Kidney androgen-regulated protein transgenic mice show hypertension and renal alterations mediated by oxidative stress.

BACKGROUND: Kidney androgen-regulated protein (KAP), a proximal tubule androgen-regulated gene, codes for a protein of unknown function. METHODS AND RESULTS: To investigate the consequences of KAP overexpression in kidney, we produced KAP transgenic mice and performed microarray expression analyses in kidneys of control and transgenic males. Downregulation of the androgen-sensitive Cyp4A14 monooxygenase gene in KAP transgenic mice prompted us to analyze blood pressure levels, and we observed that transgenic mice were hypertensive. Inhibition of 20-hydroxyeicosatetraenoic acid synthesis by N-hydroxy-N'-(4-n-butyl-2-methylphenyl) formamidine (HET0016) reduced the increased 20-hydroxyeicosatetraenoic acid levels in urine and normalized arterial pressure in transgenic mice, as did the NADPH oxidase inhibitor apocynin. Increased oxidative stress in transgenic mice was demonstrated by (1) enhanced excretion of urinary markers of oxidative stress, 8-iso-prostaglandin F2alpha, 8-hydroxydeoxyguanosine, and thiobarbituric acid-reacting substances; (2) augmented mitochondrial DNA damage and malondialdehyde levels in kidneys; and (3) diminished catalase and glutathione peroxidase activity in transgenic kidneys. Mice exhibited renal defects that included focal segmental glomerulosclerosis, proteinuria, glycosuria, and fibrosis. CONCLUSIONS: Taken together, these results indicate that KAP expression is critical for cardiovascular-renal homeostasis maintenance and that hypertension is associated with increased oxidative stress. This is the first report showing that overexpression of an androgen-regulated, proximal tubule-specific gene induces hypertension. These observations may shed light on the molecular pathophysiology of gender differences in the prevalence and severity of hypertension and chronic renal disease.

Effect of the flavonoid quercetin on cadmium-induced hepatotoxicity.

The present study was designed to evaluate whether treatment with quercetin exerts any beneficial effect on cadmium (Cd)-induced hepatotoxicity in order to establish the possible protective mechanisms of quercetin. Wistar rats were distributed in four experimental groups: control, Cd, quercetin, and Cd+quercetin. Hepatic toxicity was evaluated by measuring plasma concentrations of markers of hepatic injury. The activity of antioxidant enzymes in liver was also measured. Hepatic expression of metallothioneins (MT), and endothelial nitric oxide synthase (eNOS) was assayed by Western and Northern blot. Our results demonstrated that Cd administration induced an increased marker enzyme activity in plasma. This effect was not inhibited by quercetin. However, the administration of quercetin softened Cd-induced oxidative damage. MT levels in liver were substantially increased when the animals received Cd and quercetin. Hepatic eNOS expression was significantly increased after treatment with Cd and quercetin, being this increase higher than in animals receiving Cd alone. In conclusion, in this experimental model, quercetin was not able to prevent the Cd-induced liver damage although the animals that received both, Cd and quercetin showed a marked improvement in oxidative stress and an increase in the MT and eNOS expression. These results suggest that other mechanisms different to oxidative stress could be involved in hepatic damage.

Teaching integrative physiology using the quantitative circulatory physiology model and case discussion method: evaluation of the learning experience.

One of the problems that we have found when teaching human physiology in a Spanish medical school is that the degree of understanding by the students of the integration between organs and systems is rather poor. We attempted to remedy this problem by using a case discussion method together with the Quantitative Circulatory Physiology (QCP) program. QCP is a Windows-based computer simulation program that offers almost real-time simulation and allows users to examine the time-dependent interactions of over 750 parameters. We evaluated students' perceptions by an anonymous questionnaire. Teachers' perceptions of this teaching approach were highly positive, as it improved students' perceptions of the complexity of biological processes, their ability to differentiate between acute and chronic responses, and promoted an integrative understanding of human body function. Teachers also identified some problems with the approach, including student difficulties in adopting self-directed learning, a lack of precision in student questions during the discussion sessions, and the lack of a tradition of using several textbooks to explain the changes observed. The results of the student questionnaire revealed that >70% of the students reported that this type of learning gave them a better understanding of the complexity of physiological processes and the role of coordinated actions of several systems in the homeostatic response and enabled them to acquire a better understanding of human body functions. Thus, we conclude that this approach promotes an integrative understanding of cardiovascular and renal functions that is difficult to achieve with other methods.

