17beta-Estradiol inhibits keratinocyte-derived chemokine production following trauma-hemorrhage

Neutrophil infiltration is a key step in the development of organ dysfunction following trauma-hemorrhage (T-H). Although we have previously shown that 17beta-estradiol (E2) prevents neutrophil infiltration and organ damage following T-H, the mechanism by which E2 inhibits neutrophil transmigration remains unknown. We hypothesized that E2 prevents neutrophil infiltration via modulation of keratinocyte-derived chemokine (KC), a major attractant for neutrophils. To examine this, male C3H/HeN mice were subjected to T-H or sham operation and thereafter resuscitated with Ringer lactate and E2 (1 mg/kg body wt) or vehicle. Animals were killed 2 h after resuscitation, and Kupffer cells were isolated. Plasma levels and Kupffer cell production capacities of KC, TNF-alpha, and IL-6 were determined by BD Cytometric Bead Arrays; lung mRNA expression of KC was measured with real-time PCR; myeloperoxidase activity assays were performed to determine neutrophil infiltration, and organ damage was assessed by edema formation. Treatment with E2 decreased systemic levels and restored Kupffer cell production of KC, TNF-alpha, and IL-6, as well as KC gene expression and protein in the lung. This was accompanied with a decrease in neutrophil infiltration and edema formation in the lung. These results suggest that E2 prevents lung neutrophil infiltration and organ damage in part by decreasing KC during posttraumatic immune response.     

Evaluation of endothelin-1-induced pulmonary vasoconstriction following myocardial infarction

Endothelin (ET) levels are elevated in congestive heart failure secondary to myocardial infarction (MI) and correlate well with the severity of pulmonary hypertension (PH), suggesting that the ET peptide could contribute to the pathophysiology of venous PH. Alterations of pulmonary vasoreactivity to ET after MI and the respective roles of the ET(A) and ET(B) receptors (ET(A)-R and ET(B)-R) have never been evaluated, to our knowledge. MI was induced in rats. Three weeks later, small pulmonary resistance arteries were mounted on a microvascular myograph. Cumulative concentration-response curves to ET-1 and sarafotoxin 6c (S6c) were performed. Response to ET was also assessed in the presence of ET-R antagonists. Heterodimerization of receptors was evaluated by immunoprecipitation of the ET(B)-R, followed by western blotting for the expression of the ET(A)-R. Maximal vasoconstriction and sensitivity to ET-1 were similar in sham and MI with values of 88 +/- 3.9% and 80 +/- 3.8%, respectively. The response to S6c was similarly less in both sham (67 +/- 5.7%) and MI groups (60 +/- 6.6%). When administered alone, the ET(A)-R antagonist (10 nM A-147627.1) and the ET(B)-R antagonist (1 microM A-192621.1) had no significant effect. However, their combination markedly reduced vaso-constriction (52 +/- 5.3%; P < 0.001). The endothelial and medial distribution of ET-Rs was similar in sham and MI groups. In vitro studies demonstrated co-immunoprecipitation of the ET(A)-R and ET(B)-R. Vasoconstriction of isolated resistance pulmonary arteries to ET agonists is not altered after MI. Dual antagonism results in optimal blockade of vasoconstriction, possibly because the ET(A)-R and ET(B)-R can form functional heterodimers.     

Upregulation of hepatic prolactin receptor gene expression by 17beta-estradiol following trauma-hemorrhage.

Although studies show protective effects of 17beta-estradiol (E2) or prolactin (PRL) treatment in male rats after trauma-hemorrhage (TH), the mechanism of the salutary effects of these agents remains unknown. Because E2 modulates PRL receptor (PRL-R) expression in the liver, we examined whether E2 treatment after T-H has any effects on hepatic PLR-R gene expression. Male Sprague-Dawley rats were subjected to trauma (i.e., 5-cm midline laparotomy) and hemorrhage (35-40 mmHg for 90 min) followed by fluid resuscitation (Ringer lactate) or sham operation and then treated with E2 (50 microg/kg body wt sc) or vehicle immediately before resuscitation. Liver samples were collected at 3 h thereafter, and PRL-R mRNA expression was determined by PCR. Liver expression of PRL-R short-form gene was unaffected by T-H, whereas that of the long-form gene was suppressed. Treatment of T-H rats with E2 significantly increased PRL-R short-form gene expression and normalized PRL-R long-form gene expression to sham levels. In the isolated hepatocytes, PRL-R short-form gene expression was predominant compared with the long-form gene. In contrast, only the short form was detected in Kupffer cells. In vitro treatment by E2 demonstrated an increase in the PRL-R long-form gene in hepatocytes, but E2 had no effect on PRL-R short-form gene expression in either the Kupffer cells or hepatocytes. Thus E2 treatment after T-H in males appears to directly upregulate PRL-R long-form gene expression in hepatocytes. However, the upregulation of the PRL-R short form might involve the interaction of multiple cell types in the liver.     

