The role of STATs in apoptosis.

Signal transducers and activators of transcription (STATs) are transcription factors that mediate cytokine and growth factor induced signals that culminate in various biological responses, including proliferation and differentiation. Recent studies indicate a role for STATs in apoptosis as well. Depending upon the particular stimulus or cell type, STATs can mediate either pro-apoptotic signals or anti-apoptotic signals. STAT1 and, under some circums-tances. STAT3 are important for transducing pro-apoptotic signals whereas STAT3 and STAT5 have been implicated in promoting cell survival. Recent studies demonstrate that regulation of apoptotic pathways by STATs is largely due to transcriptional activation of genes that encode proteins that mediate or trigger the cell death process, such as Bcl-xL, caspases, Fas and TRAIL as well as those that regulate cell cycle progression, such as p21waf1. Interestingly, STAT proteins may also regulate apoptosis through a non-transcriptional mechanism by inhibiting the anti-apoptotic protein NF-kappaB. Considering that dysregulation of the STAT signaling pathway is commonly found in clinical tumor samples, understanding the mechanisms underlying STAT regulation of cell survival may lead to successful strategies for targeting STATs in cancer therapy.

CD40 activation of carcinoma cells increases expression of adhesion and major histocompatibility molecules but fails to induce either CD80/CD86 expression or T cell alloreactivity.

A major obstacle for the development of cancer immunotherapy is the poor capacity of most tumor cells to present antigen. It has previously been shown that ligation of CD40 on the surface of malignant B cells results in the induction of efficient antigen presentation primarily because of upregulated expression of MHC, costimulatory, and adhesion molecules. Ongoing clinical trials are testing the impact of CD40 ligation as immunotherapy for B cell malignancies. Because CD40 is also widely expressed in carcinomas, we studied whether CD40 activation of these cells using soluble recombinant trimeric human CD40 ligand (srhCD40L) can also induce T cell responses. Here, we show that carcinoma cells upregulate expression of CD54 and MHC molecules following in vitro exposure to srhCD40L but do not upregulate CD80 or CD86. CD40-activated carcinoma cells failed to trigger mixed lymphocyte reactions, in sharp contrast to CD40-activated lymphoma cells for which CD40 activation, as expected, resulted in increased expression of MHC, adhesion, and costimulatory molecules, and generated brisk allogeneic lymphocyte reactions. Retroviral-mediated expression of CD80 in carcinoma cells, with or without CD40 activation, triggered mixed lymphocyte reactions, provided cells were treated with IFN-gamma. Thus, the cell surface phenotype induced on carcinoma cells following CD40 activation is not fully capable of inducing T cell proliferation; however, these results support ongoing efforts to exploit costimulation in clinical efforts aimed at increasing carcinoma immunogenicity.

Evaluation of laboratory testing methods for Chlamydia trachomatis infection in the era of nucleic acid amplification.

Diagnostic tests presently available for Chlamydia trachomatis have widely varying performance characteristics. To assess evolving laboratory testing practices since the introduction of nucleic acid amplification tests (NAAT), we surveyed laboratories in Washington State about their testing practices in 1998 and compared our findings to a similar survey conducted in 1995. Laboratory directors of 61 (87%) of 70 laboratories performing chlamydial tests in 1998 returned a survey. Between 1995 and 1998, 36 laboratories discontinued chlamydial testing, and the total number of laboratories performing tests in the state decreased from 92 to 70, a 24% decline. Of the 36 laboratories that discontinued testing, 25 (69%) had previously used rapid tests. While no laboratory routinely used NAAT in 1995, ligase chain reaction (LCR) was used in 23% of laboratories in 1998 and accounted for 113,624 (36%) of the 318,133 tests performed that year. Among the remaining 204,509 tests performed in 1998, other tests employed included DNA probe (29%), enzyme immunoassay (20%), culture (12%), direct fluorescent antibody assays (3%), and rapid tests (<1%). The majority (65%) of tests performed in 1998 using technologies other than LCR or culture were done in laboratories that did more than 10,000 tests. Cost and loss of revenue to laboratories were the most frequently cited reasons for not adopting NAAT. We conclude that in Washington State, NAAT have been rapidly adopted in larger laboratories, but most patients are still tested with much less sensitive technologies. Financial constraints represent the major barrier to more widespread use of DNA amplification tests.

