Microvascular regulatory role and increased expression of vascular endothelial growth factor receptor type 2 in experimental gingivitis.

OBJECTIVE: The aim of the present study was to investigate the possible microvascular regulatory role of vascular endothelial growth factor receptor type 2 (VEGFR2) in experimental gingivitis in rats. BACKGROUND: Our previous results demonstrated that functionally active VEGFR2s are located in the venules of rat gingiva. While there is no remarkable endogenous gingival VEGF production under normal circumstances, exogenous VEGF, via VEGFR2, shows venodilatory effects. We assumed that VEGF plays an important role in vasoregulatory processes (vasodilation, increased permeability, angiogenesis) of gingival inflammation. METHODS: Gingivitis was induced by placing ligatures and composite material around and between the lower incisors of anesthetized Wistar rats next to the gingival margin. Seven days later, VEGFR2 antagonist (ZM323881), was dripped upon the labial gingiva next to the lower incisors. Diameter changes of the selected gingival venules were measured by vital microscopy. Animals with healthy gingiva served as controls. Venule diameter changes were compared to the baseline and to control groups (no ligature). Immunohistochemical and Western blot analysis for VEGFR2 were utilized. RESULTS: After 15, 30 and 60 min of local application of ZM323881, there was a significant venoconstriction in the inflamed gingiva compared to the baseline, while no change was recorded in controls. Endothelium, smooth muscle cells and pericytes of the gingivitis group showed increased VEGFR2 expression. CONCLUSION: Our findings suggest that there is an increased VEGF production in gingivitis, which may play an important role in vasodilation of rat gingival venules.

Psychosocial characteristics and self-reported functional status in patients on maintenance dialysis in Hungary.

AIMS: This survey was conducted to assess psychosocial problems and functional status among patients on maintenance dialysis in Hungary. METHODS: All adult patients (n = 4,321) receiving maintenance dialysis in the 56 dialysis centers in Hungary in 2006 were approached to participate in a national, cross-sectional survey. Patients completed a brief self-reported questionnaire. Socio-demographic parameters, disease-related information and data about functional status were collected. Self-rated health and depressive symptoms were also assessed. RESULTS: Mean age was 62 ± 14 y; 52% were males. The prevalence of diabetes was 30%. 46% of participants reported having depressive symptoms. Significant functional limitation was frequent. In multivariable regression models, female gender, poor self-reported finances, less education, history of acute myocardial infarction (AMI) or cerebrovascular disease, the presence of visual or hearing impairment and difficulties with basic activities of daily living were independently associated with the presence of depressive symptoms. In a separate model, age, dialysis vintage, history of AMI or cerebrovascular disease, the presence of visual or hearing impairments, difficulties with basic activities of daily living and also having depressive symptoms were independently associated with self-rated health score. CONCLUSIONS: Chronic dialysis patients in Hungary have disadvantaged socioeconomic status, frequent depressive symptoms and many functional limitations. Professional psychosocial help would be particularly important for this underprivileged patient population in addition to high quality dialysis to optimize outcomes.

The importance of body composition and dry weight assessments in patients with chronic kidney disease.

Chronic volume overload is the major cause of hypertension and other cardiovascular morbidity in dialysis patients. One of the most important goals of physicians who take care of patients with chronic renal failure is to obtain near euvolemia or "dry body weight" in order to maintain or normalize blood pressure and prevent further cardiovascular events. In clinical practice, exact estimation of dry weight in hemodialysis patients remains a major challenge. Alterations in body composition, particularly malnutrition, are common in patients receiving long-term hemodialysis and contribute to a high mortality rate. In contrast, obesity - a known risk factor for cardiovascular morbidity and mortality - is prevalent amongst kidney allograft recipients in - long term after renal transplantation. Several technological tools and biochemical markers for estimation of plasma volume and body composition are available for clinical use. Our aim was to highlight the importance of control of body fluid volume and body composition in patients with chronic kidney disease and to describe the different methods available for such measurements.

Body mass index, waist circumference and mortality in kidney transplant recipients.

