823
Effects of endothelin receptor blockade on hypervasoreactivity in streptozotocindiabetic rats: vessel-specific involvement of thromboxane A2 Emi Arikawa, Claudia Cheung, Inna Sekirov, Mary L. Battell, Violet G. Yuen, and John H. McNeill
Abstract: Increased vasoconstrictor response to norepinephrine (NE) and endothelin (ET)-1 in arteries from diabetic animals is ameliorated by chronic endothelin receptor blockade with bosentan and was absent in endothelium-denuded arteries, suggesting the involvement of ET-1 and an endothelium-derived contracting factor such as thromboxane A2 (TxA2). To examine this possibility, we determined the effects of acute blockade of ET receptors or inhibition of TxA2 synthesis on the vascular function of superior mesenteric arteries (SMA) and renal arteries (RA) isolated from nondiabetic and 11week streptozotocin (STZ) diabetic rats chronically treated with either bosentan or vehicle. Both in vitro incubation with bosentan and a selective ETA receptor blocker, BQ123, eradicated the increase in NE contractile responses in diabetic SMA. Additionally, in vitro incubation with the thromboxane synthase inhibitor, dazmegrel, abrogated the exaggerated NE and ET-1 contractile responses in diabetic SMA. Conversely, in RA, no significant acute effect of bosentan, BQ123, nor dazmegrel on vascular responses to NE was observed. Dazmegrel incubation attenuated the maximum contractile responses to ET-1 in diabetic RA; however, these responses in diabetic RA remained significantly greater than those of other groups. Diabetic RA but not SMA exhibited an enhanced contractile response to the TxA2 analogue U46619, which was corrected by chronic bosentan treatment. Immunohistochemical analyses in diabetic SMA revealed an increase in ETA receptor level that was normalized by chronic bosentan treatment. These data indicate that an interaction between ET-1 and TxA2 may be involved in mediating the exaggerated vasoconstrictor responses in diabetic arteries. Furthermore, the underlying mechanisms appear to be vessel specific. Key words: endothelin-1, thromboxane A2, streptozotocin, diabetic rats, vascular reactivity, bosentan, dazmegrel, superior mesenteric arteries, renal arteries. Re´sume´ : La re´ponse vasoconstrictrice accrue a` la nore´pine´phrine (NE) et a` l’endothe´line (ET)-1 dans les arte`res d’animaux diabe´tiques a e´te´ normalize´e par le blocage chronique des re´cepteurs de l’endothe´line avec du bosentan et absente dans les arte`res de´nude´es d’endothe´lium, laissant croire a` l’intervention de l’ET-1 et d’un facteur contractant d’origine endothe´liale comme la thromboxane A2 (TxA2). Pour examiner cette possibilite´, nous avons de´termine´ les effets du blocage aigu des re´cepteurs ET ou ceux de l’inhibition de la synthe`se de la TxA2 sur la fonction vasculaire d’arte`res me´sente´riques supe´rieures (AMS) et d’arte`res re´nales (AR) isole´es provenant de rats non diabe´tiques et de rats rendus diabe´tiques par l’administration (11 semaines) de streptozotocine, traite´s de fac¸on chronique avec du bosentan ou un ve´hicule. Tant l’incubation in vitro avec le bosentan qu’avec un bloqueur se´lectif du re´cepteur ETA, BQ123, a supprime´ l’augmentation des re´ponses contractiles a` la NE dans les AMS diabe´tiques. De plus, l’incubation in vitro avec l’inhibiteur de la thromboxane synthase, dazmegrel, a supprime´ les re´ponses contractiles exage´re´es a` la NE et a` l’ET-1 dans les AMS diabe´tiques. Re´ciproquement, dans les AR, aucun effet aigu notable du bosentan, de BQ123 ou du dazmegrel sur les re´ponses vasculaires a` la NE n’a e´te´ observe´. L’incubation avec le dazmegrel a atte´nue´ les re´ponses contractiles maximales a` l’ET1 dans les AR diabe´tiques; toutefois, ces re´ponses sont demeure´es conside´rablement plus e´leve´es que celles des autres groupes. Les AR, mais pas les AMS, diabe´tiques ont pre´sente´ une re´ponse contractile accrue a` l’analogue de la TxA2, U46619, qui a e´te´ corrige´e par le traitement chronique au bosentan. Des analyses immunohistochimiques dans les AMS diabe´tiques ont re´ve´le´ une augmentation du taux du re´cepteur ETA, qui a e´te´ normalize´e par le traitement chronique au bosentan. Ces re´sultats indiquent qu’une interaction entre l’ET-1 et la TxA2 pourrait eˆtre jouer un roˆle dans la me´diation des re´ponses vasoconstrictrices exage´re´es dans les arte`res me´sente´riques. Le ou les me´canismes sous-jacents semblent eˆtre spe´cifiques aux vaisseaux.
Received 6 January 2006. Published on the NRC Research Press Web site at http://cjpp.nrc.ca on 27 October 2006. E. Arikawa, C. Cheung, I. Sekirov, M.L. Battell, V.G. Yuen, and J.H. McNeill.1 Division of Pharmacology and Toxicology, Faculty of Pharmaceutical Sciences, The University of British Columbia, Vancouver, BC V6T 1Z3, Canada. 1Corresponding
author (e-mail:
[email protected]).