Retinoic acid-induced blr1 expression requires RARalpha, RXR, and MAPK activation and uses ERK2 but not JNK/SAPK to accelerate cell differentiation.

Upstream signaling requirements of retinoic acid (RA)-induced blr1 expression and downstream signaling consequences of blr1 over-expression in a human myeloid leukemia cell line demonstrate that mitogen-activated protein kinase (MAPK) signaling complexes are involved in both avenues. RA-induced myeloid differentiation and G1/G0 growth arrest of HL-60 cells is known to require the activation of the RARalpha and RXR retinoid receptors, as well as activation of the MAPK, ERK2. Transcriptional activation of the Burkitt's lymphoma receptor 1 (blr1) gene occurs early during RA-induced differentiation of HL-60 cells and requires these same three activating processes. The use of retinoid ligands that activate either the RARalpha or the RXR retinoid receptors revealed that blr1 mRNA induction was detectable only when both RARalpha and RXR were activated. Neither the RARalpha nor RXR selective ligands alone induced expression of blr1, but the combination of the two ligands induced the expression of blr1 to the same extent as RA. The MAPKK (MEK) inhibitor, PD98059, was used to determine whether extracellular signal-regulated kinase (ERK2) activation was necessary for induction of blr1 mRNA. PD98059 inhibited induced blr1 mRNA expression, due to RA or activated RARalpha plus RXR ligands, indicating that ERK2 activation is necessary for blr1 mRNA expression. Previous studies showed that ectopic expression of blr1 also caused increased MAPK activation, in particular ERK2, and subsequently accelerated RA-induced differentiation and G1/G0 growth arrest. Inhibition of ERK2 activation inhibited differentiation of blr1 transfectants, suggesting that the accelerated differentiation reflected blr1-enhanced ERK2 activation. The present data also demonstrate that ectopic expression of blr1 increased JNK/SAPK activity, but JNK/ SAPK activation was not needed for accelerated RA-induced differentiation and growth arrest. The results show that the signals known to be required for HL-60 differentiation, activated RARalpha, RXR, and ERK2, are necessary for blr1 mRNA expression. Downstream consequences of blr1 overexpression include enhanced MAPK signaling.

Vasopressin V1a receptor signaling in a rat choroid plexus cell line.

A new cell line was derived from primary culture of rat choroid plexus (RCP) by immortalization with the TSOri minus adenovirus. The selected clone expressed vasopressin V1a receptors at a density of 64,000 sites per cell, and a K(d) of 7.2 nM. Addition of vasopressin to the RCP cells induced a transient calcium peak comparable to V1a receptor signalling in different expression systems. This [Ca(2+)](i) increase was dose-dependent with an EC(50) of 22 nM vasopressin. Similar [Ca(2+)](i) increase was elicited by addition of serotonin, angiotensin II, endothelin-1, and bradykinin. Heterologous desensitization of V1a receptor was observed in RCP cells exposed to the phorbol ester PMA or following stimulation of other receptors coupled to the phosphoinositide pathway. Positive immunolabelling with Factor VIII, Flt1 and CD 34 antibodies suggests that this new RCP cell line originated from endothelial cells of rat choroid plexus.

Shear stress induces angiotensin converting enzyme expression in cultured smooth muscle cells: possible involvement of bFGF.