Protective effect of quercetin on experimental chronic cadmium nephrotoxicity in rats is based on its antioxidant properties.

Oxidative stress can play a key role in Cd-induced dysfunctions. Quercetin is a potent oxygen free radicals scavenger and a metal chelator. Our aim was to study the effect of quercetin on Cd-induced kidney damage and oxidative stress as well as its mechanism of action. Wistar rats were distributed in four experimental groups: control rats; Cd; quercetin and Cd+quercetin. Renal toxicity was evaluated by measuring urinary excretion of proteins, albumin, glucose and enzymes markers of tubular necrosis, as well as plasma concentration of creatinine. Plasma TBARS concentration and activity of antioxidant enzymes in kidney were also measured. Renal cell damage was assessed by electron microscopy. Animals that received both Cd and quercetin showed a better renal function than those receiving Cd alone. Cd-induced tubular lesions were markedly reduced in rats that also received quercetin. Cd-induced increase in plasma TBARS was prevented by the administration of quercetin. Total plasma antioxidants and renal superoxide dismutase and glutathione-reductase activities were higher in the group that received Cd and quercetin than in rats that received Cd alone. Quercetin administration does not modify the renal content or the urinary excretion of Cd. In conclusion, quercetin treatment prevents renal tubular damage and increased oxidative stress induced by chronic Cd administration, most probably throughout its antioxidant properties.

Endoglin regulates nitric oxide-dependent vasodilatation.

Endoglin is a membrane glycoprotein that plays an important role in cardiovascular development and angiogenesis. We examined the role of endoglin in the control of vascular tone by measuring nitric oxide (NO)-dependent vasodilation in haploinsufficient mice (Eng+/-) and their Eng+/+ littermates. The vasodilatory effect of acetylcholine, bradykinin, and sodium nitroprusside was assessed in anesthetized mice; in isolated, perfused hindlimbs; and in aortic rings. The substantial hypotensive and vasodilatory response induced by acetylcholine and bradykinin in Eng+/+ was markedly reduced in Eng+/- mice. Both kinds of animals had similar responses to sodium nitroprusside, suggesting that the deficient vasodilatory effect is not due to a NO response impairment. Urinary and plasma concentrations of nitrites, a NO metabolite, were lower in Eng+/- than in Eng+/+ mice. The levels of endothelial nitric oxide synthase (eNOS) in kidneys and femoral arteries were about half in Eng+/- than in Eng+/+ mice and were also reduced in primary cultures of aortic endothelial cells from Eng+/- compared with those from Eng+/+ mice. Furthermore, overexpression or suppression of endoglin in cultured cells induced a marked increase or decrease in the protein levels of eNOS, respectively. Thus, our results in vivo and in vitro demonstrate a relationship between endoglin and NO-dependent vasodilation mediated by the regulation of eNOS expression.

Adenosine activates mesangial cell proliferation.

This study investigates the proliferative effect of adenosine (ADO) in cultured mesangial cells, and the possible mediation of A1 and/or A2 receptors in this proliferative effect of ADO. ADO (10(-5) M) induced a significant increase in the [3H]thymidine incorporation into DNA with respect to quiescent cells. This increase was similar to that obtained with the ADO A1 receptor agonist, R-PIA (10(-5) M), and with the ADO A2 receptor agonist, NECA (10(-5) M). Theophylline (10(-4) M), and ADO receptors inhibitor, completely inhibits the ADO-induced proliferation. The combinations NECA + A2 receptor antagonist, PD 116,948 (AT1, 10(-6) M) and PIA + A2 receptor antagonists, PD 115,199 (AT2, 10(-2) M) did not induce any significant difference with respect to cells maintained in control conditions. These findings demonstrate the proliferative effect of ADO in cultured mesangial cells, and that this effect is not specific to either of A1 or A2 receptors activation.

Nitric oxide production by mononuclear leukocytes in alcoholic cirrhosis.