Can. J. Physiol. Pharmacol. 84: 823–833 (2006)
doi:10.1139/Y06-042
#
2006 NRC Canada
824
Can. J. Physiol. Pharmacol. Vol. 84, 2006
Mots cle´s : endothe´line-1, thromboxane A2, streptozotocine, rats diabe´tiques, re´activite´ vasculaire, bosentan, dazmegrel, arte`res me´sente´riques supe´rieures, arte`res re´nales. [Traduit par la Re´daction]
______________________________________________________________________________________ Introduction Endothelin-1 (ET-1) is a potent endothelium-derived contracting factor that is established as a potential candidate mediator in the development of several diabetic complications including retinopathy (Bursell et al. 1995; Takagi et al. 1996; Deng et al. 1999), nephropathy (Fukui et al. 1993; Benigni et al. 1998; Peppa-Patrikiou et al. 1998), neuropathy (Jarvis et al. 2000), and cardiomyopathy (Chen et al. 2000; Verma et al. 2001). Indeed, several reports, including ours, have demonstrated that blockade of the endothelin system has favourable effects on the cardiovascular system in experimental diabetes (Deng et al. 1999; Gilbert et al. 2000; Chen et al. 2000; Verma et al. 2001; Verma et al. 2002; Arikawa et al. 2001). In previous work, we demonstrated that isolated arteries from 11-week streptozotocin diabetic rats showed an increased tissue level of ET-1 and an augmented contractile response to norepinephrine (NE) and ET-1 (Verma et al. 2001; Arikawa et al. 2001). The hyper-reactivity was ameliorated by chronic treatment with the endothelin receptor antagonist bosentan, suggesting that ET-1 could contribute to this vascular derangement. This enhanced vasoconstrictor response was shown to be dependent on the presence of an intact endothelium, but it did not appear to be related to decreased endothelium-derived nitric oxide (NO) production (Arikawa et al. 2001). The endothelium dependence of the increased response to vasoconstrictors in the diabetic rats suggests that an endothelium-derived vasoconstrictor such as thromboxane A2 (TxA2) and the endoperoxide prostaglandin H2 (PGH2) may be important in mediating this effect (Agrawal and McNeill 1987; Cohen 1993). Earlier work from our laboratory demonstrated that the hyper-responsiveness to NE in superior mesenteric arteries from diabetic rats was corrected by in vitro inhibition of cyclooxygenase with indomethacin (Agrawal and McNeill 1987), implying that an altered synthesis of endogenous vasoconstrictor eicosanoids could be a factor in the enhanced NE responses of the diabetic vessels. Indeed, an increase in TxA2 release from diabetic aorta has been previously demonstrated (Tesfamariam et al. 1989; Cohen 1993). Additionally, experimental diabetes has been shown to increase contractile responses to U46619, a TxA2 analogue (Hattori et al. 1999). Multiple lines of evidence have shown an interaction between the ET-1 and TxA2 systems. For example, ET-1 has been shown to stimulate the production of TxA2 (Reynolds and Mok 1990; Zaugg et al. 1996; Muck et al. 1993; Galipeau et al. 2001). Furthermore, ET-1-induced vasoconstriction is shown to be dependent on TxA2 in human placental vessels (Howarth et al. 1995). Thromboxane A2 appears to play a role in the vasoconstrictor effect of ET-1 in the animal models of 2 different diseases, the spontaneously hypertensive rat and the post-ischemic rat heart, but not in their control counterparts (i.e., the Wistar–Kyoto rat and the nonischemic rat heart, respectively) (Taddei and Vanhoutte
1993a; Howarth et al. 1995). Additionally, Moreau et al. (1996) demonstrated that bosentan was able to produce a parallel rightward shift of the contractions to U46619, indicating the antagonism of the direct stimulation of TxA2 receptors by the mixed ET receptor blocker. Based on the above discussion, it is plausible that bosentan improved vascular responses in diabetes by blocking ET-1-induced TxA2 synthesis and (or) release and (or) action. Therefore, experiments were conducted to elucidate (i) whether hyper-reactivity in diabetic arteries was mediated by direct vascular actions of ET-1 through its receptors, (ii) whether TxA2 is involved in this hyper-responsiveness to vasoconstrictors in diabetes, and (iii) whether the beneficial effect of chronic bosentan treatment observed in diabetic arteries was related to its ability to block the direct effects of ET-1 and (or) its effects on the interaction between the ET-1 and TxA2 systems.
Materials and methods Materials BQ123, 3,3’-diaminobenzidine tetrahydrochloride with cobalt enhancer (Sigma FastTM DAB with Metal Enhancer), human/porcine endothelin-1 (ET-1), gum arabic, disodium EDTA (ethylenediaminetetraacetic acid), norepinephrine (NE) hydrochloride (arterenol hydrochloride), streptozotocin (STZ), U46619 (9,11-dideoxy-11a,9a-epoxymethanoprostaglandin F2a) were obtained from Sigma, St. Louis, Mo. Sheep polyclonal anti-ETA receptor antibodies were obtained from Research Diagnostics, Inc., Flanders, N.J. Sheep polyclonal anti-ETB receptor antibodies were from Maine Biotechnology Services, Inc., Portland, Me. Vectastain Elite kits were purchased from Vector Laboratories, Inc., Burlingame, Calif. Radioimmunoassay kits for insulin were from Linco Research Inc., St. Charles, Mo. Bosentan was a generous gift from Dr. M. Clozel, Actelion, Allschwil, Switzerland. Dazmegrel (UK-038485) was generously provided by Pfizer, Ltd., Sandwich, Kent, UK. Unless otherwise stated, all other chemicals were reagent grade and obtained from either Sigma, BDH Inc., Toronto, Ont., or Fisher Scientific, Nepean, Ont. Experimental design Male Wistar rats of approximately 190–220 g were obtained from Charles River Laboratories Inc., St. Charles, Que. They were cared for in accordance with the principles and guidelines of the Canadian Council on Animal Care and protocol for experimental animal use were reviewed by the Animal Care Committee, The University of British Columbia. The rats were housed on a 12-h light (06:00–18:00): 12-h dark cycle and received rat chow and water ad libitum. Rats were randomly assigned to 2 groups. One group received a single caudal vein injection of STZ at a dose of 60 mg/kg body mass (under halothane anesthesia) and served as the diabetic group. The other group was injected #
2006 NRC Canada
Arikawa et al.
with the vehicle saline solution (0.9% sodium chloride) and served as the age- and mass-matched control group. The rats injected with STZ were checked for hyperglycemia (set at ‡15 mmol/L in our laboratory) at 72 h using Accu-Chek Advantage reagent strips read by an Accu-Chek glucose meter (Roche Diagnostics Corporation, Indianapolis, Ind.). The control and diabetic rats were further divided into control (C), control bosentan-treated (CT), diabetic (D), and diabetic bosentan-treated (DT). One week after STZ injection, oral bosentan treatment (100 mgkg–1day–1 by daily oral gavage; bosentan suspended in 1% gum arabic) was administered to the CT and DT groups for 10 weeks whereas the C and D rats received the vehicle (1% gum arabic). Five-hour fasted blood samples were collected from the tail vein for determination of plasma glucose and insulin levels. Immunohistochemistry Relative tissue ET receptor protein expression in the arteries was assessed by immunohistochemical analyses of ETA receptor-immunoreactivity (ETA-ir) and ETB receptorir (ETB-ir) as described previously (Verma et al. 2001) with a few modifications. Superior mesenteric arteries and renal arteries isolated from the 4 experimental rat groups were immersed in 10% formalin (for 8 h) and embedded in paraffin. Slide sections of paraffin-embedded tissue were dewaxed in xylene. Endogenous peroxidase activity was blocked by 1.5h incubation in methanol containing 0.6% H2O2, and the slides were blocked with 5% normal rabbit serum for 1 h at room temperature. The slides were then subsequently incubated with either sheep polyclonal anti-ETA receptor antibodies (at a 1:180 dilution) or sheep polyclonal anti-ETB receptor antibodies (at a 1:250 dilution) at 4 8C for 45 h. Nonimmune sheep serum was used as control for verifying the specificity of staining. The slides were then washed in PBS for 10 min and immunostained with an avidin–biotin– peroxidase system (Vectastain Elite kit, Vector Laboratories, Inc.) using diaminobenzidine (with cobalt enhancer, Sigma) as the chromagen. A digital imaging system was used for assessing the intensity of staining. Slides were viewed through a Nikon Diaphot TMD inverted microscope. A video camera connected to an IBM compatible computer with Northern Eclipse Software (Empix Imaging Inc., Mississauga, Ont.) converted the data to a digital image, each with a gray value ranging from 0 to 255. For each tissue, 2 (for renal arteries) or 4 (for superior mesenteric arteries) different images were taken. Areas with an equal size were selected for each image. A threshold above which pixels were counted was established. Any areas darker than the threshold would be recognized as ‘‘objects’’. Percent object area (= total object area/ selected area 100%) from each image was obtained. Values from the images were averaged for each rat tissue. Vascular reactivity study protocols Following 10 weeks of bosentan treatment, superior mesenteric (SMA) and renal arteries (RA) were isolated for the assessment of vascular reactivity as previously described (Verma et al. 2001; Arikawa et al. 2001). Each artery was cut into 2 rings (2–3 mm in length) and suspended on wire hooks in individual tissue baths containing modified Krebs– Ringer bicarbonate solution maintained at 37 8C, and oxygenated with 95% O2 and 5% CO2. Following an initial
825
equilibration period (60 min), the tissues were exposed to agonists and isometric vascular responses recorded according to the following order: (i) a cumulative concentration response curve (CRC) to NE alone (10–9–10–4 mol/L), (ii) a CRC to NE constructed in the presence of either BQ123 (1 mmol/L incubated for 30 min prior to CRC) or bosentan (1 mmol/L preincubated for 30 min), (iii) a CRC to NE in the presence of dazmegrel (1 mmol/L for 30 min), (iv) a CRC to U46619 (10–12–10–6 mol/L), or (v) a CRC to ET-1 (10–10 – 3 10–8 mol/L) constructed either in the absence or in the presence of dazmegrel (1 mmol/L for 30 min). At the end of the experiment, the tissues were removed and blotted dry, and the cross-sectional area of each vascular ring was calculated: cross-sectional area (mm2) = mass (mg)/(length (mm) density (mg/mm3)). The density of the arteries was assumed to be 1.05 mg/mm3 (Verma et al. 1996). The absolute tension generated was corrected for cross-sectional area and expressed as g/mm2. Agonist pD2 values (–log ED50) were calculated by nonlinear regression analysis of the individual CRCs and used as an index of sensitivity (Verma et al. 1996). Biochemical measurements Plasma glucose was measured with an automatic Beckman Glucose Analyzer 2 (Beckman Instruments, Inc., Diagnostic Systems Group, Brea, Calif.). Plasma insulin levels were measured using double antibody radioimmunoassay kits from Linco Research Inc. Statistical analyses Values are expressed as mean ± SE; n indicates the number of rats in each group. Statistical analyses were performed using a 1-way analysis of variance (ANOVA) or a repeated measures ANOVA (general linear models ANOVA), followed by a Newman–Keuls test, with the Number Cruncher Statistical System (Kaysville, Utah). The level of significance was set at p < 0.05.
Results General characteristics of untreated and bosentantreated control and STZ-diabetic rats The general characteristics of the untreated and bosentantreated rats at basal and week 10 are summarized in Table 1. Induction of STZ-diabetes in D rats resulted in characteristic symptoms of diabetes including hyperglycemia, hypoinsulinemia, decreased body mass gain, increased food and fluid intake when compared with age-matched controls (C rats). Bosentan treatment did not affect the plasma glucose or insulin levels in either control (CT) or diabetic (DT) rats, nor did it affect body mass gain and food and fluid intake in these rats. Effects of chronic bosentan treatment on the expression of ETA and ETB receptors in diabetic rat arteries The protein expression of ETA receptors, as analyzed by immunohistochemistry, was increased in the superior mesenteric arteries from diabetic (D) rats that exhibited a higher intensity of anti-ETA receptor immunostaining than those of control (C) rats (Figs. 1 and 2), whereas there was no change in ETA receptor expression in the renal arteries after #
2006 NRC Canada
826
Can. J. Physiol. Pharmacol. Vol. 84, 2006 Table 1. Representative general characteristics of the rats before and after 10 weeks of bosentan treatment. C (n = 8) Body mass (g) Food intake (g/day) Fluid intake (mL/day) Plasma insulin (ng/mL) Plasma glucose (mmol/L)
Basal 227±2 30±1 71±2 2.5±0.3 7.9±0.2
CT (n = 8) Week 10 482±14 38±5 78±5 2.7±0.3 7.1±0.1
Basal 223±3 34±0 75±3 2.5±0.3 8.2±0.1
D (n = 9) Week 10 482±6 34±1 79±3 3.2±0.4 7.3±0.1
Basal 222±4 54±1* 287±8* 1.0±0.1* 24.0±0.9*
DT (n = 8) Week 10 366±15* 50±1* 259±14* 0.4±0.1* 28.4±1.5*
Basal 225±3 51±2* 284±11* 1.0±0.1* 23.1±1.2*
Week 10 371±13* 50±4* 253±10* 0.4±0.1* 24.6±2.6*
Note: Values are expressed as mean ± SE. C, CT, D, and DT denote control, control bosentan-treated, diabetic, and diabetic bosentan-treated rats, respectively. *p < 0.05 vs. C and CT.
Fig. 1. ETA receptor-like immunoreactivity in superior mesenteric arteries. Representative figures showing ETA receptor-like immunoreactivity (examined by immunohistochemistry) in superior mesenteric arteries from untreated control, control bosentan-treated, diabetic, and diabetic bosentan-treated rats.
11 weeks of STZ-diabetes (data not shown). Interestingly, treatment with bosentan restored the ETA receptor levels in diabetic SMA to control levels (Figs. 1 and 2). No effect of bosentan treatment on the expression of RA ETA receptor levels was seen in either control or diabetic rats (not shown). The immunoreactivity for ETB receptors in the SMA and RA did not differ among rat treatment groups (data not shown). The specificity of the polyclonal antibodies against ETA and ETB receptors was demonstrated by the absence of immunostaining in tissue sections incubated with nonimmune sheep serum (data not shown). Vascular reactivity in SMA In agreement with our published results (Verma et al. 2001), the maximum contractile responses of SMA to NE and ET-1 were markedly increased in the untreated diabetic rats when compared with the control rats (Fig. 3). This hyper-reactivity towards NE and ET-1 was completely corrected after long-term bosentan treatment in the DT rats (Fig. 3). Similarly to the effects produced by the long-term bo-
Fig. 2. ETA receptor protein levels examined by immunohistochemistry in superior mesenteric arteries from control (C) (n = 8), control bosentan-treated (CT) (n = 9), diabetic (D) (n = 8), and diabetic bosentan-treated (DT) (n = 6) rats. Each bar represents the mean ± SE. *p < 0.05 vs. C, CT, and DT.