Higher body mass index (BMI) appears paradoxically associated with better outcomes in patients with chronic kidney disease. Whereas higher BMI reflects both increased visceral and subcutaneous fat and/or muscle mass, a combined assessment of BMI and waist circumference may enable differentiation of visceral adiposity from muscle and/or nonvisceral fat mass. We examined the association of BMI and waist circumference with all-cause mortality in a prospective cohort of 993 kidney transplant recipients. Associations were examined in Cox models with adjustment for demographic and comorbid conditions and for inflammatory markers. Unadjusted death hazard ratios (95%CI) associated with one standard deviation higher BMI and waist circumference were 0.94 (0.78, 1.13), p = 0.5 and 1.20 (1.00, 1.45), p = 0.05, respectively. Higher BMI was associated with lower mortality after adjustment for waist circumference (0.48 [0.34, 0.69], p < 0.001), and higher waist circumference was more strongly associated with higher mortality after adjustment for BMI (2.18 [1.55-3.08], p < 0.001). The associations of waist circumference with mortality remained significant after additional multivariable adjustments. Higher BMI and waist circumference display opposite associations with mortality in kidney transplant recipients. Waist circumference appears to be a better prognostic marker for obesity than BMI.

Sugar-free, glycine-stabilized intravenous immunoglobulin prevents skin but not renal disease in the MRL/lpr mouse model of systemic lupus.

Intravenous immunoglobulin (IVIG) has a therapeutic potential in many autoimmune diseases. Based on its immune modulating and complement inhibiting effects, IVIG has been tested in systemic lupus erythematosus (SLE), but due to osmotic tubular injury caused by immunoglobulin-stabilizing sugar components, lupus nephritis had been accelerated in some patients, thus IVIG use in SLE has been abandoned. The availability of non-sugar-stabilized IVIG raised the possible re-evaluation of IVIG for SLE. We investigated high-dose, long-term non-sugar-stabilized IVIG treatment on skin and renal SLE manifestations in the MRL/lpr mouse model. Animals were treated once a week with glycine-stabilized IVIG or saline (0.2 ml/ 10 g BW) from 6 weeks until they were humanely killed at 5 months of age. IVIG diminished macroscopic cutaneous lupus compared with saline treated mice. Histology and complement-3 immunostaining also demonstrated a significant reduction of skin disease after IVIG treatment. However, renal histology and function were similar in both groups. Compared with typical osmotic tubular damage induced by 5% sucrose and 10% maltose (used for IVIG stabilization), we did not observe any osmotic tubular injury in the glycine-stabilized IVIG treated mice. Our data demonstrate a beneficial effect of IVIG on skin lupus without renal side-effects. Deeper understanding of the organ-specific pathomechanism may aid an individualized SLE therapy.

Lupus nephritis reoccurs following transplantation in the lupus prone mouse.

The incidence and pathomechanism of recurrent lupus nephritis (RLN) after transplantation is not clearly understood. Burning out of the autoimmune process or local immunoregulatory mechanisms in the kidney may be responsible for the low incidence of recurrence. These mechanisms cannot be investigated in human subjects, due to post-transplant immunosuppression. To investigate the pathomechanisms of RLN, male and female kidneys were transplanted from FAS deficient lupus prone (LPR) or control (FAS intact) MRL mice into either LPR or MRL recipients. Urinary protein and blood urea were assessed. Double negative (DN) lymphocyte proliferation was determined by flow cytometry. Two months after transplantation inflammatory infiltration of the glomerular, vascular and interstitial compartments were determined. Renal function as demonstrated by blood urea levels was normal in MRL recipients, but elevated in LPR recipients, independent of the donor strain. Paralleling functional results, inflammatory infiltration was mild or absent in MRL recipients of MRL grafts, and mild to moderate in MRL recipients of LPR grafts, suggesting that kidney removal from the autoimmune (LPR) environment significantly reduced inflammation. Graft infiltration was most severe in LPR recipients: grafts were similarly inflamed independent of the donor. All LPR recipients had significantly less CD4+ Th cells versus MRL mice. Transplantation of LPR grafts into MRL recipients reduced CD4+ Th cell percentage, accompanied by a slight induction of lupus autoantibody production. Our results demonstrate that lupus nephritis is not kidney specific in the LPR model with recurrence after transplantation in the absence of immunosuppression.

Advances in renal (patho)physiology using multiphoton microscopy.