Nitric oxide is now established as a biological mediator of clinical relevance. The present study investigated the production of nitric oxide by lympho-mononuclear leukocytes from alcoholic patients with either hepatitis or cirrhosis. The study included 42 patients, 12 without any liver disease and 30 alcoholic patients, 13 of whom had histologically confirmed cirrhosis and 17 alcoholic hepatitis. Cells were obtained from peripheral blood by density gradient and incubated in sterile conditions in RPMI 1640 for 6 h at 37 degrees C. Culture supernatants were assayed for nitrite concentration using the Griess reaction. Cells from cirrhotic but not from hepatopathic patients showed significantly higher nitrite production than controls (cirrhotic, 0.36 +/- 0.07; hepatopathic, 0.13 +/- 0.02; control: 0.25 +/- 0.05 nmol/10(6) cells/6 h). In cirrhotic patients L-Nitro-arginine methylester inhibited nitrite production (0.18 +/- 0.05). These data suggest that alcoholic cirrhotic but nonhepatopathic patients show an increased nitric oxide production by blood lymphomononuclear cells. This production could be involved in the systemic vasodilation in cirrhotic patients.

Temporal changes in renal endoglin and TGF-beta1 expression following ureteral obstruction in rats.

Chronic renal disease is characterized by the accumulation of extracellular matrix proteins in the kidney and a loss of renal function. Tubulointerstitial fibrosis has been reported to play an important role in the progression of chronic renal diseases. Transforming growth factor-beta1 (TGF-beta1) is a profibrotic cytokine playing a major contribution to fibrotic kidney disease. Endoglin is a membrane glycoprotein of the TGF-beta1 receptor system. The aim of this work was to determine the time-course expression of renal type I and IV collagens, endoglin and TGF-beta1 in a rat model of induced tubulointerstitial fibrosis at 1, 3, 10 and 17 days after unilateral ureteral obstruction (UUO). In 17 days-ligated (L)-renal samples, a marked interstitial fibrosis was detected by Masson's trichromic and Sirius red staining, accompanied by an increase in type I collagen expression as shown by immunohistochemical analysis. Northern blot studies revealed a progressive increase in collagen alpha2(I), TGF-beta1 and endoglin mRNA expression in L kidneys when compared with the corresponding non-ligated (NL) kidneys from the animals subjected to left UUO. Seventeen days after UUO, significant increases in collagen alpha2(I), collagen alpha1(IV), TGF-beta1 and endoglin mRNA levels were detected in L kidneys vs NL kidneys. Significantly higher levels of the protein endoglin were found in L kidneys than in NL kidneys 10 and 17 days following obstruction. A marked increase expression for endoglin and TGF-beta1 was localized in renal interstitium by immunohistochemical studies 17 days after obstruction. In conclusion, this work reports the upregulation of endoglin coincident to that of its ligand TGF-beta1 in the kidneys of rats with progressive tubulointerstitial fibrosis induced by UUO.

Atrial natriuretic peptide in rats with experimental cirrhosis of the liver without ascites.

Plasma levels of atrial natriuretic peptide (ANP) have been measured in eight sodium-retaining cirrhotic nonascitic rats and eight control animals before and after extracellular volume expansion by isotonic saline infusion (30 ml/kg BW, 20 min). In addition, disappearance of [125I]ANP was studied in six control and six cirrhotic rats. The effect of infusing synthetic rat ANP-(1-28) (1 microgram) on mean arterial pressure and renal function has been also studied. In basal conditions, cirrhotic rats showed higher ANP plasma levels than control animals (71.1 +/- 16.6 vs. 43.9 +/- 5.1 pg/ml; P less than 0.05). After extracellular volume expansion, ANP increased in both control and cirrhotic rats; the increase in cirrhotic was higher than that in control rats (88 +/- 27% vs. 33 +/- 8%; P less than 0.05). Disappearance of iodinated ANP from plasma was identical in control and cirrhotic rats. ANP infusion induced a larger decrease in mean arterial pressure in control (21 +/- 5%) than in cirrhotic rats (9 +/- 2.5%). ANP induced comparable increases in glomerular filtration rate and renal plasma flow in both groups of animals. However, diuretic and natriuretic effects were markedly impaired in cirrhotic animals. Thus, urinary flow increased by 91 +/- 18 microliters/min in control animals, but only by 37 +/- 7 microliters/min in cirrhotic animals. Fractional sodium excretion increased to 1.7 +/- 0.44% in controls and to 0.54 +/- 0.12% in cirrhotic rats (P less than 0.05). It is concluded that the defect in renal handling of sodium in cirrhotic rats is not due to a lack of ANP synthesis or release. In addition, these animals show an impaired renal response to ANP.