sentan treatment, in vitro incubation with bosentan in untreated diabetic SMA also abolished the exaggerated NE response, in that the diabetic arteries exhibited similar contractile responses towards NE as the arteries from the control rats (maximum tension in g/mm2: C, 12.67 ± 1.20 vs. D, 14.90 ± 1.20, p > 0.05; Fig. 4A). Similarly, following the in vitro incubation with the selective ETA blocker, BQ123, the hyper-reactivity of the D group towards NE was normalized, with the maximum contractile response to NE in the diabetic group being restored to the control level (maximum tension in g/mm2: C, 11.54 ± 1.26, D, 11.36 ± 0.20, DT, 12.50 ± 1.61, p > 0.05; Fig. 4A). Figures 5A and 5B illustrate the effects of the in vitro incubation with the thromboxane synthase inhibitor dazmegrel on the vasoconstrictor responses to NE and ET-1, respectively, in SMA. Dazmegrel incubation resulted in the normalization of the increased response to NE in the untreated diabetic arteries to the control values (maximum tension in g/mm2: C, 12.51 ± 1.20 vs. D, 14.68 ± 2.73, p > 0.05; Fig. 5A). As with NE responses, the exaggerated contractile response towards ET-1 of the untreated diabetic SMA was abolished following in vitro incubation with dazmegrel when compared with control arteries (maximum tension in g/mm2: C, 13.42 ± 1.21 vs. D, 16.33 ± 2.79, p > 0.05; Fig. 5B). No difference in the maximum contractile responses was #
2006 NRC Canada
Arikawa et al.
827
Fig. 3. Contractile responses to norepinephrine (NE) and endothelin-1 (ET-1) in arteries from control and diabetic rats. (A) NE and (B) ET1 concentration response curve in superior mesenteric arteries and renal arteries from untreated control (C), control bosentan-treated (CT), diabetic (D), and diabetic bosentan-treated (DT) rats. n = 5–8 per group. Each point represents the mean ± SE. *p < 0.05 vs. C; #p < 0.05 vs. CT; {p < 0.05 vs. DT.
observed between the untreated diabetic and control arteries in the contractile responses towards the TxA2 analogue, U46619 (Fig. 6). Moreover, chronic bosentan treatment did not affect U46619 responses in control or diabetic SMA. Diabetic arteries exhibited a greater response to U46619 at concentrations higher than 3 10–7 mol/L when compared with CT arteries (Fig. 6). Chronic bosentan treatment did not affect the contractile response to NE, ET-1, or U46619 in control rats (i.e., CT rats). The sensitivity to NE and ET-1 did not differ among the SMA from different rat groups (Table 2A). Intriguingly, in contrast to the results on the maximum contractile responses, data from the nonlinear regression analysis revealed a lower sensitivity to the thromboxane analogue, U46619, in D than in CT arteries (pD2 values for U46619: CT, 9.51 ± 0.35 vs. D, 8.14 ± 0.26, p < 0.05; Table 2A). Chronic bosen-
tan treatment did not affect the agonist sensitivity to NE, U46619, nor ET-1 in the SMA from all rats. As well, in vitro incubation with bosentan, BQ123, or dazmegrel did not appear to have any effect on the sensitivity to NE or ET-1 in the arteries from control and diabeitc groups. Vascular reactivity in RA Similarly to the observations made in the SMA, RA from the untreated diabetic rats exhibited an exaggerated contractile response to NE and ET-1 and chronic bosentan treatment normalized this hyper-reactivity in the diabetic treated group (Fig. 3). However, unlike the results obtained in the SMA, in vitro incubation with bosentan or BQ123 did not normalize the exaggerated response to NE in the untreated diabetic RA (maximum tension in g/mm2: NE+bosentan: C, 11.80 ± #
2006 NRC Canada
828
Can. J. Physiol. Pharmacol. Vol. 84, 2006
Fig. 4. Norepinephrine (NE) concentration response curve (CRC) in the presence of bosentan or BQ123 (1 mmol/L for 30 min) in (A) superior mesenteric arteries and (B) renal arteries from untreated control (C) and diabetic (D) rats. Each point represents the mean ± SE; n = 6–8 per group. *p < 0.05 vs. C.
1.63 vs. D, 24.11 ± 3.53, p < 0.05; and NE + BQ123: C, 18.64 ± 2.82 vs. D, 31.35 ± 5.50, p < 0.05; Fig. 4B). In vitro incubation with dazmegrel also did not correct the maximum contractile response to NE in the untreated diabetic arteries (maximum tension in g/mm2: C, 10.75 ± 1.19 vs. D, 23.43 ± 3.79, p < 0.05; Fig. 5A). Inhibition of thromboxane synthase with dazmegrel resulted in a slight but significant decrease in the maximum contractile responses to ET-1 in diabetic RA (maximum tension of D renal arteries in g/mm2: ET-1, 30.54 ± 4.97 vs. ET-1 + dazmegrel, 21.18 ± 4.28, p < 0.05; Figs. 3B and 5B); however, these responses in diabetic arteries remained significantly greater than those of the control group in the presence of dazmegrel (maximum tension in g/mm2: C, 8.65 ± 1.04 vs. D, 21.18 ± 4.28, p < 0.05; Fig. 5B). In response to U46619, the untreated diabetic arteries had a significantly higher maximum contraction compared with the control, and this exaggerated response was completely normalized with chronic bosentan treatment in the DT group
(maximum tension in g/mm2: C, 11.54 ± 2.29; D, 21.62 ± 2.81*; DT, 6.65 ± 1.40, *p < 0.05 vs. C and DT; Fig. 6A). Chronic bosentan treatment did not cause any changes to the contractile responses to NE, ET-1, or U46619 in the control rats. The agonist sensitivity to NE and U46619 did not differ among the RA from different rat groups, indicating that chronic bosentan treatment did not affect the sensitivity to these vasoactive substances in the RA (Table 2B). In vitro incubation with bosentan or BQ123 elicited no effect on the sensitivity of control or diabetic arteries to NE. However, dazmegrel incubation resulted in a small decrease in the sensitivity to NE in control RA (pD2 values: in C: NE, 6.65 ± 0.09 vs. NE + dazmegrel, 6.35 ± 0.08, p < 0.05; Table 2B). Data from the nonlinear regression analysis indicated a higher sensitivity to ET-1 in untreated diabetic arteries as compared with control arteries; this increase in the sensitivity remained significant after chronic bosentan treatment in DT rats (pD2 values for ET-1: C, 8.25 ± 0.04; D, 8.66 ± 0.11*; DT, 8.65 ± 0.07*; p < 0.05 vs. C; Table 2B). #
2006 NRC Canada
Arikawa et al.
829
Fig. 5. (A) Norepinephrine (NE) and (B) endothelin-1 (ET-1) concentration response curve in the presence of dazmegrel (1 mmol/L for 30 min) in superior mesenteric arteries and renal arteries from untreated control (C) and diabetic (D) rats. Each point represents the mean ± SE; n = 6–8 per group. *p < 0.05 vs. C.
Fig. 6. U46619 concentration response curve in superior mesenteric arteries and renal arteries from untreated control (C), control bosentantreated (CT), diabetic (D), and diabetic bosentan-treated (DT) rats. Each point represents the mean ± SE from n = 5–8 per group. *p < 0.05 vs. C; #p < 0.05 vs. CT; {p < 0.05 vs. DT.