Multiphoton excitation fluorescence microscopy is a state-of-the-art confocal imaging technique ideal for deep optical sectioning of living tissues. It is capable of performing ultrasensitive, quantitative imaging of organ functions in health and disease with high spatial and temporal resolution which other imaging modalities cannot achieve. For more than a decade, multiphoton microscopy has been successfully used with various in vitro and in vivo experimental approaches to study many functions of different organs, including the kidney. This study focuses on recent advances in our knowledge of renal (patho)physiological processes made possible by the use of this imaging technology. Visualization of cellular variables like cytosolic calcium, pH, cell-to-cell communication and signal propagation, interstitial fluid flow in the juxtaglomerular apparatus (JGA), real-time imaging of tubuloglomerular feedback (TGF), and renin release mechanisms are reviewed. A brief summary is provided of kidney functions that can be measured by in vivo quantitative multiphoton imaging including glomerular filtration and permeability, concentration, dilution, and activity of the intrarenal renin-angiotensin system using this minimally invasive approach. New visual data challenge a number of existing paradigms in renal (patho)physiology. Also, quantitative imaging of kidney function with multiphoton microscopy has tremendous potential to eventually provide novel non-invasive diagnostic and therapeutic tools for future applications in clinical nephrology.

Skin disease is prevented but nephritis is accelerated by multiple pregnancies in autoimmune MRL/LPR mice.

The role of pregnancy in the progression of systemic lupus erythematosus (SLE) is still poorly understood. We analysed the effect of repeated pregnancies in MRL/lpr mice, a murine model of SLE. Seven-week old female mice were used: multiparous mice underwent three consecutive pregnancies (M); age-matched virgin mice served as controls (V). Animals were harvested at 20 weeks of age. Skin lesions were characterized by hair loss and scabs in the dorsum of the neck. Virgin skins showed thickened dermis, fibrosis and mononuclear cell infiltrates, which were practically absent in M. This was accompanied by higher IFN-gamma and lower IL-10 mRNA expression levels in V compared to M skin. Plasma IFN-gamma protein levels were also upregulated in V versus M. However, survival and kidney function were dramatically reduced and accompanied by hypertension after multiple pregnancies. Kidney histology also showed markedly increased renal lesions in M. In contrast to plasma and skin levels, both IL-10 and IFN-gamma mRNA were lower in the kidneys of V versus M mice. Concluding our findings, the pathomechanisms of lupus kidney and skin disease may be regulated differently at the organ level during pregnancy. Both IFN-gamma and IL-10 may be important regulatory cytokines at the local level.

Renin-angiotensin system affects endothelial morphology and permeability of renal afferent arteriole.

The afferent arteriole (AA) is an important regulatory site of renal function and blood pressure. We have demonstrated endothelial fenestration and high permeability in the vicinity of renin granulated epithelioid cells in the juxtaglomerular portion of the afferent arteriole in different mammals. The permeability of fenestrated endothelium of afferent arteriole may be important in connection to various physiologic and pathophysiologic processes. We have assumed that the permeable fenestration may serve as a communication channel between the intravascular circulation and a pathway for renin secretion. Utilising the multiphoton image technique we were able to visualise the endothelial fenestration and renin granules of the in vitro microperfused AA and in vivo AA. We demonstrated that ferritin-positive, i.e., permeable portion of the afferent arteriole, under control conditions is on average 45 microm, which is about one-third to half of the total length of the afferent arteriole. The length of this portion is not constant and can change by physiologic and pharmacologic manipulation of renin formation. The permeability of the afferent arteriole is not changing only parallel with the pharmacologically stimulated renin secretion as already demonstrated in adult rats, but also with the change of renin appearance in afferent arteriole within the very first few days of life after birth. Independently from the age there is a significant correlation between the renin-positive and permeable portion of the AA. Further studies are necessary to clarify the physiological significance of afferent arteriolar permeability and its changes in the postnatal development of the kidney, as well as in correlation with activity of renin- angiotensin system.

Angiotensin II activates plasminogen activator inhibitor-I promoter in renal tubular epithelial cells via the AT1 receptor.

BACKGROUND: Plasminogen activator inhibitor-1 (PAI-1) regulates normal extracellular matrix (ECM) metabolism and it is a key regulator of the fibrotic process. Both angiotensin II (Ang II) and angiotensin IV (Ang IV) have been reported to stimulate PAI-1 expression. It is not known how PAI-1 expression is regulated by the renin-angiotensin system (RAS) in renal tubular cells. METHODS: To dissect signaling mechanisms contributing to the up-regulation of the PAI-1 promoter, porcine proximal tubular cells stably expressing the rabbit AT1 receptor (LLC-PK/AT1) were transiently transfected with a luciferase reporter construct containing the PAI-1 promoter. Promoter activation was assessed by measuring luciferase activity from cell lysates. RESULTS: Ang II dose-dependently stimulated the transcriptional activity of the PAI-1 promoter in renal proximal tubular cells whereas Ang IV had no consistent effect on the promoter activity. Neither inhibition of the Extracellular Signal Regulated Kinase (ERK) cascade nor inhibition of the c-Jun-N-terminal Kinase (JNK) pathway did reduce the stimulation of the PAI-1 promoter by Ang II. However, genistein, a tyrosine kinase inhibitor blocked the effect of Ang II. CONCLUSION: Ang II but not Ang IV activates the PAI-1 promoter in renal proximal tubular cells and this effect is mediated by tyrosine kinases.

