The Expression of RAC1 and Mineralocorticoid Pathway-Dependent Genes are Associated With Different Responses to Salt Intake.

BACKGROUND: Rac1 upregulation has been implicated in salt-sensitive hypertension as a modulator of mineralocorticoid receptor (MR) activity. Rac1 could affect the expression of oxidative stress markers, such as hemoxigenase-1 (HO-1) or nuclear factor-B (NF-κB), and the expression of neutrophil gelatinase-associated lipocalin (NGAL), a cytokine upregulated upon MR activation. AIM: We evaluated RAC1 expression in relation of high salt intake and association with MR, NGAL, HO-1, and NF-κB expression, mineralo- and glucocorticoids levels, and inflammatory parameters. SUBJECTS AND METHODS: We studied 147 adult subjects. A food survey identified the dietary sodium (Na) intake. RAC1 expression was considered high or low according to the value found in normotensive subjects with low salt intake. We determined the gene expression of RAC1, MR, NGAL, HO-1, NF-κB, and 18S, isolated from peripheral leukocytes. We measured aldosterone, cortisol, sodium, potassium excretion, metalloproteinase (MMP9 y MMP2), and C-reactive protein. RESULTS: We identified 126 subjects with high Na-intake, 18 subjects had high, and 108 low-RAC1 expression. The subjects with high-RAC1 expression showed a significant increase in MR (P = 0.0002), NGAL (P < 0.0001) HO-1 (P = 0.0004), and NF-κB (P < 0.0001) gene expression. We demonstrated an association between RAC1 expression and MR (R sp 0.64; P < 0.0001), NGAL (R sp 0.48; P < 0.0001), HO-1 (R sp 0.53; P < 0.0001), and NF-κB (R sp0.52; P < 0.0001). We did not identify any association between RAC1 and clinical or biochemical variables. CONCLUSIONS: RAC1 expression was associated with an increase in MR, NGAL, NF-κB, and HO-1 expression, suggesting that RAC1 could be a mediator of cardiovascular damage induced by sodium, and may also useful to identify subjects with different responses to salt intake.

Aldosterone as a modulator of immunity: implications in the organ damage.

High plasmatic levels of aldosterone cause hypertension and contribute to progressive organ damage to the heart, vasculature, and kidneys. Recent studies have demonstrated a role for the immune system in these pathological processes. Aldosterone promotes an inflammatory state characterized by vascular infiltration of immune cells, reactive oxidative stress, and proinflammatory cytokine production. Further, cells of the adaptive immune system, such as T cells, seem to participate in the genesis of mineralocorticoid hormone-induced hypertension. In addition, the observation that aldosterone can promote CD4⁺ T-cell activation and Th17 polarization suggests that this hormone could contribute to the onset of autoimmunity. Here we discuss recent evidence supporting a significant involvement of the immune system, especially adaptive immunity, in the genesis of hypertension and organ damage induced by primary aldosteronism. In addition, possible new therapeutic approaches consisting of immunomodulator drugs to control exacerbated immune responses triggered by elevated aldosterone concentrations will be described.

Aldosterone promotes autoimmune damage by enhancing Th17-mediated immunity.

Excessive production of aldosterone leads to the development of hypertension and cardiovascular disease by generating an inflammatory state that can be promoted by T cell immunity. Because nature and intensity of T cell responses is controlled by dendritic cells (DCs), it is important to evaluate whether the function of these cells can be modulated by aldosterone. In this study we show that aldosterone augmented the activation of CD8(+) T cells in a DC-dependent fashion. Consistently, the mineralocorticoid receptor was expressed by DCs, which showed activation of MAPK pathway and secreted IL-6 and TGF-beta in response to aldosterone. In addition, DCs stimulated with aldosterone impose a Th17 phenotype to CD4(+) T cells, which have recently been associated with the promotion of inflammatory and autoimmune diseases. Accordingly, we observed that aldosterone enhances the progression of experimental autoimmune encephalomyelitis, an autoimmune disease promoted by Th17 cells. In addition, blockade of the mineralocorticoid receptor prevented all aldosterone effects on DCs and attenuated experimental autoimmune encephalomyelitis development in aldosterone-treated mice. Our data suggest that modulation of DC function by aldosterone enhances CD8(+) T cell activation and promotes Th17-polarized immune responses, which might contribute to the inflammatory damage leading to hypertension and cardiovascular disease.

