The primary physiological function of the renin-angiotensin system is to maintain vascular resistance and extracellular fluid volume homeostasis. This is mainly accomplished by the regulatory actions of Ang II on the peripheral vasculature, heart, CNS, kidney, and adrenal glands. As the rate-limiting component of the renin-angiotensin cascade, renin secretion and production is mostly stimulated by volume or salt depletion, reduction in renal vascular perfusion pressure, and sympathetic nerve activity (1
). In the present study we demonstrate that VDR-null mice have a sustained elevation of renin expression while still maintaining a normal level of blood electrolytes. The augmentation of renin synthesis leads to increased plasma Ang II production from angiotensinogen, which drives VDR-null mice to increase water intake and intestinal salt absorption, since Ang II is a very potent thirst-inducing agent that acts on the CNS, as well as a potent stimulator of intestinal sodium absorption (1
). As a result, the mutant mice have to excrete more urine and salt to maintain volume and electrolyte homeostasis. Since Ang II is a potent vasoconstrictor, its augmentation also leads to the development of hypertension and cardiac hypertrophy in VDR null mice, although the latter effect still needs more experimental verification. This is not unexpected, as hypertension and cardiac hypertrophy have been well documented in patients and animals with high renin and Ang II (37
). Thus, a new steady state of the renin-angiotensin system is established in VDR-null mice, in which the basal renin expression is higher but still responds appropriately to the same tubular salt load and volume stimuli as in the normal state. Based on these assessments, it is believed that the upregulation of renin expression is a primary defect in VDR-null mice.
That VDR-null mice maintain a high level of renin expression is, to our knowledge, a novel finding, but the underlying physiological cause can be complicated. The observation that renin expression in VDR-null mice reacts properly to high salt load or dehydration indicates that the mechanism underlying the sustained renin elevation is independent of the pathways activated by tubular salt load or volume depletion. In fact, the involvement of cyclooxygenase-2 (COX-2), which may play an important role in macula densa–mediating renin release (39
), in renin elevation in VDR-null mice is unlikely, since we observed the same low COX-2 protein level in the kidneys of both VDR-null and wild-type mice (data not shown). Since adult VDR-null mice develop hypocalcemia and secondary hyperparathyroidism, the upregulation of renin expression could be due to VDR inactivation per se, hypocalcemia, and/or high PTH. However, several lines of evidence from our study strongly suggest that vitamin D regulation of renin gene expression is direct and independent of the calcium status: (a) Preweaned VDR-null mice that have not yet developed hypocalcemia already show an elevated renin expression; (b) when the blood ionized calcium of adult VDR-null mice is normalized by the HCa-Lac diet, their renin expression and Ang II level are still elevated; (c) conversely, Gcm2-null mice, which are as hypocalcemic as VDR-null mice, do not manifest elevated renin expression; (d) in wild-type mice, reduction of 1,25(OH)2
biosynthesis also results in elevated renin expression, whereas injection of 1,25(OH)2
leads to reduced renin expression; and (e) 1,25(OH)2
directly suppresses renin gene transcription in As4.1 cells by a VDR-mediated mechanism. Therefore, our data provide very compelling evidence to establish that vitamin D is a potent negative endocrine regulator of renin expression in vivo.
Although we did not observe a stimulation of renin expression in As4.1 cells either treated with PTH, or transfected with the PTH/PTHrP receptor and then treated with PTH, we cannot, at this time, completely exclude the possibility that, in vivo, secondary hyperparathyroidism may also contribute to the renin upregulation in VDR-null mice. This is because the serum PTH level in the normocalcemic preweaned or HCa-Lac diet–treated VDR-null mice is still significantly higher than that of the wild-type mice (even though it is much lower than that of the untreated adult VDR-null mice). Previous studies have shown that intravenous infusion of PTH increases plasma renin activity and renin release in humans and animals (33
), but the molecular mechanism whereby PTH regulates renin expression in vivo remains unknown. PTH may indirectly regulate renin expression in vivo.
exerts its actions by binding to the VDR. In most cases where 1,25(OH)2
acts as a positive regulator, the liganded VDR heterodimerizes with the RXR and binds to specific DNA sequences (VDRE) in the promoter of target genes to regulate gene expression. On the other hand, 1,25(OH)2
can also act as a negative regulator, but the mechanism of the negative regulation is more complicated and only partially understood. For instance, inhibition of other transcriptional complexes by VDR-RXR heterodimer or VDR homodimer (41
), interaction of VDR-RXR heterodimer with corepressors (43
), and binding of VDR to a negative VDRE (44
) have been reported for the VDR-mediated transcriptional repression. We postulate that 1,25(OH)2
suppresses renin gene expression through a cis
-DNA element(s) in the renin gene promoter. Analysis of the renin gene promoter is underway to elucidate the molecular mechanism.
In summary, we have demonstrated that vitamin D functions as a novel negative endocrine regulator of the renin-angiotensin system in animals. Our data indicate that maintaining a normal level of serum 1,25(OH)2D3 is important not only for calcium homeostasis, but also for the homeostasis of electrolytes, volume, and blood pressure.