Epithelial-mesenchymal transition in renal tubular cells in the pathogenesis of progressive tubulo-interstitial fibrosis.

Epithelial-mesenchymal transition (EMT) plays an important role in embryogenesis and organ formation. Over the last 10-15 years it has been established that EMT is a significant mechanism of tumor progression and metastasis formation and also of progressive tissue fibrosis in the kidney, liver and lung. EMT seen in these diverse physiological and pathophysiological contexts shares a number of stages and modules, but also carries distinct, context specific characteristics. EMT in tissue fibrosis is a form of reverse embryogenesis, when highly specialized epithelial cells in the specific organs will respond to injury with loosing their epithelial characteristics and functions and regaining characteristics of the cells from which they originated. EMT in the context of tissue fibrosis can be induced by different forms of injury or a set of humoral factors. The process is regulated by a complex balance of humoral and microenvironmental stimuli, in which cell-cell contacts and interaction of the transitioning cell with the extracellular matrix components is very important. Intense research in this exciting field yielded good understanding of many of the details of this fascinating process, although numerous questions still await proper answers. There is indication that understanding of the molecular mechanisms underlying "fibrotic" EMT may lead to the design of specific and effective therapeutic measures for progressive tissue fibrosis.

Anemia is associated with mortality in kidney-transplanted patients--a prospective cohort study.

Although anemia is a known risk factor of mortality in several patient populations, no prospective study to date has demonstrated association between anemia and mortality in kidney-transplanted patients. In our prospective cohort study (TransQol-HU Study), we tested the hypothesis that anemia is associated with mortality and graft failure (return to dialysis) in transplanted patients. Data from 938 transplanted patients, followed at a single outpatient transplant center, were analyzed. Sociodemographic parameters, laboratory data, medical history and information on comorbidity were collected at baseline. Data on 4-year outcome (graft failure, mortality or combination of both) were collected prospectively from the patients' charts. Both mortality and graft failure rate during the 4-year follow-up was significantly higher in patients who were anemic at baseline (for anemic vs nonanemic patients, respectively: mortality 18% vs. 10%; p < 0.001; graft failure 17% vs 6%; p < 0.001). In multivariate Cox proportional hazard models the presence of anemia significantly predicted mortality (HR = 1.690; 95% CI: 1.115-2.560) and also graft failure (HR = 2.465; 95% CI: 1.485-4.090) after adjustment for several covariables. Anemia, which is a treatable complication, is significantly and independently associated with mortality and graft failure in kidney-transplanted patients.

The effect of local nitric oxide synthase inhibition on the diameter of pulpal arteriole in dental bond material-induced vasodilation in rat.

Local application of dental bond materials can cause pulpal vasodilation and hyperemia. Such local hemodynamic changes may be mediated by alterations in the levels of locally generated nitric oxide (NO). In different species systemic administration of NO synthase inhibitors leads to a decrease in pulpal blood flow. In contrast, the local administration of these inhibitors has not been tested yet. Therefore, the effect of locally blocked NO synthase on the internal diameter of rat pulpal arterioles under basal conditions and immediately after dental bond material application was studied by using vitalmicroscopic technique. The NO synthase blocker (L-NAME) was locally administered on a thinned dentine layer of the left lower incisor. L-NAME reduced the diameter of the pulpal arteriole both in basal and after bond material-induced hyperaemic conditions. These data suggest that the local formation of NO may have a significant role in the acute vasodilation induced by bond material application and also in maintenance of basal pulpal arteriolar tone.

Comparative analysis of pulpal circulatory reaction to an acetone-containing and an acetone-free bonding agent as measured by vitalmicroscopy.