Pathogenesis of hyperprolactinemia in uremic rats.

Experiments have been carried out in order to clarify to what extent the absence of PRL renal catabolism in experimental renal insufficiency is responsible for the high PRL circulating levels. Furthermore, the relative contribution of the glomerular filtration rate and peritubular degradation to PRL renal clearance have been assessed. Circulating PRL basal levels were measured by RIA in sham-operated and intact control rats and in three uremic rat models: urine autoinfusion, bilateral ureteral ligation, and bilateral nephrectomy. Plasma PRL basal levels (nanograms per ml; mean +/- SEM) were increased in sham-operated rats (30.3 +/- 5.1) with respect to control animals (18.5 +/- 2.7; P less than 0.05). Bilaterally nephrectomized animals (66.4 +/- 16.4) and those with bilateral ureteral ligation (69.3 +/- 15.9) developed similar hyperprolactinemia, in contrast to urine-autoinfused rats (20.2 +/- 2.1; P less than 0.005) whose hormone levels were similar to those of control animals. Creatinine levels were markedly elevated and comparable in the three uremic rat groups. The results suggest that: 1) hyperprolactinemia in rats in acute renal insufficiency is due primarily to reduced renal function; 2) PRL renal catabolism in the rat requires a certain rate of glomerular filtration; and 3) PRL peritubular degradation does not seem to be relevant in PRL catabolism by rat kidney.

Renal metabolism of gut glucagon-like immunoreactivity.

To examine the role of the kidney in the catabolism of gut glucagon-like immunoreactivity (GLI), we compared the plasma GLI responses of normal and nephrectomized dogs given intraduodenal glucose loads and studied the clearance of gut GLI by the isolated perfused rat kidney. Both basal and postload plasma samples were analyzed with a glucagon C-terminal specific antibody and a GLI-reacting N-terminal antibody. The GLI response was taken to be the difference between the increments seen with these two antibodies after glucose loading. Although glucose-induced GLI increments could not be detected in untreated plasma from nephrectomized dogs, chromatographed plasma revealed a significant rise in GLI-fraction II (7000--12000 daltons) in both nephrectomized and normal dogs (732 +/- 200 and 586 +/- 111 pg/ml, respectively). We also found that crystalline glucagon was cleared by the isolated closed-circuit perfused rat kidney, but gut-GLI either as crude extract or as peak I (7000--12000 daltons) was not. Our data suggest that the kidney may not play an important role in gut-GLI catabolism.

Effect of hypothalamic-hypophysary inhibitory factor on mesangial cell activation.

We examined the effect of a sodium pump inhibitor isolated from bovine hypothalamus and pituitary tissues on contraction, proliferation, and calcium mobilization in primary cultures of rat mesangial cells. Hypothalamic-hypophysary inhibitory factor (HHIF) inhibited rubidium uptake in a concentration-dependent manner (0.2 U/mL: 56.8 +/- 6.3% inhibition). It also induced a concentration- and time-dependent decrease in planar cell surface area. Maximal contraction (25 +/- 5% reduction in cell size) was reached at 60 minutes with a concentration of 0.2 U/mL. This effect was inhibited by both verapamil and TMB-8 (10(-5) mol/L). HHIF was also observed to increase DNA synthesis (0.2 U/mL: 4361 +/- 168 versus 2129 +/- 162 cpm per well under control conditions) and cell proliferation (0.2 U/mL: 52,290 +/- 1931 versus 10,512 +/- 121 cells per well under control conditions). Both effects were also inhibited by verapamil and TMB-8. Moreover, HHIF induced the expression of immediate early genes c-fos and c-jun mRNA. HHIF-induced effects were accompanied by an increase in cytosolic free calcium (203 +/- 58 versus 101 +/- 2 nmol/L under control conditions), which was inhibited by verapamil and TMB-8. In summary, HHIF induces mesangial cell contraction and proliferation; these effects seem to be mediated by an increase in cytosolic free calcium levels.