#
2006 NRC Canada
830
Can. J. Physiol. Pharmacol. Vol. 84, 2006 Table 2. Sensitivities to various agents. Sensitivity (pD2) C
CT
D
DT
(A) Sensitivities to various agents in the superior mesenteric arteries from the 4 rat groups NE 6.84±0.06 6.74±0.17 6.87±0.16 6.98±0.11 NE + Bos 6.78±0.10 7.00±0.22 NE + BQ123 6.59±0.09 6.82±0.11 NE + Daz 6.58±0.07 6.71±0.19 9.22±0.34 U46619 9.09±0.38 9.51±0.35 8.14±0.26{ ET-1 8.40±0.06 8.37±0.09 8.33±0.09 8.45±0.09 ET-1 + Daz 8.37±0.20 8.41±0.06 (B) Sensitivities to various agents in the renal arteries from the 4 rat groups NE 6.65±0.09 6.54±0.09 6.81±0.10 6.84±0.10 NE + Bos 6.55±0.05 6.61±0.10 NE + BQ123 6.42±0.07 6.55±0.07 NE + Daz 6.35±0.08{ 6.48±0.06 U46619 7.40±0.74 8.20±0.26 8.54±0.53 7.65±0.42 ET-1 8.25±0.04 8.30±0.12 8.66±0.11*{ 8.65±0.07*{ ET-1 + Daz 8.29±0.06 8.55±0.11 Note: Values are expressed as mean ± SE. pD2 represents –log ED50. n = 5–8 per group. *p < 0.05 vs. C; {p < 0.05 vs. CT; {p < 0.05 vs. NE alone in the same rat group. C, control rats; CT, control bosentan-treated rats; D, diabetic; DT, diabetic bosentan-treated rats; NE, norepinephrine; Bos, bosentan; Daz, dazmegrel; ET-1, endothelin-1.
Discussion We have previously shown that increased responses to vasoconstrictors in isolated arteries from diabetic animals were ameliorated by chronic endothelin receptor blockade with bosentan and were absent in endothelium-denuded arteries (Arikawa et al. 2001; Verma et al. 2001), which suggests the involvement of ET-1 and an endothelium-derived contracting factor such as thromboxane A2. Confirming our previous findings, in the present study we demonstrated that the augmented NE and ET-1 contractile responses in both endothelium-intact SMA and RA from the diabetic rats were normalized with chronic treatment with the mixed ETA and ETB receptor antagonist, bosentan. Effects of in vitro incubation with bosentan, BQ123, and (or) dazmegrel on vascular responses to vasoconstrictors in SMA In SMA, the enhanced vasoconstrictor effects of NE in diabetes could also be abolished by acute blockade with either bosentan or BQ123. Hence, the diabetic-induced increase in the contractile response to NE appears to be mediated by the ETA receptor in this vessel. This finding can be reconciled with the increase in immunoreactive ETA receptor levels that we observed in the SMA from diabetic rats (Figs. 1 and 2). Moreover, the amelioration of the enhanced NE response in diabetes after long-term treatment with bosentan may be attributable to the normalization of the ETA receptor level as seen in the mesenteric arteries from bosentan-treated diabetic rats. As mentioned above, we proposed that the enhanced contractile response to NE in diabetes may be mediated through an increase in ET-1-induced TxA2 synthesis and (or) action. Indeed, this hypothesis is supported by our findings that in-
hibition of thromboxane synthase with dazmegrel abrogated the differences seen in the NE response between the diabetic and control SMA (Fig. 5A). Hence, the above results suggest that in this vessel diabetes-associated hyper-reactivity to NE may be mediated via an increase in ETA receptor activation and subsequent stimulation of thromboxane synthesis in the endothelium. In parallel with the findings on NE responses, the exaggerated ET-1-induced contractile responses in diabetic arteries, which was corrected by chronic bosentan treatment, was also shown to be alleviated by inhibiting thromboxane synthesis with dazmegrel. Hence, similarly to the responses to NE, the enhanced response to ET-1 in diabetic arteries may be due to an increase in ET-1-induced thromboxane synthesis. Similar interactions between NE, ET-1, and TxA2 has been noted in another rodent model, the spontaneously hypertensive rats (SHR) (Zerrouk et al. 1997). Zerrouk et al. (1997) have shown that in SHR aortae (but not in Wistar– Kyoto (WKY) rat aortae), ET-1 at a subthreshold concentration (3 10–10 mol/L) potentiated the contractile responses to NE. This potentiation was endothelium-dependent and was suppressed by a cyclooxygenase inhibitor (piroxicam) as well as by a PGH2–TxA2 receptor antagonist (SQ29548). As well, this effect of ET-1 was elicited by the ETA receptor since it was blocked by BQ123, but not BQ788. Furthermore, they have demonstrated that the TxA2 mimetic U46619 also amplified the contractile responses to NE in denuded aortae from both SHR and WKY rats (Zerrouk et al. 1997), thus providing evidence for this intricate relationship between NE, ET-1, and TxA2. Taddei and Vanhoutte (1993a, 1993b) also reported that the contractile response to ET-1 in SHR aortae involved the release of #
2006 NRC Canada
Arikawa et al.