OBJECTIVE: Hemodynamic stresses are considered to be important regulators of gene expression in vascular cells. In this study, we have investigated the role of shear stress on ACE expression in cultured rat vascular cells, and focused on the regulation of ACE expression in smooth muscle cells. METHODS: Rat aortic endothelial cells, smooth muscle cells and fibroblasts isolated from Wistar rats were submitted to shear stress using a laminar shear flow parallel chamber. RESULTS: A 10 dynes/cm2 shear rate for 24 h increased ACE activity in the three vascular cell types (x 2.14 in endothelial cells, x 2.9 in smooth muscle cells, x 3.33 in fibroblasts). This induction was blocked by a 24 h pre-incubation with a translation blocker (10(-4)M cycloheximide) showing the role of protein neosynthesis. Therefore the study was focused on smooth muscle cells and we demonstrated that the increase in ACE activity was due to an elevation in ACE mRNA level in response to a 10 dynes/cm2 shear stress for 24 h. This induction was dependent on the shear intensity (P < 0.0001). Six hours of a 15 dynes/cm2 shear stress showed no effect on ACE activity or mRNA expression. In contrast, the same duration of shear significantly increased bFGF mRNA level (x 3.7). Conversely, bFGF dose dependently increased ACE mRNA expression and activity in smooth muscle cells. This result suggests that bFGF could be one of the potential inductors of ACE expression in the stressed smooth muscle cells. CONCLUSIONS: Mechanical stress increases ACE expression in vascular cells. bFGF could be one of the potential factors involved in this activation. This phenomenon could participate in the role of ACE activity in vascular wall remodeling.

Retinoic acid-induced blr1 expression promotes ERK2 activation and cell differentiation in HL-60 cells.

Retinoids are known to induce the differentiation and cell cycle arrest of human myeloid leukemia cells in vitro. Differential display was used to identify putative early regulatory genes that are differentially expressed in HL-60 human promyelocytic leukemia cells treated with retinoic acid. One of the cDNAs cloned encodes sequences identifying Burkitt's lymphoma receptor 1 (BLR1), a recently described chemokine receptor. Northern blot analysis demonstrates that blr1 mRNA expression increases within 9 h of retinoic acid treatment, well before functional differentiation or G(1)/G(0) growth arrest at 48 h or onset of morphological changes, suggesting a possible regulatory function. The expression of blr1 mRNA is transient, peaking at 72 h when cells are differentiated. blr1 mRNA also is induced by other differentiation-inducing agents, 1alpha,25-dihydroxyvitamin D(3) and DMSO. Induction of blr1 mRNA by retinoic acid is not blocked by the protein synthesis inhibitor cycloheximide. In HL-60 cells stably transfected with blr1 cDNA, ectopic expression of blr1 causes an increase in ERK2 MAPK activation and promotes retinoic acid-induced G(1)/G(0) growth arrest and cell differentiation. The early expression of blr1 mRNA during differentiation, its ability to increase ERK2 activation, and its enhancement of retinoic acid-induced differentiation suggest that blr1 expression may be involved in retinoic acid-induced HL-60 differentiation.

Isolation of human hepatocytes after hepatic warm and cold ischemia: a practical approach using University of Wisconsin solution.

A method is described for isolating human hepatocytes from tissue fragments after warm and cold ischemia as experienced during hepatic resections. Cells with a high trypan blue dye exclusion and good culture characteristics were isolated by employing an initial tissue perfusion with UW solution. The method could facilitate transfer of liver tissues between distant centers for cell isolation studies.

Progressive maturation resistance to microcystin-LR cytotoxicity in two different hepatospheroidal models.