Molecular heterogeneity of circulating prolactin in chronic uremic men and renal transplant recipients.

Serum PRL levels and its molecular heterogeneity were analyzed, basally and after 500 micrograms TRH given acutely, in four groups of men: normal (C, n = 12), chronic renal failure (CRF, n = 11), hemodialysis (HD, n = 12), and transplant recipients (T, n = 11). The mean basal PRL level was higher in group CRF than in group C and even higher in group HD. The basal hyperprolactinemia was due to increased concentrations of little PRL. The absolute levels of total and little PRL 20 min after TRH were comparable in the four groups. The disappearance index (DI = PRL20-PRL120/PRL20) for total and little PRL was lower in CRF than in C and even lower in HD. A positive correlation was found between the DIs of total and little PRL and creatinine clearance in group CRF. Group T had basal and 20 min serum PRL levels and a pattern of molecular distribution similar to those of group C but total and little PRL DI was lower. These results demonstrate that uremic hyperprolactinemia is due to increases in little PRL without major changes in big and big-big forms of PRL. The reduction of glomerular filtration rate seems to be one of the most important mechanisms responsible for little PRL accumulation.

Endoglin is expressed in the chicken vasculature and is involved in angiogenesis.

Endoglin is a component of the transforming growth factor beta (TGF-beta) receptor complex, highly expressed by endothelial cells. Mutations in the endoglin gene are responsible for hereditary hemorrhagic telangiectasia type 1 (HHT1), an autosomal dominant vascular disorder caused by a haploinsufficiency mechanism. Vascular lesions (telangiectasia and arteriovenous malformations) in HHT1 are associated with loss of the capillary network, suggesting the involvement of endoglin in vascular repair processes. Using the chick chorioallantoic membrane (CAM) as an angiogenic model, we have analyzed the expression and function of chicken endoglin. A pan-specific polyclonal antibody (pAb) recognized chicken endoglin as demonstrated by immunostaining and Western blot analysis. In ovo treatment of chicken embryos with this pAb resulted in a significantly increased area of CAM. This effect was likely mediated by modulation of the ligand binding to endoglin as this pAb was able to inhibit TGF-beta1 binding. These results support the involvement of endoglin in the angiogenic process.

Transforming growth factor-beta1 induces collagen synthesis and accumulation via p38 mitogen-activated protein kinase (MAPK) pathway in cultured L(6)E(9) myoblasts.

Transforming growth factor-beta (TGF-beta) plays a pivotal role in the extracellular matrix accumulation observed in chronic progressive tissue fibrosis, but the intracellular signaling mechanism by which TGF-beta stimulates this process remains poorly understood. We examined whether mitogen-activated protein kinase (MAPK) routes were involved in TGF-beta1-induced collagen expression in L(6)E(9) myoblasts. TGF-beta1 induced p38 and extracellular signal-regulated kinase (ERK) 1/2 phosphorylation whereas no effect on Jun N-terminal kinase phosphorylation was observed. Biochemical blockade of p38 but not of the ERK MAPK pathway abolished TGF-beta1-induced alpha(2)(I) collagen mRNA expression and accumulation. These data indicate that TGF-beta1-induced p38 activation is involved in TGF-beta1-stimulated collagen synthesis.

Prostanoid production in post-gastrectomy gastritis. Influence of sucralfate.