endothelium-derived TxA2 evoked by stimulation of the ETA receptor; this phenomenon was not observed in WKY rat aortae (Taddei and Vanhoutte 1993a, 1993b). The notion that the hyper-reactivity to vasoconstrictors in diabetic SMA is dependent on an ETA receptor-activated process in the endothelium is intriguing, since it would suggest that ETA receptors, which are considered to be primarily vascular smooth muscle receptors, may also be present in the endothelium. The above findings from SHR aortae support this idea since the ET-1-induced effects noted in those aortae (including the stimulation of the release of TxA2) were abrogated by the blockade of ETA receptors as well as removal of the endothelium (Taddei and Vanhoutte 1993a, 1993b; Zerrouk et al. 1997). Data from another study also provided evidence for the presence of functioning ETA receptors and their mRNA expression in the endothelial cells from porcine aortic valve in situ (Nishimura et al. 1995). Since our studies for the expression of ET receptors in control and diabetic rat arteries had not been designed to assess the distribution of ET receptors in different cell types, further studies will be required to clarify whether ETA receptors are present in the endothelium of diabetic SMA. Effects of in vitro incubation with bosentan, BQ123, and (or) dazmegrel on vascular responses to vasoconstrictors in RA Although chronic treatment with bosentan equally ameliorated the abnormal vasoconstrictor responses in both the SMA and RA from diabetic rats, results obtained from SMA and RA were different when acute effects of different blockers were examined. In contrast to the observations in the diabetic SMA, the contractile response to NE in diabetic rat RA remained significantly greater than that of the control rats after the incubation with bosentan, BQ123, or dazmegrel. In addition, in the RA from diabetic rats, the exaggerated response to ET-1 remained apparent after in vitro incubation with dazmegrel. However, dazmegrel incubation partially attenuated the maximum contractile responses to ET-1 in diabetic RA, indicating the presence of an interaction between ET-1 and TxA2 in those arteries. This difference observed between the different arteries may possibly be due to a difference in ETA and (or) ETB receptor density in the RA and the SMA. This notion is supported by the observation of the regional differences in vascular response to ET-1 (Gardiner et al. 1989). In addition, diabetes has been shown to result in a relative decrease in the ETB receptor level in the endothelium compared with that of vascular smooth muscle cell in rat kidney, indicating that in diabetic states there may be a preferential increase in ETB receptor-induced vasoconstriction over vasodilation (Kakoki et al. 1999); however, whether such change also occurs in the mesenteric arteries during diabetes remains to be elucidated. Vascular responses to U46619 In diabetes, prostanoid metabolism as well as vascular responsiveness to prostanoids appears to be altered. There seems to be a shift from vasodilator to vasoconstrictor prostanoid production in the diabetic state (Halushka et al. 1985). Vascular production of prostacyclin in diabetic animals has been shown to be decreased (Roth et al. 1983;
831
Rogers and Larkins 1981; Wakasugi et al. 1991). Conversely, both the plasma levels and platelet synthesis of TxA2 have been reported to be increased in diabetic patients (Jain et al. 1998; Chen et al. 1990). An increase in TxA2 release from the endothelium of diabetic aorta has also been previously demonstrated (Tesfamariam et al. 1989; Cohen 1993). As well, femoral arterial rings from diabetic dogs exhibited an increased production of TxA2 that was proposed to be responsible for the increased vasoconstrictor response to NE in seen in those animals (Koltai et al. 1990). Additionally, experimental diabetes has been demonstrated to increase contractile responses to the TxA2 analogue U46619 (Quilley and McGiff 1990; Roth et al. 1983; Hattori et al. 1999). Thus, it is possible that vasoconstrictor prostanoid such as TxA2 may contribute to vascular dysfunction in diabetes. To determine whether the vasoreactivity to TxA2 was altered in our diabetic rats, we examined the contractile responses to U46619, a TxA2 mimetic. In rat SMA, there was no significant change in the contractile response to U46619 after long-term diabetes, and chronic bosentan treatment did not affect this response in either control or diabetic groups. On the contrary, in RA, the contractile response to U46619 in diabetic rats was enhanced, and chronic bosentan treatment was able to correct this abnormality. How diabetes causes a greater renal vasoconstrictor response to TxA2 and how bosentan treatment corrects this vascular defect is not clear. One possibility is that there may be an increase in TxA2 receptor levels in diabetic RA, and chronic bosentan treatment may act to counter this increase, although this is speculative at present. Taken together, our results indicate that an increase in ETA receptor-mediated synthesis and (or) release of TxA2 may account for the diabetes-induced enhancement of vasoconstrictor responses to NE and ET-1 in the SMA. Bosentan treatment may exert its beneficial effects on diabetic vascular function by normalizing the ETA receptor levels as well as blocking the ETA receptor-induced effect on TxA2 synthesis in this vessel. The exact mechanism by which ET-1 stimulates the production of TxA2 in the vascular endothelium is not clearly defined. Several in vitro studies have shown that ET-1 increases the release of arachidonate metabolites by increasing both the activity and expression of PLA2 (Resink et al. 1989; Schramek et al. 1994a, 1994b; Husain and Abdel-Latif 1998). There is also evidence that ET-1 stimulates the expression of cyclooxygenase 2 (COX2) in rat mesangial cells (Kester et al. 1994; Hughes et al. 1995; Sugimoto et al. 2001). Whereas it has been shown in human endothelial cells that an up-regulation of COX-2 results in an increased production of PGI2 and PGE2 and that TxA2 synthesis appears to be solely mediated by cyclooxygenase 1 (COX-1) (Caughey et al. 2001), it is not known exactly how the activity of COX-1 and COX-2 is related to the production of each individual eicosanoid in the vascular endothelium under the pathological conditions of diabetes. Lu¨scher’s group has recently reported intriguing findings that high glucose selectively upregulated COX-2 expression in endothelial cells, which was associated with an enhanced thromboxane A2 but an impaired prostacyclin release (Cosentino et al. 2003), and that this glucose effect appeared to be dependent on PKC signaling, thus providing #
2006 NRC Canada
832
a potential mechanism for ET-1, which activates PKC, to increase TxA2 and consequently vasoconstrictor effects. In contrast to the SMA, in the RA, an increase in vascular responses to TxA2, rather than a change in ET-1-induced TxA2 synthesis, may contribute to the augmented responses to NE and ET-1 in diabetes. Chronic bosentan treatment may normalize the responses to NE and ET-1 in diabetic RA by correcting TxA2 responses. How bosentan attenuates the increased vasoconstrictor responses to TxA2 in diabetic renal arteries is not known. It has been suggested that the alterations in TxA2 response in diabetes may be due to changes in the affinity and number of TxA2 receptors (Morinelli et al. 1993; Ozturk et al. 1996); hence, we propose that bosentan may prevent diabetic-induced changes in TxA2 receptors. Further studies aiming at examining the effects of TxA2 receptor blockade on NE and ET-1 responses and determining the expression and properties of TxA2 receptors in untreated and bosentan-treated diabetic RA are necessary to validate such a hypothesis. In conclusion, the above study demonstrated a role for ET-1 in mediating the exaggerated vascular responses to vasoconstrictors in diabetic SMA and RA. The data indicate that an interaction between ET-1 and TxA2 is involved in this vascular defect in diabetes. Furthermore, the underlying mechanisms appear to be tissue specific as the ET receptor blockers and the thromboxane synthase inhibitor exerted different effects on vascular responses in arteries from different vascular beds. Also, both long- and short-term effects of bosentan may contribute to the beneficial effects of the blocker on the diabetic vasculature.
Acknowledgements This study was supported by funding from the Canadian Diabetes Association. Emi Arikawa and Inna Sekirov were the recipients of a Heart and Stroke Foundation of Canada Research Traineeship and a Heart and Stroke Foundation of BC and Yukon Summer Studentship, respectively, at the time of the study. The gift of bosentan from Dr. M. Clozel, Actelion Ltd., Switzerland is gratefully acknowledged. Dazmegrel was a generous gift provided by Pfizer, UK. We thank Dr. Jian Wang for his technical advice and assistance in this study. We thank Dr. Robert A. Harris and Dr. Sid Katz for the use of their computerized imaging system.
