in BP. To investigate the genes responsible for the circadian variation in BP in hypertension, hypothalamic tissue was collected from BPH/2J and normotensive BPN/3J mice at the ‘peak’ (n = 12) and ‘trough’ (n = 6) of diurnal BP. Using Affymetrix GeneChip® Mouse Gene 1.0 ST Arrays, validation by quantitative real-time PCR and a statistical method that adjusted for clock genes, we identified 212 hypothalamic genes whose expression differed between ‘peak’ and ‘trough’ BP in the hypertensive strain. These included genes with known roles in BP regulation, such as vasopressin, oxytocin and thyrotropin releasing hormone, as well as genes not recognized previously as regulators of BP, including chemokine (C-C motif) ligand 19, hypocretin and zinc finger and BTB domain containing 16. Gene ontology analysis showed an enrichment of terms for inflammatory response, mitochondrial proton-transporting ATP synthase complex, structural constituent of ribosome, amongst others. In conclusion, we have identified genes whose expression differs between the peak and trough of 24-hour circadian BP in BPH/2J mice, pointing to mechanisms responsible for diurnal variation in BP. The findings may assist in the elucidation of the mechanism for the morning surge in BP in essential hypertension

Far-Infrared Therapy Induces the Nuclear Translocation of PLZF Which Inhibits VEGF-Induced Proliferation in Human Umbilical Vein Endothelial Cells

Many studies suggest that far-infrared (FIR) therapy can reduce the frequency of some vascular-related diseases. The non-thermal effect of FIR was recently found to play a role in the long-term protective effect on vascular function, but its molecular mechanism is still unknown. In the present study, we evaluated the biological effect of FIR on vascular endothelial growth factor (VEGF)-induced proliferation in human umbilical vein endothelial cells (HUVECs). We found that FIR ranging 3∼10 µm significantly inhibited VEGF-induced proliferation in HUVECs. According to intensity and time course analyses, the inhibitory effect of FIR peaked at an effective intensity of 0.13 mW/cm2 at 30 min. On the other hand, a thermal effect did not inhibit VEGF-induced proliferation in HUVECs. FIR exposure also inhibited the VEGF-induced phosphorylation of extracellular signal-regulated kinases in HUVECs. FIR exposure further induced the phosphorylation of endothelial nitric oxide (NO) synthase (eNOS) and NO generation in VEGF-treated HUVECs. Both VEGF-induced NO and reactive oxygen species generation was involved in the inhibitory effect of FIR. Nitrotyrosine formation significantly increased in HUVECs treated with VEGF and FIR together. Inhibition of phosphoinositide 3-kinase (PI3K) by wortmannin abolished the FIR-induced phosphorylation of eNOS and Akt in HUVECs. FIR exposure upregulated the expression of PI3K p85 at the transcriptional level. We further found that FIR exposure induced the nuclear translocation of promyelocytic leukemia zinc finger protein (PLZF) in HUVECs. This induction was independent of a thermal effect. The small interfering RNA transfection of PLZF blocked FIR-increased PI3K levels and the inhibitory effect of FIR. These data suggest that FIR induces the nuclear translocation of PLZF which inhibits VEGF-induced proliferation in HUVECs

Insights into angiotensin II receptor function through AT2 receptor knockout mice

Angiotensin II signals via at least two receptors termed AT1 and AT2. The function of the AT1 receptor is well defined, while that of the AT2 receptor is still shrouded in uncertainty. AT2 gene-deficient (−/−) mice have been helpful in unravelling the function of the AT2 receptor. We have studied AT2−/− and AT2+/+ mice with classical physiological techniques developed for the rat. We found that although AT2−/− mice have normal glomerular filtration rate, the pressure–natriuresis relationship in these mice, compared with AT2+/+ mice, is shifted rightward. Moreover, medullary blood flow fails to increase with increased perfusion pressure while the AT1 receptor expression in the kidneys is increased. We used telemetry and found that AT2−/− mice have about 10 mmHg higher blood pressures than AT2+/+ mice and that their circadian rhythm is disturbed. Moreover, their baroreflexes, as measured by spectral analyses, differs from AT2+/+ controls. The cardiac function of AT2−/− mice is remarkably preserved and the differences are subtle. However, if the mice are given l-NAME hypertension, they exhibit an end-systolic pressure–volume relationship that reveals decreased contractility and probable increased vascular stiffness. Furthermore, the hearts of AT2−/− mice hypertrophy more in response to l-NAME than those of AT2+/+ mice and perivascular fibrosis is increased. DOCA-salt treatment also shows a more rightward pressure–natriuresis relationship in AT2−/− compared with AT2+/+ mice. The renal iNOS expression is increased with DOCA-salt treatment. Our findings support the notion that the AT2 receptor signals antiproliferative and antifibrotic effects and that its presence results in lower blood pressures and lesser responses to secondary forms of hypertension. Technical advances that have allowed us to adapt methods for the rat to the much smaller mouse have facilitated our studies

A novel angiotensin II type 2 receptor signaling pathway: possible role in cardiac hypertrophy

We describe a novel signaling mechanism mediated by the G-protein-coupled receptor (GPCR) angiotensin II (Ang II) type 2 receptor (AT(2)). Yeast two-hybrid studies and affinity column binding assay show that the isolated AT(2) C-terminus binds to the transcription factor promyelocytic zinc finger protein (PLZF). Cellular studies employing confocal microscopy show that Ang II stimulation induces cytosolic PLZF to co-localize with AT(2) at the plasma membrane, then drives AT(2) and PLZF to internalize. PLZF slowly emerges in the nucleus whereas AT(2) accumulates in the perinuclear region. Nuclear PLZF binds to a consensus sequence of the phosphatidylinositol-3 kinase p85α subunit (p85α PI3K) gene. AT(2) enhances expression of p85α PI3K followed by enhanced p70(S6) kinase, essential to protein synthesis. An inactive mutant of PLZF abolishes this effect. PLZF is expressed robustly in the heart in contrast to many other tissues. This cardiac selective pathway involving AT(2), PLZF and p85α PI3K may explain the absence of a cardiac hypertrophic response in AT(2) gene-deleted mice