Despite great progress in the production of new dental polymers, application of these products is still controversial. The unlined utilization of cytotoxic adhesive materials on pulpal dentin can adversely influence the pulp, leading to alterations in local microcirculation that can be an early sign of pathological changes. In a previous study by the authors, the effect of an acetone-free bondmaterial was examined on the vascular diameter of pulpal vessels by means of vitalmicroscopy. In this study, experiments comparing experimental data provided by an acetone-containing bondmaterial to these earlier findings with acetone-free ones have been performed. Thirty male Sprague-Dawley rats (weighing 333+/-9 g) were used for this investigation. The first lower incisor was prepared for vitalmicroscopy. Changes in vessel diameter were recorded prior to and 5, 15, 30 and 60 minutes after the investigated materials (Scotchbond Multi-Purpose Dental Adhesive System or Prime & Bond 2.1) were administered on dentin as recommended by the manufacturer. In control rats (saline administration), the vessel diameter was stable during the experiment. In the presence of acetone-free bondmaterial (Scotchbond), the vessel diameter was increased during the experimental period in relation to the baseline (12.15+/-2.85%; 16.36+/-2.39%; 14.16+/-3.48%; 12.12+/-3.72%). In the presence of acetone-containing bondmaterial (Prime & Bond 2.1), a similar result was observed (10.56+/-2.27%; 16.13+/-2.94%; 17.88+/-2.54%; 14.54+/-3.16%). The differences between the control values and those registered with test groups were significant (p<0.05; ANOVA). There was no significant difference among the test groups. The results of this study suggest that dental bond materials applied on a very thin layer of dentin may affect the blood supply to the dental pulp. However, no stasis or prestasis has been detected, indicating a possible reversible effect. The authors could not show any statistical difference between the vasodilatation caused by the acetone-containing and the acetone-free bond material.

[Acute effect of calcium hydroxide-containing material in the rat's dental pulp microcirculation]. / Kálciumhidroxid-tartalmú pulpasapkázó anyag akut hatása patkányfogak pulpális keringésére.

Experimental stimulation and clinical procedures applied on the crown of the tooth cause vascular reaction in the dental pulp. Laser-Doppler flowmetry is a good method for determining the blood flow of the dental pulp. The aim of these experiments was to study the acute effect of a calcium hydroxide-containing pulp-capping material (Dycal, DeTrey) on blood flow of the dental pulp after the application into a deep test-cavity. Two groups of male Sprague-Dawley rats (308 g +/- 50 S.E.) were used: control- and test-group, n = 10. A standardised deep class five cavity was prepared in the left lower incisor of each rat. Laser-Doppler flowmeter was used to measure the vascular reaction of the pulp. The levels of blood flow were recorded prior to (0 min.) and after the application (1, 5, 15, 30, 60 min.) of Dycal. Results were evaluated with one-way ANOVA. Significantly higher pulpal blood flow was found only in the first minute after the application of Dycal. In any other time no significant difference was found between the results before and after (5, 15, 30, 60 min.) the application in test- and control-group.

The use of a second reporter plasmid as an internal standard to normalize luciferase activity in transient transfection experiments may lead to a systematic error.

beta-galactosidase reporter plasmids containing different viral or minimal promoters are commonly used to correct variable transfection efficiencies in transient transfection experiments. The transcriptional activity of these promoters is thought to be stable under most circumstances. To determine if expression of beta-galactosidase from the commonly used beta-galactosidase plasmids remains stable upon stimulation of the cells with agonists we performed transient transfection experiments. CHO cells stably expressing the rat AT(1A) receptor were transfected with RSVbeta- or CMVbeta- or pTKbeta plasmids alone or together with a reporter construct in which luciferase transcription is driven by the c-fos promoter. Luciferase and/or beta-galactosidase activity was measured from the lysate of cells treated with angiotensin II or serum. We found that agonists increased the transcriptional activity of the different beta-galactosidase plasmids. The effect of angiotensin II and serum was different on the different promoters. Finally, cotransfection of other plasmids also modulated beta-galactosidase activity. These agonist induced variations of beta-galactosidase activity may influence the analysis and interpretation of the results in a systematic manner. Consequently we conclude that the use of a second reporter system to control for transfection efficiency in certain types of experiments may lead to a systematic error and is questionable as a general procedure.

[Effect of pentoxifylline on the healing of experimental anastomosis of the left colon in rats]. / A pentoxifyllinum hatása a kísérletes bal colonfél anasztomózis gyógyulására.