References Agrawal, D.K., and McNeill, J.H. 1987. Effect of diabetes on vascular smooth muscle function in normotensive and spontaneously hypertensive rat mesenteric artery. Can. J. Physiol. Pharmacol. 65: 2274–2280. PMID:3449189. Arikawa, E., Verma, S., Dumont, A.S., and McNeill, J.H. 2001. Chronic bosentan treatment improves renal artery vascular function in diabetes. J. Hypertens. 19: 803–812. doi:10.1097/ 00004872-200104000-00018. PMID:11330884. Benigni, A., Colosio, V., Brena, C., Bruzzi, I., Bertani, T., and Remuzzi, G. 1998. Unselective inhibition of endothelin receptors reduces renal dysfunction in experimental diabetes. Diabetes, 47: 450–456. PMID:9519753. Bursell, S.E., Clermont, A.C., Oren, B., and King, G.L. 1995. The in vivo effect of endothelins on retinal circulation in nondiabetic and diabetic rats. Invest. Ophthalmol. Vis. Sci. 36: 596–607. PMID:7890491.
Can. J. Physiol. Pharmacol. Vol. 84, 2006 Caughey, G.E., Cleland, L.G., Penglis, P.S., Gamble, J.R., and James, M.J. 2001. Roles of cyclooxygenase (cox)-1 and cox-2 in prostanoid production by human endothelial cells: selective up-regulation of prostacyclin synthesis by cox-2. J. Immunol. 167: 2831–2838. PMID:11509629. Chen, S., Evans, T., Mukherjee, K., Karmazyn, M., and Chakrabarti, S. 2000. Diabetes-induced myocardial structural changes: Role of endothelin-1 and its receptors. J. Mol. Cell. Cardiol. 32: 1621–1629. doi:10.1006/jmcc.2000.1197. PMID:10966825. Chen, S.Y., Yu, B.J., Liang, Y.Q., and Lin, W.D. 1990. Platelet aggregation, platelet camp levels and thromboxane b2 synthesis in patients with diabetes mellitus. Chin. Med. J. (Engl.), 103: 312–318. PMID:2167823. Cohen, R.A. 1993. Dysfunction of vascular endothelium in diabetes mellitus. Circulation, 87: V67–V76. Cosentino, F., Eto, M., De Paolis, P., van der Loo, B., Bachschmid, M., Ullrich, V., et al. 2003. High glucose causes upregulation of cyclooxygenase-2 and alters prostanoid profile in human endothelial cells: role of protein kinase c and reactive oxygen species. Circulation, 107: 1017–1023. doi:10.1161/01.CIR. 0000051367.92927.07. PMID:12600916. Deng, D., Evans, T., Mukherjee, K., Downey, D., and Chakrabarti, S. 1999. Diabetes-induced vascular dysfunction in the retina: role of endothelins. Diabetologia, 42: 1228–1234. doi:10.1007/ s001250051296. PMID:10525664. Fukui, M., Nakamura, T., Ebihara, I., Osada, S., Tomino, Y., Masaki, T., et al. 1993. Gene expression for endothelins and their receptors in glomeruli of diabetic rats. J. Lab. Clin. Med. 122: 149–156. PMID:8340699. Galipeau, D., Arikawa, E., Sekirov, I., and McNeill, J.H. 2001. Chronic thromboxane synthase inhibition prevents fructose-induced hypertension. Hypertension, 38: 872–876. PMID:11641301. Gardiner, S.M., Compton, A.M., and Bennett, T. 1989. Regional hemodynamic effects of endothelin-1 in conscious, unrestrained, wistar rats. J. Cardiovasc. Pharmacol. 13: S202–S204. PMID:2473311. Gilbert, R.E., Rumble, J.R., Cao, Z., Cox, A.J., van Eeden, P., Allen, T.J., Kelly, D.J., and Cooper, M.E. 2000. Endothelin receptor antagonism ameliorates mast cell infiltration, vascular hypertrophy, and epidermal growth factor expression in experimental diabetes. Circ. Res. 86: 158–165. PMID:10666411. Halushka, P.V., Mayfield, R., and Colwell, J.A. 1985. Insulin and arachidonic acid metabolism in diabetes mellitus. Metabolism, 34: 32–36. PMID:3934501. Hattori, Y., Kawasaki, H., and Kanno, M. 1999. Increased contractile responses to endothelin-1 and u46619 via a protein kinase cmediated nifedipine-sensitive pathway in diabetic rat aorta. Res. Commun. Mol. Pathol. Pharmacol. 104: 73–80. PMID:10604280. Howarth, S.R., Vallance, P., and Wilson, C.A. 1995. Role of thromboxane a2 in the vasoconstrictor response to endothelin-1, angiotensin ii and 5-hydroxytryptamine in human placental vessels. Placenta, 16: 679–689. doi:10.1016/0143-4004(95)90012-8. PMID:8710799. Hughes, A.K., Padilla, E., Kutchera, W.A., Michael, J.R., and Kohan, D.E. 1995. Endothelin-1 induction of cyclooxygenase-2 expression in rat mesangial cells. Kidney Int. 47: 53–61. PMID:7731170. Husain, S., and Abdel-Latif, A.A. 1998. Role of protein kinase c alpha in endothelin-1 stimulation of cytosolic phospholipase a2 and arachidonic acid release in cultured cat iris sphincter smooth muscle cells. Biochim. Biophys. Acta, 1392: 127–144. PMID:9593858. #
2006 NRC Canada
Arikawa et al. Jain, S.K., Krueger, K.S., McVie, R., Jaramillo, J.J., Palmer, M., and Smith, T. 1998. Relationship of blood thromboxane-b2 (txb2) with lipid peroxides and effect of vitamin e and placebo supplementation on txb2 and lipid peroxide levels in type 1 diabetic patients. Diabetes Care, 21: 1511–1516. PMID:9727900. Jarvis, M.F., Wessale, J.L., Zhu, C.Z., Lynch, J.J., Dayton, B.D., Calzadilla, S.V., et al. 2000. Abt-627, an endothelin et(a) receptorselective antagonist, attenuates tactile allodynia in a diabetic rat model of neuropathic pain. Eur. J. Pharmacol. 388: 29–35. doi:10.1016/S0014-2999(99)00865-1. PMID:10657544. Kakoki, M., Hirata, Y., Hayakawa, H., Tojo, A., Nagata, D., Suzuki, E., et al. 1999. Effects of hypertension, diabetes mellitus, and hypercholesterolemia on endothelin type b receptormediated nitric oxide release from rat kidney. Circulation, 99: 1242–1248. PMID:10069794. Kester, M., Coroneos, E., Thomas, P.J., and Dunn, M.J. 1994. Endothelin stimulates prostaglandin endoperoxide synthase-2 mrna expression and protein synthesis through a tyrosine kinasesignaling pathway in rat mesangial cells. J. Biol. Chem. 269: 22574–22580. PMID:8077206. Koltai, M.Z., Rosen, P., Ballagi-Pordany, G., Hadhazy, P., and Pogatsa, G. 1990. Increased vasoconstrictor response to noradrenaline in femoral vascular bed of diabetic dogs. Is thromboxane a2 involved? Cardiovasc. Res. 24: 707–710. PMID:2121358. Moreau, P., Takase, H., and Luscher, T.F. 1996. Effect of endothelin antagonists on the responses to prostanoid endotheliumderived contracting factor. Br. J. Pharmacol. 