Effects of expression of p53 and Gadd45 on osmotic tolerance of renal inner medullary cells.

The response of renal inner medullary (IM) collecting duct cells (mIMCD3) to high NaCl involves increased expression of Gadd45 and p53, both of which have important effects on growth and survival of the cells. However, mIMCD3 cells, being immortalized by SV40, proliferate rapidly, which is known to sensitize cells to high NaCl, whereas IM cells in situ proliferate very slowly and survive much higher levels of NaCl. In the present studies, we have examined the importance of Gadd45 and p53 for survival of normal IM cells in their usual high-NaCl environment by using more slowly proliferating second-passage mouse inner medullary epithelial (p2mIME) cells and comparing cells from wild-type and gene knockout mice. Acutely elevating NaCl (and/or urea) reduces Gadd45a, but increases Gadd45b and Gadd45g mRNA, depending on the mix of NaCl and urea and the rate of increase of osmolality. Nevertheless, p2mIME cells from Gadd45b(-/-), Gadd45g(-/-), and Gadd45bg(-/-) mice survive elevation of NaCl (or urea) essentially the same as do wild-type cells. p53(-/-) Cells do not tolerate as high a concentration of NaCl (or urea) as p53(+/+) cells, but urinary concentrating ability of p53(-/-) mice is normal, as is the histology of inner medullas from p53(-/-) and Gadd45abg(-/-) mice. Thus although Gadd45 and p53 may play roles in osmotically stressed mIMCD3 cells, we do not find that their expression makes an important difference, either for Gadd45 in slower proliferating p2mIME cells or for Gadd45 or p53 in normal inner medullary epithelial cells in situ.

Toxicity of acetaminophen, salicylic acid, and caffeine for first-passage rat renal inner medullary collecting duct cells.

Chronic excess ingestion of nonsteroid anti-inflammatory drugs causes renal medullary necrosis. Previously, using an immortalized line of mouse inner medullary collecting ducts cells (mIMCD3), we found that acetaminophen, salicylic acid, and caffeine are toxic, and the effects of acetaminophen and caffeine are strongly additive. Furthermore, toxicity was greater in proliferating than in nonproliferating cells. Important limitations were that mIMCD3 cells do not readily tolerate the high concentrations of salt and urea normally present in renal inner medullas and proliferate much more rapidly than inner medullary cells in vivo. Thus, these cells may not serve as an appropriate model for the in vivo IMCD. The present studies address these limitations by using passage-1 rat inner medullary collecting duct (p1rIMCD) cells, which tolerate high salt and urea and become contact inhibited when confluent. At 640 mOsmol/kg (the lowest normal inner medullary osmolality), the drugs, singly and in combination, reduce the number of proliferating (i.e., subconfluent) p1rIMCD cells more than they do confluent cells. Effects of acetaminophen and caffeine are strongly additive. Addition of as little as 0.1 mM caffeine significantly enhances the toxicity of acetaminophen plus salicylic acid. With confluent cells at 640 mOsmol/kg and very slowly growing cells at 1370 mOsmol/kg, combinations of drugs that include acetaminophen increase proliferation, accompanied by DNA damage and apoptosis. We conclude that these drugs are toxic to renal inner medullary collecting duct cells under the conditions of high osmolality normally present in the inner medulla, that combinations of the drugs are more toxic than are the drugs individually, and that the toxicity includes induction of proliferation of these cells that are otherwise quiescent in the presence of high osmolality.