The healing of colonic anastomoses is determined by several factors such as microcirculation, the strength of the inflammatory response, and the time required for regeneration. We investigated the effects of pentoxifylline--a drug which improves microcirculation and modulates leukocyte functions--on the healing of experimental anastomosis on the left colon of rats. As a result of drug treatment (0.25 mg/100 g, i.p.) in Group I anastomosis bursting pressure (ABP) was by 56 +/- 17% higher at day 2 than in controls with no pentoxifylline treatment. On the 5th postoperative day in Group I, ABP reached 80 +/- 8% the value for the intact colon (218 +/- 21 mmHg), whereas respective value in the control (untreated) group was only 47 +/- 7%. In Group II (pentoxifylline: 2 mg/100 g, i.p.) ABP was by 55 +/- 10% and by 73 +/- 8% higher than control values at postoperative days 1 and 2, respectively. At day 2, in Group I colonic blood flow measured at the anastomosis line by 86Rb uptake technique was significantly higher than in the untreated controls (0.18 +/- 0.01 ml/min vs. 0.14 +/- 0.01 ml/min, (p < 0.02). Blood flow measured in colon tissue above and below the anastomosis changed differently. Pentoxifylline treatment also suppressed the peritoneal inflammatory response assessed with peritoneal reaction index (2.0 +/- 0.3 vs. 1.1 +/- 0.2, p < 0.01). The results of the present study show that pentoxifylline treatment shortens the time needed for the healing of colonic anastomosis. These observations suggest that pentoxifylline medication can prevent failure of colonic anastomoses.

The effect of acid etching on vascular diameter of pulp-vessels in rat incisor (vitalmicroscopic study).

Conditioning agents used on dentin with composite materials are biologically active and may have deleterious effects on the pulpal microcirculation. No data are available on the immediate vascular effect of etching materials applied on a constant thin pulpal dentin. In this study the authors examined whether the application of 36% phosphoric acid (Conditioner 36, 15 seconds) or itakonic acid with 10% maleic acid (NRC Non-Rinse Conditioner, 20 seconds), as recommended by the manufacturers, alters the blood circulation in the pulp of the rat's lower incisors. The effect of prolonged etching time (60 seconds) was also assessed (Conditioner 36). The application of saline served as the untreated control. The technique of vitalmicroscopy was used on the first lower incisor of 40 (10-10 in each group) male Sprague-Dawley rats (weighing 350 +/- 8 g SE) to record the changes in vessel diameter prior to and at 5, 15, 30 and 60 minutes after the test materials were administered on the dentin. In the control rats, the vessel diameter was stable during the entire experiment. Acid conditioning as recommended by the manufacturers tended to cause vasodilatation, though these alterations were statistically not significant when compared to the control group (ANOVA, p > 0.05). After prolonged etching time (Conditioner 36, 60 seconds) significant vasoconstriction (-14.4 +/- 6.13; -10.59 +/- 4.2; -11.96 +/- 6.75; -5.49 +/- 5.78%) was observed (ANOVA, p < 0.05). In this group, stasis developed in pulpal blood circulation in 40% of rats (Cochran's-Q test, p < 0.05), gas-bubble formation was observed in 30% and the disappearance of the pulpal wall occurred in 20%. These results suggest that exposition time with acid is crucial to the pulpal microcirculation. That is, acid conditioning applied as indicated (for 15-20 seconds) onto a very thin layer of dentin only slightly affects the blood supply to the dental pulp; however, prolonged etching time (for 60 seconds) results in immediate failure of microcirculation in the dental pulp of rats.

Extracellular signal-regulated kinase and the small GTP-binding protein p21Rac1 are involved in the regulation of gene transcription by angiotensin II.

To study the role of extracellular-signal-regulated kinase (ERK) cascade and the small GTP-ase proteins in the activation of the c-fos promoter by angiotensin II (AII), transient transfection experiments were performed in CHO cells stably expressing the rat AT(1A) receptor. In this system AII activated ERK in 1 min and also increased the transcriptional activity of the c-fos promoter-luciferase reporter gene construct. The activation of the promoter proved to be dependent on the Ras-Raf-ERK cascade as cotransfection of expression vectors known to specifically inhibit this cascade blocked the effect of AII. Dominant-negative p21Rac1 mutant partially blocked the activation of the c-fos promoter by AII. However, activation of the c-fos promoter was independent of protein kinase C (PKC) as bisindolylmaleimide I, a specific PKC inhibitor did not block the effect of AII. These results suggest that AII activates the transcription of the c-fos through the Ras-Raf-ERK cascade. Furthermore, p21Rac1 is involved in the modulation of the c-fos promoter by AII.