118: 1429–1432. PMID:8832068. Morinelli, T.A., Tempel, G.E., Jaffa, A.A., Silva, R.H., Naka, M., Folger, W., and Halushka, P.V. 1993. Thromboxane a2/prostaglandin h2 receptors in streptozotocin-induced diabetes: effects of insulin therapy in the rat. Prostaglandins, 45: 427–438. doi:10.1016/0090-6980(93)90119-R. PMID:8321912. Muck, A.O., Seeger, H., Korte, K., and Lippert, T.H. 1993. The effect of 17 beta-estradiol and endothelin 1 on prostacyclin and thromboxane production in human endothelial cell cultures. Clin. Exp. Obstet. Gynecol. 20: 203–206. PMID:8281701. Nishimura, J., Aoki, H., Chen, X., Shikasho, T., Kobayashi, S., and Kanaide, H. 1995. Evidence for the presence of endothelin eta receptors in endothelial cells in situ on the aortic side of porcine aortic valve. Br. J. Pharmacol. 115: 1369–1376. PMID:8564193. Ozturk, Y., Altan, V.M., and Yildizoglu-Ari, N. 1996. Effects of experimental diabetes and insulin on smooth muscle functions. Pharmacol. Rev. 48: 69–112. PMID:8685248. Peppa-Patrikiou, M., Dracopoulou, M., and Dacou-Voutetakis, C. 1998. Urinary endothelin in adolescents and young adults with insulin- dependent diabetes mellitus: relation to urinary albumin, blood pressure, and other factors. Metabolism, 47: 1408–1412. PMID:9826222. Quilley, J., and McGiff, J.C. 1990. Renal vascular responsiveness to arachidonic acid in experimental diabetes. Br. J. Pharmacol. 100: 336–340. PMID:2116203. Resink, T.J., Scott-Burden, T., and Buhler, F.R. 1989. Activation of phospholipase a2 by endothelin in cultured vascular smooth muscle cells. Biochem. Biophys. Res. Commun. 158: 279–286. doi:10.1016/S0006-291X(89)80209-8. PMID:2492195. Reynolds, E.E., and Mok, L.L. 1990. Role of thromboxane a2/prostaglandin h2 receptor in the vasoconstrictor response of rat aorta to endothelin. J. Pharmacol. Exp. Ther. 252: 915–921. PMID:2138670. Rogers, S.P., and Larkins, R.G. 1981. Production of 6-oxoprostaglandin f1alpha by rat aorta: influence of diabetes, insulin
833 treatment, and caloric deprivation. Diabetes, 30: 935–939. PMID:7028546. Roth, D.M., Reibel, D.K., and Lefer, A.M. 1983. Vascular responsiveness and eicosanoid production in diabetic rats. Diabetologia, 24: 372–376. PMID:6347785. Schramek, H., Wang, Y., Konieczkowski, M., Rose, P.M., Sedor, J.R., and Dunn, M.J. 1994a. Endothelin-1 stimulates cytosolic phospholipase a2 in chinese hamster ovary cells stably expressing the human eta or etb receptor subtype. Biochem. Biophys. Res. Commun. 199: 992–997. doi:10.1006/bbrc.1994.1327. PMID:8135848. Schramek, H., Wang, Y., Konieczkowski, M., Simonson, M.S., and Dunn, M.J. 1994b. Endothelin-1 stimulates cytosolic phospholipase a2 activity and gene expression in rat glomerular mesangial cells. Kidney Int. 46: 1644–1652. PMID:7700023. Sugimoto, T., Haneda, M., Sawano, H., Isshiki, K., Maeda, S., Koya, D., et al. 2001. Endothelin-1 induces cyclooxygenase-2 expression via nuclear factor of activated t-cell transcription factor in glomerular mesangial cells. J. Am. Soc. Nephrol. 12: 1359–1368. PMID:11423565. Taddei, S., and Vanhoutte, P.M. 1993a. Endothelium-dependent contractions to endothelin in the rat aorta are mediated by thromboxane a2. J. Cardiovasc. Pharmacol. 22: S328–S331. PMID:7509978. Taddei, S., and Vanhoutte, P.M. 1993b. Role of endothelium in endothelin-evoked contractions in the rat aorta. Hypertension, 21: 9–15. PMID:8418029. Takagi, C., Bursell, S.E., Lin, Y.W., Takagi, H., Duh, E., Jiang, Z., et al. 1996. Regulation of retinal hemodynamics in diabetic rats by increased expression and action of endothelin-1. Invest. Ophthalmol. Vis. Sci. 37: 2504–2518. PMID:8933767. Tesfamariam, B., Jakubowski, J.A., and Cohen, R.A. 1989. Contraction of diabetic rabbit aorta caused by endothelium-derived pgh2- txa2. Am. J. Physiol. 257: H1327–H1333. PMID:2511765. Verma, S., Bhanot, S., Yao, L., and McNeill, J.H. 1996. Defective endothelium-dependent relaxation in fructose-hypertensive rats. Am. J. Hypertens. 9: 370–376. doi:10.1016/0895-7061(95) 00392-4. PMID:8722439. Verma, S., Arikawa, E., and McNeill, J.H. 2001. Long-term endothelin receptor blockade improves cardiovascular function in diabetes. Am. J. Hypertens. 14: 679–687. doi:10.1016/S08957061(01)01302-4. PMID:11465653. Verma, S., Arikawa, E., Lee, S., Dumont, A.S., Yao, L., and McNeill, J.H. 2002. Exaggerated coronary reactivity to endothelin-1 in diabetes: reversal with bosentan. Can. J. Physiol. Pharmacol. 80: 980–986. doi:10.1139/y02-122. PMID:12450064. Wakasugi, M., Noguchi, T., Inoue, M., Tawata, M., Shindo, H., and Onaya, T. 1991. Effects of aldose reductase inhibitors on prostacyclin (pgi2) synthesis by aortic rings from rats with streptozotocin-induced diabetes. Prostaglandins Leukot. Essent. Fatty Acids, 44: 233–236. doi:10.1016/0952-3278(91)90022-W. PMID:1840007. Zaugg, C.E., Hornstein, P.S., Zhu, P., Simper, D., Luscher, T.F., Allegrini, P.R., and Buser, P.T. 1996. Endothelin-1-induced release of thromboxane a2 increases the vasoconstrictor effect of endothelin-1 in postischemic reperfused rat hearts. Circulation, 94: 742–747. PMID:8772697. Zerrouk, A., Champeroux, P., Safar, M., and Brisac, A.M. 1997. Role of endothelium in the endothelin-1-mediated potentiation of the norepinephrine response in the aorta of hypertensive rats. J. Hypertens. 15: 1101–1111. doi:10.1097/00004872-19971510000008. PMID:9350584.
#
2006 NRC Canada