Prostaglandins seem to play an important role in the protection of the gastric mucosa, but the effect of cytoprotective drugs such as sucralfate on prostaglandin synthesis and in the prevention of gastritis induced by gastric surgery has not been definitely established. The purpose of this study was to determine: (1) the prostanoid production and the histologic changes occurring after various surgical techniques in rat gastric mucosa; and (2) the influence of sucralfate treatment on prostanoid levels and gastric damage induced by gastric surgery. Animals included in the study had undergone one of three surgical procedures: Billroth I, Billroth II, or truncal vagotomy plus pyleroplasty. Animals that had no surgery and "sham-operated" animals were used as controls. One third of the animals in each group received sucralfate treatment (an average of 100 mg/kg per day). Samples of gastric mucosa were taken after one year, and prostaglandin E2 (PGE2), 6-keto-PGF1 alpha, and thromboxane B2 synthesis were measured by radioimmunoassay. The sucralfate-treated group consistently showed higher PGE2 and 6-keto-PGF1 alpha values and a higher 6-keto-PGF1 alpha:thromboxane B2 ratio, as well as lower thromboxane B2 levels than untreated animals, although the differences were statistically significant only in the 6-keto-PGF1 alpha:thromboxane B2 ratio. Similarly, gastritis was more frequent and more severe in the untreated animals. In conclusion, sucralfate seems to provide protection against gastritis induced by gastric surgery and increases the 6-keto-PGF1 alpha:thromboxane B2 ratio.

Protective effect of exogenous nitric oxide on the renal function and inflammatory response in a model of ischemia-reperfusion.

BACKGROUND: Tissue subjected to a period of ischemia undergoes morphological and functional damage that increases during the reperfusion phase. The aim of the present work was to assess the possible improvement induced by exogenous administration of nitric oxide (NO) on renal injury and inflammatory reaction in an experimental animal model of renal ischemia-reperfusion (I-R). METHODS: Ischemia was achieved by ligation of the left arteria and vein for 60 min, followed first by contralateral nephrectomy and then reestablishment of blood flow. Molsidomine, used as an NO donor, was administered by systemic injection 30 min before reperfusion. The effect of molsidomine was compared with the effect of hydralazine, a non-NO donor hypotensive agent. RESULTS: Treatment with molsidomine improved the renal dysfunction (increase in plasma creatinine and urea levels) caused by I-R. Moreover, molsidomine blunted the enhanced production of proinflammatory cytokines (tumor necrosis factor [TNF]-alpha and interleukin [IL] 1alpha), the increase in tissular levels of superoxide anions and oxygen free radical scavengers, and the neutrophilic infiltration observed in the ischemic kidney. One hundred percent survival was achieved in the group of animals treated with the NO donor, whereas the groups of animals undergoing I-R that did not receive molsidomine showed a 40% mortality from the second day after reperfusion. CONCLUSIONS: The present work demonstrated that systemic treatment with an NO donor before reperfusion improved renal function and diminished inflammatory responses in a kidney subjected to an I-R process.

Effect of adenosine and adenosine analogues on cyclic AMP accumulation in cultured mesangial cells and isolated glomeruli of the rat.

1. Changes in intracellular levels of adenosine 3':5'-cyclic monophosphate (cyclic AMP) were studied in rat isolated glomeruli and cultured glomerular mesangial cells exposed to adenosine and to the preferential A1 receptor agonist N6-R-1-methyl-2-phenylethyl adenosine (R-PIA), or the potent A2 adenosine receptor agonist 5-(N-ethylcarboxamide)adenosine (NECA). 2. Whereas NECA and adenosine triggered a dose-dependent increase in cyclic AMP values with EC50 values of approximately 10(-6) M and 3 x 10(-5) M respectively, R-PIA lowered cyclic AMP levels at concentrations of 10(-6) M or less and increased them at higher concentrations. 3. The time-course of the increase induced by 10(-6) M NECA was slower than that induced by 10(-4) M adenosine. Adenosine produced a maximal stimulation within the first minute, whereas the effect of NECA in both glomeruli and mesangial cells was noticeable only from the second minute of incubation. 4. The effects of the agonists R-PIA and NECA on the cyclic AMP system were blocked respectively by the A1 adenosine receptor antagonist, 8-cyclopentyl-1, 3-dipropylxanthihe (DPCPX) at 10(-6) M and the A2 antagonist N-(2-dimethylaminoethyl)-N-methyl-4-(2, 3, 6, 7-tetrahydro-2,b-dioxo-1, 3-dipropyl-1H-purin-8-yl) benzene sulphonamide (PD115,199) at 10(-6) M. Theophylline, a known antagonist of adenosine receptors, inhibited the action of adenosine on cyclic AMP in mesangial cells. Dipyridamole, an inhibitor of the uptake of adenosine by the cells, enhanced the response to adenosine.5. These results suggest the existence of Al and A2 adenosine receptors with opposite actions on intracellular levels of cyclic AMP in both glomeruli and mesangial cells. Adenosine seems to increase cyclic AMP through the activation of a surface adenosine receptor with pharmacological properties distinct from those exhibited by the A2 adenosine receptor.