To develop three-dimensional (3D) cytotoxicity models further, microcystin-induced cytoskeletal disruption was tested in two different models of multicellular hepatocyte aggregate formation (hepatospheroids). Rat hepatocyte suspensions were seeded either onto poly(2-hydroxyethylmethacrylate)-treated culture wells (poly-HEMA) or in a rotating wall vessel (RWV) device which provides minimal shear forces and enhances differentiated 3D growth. Ninety percent of spheroids forming on poly-HEMA tended to fuse and form nonhomogeneous multilobular structures by day 4 of incubation. In contrast, spheroids cultured in the low-shear environment formed homogeneous aggregates that averaged 126 + 10 microm diameter in size at day 7. Microcystin-LR (10(-6) mol/L) was put into contact (90 min in serum-free medium) with hepatocyte suspensions and spheroids formed in both systems for 1, 4 or 7 days. As already described, microcystin-LR (after 90 min), induced cytoskeletal disruptions (blebs) in 98% of the isolated primary hepatocytes maintained in suspension. In 3D cultures, blebs were detected only on poly-HEMA nonhomogeneous early prespheroids. All other mature spheroids (poly-HEMA or RWV) exposed to the toxin did not exhibit obvious morphological signs of toxicity. Moreover, microcystin-LR pre-incubation with hepatocyte suspension prevented the formation of conventional spheroids. In conclusion, the low-shear, simulated-microgravity environment generated high yields of regularly engineered spheroids. In both models, progressive resistance of mature spheroids to microcystin-LR-induced cell deformation developed with time in culture. Microcystin-LR inhibition of the formation of rat hepatospheroids in isolated hepatocyte suspension could be used as a complementary biological assay for detection of the presence of biologically active microcystin-LR in water samples.

Circulating and cellular markers of endothelial dysfunction with aging in rats.

The influence of age on endothelial functional markers was investigated in rats. Angiotensin I converting enzyme (ACE) activity and nitric oxide synthase (NOS) mRNA expressions were examined in the lung and aorta of 10-, 20-, and 30-mo-old normotensive rats. These data were extended by the measurement of circulating endothelial cells. ACE activity was significantly decreased in plasma (P < 0.01) and lungs (P < 0.01) at 30 mo, whereas it was significantly increased in the aorta (P < 0.001) at this age. Conversely, ACE mRNA levels decreased with age in the lung (P < 0.05). The level of constitutive endothelial NOS (eNOS) mRNA was significantly reduced in the aorta of 30-mo-old rats (P < 0.05), but no changes were observed in the lungs. The level of inducible NOS (iNOS) mRNA in the aorta was significantly decreased in 20- and 30-mo-old rats (P < 0.01), whereas it was significantly increased in the lung at 30 mo (P < 0.01). Interestingly, eNOS was expressed approximately 30 times more (P < 0.001) in the aorta than iNOS, whereas in the lung it was only slightly higher than iNOS (35%; P < 0.001). Neuronal NOS mRNA expression was not modified with aging. In the aorta, guanosine 3',5'-cyclic monophosphate concentration followed NOS expressions and showed a significant decrease at 30 mo (P < 0.001). An increase in the number of circulating endothelial cells was observed in the oldest rats, possibly reflecting an increase in endothelial cell turnover with aging. The present results demonstrate that aging modifies the expression of endothelial markers implicated in the regulation of vasomotor tone. This age-dependent impairment of endothelial functions could contribute to the increased risk of pathological processes within the arterial wall associated with aging.

Conversion of big-endothelin-1 elicits an endothelin ETA receptor-mediated response in endothelial cells.

Functional conversion of big-endothelin-1 to endothelin-1 and characterization of endothelin receptor subtype were investigated in cultured rat aortic endothelial cells. Exogenous endothelin-1 and big-endothelin-1 both increased arachidonic acid release and inositol phosphate production dose dependently. Endothelin-1 was more potent than big-endothelin-1 as indicated by EC50 values: 0.5 +/- 0.1 nM and 10.0 +/- 2.0 nM for endothelin-1-induced arachidonic acid release and inositol phosphate formation, respectively, versus 1.0 +/- 0.4 nM and 35.0 +/- 6.0 nM for big-endothelin-1-induced responses. Big-endothelin-1, but not endothelin-1 actions were inhibited by phosphoramidon. Comparative studies of endothelin receptor agonists and antagonists showed that endothelin-3 but not sarafotoxin S6c stimulated arachidonic acid release and inositol phosphate formation. The responses to big-endothelin-1 and endothelin-1 were specifically inhibited by the selective endothelin ETA receptor antagonist, [cyclo-D-Trp-D-Asp-Pro-D-Val-Leu] (BQ-123) but not by the selective endothelin ETB receptor antagonist [N-cis-2,6-dimethylpiperidinocarbonyl-L-gamma- methyl-Leu-D-Trp-(COMe)-D-NLeu-ONa] (BQ-788). [125I]Endothelin-1 binding was inhibited by endothelin-1, endothelin-3 and BQ-123 but not by BQ-788. These results indicate that the pharmacological responses to big-endothelin-1 in aortic endothelial cells are due to the extracellular phosphoramidon-sensitive conversion to endothelin-1. Endothelin effects are mediated through endothelin ETA receptors in these cells.

