Angiotensin II type 2 receptor-mediated gene expression profiling in human coronary artery endothelial cells.

Despite intensive investigation, the molecular mechanism by which the angiotensin II type 2 (AT2) receptor exerts its cellular and physiological actions remains elusive. In the present study, we have used microarray expression analysis to identify genes whose expression was regulated by this receptor and to determine its cellular consequences. Lentiviral vector was used to express the AT2 receptor in human coronary artery endothelial cells (HCAECs), followed by analysis of expression profiles. We observed approximately 5224 genes regulated in an AT2 receptor ligand-independent manner in HCAECs expressing the AT2 receptor. In addition, 1235 genes were differentially expressed in response to the AT2 receptor-specific ligand, CGP42112A. Validity of the expression profiles was demonstrated by real-time reverse-transcriptase polymerase chain reaction quantitation of 5 genes. Because some of these genes could be linked to the regulation of extracellular matrix association, we studied the effect of the AT2 receptor on cell migration. Expression of the AT2 receptor resulted in a 2-fold inhibition of HCAEC migration. Taken together, these observations demonstrate that the AT2 receptor regulates expression of genes relevant to cell migration, protein processing, intracellular signaling, and DNA repair in both ligand-dependent and ligand-independent manners.

Increased PI3-kinase in presympathetic brain areas of the spontaneously hypertensive rat.

Existing evidence led us to hypothesize that increases in p85alpha, a regulatory subunit of PI3-kinase, in presympathetic brain areas contribute to hypertension. PI3-kinase p85alpha, p110alpha, and p110delta mRNA was 1.5- to 2-fold higher in the paraventricular nucleus (PVN) of spontaneously hypertensive rats (SHR) compared with their controls, Wistar Kyoto rats (WKY). The increase in p85alpha/p110delta was attenuated in SHR treated with captopril, an angiotensin (Ang)-converting enzyme inhibitor, from in utero to 6 months of age. In the rostral ventrolateral medulla (RVLM), p110delta mRNA was approximately 2-fold higher in SHR than in WKY. Moreover, the increases in mRNA were associated with higher PI3-kinase activity in both nuclei. The functional relevance was studied in neuronal cultures because SHR neurons reflect the augmented p85alpha mRNA and PI3-kinase activity. Expression of a p85 dominant-negative mutant decreased norepinephrine (NE) transporter mRNA and [3H]NE uptake by approximately 60% selectively in SHR neurons. In summary, increased p85alpha/p110delta expression in the PVN and RVLM is associated with increased PI3-kinase activity in the SHR. Furthermore, normalized PI3-kinase p85alpha/p110delta expression within the PVN might contribute to the overall effect of captopril, perhaps attributable to a consequent decrease in NE availability.

Functional genomics as an emerging strategy for the investigation of central mechanisms in experimental hypertension.

Centrally mediated increases in sympathetic nerve activity and attenuated arterial baroreflexes contribute to the pathogenesis of hypertension. Despite the characterization of cellular and physiological mechanisms that regulate blood pressure and alterations that contribute to hypertension, the genetic and molecular basis of this pathophysiology remains poorly understood. Strategies to identify genes that contribute to central pathophysiologic mechanisms in hypertension include integrative biochemistry and physiology as well as functional genomics. This article summarizes recent progress in applying functional genomics to elucidate the genetic basis of altered central blood pressure regulatory mechanisms in hypertension. We describe approaches others and we have undertaken to investigate gene expression profiles in hypertensive models in order to identify genes that contribute to the pathogenesis of hypertension. Finally, we provide the readers a roadmap for negotiating the route from experimental findings of gene expression profiling to translating their therapeutic potential. The combination of gene expression profiling and the phenotypic characterization of in vitro and in vivo loss or gain of function experiments for candidate genes have the potential to identify genes involved in the pathogenesis of hypertension and may present novel targets for therapy.

Effects of high salt intake on brain AT1 receptor densities in Dahl rats.

To assess effects of dietary salt on brain AT1 receptor densities, 4-wk-old Dahl salt-sensitive (Dahl S) and salt-resistant (Dahl R) rats were fed a regular (101 mumol Na/g) or high (1,370 mumol Na/g)-salt diet for 1, 2, or 4 wk. AT1 receptors were assessed by quantitative in vitro autoradiography. AT1 receptor densities did not differ significantly between strains on the regular salt diet. The high-salt diet for 1 or 2 wk increased AT1 receptor binding by 21-64% in the Dahl S rats in the subfornical organ, median preoptic nucleus, paraventricular nucleus, and suprachiasmatic nucleus. No changes were noted in the Dahl R rats. After 4 wk on a high-salt diet, increases in AT1 receptor binding persisted in Dahl S rats but were now also noted in the paraventricular nucleus, median preoptic nucleus, and suprachiasmatic nucleus of Dahl R rats. At 4 wk on the diet, intracerebroventricular captopril caused clear decreases in blood pressure only in the Dahl S on the high-salt diet but caused largely similar relative increases in brain AT1 receptor densities in Dahl S and R on the high-salt diet versus regular salt diet. These data demonstrate that high salt intake rapidly (within 1 wk) increases AT1 receptor densities in specific brain nuclei in Dahl S and later (by 4 wk) also in Dahl R rats. Because the brain renin-angiotensin system only contributes to salt-induced hypertension in Dahl S rats, further studies are needed to determine which of the salt-induced increases in brain AT1 receptor densities contribute to the hypertension and which to other aspects of body homeostasis.

Brain renin-angiotensin system dysfunction in hypertension: recent advances and perspectives.

This review focuses on the dysfunction of the intrinsic brain renin-angiotensin system (RAS) in the pathogenesis of hypertension. Hyperactivity of the brain RAS plays a critical role in mediating hypertension in both humans and animal models of hypertension, including the spontaneously hypertensive rat (SHR). The specific mechanisms by which increased brain RAS activity results in hypertension are not well understood but include increases in sympathetic vasomotor tone and impaired arterial baroreflex function. We discuss the contribution of endogenous angiotensin (Ang) II actions on presympathetic vasomotor rostral ventrolateral medulla neurons to enhance sympathetic activity and maintain hypertension. In addition, we discuss Ang II-induced attenuation of afferent baroreceptor feedback within the nucleus tractus solitarius and its relevance to the development of hypertension. We also outline the cellular and molecular mechanisms of Ang II signal transduction that may be critical for the initiation and establishment of hypertension. In particular, we present evidence for a phosphoinositide-3-kinase-dependent signaling pathway that appears to contribute to hypertension in the SHR, possibly via augmented Ang II-induced increases in neuronal firing rate and enhanced transcriptional noradrenaline neuromodulation. Finally, we outline future directions in utilizing our understanding of the brain RAS dysfunction in hypertension for the development of improved therapeutic intervention in hypertension.