Effects of captopril, losartan, and nifedipine on cell hypertrophy of cultured vascular smooth muscle from hypertensive Ren-2 transgenic rats.

1. We hypothesized that tissular renin-angotensin system (RAS) induces vascular hypertrophy in hypertensive Ren-2 transgenic rats (TGR; strain name TGR(mRen2)L27). This assumption was tested in cell cultures of vascular smooth muscle (VSMC) from both hypertensive TGR and control normotensive Sprague-Dawley (SD) rats. Planar cell surface area, protein synthesis, and protein content per cell were studied, the role for locally produced angiotensin II (AII) was evaluated and the possible pharmacological interference by different drugs was analysed. 2. By use of radioimmunoassay techniques, AII could be determined in TGR cultures (10.25 +/- 0.12 pg per 10(7) cells) while it could not be detected in SD ones. 3. Under serum-free conditions, VSMC from hypertensive TGR were hypertrophic when compared to SD VSMC, as they presented a higher protein content per cell (335 +/-18 and 288 +/- 7 pg per cell respectively; P<0.05) and increased mean planar cell surface area, as determined by image analysis (4,074 +/- 238 and 4,764 +/- 204 microm2, respectively; P < 0.05). 4. When exogenously added to cultured SD and TGR VSMC, AII (100 pM to 1 microM) promoted protein synthesis and protein content in a concentration-dependent manner without affecting DNA synthesis. Maximal effects were observed at 100 nM. At this concentration, AII effectively increased planar cell surface area in both SD and TGR cultures by approximately 20%. 5. Treatment of TGR cultures, in the absence of exogenous AII, with the angiotensin-converting enzyme inhibitor captopril or the angiotensin AT1 receptors antagonist losartan (100 nM to 10 microM) reduced planar cell surface area in a concentration-dependent manner. In addition, both captopril and losartan (10 microM), decreased protein synthesis by approximately 15%. 6. Treatment of SD VSMC, in the absence of exogenous AII, with both captopril and losartan had no effect either on planar cell surface area or protein synthesis. 7. Treatment with the Ca2+ antagonist nifedipine (100 nM to 10 microM) reduced cell size in both SD and TGR cultures. Maximal cell reduction reached by nifedipine averaged 906 +/- 58 and 1,292 +/- 57 microm2, in SD and TGR, respectively (P<0.05). In addition, nifedipine, nitrendipine and nisoldipine (all at 10 microM) decreased protein synthesis in both cell types by 15-25%. 8. We concluded that cultured VSMC from TGR are hypertrophic in comparison with those from SD. This cell hypertrophy can be the consequence of the expression of the transgene Ren-2 that activates a tissular RAS and locally produces AII, which acts in a paracrine, autocrine, or intracrine manner. Cell hypertrophy in TGR cultures could be selectively reduced by RAS blockade, while nifedipine decreased cell size and protein synthesis in both hypertrophic and non hypertrophic cells.

Is renal protection with calcium antagonists possible?

Organ protection is the main goal in the treatment of high blood pressure. Consequently, this protective capacity should be one of the main characteristics of any drug used in the treatment of hypertension. A renal protective agent should protect the kidney from intrinsic renal vasoconstrictors and exogenous agents, and should also protect, or at least delay, the decline in renal function in the presence of renal insufficiency, by mechanisms other than increasing glomerular filtration pressure. Verapamil protects mesangial cells from the reduction in surface area induced by endothelin in vitro. In human subjects, it minimises the renal impairment provoked by the administration of cisplatin, and in mice it protects superficial cortical blood flow from the vasoconstriction elicited by cyclosporin. Finally, verapamil may protect from glomerulosclerosis as a result of its capacity to inhibit mesangial cell replication.