New cell substrates for in vitro evaluation of microcystin hepato-cytotoxicity.

Algal toxins, as Microcystins released in water supplies, may represent a serious health hazard. The behaviour of primary hepatocytes was compared with that of immortalized liver cells, with the intention of providing a new test on Microcystin cellular toxicity. Immortalized liver cells were obtained by transfection with SV40 Large T antigen-bearing plasmids. Primary hepatocytes were used as a reference. Microcystin-LR at 10(-6), 10(-7)and 10(-9)m was added to hepatocytes maintained in suspension or cultured as three-dimensional hepatospheroids for 20 and 90 min at 37 degrees C. Toxic effects were monitored by cytoskeletal disruption ('blebs') using both light and scanning electron microscopy (SEM), lactate dehydrogenase release (LDH) and trypan blue dye exclusion test. Microcystin-LR at all doses induced bleb formation and a loss of microvilli in both primary hepatocytes and immortalized cell suspensions in comparison with controls. A high level of blebbed cells was detected in the absence of increased LDH release. The blebbing phenomenon was readily detectable by light microscopy but its morpho-complexity was unmasked by SEM, with early toxic events being indentified as loss of microvilli prior to bleb formation. Cells of primary hepatospheroids appeared to be less tightly attached to each other and more likely to bleb than immortalized ones. These immortalized cells could limit the use of primary cells and increase the reproducibility of the assay.

Selective isolation of rat aortic wall layers and their cell types in culture--application to converting enzyme activity measurement.

The rat aorta, whose three wall layers can be separated by microdissection offers the rare possibility of comparing physiological characteristics of in vivo tissular cell components and corresponding cells after culture. We developed a technique allowing the dissociation of the three tunicae (intima, media and adventitia) of the rat aorta and the culture of their main cell types, i.e.: endothelial cells (EC) from intima, smooth muscle cells (SMC) from media and fibroblasts (Fib) from adventitia. Comparison between selected tunicae in vivo and their corresponding cells in vitro was performed via arterial angiotensin converting enzyme (ACE) activity measurements in Wistar rats. In vivo microsomial ACE activity for each tunica was as follows: 368.9 +/- 34.3 (endothelium), 10.5 +/- 1.9 (media) and 10.2 +/- 4.9 (adventitia) pmol/mg protein/min. Corresponding cell primary culture values were 1.2 +/- 0.1 (EC), 0.06 +/- 0.02 (SMC) and 0.24 +/- 0.01 (Fib) pmol/mg protein/min. Incubation of serum-deprived cells with Dexamethasone (10(-7) M) over 48 hr induced a statistically significant shift of total ACE activity from controls to stimulated cells of 2.9 +/- 0.3 to 9.7 +/- 1.0 in EC, 0.8 +/- 0.1 to 32.1 +/- 4.9 in SMC and 1.03 +/- 0.65 to 57.2 +/- 2.1 pmol/mg prot/min in fibroblasts. In the rat aorta, ACE was present not only in the intimal endothelial cell lining, but also in the media and the adventitia. ACE activity levels in primary cultured vascular cells were about 100-fold less than those found in the ex vivo tissues. Nevertheless, ACE expression seems to be more constitutive in endothelial cells and more inducible in smooth muscle cells and fibroblasts. This methodological approach should be of interest in studying environmental or genetic regulation of protein expression in the three layers/three cell types of the vascular wall.

Hyperproliferation of aortic smooth muscle cells and fibroblasts from young SHR rats is not shared by endothelial cells.

1. To study the hypertensive genotypic influence on growth kinetics of the three aortic wall cell types. 2. Using young spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats weighing 160-180 g, we compared the proliferative properties of endothelial cells (EC), smooth muscle cells (SMC) and fibroblasts that were isolated from the thoracic aorta of each strain and cultured. Growth-arrested cells were exposed to P < -thymidine after stimulation with 150 micrograms/mL endothelial cell growth supplement. Proliferation assays were performed by cell seeding on decellularized aortic explants and cell counting 2, 4, 5, 6 and 7 days after seeding. The influence of SMC from SHR on the growth kinetics of EC was evaluated by co-cultures in transwell systems. 3. After stimulation, SMC from SHR exhibited a greater P < -thymidine incorporation rate than those from WKY rats (ratios over controls: 3.90 +/- 0.48 [7] vs 1.85 +/- 0.25 [7] respectively, P < 0.05). This was also true for adventitial SHR fibroblasts: (13.1 +/- 0.6 [6] vs 9.9 +/- 1.0 [6] WKY P < 0.05). On the contrary, there was no difference in the P < -thymidine incorporation rates between EC of each strain, regardless of the passage and the time considered. Cell proliferation on matrix explants confirmed the hyperproliferation of SMC and fibroblasts from SHR, while EC of each strain proliferated equally. Smooth muscle cells from SHR did not influence the growth kinetics of EC in co-culture and vice versa. 4. The changes in growth patterns of aortic cells isolated from young prehypertensive SHR seem to be restricted to SMC and fibroblasts.

ACE in three tunicae of rat aorta: expression in smooth muscle and effect of renovascular hypertension.

Angiotensin I-converting enzyme (ACE) is known to be present at the surface of endothelial cells and also in the adventitia in large vessels. The presence of ACE in the vascular smooth muscle remains controversial. We microdissected segments of adventitia and media with or without endothelium from a region devoid of collateral arteries. The membrane-bound ACE activity in the media averaged 41% (pmol [glycine-1-14C]hippuryl-L-histidyl-L-leucine hydrolyzed.g tissue-1.min-1) of the values found in the whole aorta, whereas the adventitia contained only 6%. Immunoreactive ACE in media was characterized by Western blotting. ACE mRNAs were detected and characterized after polymerase chain amplification in isolated media. Angiotensin I and angiotensin II were equally able to contract medial rings, and the response to angiotensin I was blocked by enalaprilat. In aortas of two-kidney, one-clip hypertensive rats, there was an increase in ACE mRNA estimated by ribonuclease protection assay (P = 0.02) and in ACE activity at 15 days and 1 and 3 mo after clipping. This corresponded to a 1.5- to 2-fold increase in the ACE activity of both the media and the adventitia compared with sham-operated rats (P < or = 0.02). Thus ACE gene expression occurs in smooth muscle of rat aorta, which contains roughly the same amount of enzyme as the endothelium and readily converts angiotensin I to angiotensin II. ACE in the medial layer and the adventitia is upregulated in renovascular hypertension.

The vasodilatory effect of endogenous nitric oxide is a major counter-regulatory mechanism in the spontaneously hypertensive rat.

OBJECTIVE: A decreased responsiveness to endothelium-dependent vasodilatory substances is characteristically seen in isolated arteries from spontaneously hypertensive rats (SHR). However, the precise status and role of nitric oxide (NO), which is, at least in part, the endothelium-derived relaxing factor, remains unclear in SHR. The objective of the present study was to evaluate the importance of NO release in vivo. METHODS: The effect on systolic blood pressure of chronic administration of NG-nitro-L-arginine methyl ester (L-NAME, an NO-synthase inhibitor) was studied. Twenty SHR and 10 Wistar-Kyoto (WKY) rats were given 25 mg/kg per day L-NAME by gavage. Thirteen SHR and 14 WKY rats given water for the same period were used as controls. Rats were killed after 15 days and the aortic wall cyclic GMP (cGMP, the second messenger of NO) and cGMP-dependent kinase (the effector of cGMP) concentrations were assessed. RESULTS: During the trial, 11 of the 20 SHR given L-NAME died. Mean +/- SD systolic blood pressure increased from 131 +/- 8 to 171 +/- 10 mmHg in WKY rats given L-NAME and from 185 +/- 10 to 249 +/- 22 mmHg in SHR given L-NAME. Aortic cGMP content was similar in control WKY rats (2122 +/- 707 fmol/mg protein) and in control SHR (2098 +/- 704 fmol/mg protein), and was decreased in the L-NAME-treated WKY rats and SHR to 308 +/- 87 and 644 +/- 222 fmol/mg protein, respectively. The aortic concentrations of cGMP-dependent kinase were not different in any group. CONCLUSIONS: Basal release of NO does not appear to be impaired in SHR, but represents a major counter-regulatory mechanism in this genetic model of arterial hypertension.

Continuous versus intermittent angiotensin converting enzyme inhibition in renal hypertensive rats.

Converting enzyme inhibitors impair renal function of the kidney beyond a stenosis of the renal artery in humans and induce histological lesions in the clipped kidney of renal hypertensive rats. In two-kidney, one clip hypertensive rats, we compared the time course and magnitude of the biochemical effects of angiotensin converting enzyme inhibition on the plasma renin-angiotensin system, cardiac hypertrophy, renal lesions, and 24-hour blood pressure decrease induced by either intermittent angiotensin converting enzyme inhibition administration (benazepril PO, 10 mg/kg once a day, n = 93) or continuous administration (benazeprilat, 3 mg/kg per day via osmotic pumps, n = 92). Control rats (n = 91) received the drug vehicle intermittently or continuously. Mortality was significantly reduced by both intermittent (n = 3/93) and continuous (n = 3/92) inhibition compared with controls (n = 18/91) (P < .001). Changes in the plasma renin-angiotensin system and blood pressure were parallel. A continuous suppression of the activity of the plasma renin-angiotensin system was associated with a 24-hour decrease in blood pressure with continuous inhibition, whereas intermittent inhibition induced a similar fall in blood pressure only for the first hours after gavage.(ABSTRACT TRUNCATED AT 250 WORDS)

[Vasodilator effect of nitric oxide is a necessary counter-regulation in the spontaneously hypertensive rat]. / L'effet vasodilatateur de l'oxyde d'azote est une contre-régulation indispensable chez le rat spontanément hypertendu.

The chronic inhibition of NO-synthase by NG-nitro-L-arginine methyl ester (L-NAME, a L-arginine analogue) induces a dose-dependent decrease in aortic cGMP and an increase in blood pressure. We used this pharmacological approach to evaluate the release of NO in vivo in spontaneously hypertensive rats (SHR); 15 SHR and 10 Wistar-Kyoto rats (WKY) were given 25 mg L-NAME/kg/d by gavage for 15 days; 10 SHR and 10 WKY rats given water for the same period were used as control. During the trial, 10/15 SHR given L-NAME died. Systolic blood pressure (mmHg) increased from 132 +/- 6 to 170 +/- 4 in WKY given L-NAME and from 169 +/- 4 to 242 +/- 6 in SHR given L-NAME. Aortic cGMP content (fmol/mg protein) was 2,204 +/- 382 and 2,076 +/- 461 fmol/mg control WKY and SHR (NS), and was decreased to 324 +/- 44 and 641 +/- 70 in WKY and SHR given L-NAME respectively (p < 0.0001 each). L-NAME increased plasma atrial natriuretic factor only in SHR. In summary, basal aortic cGMP content, reflecting the basal release of NO, was similar in WKY and SHR. The decrease in aortic cGMP content of SHR given L-NAME, due to the blockade of NO-synthase, was accompanied by a large increase in systolic blood pressure and a tremendous mortality rate. Thus, basal release of NO is probably not impaired in SHR, but represents a major counterregulatory mechanism in this genetic model of arterial hypertension.