Resveratrol improves cardiac contractility following trauma-hemorrhage by modulating Sirt1

Mitochondria play a critical role in metabolic homeostasis of a cell. Our recent studies, based on the reported interrelationship between c-Myc and Sirt1 (mammalian orthologue of yeast sir2 [silent information regulator 2]) expression and their role in mitochondrial biogenesis and function, demonstrated a significant downregulation of Sirt1 protein expression and an upregulation of c-Myc following trauma-hemorrhage (T-H). Activators of Sirt1 are known to improve mitochondrial function and the naturally occurring polyphenol resveratrol (RSV) has been shown to significantly increase Sirt1 activity by increasing its affinity to both NAD+ and the acetylated substrate. In this study we tested the salutary effect of RSV following T-H and its influence on Sirt1 expression. Rats were subjected to T-H or sham operation. RSV (8 mg/kg body weight, intravenously) or vehicle was administered 10 min after the onset of resuscitation, and the rats were killed 2 h following resuscitation. Sirtinol, a Sirt1 inhibitor, was administered 5 min prior to RSV administration. Cardiac contractility (±dP/dt) was measured and heart tissue was tested for Sirt1, Pgc-1α, c-Myc, cytosolic cytochrome C expression and ATP level. Left ventricular function, after T-H, was improved (P < 0.05) following RSV treatment, with significantly elevated expression of Sirt1 (P < 0.05) and Pgc-1α (P < 0.05), and decreased c-Myc (P < 0.05). We also observed significantly higher cardiac ATP content, declined cytosolic cytochrome C and decreased plasma tumor necrosis factor-α in the T-H-RSV group. The salutary effect due to RSV was abolished by sirtinol, indicating a Sirt1-mediated effect. We conclude that RSV may be a useful adjunct to resuscitation fluid following T-H.     

p38 MAPK-dependent eNOS upregulation is critical for 17beta-estradiol-mediated cardioprotection following trauma-hemorrhage

Studies have shown that p38 MAPK and nitric oxide (NO), generated by endothelial NO synthase (eNOS), play key roles under physiological and pathophysiological conditions. Although administration of 17beta-estradiol (E2) protects cardiovascular injury from trauma-hemorrhage, the mechanism by which E2 produces those effects remains unknown. Our objective was to determine whether the E2-mediated activation of myocardial p38 MAPK and subsequent eNOS expression/phosphorylation would protect the heart following trauma-hemorrhage. To study this, male Sprague-Dawley rats underwent soft-tissue trauma (midline laparatomy) and hemorrhagic shock (mean blood pressure 35-40 mmHg for 90 min), followed by fluid resuscitation. Animals were pretreated with specific p38 MAPK inhibitor SB-203580 (SB; 2 mg/kg), and nonselective NO synthase inhibitor NG-nitro-l-arginine methyl ester (l-NAME; 30 mg/kg) 30 min before vehicle (cyclodextrin) or E2 (100 microg/kg) treatment, followed by resuscitation, and were killed 2 h thereafter. Cardiovascular performance and other parameters were measured. E2 administration following trauma-hemorrhage increased cardiac p38 MAPK activity, eNOS expression and phosphorylation at Ser(1177), and nitrate/nitrite levels in plasma and heart tissues; these were associated with normalized cardiac performance, which was reversed by SB administration. In addition, E2 also prevented trauma-hemorrhage-induced increase in cytokines (IL-6 and TNF-alpha), chemokines (macrophage inflammatory protein-2 and cytokine-induced neutrophil chemoattractant-1), and ICAM-1, which was reversed by l-NAME administration. Administration of E2 following trauma-hemorrhage attenuated cardiac tissue injury markers, myeloperoxidase activity, and nitrotyrosine level, which were reversed by treatment with SB and l-NAME. The salutary effects of E2 on cardiac functions and tissue protection following trauma-hemorrhage are mediated, in part, through activation of p38 MAPK and subsequent eNOS expression and phosphorylation.     

17beta-Estradiol and the phytoestrogen genistein attenuate neuronal apoptosis induced by the endoplasmic reticulum calcium-ATPase inhibitor thapsigargin

Estrogenic compounds have been shown to protect neurons from a variety of toxic stimuli in vitro and in vivo and depletion of estrogen at menopause has been associated with increased risk of neurodegenerative diseases. Genistein is an isoflavone soy derivative that binds to estrogen receptors with selective estrogen receptor modulator (SERM) properties. Recent FDA recommendations of soy intake for cholesterol reduction have prompted investigation into the potentially estrogenic role of dietary soy phytochemicals in the brain. In this study, we have shown that 50nM genistein significantly reduces neuronal apoptosis in an estrogen receptor-dependent manner. The importance of apoptosis in the brain has been recognized with regard to organization of the developing brain as well as degeneration in response to disease or stroke; however, the effects of estrogenic compounds on neuronal apoptosis have not been thoroughly examined. We developed a model of apoptotic toxicity in primary cortical neurons by using the endoplasmic reticulum (ER) calcium-ATPase inhibitor, thapsigargin, to test potential anti-apoptotic effects of 17beta-estradiol and genistein. Estrogen receptor beta, but not estrogen receptor alpha, was detected in our primary neuron cultures. Thapsigargin-induced apoptosis was confirmed by loss of mitochondrial function, DNA laddering, nuclear condensation and fragmentation, and caspase activation. Both 17beta-estradiol and genistein reduced the number of apoptotic neurons and reduced the number of neurons containing active caspase-3. This effect was blocked by co-addition of ICI 182780. Our results demonstrate that genistein and 17beta-estradiol have comparable anti-apoptotic properties in primary cortical neurons and that these properties are mediated through estrogen receptors.     

Cholesterol modulates vascular reactivity to endothelin-1 by stimulating a pro-inflammatory pathway

Hypercholesterolemia (HC) is associated with coronary endothelial dysfunction and increased circulating levels of endothelin-1. We show that pre-treatment of intact rat aortic rings with cholesterol synergistically enhances the vasoconstriction induced by endothelin-1 suggesting that elevated levels of cholesterol may predispose to hypertension by modulating the vascular reactivity to endogenous vasoconstrictors. Moreover, we report that SB202190, a selective inhibitor of p38 MAPK, and PD98059 an inhibitor of MEK1/2 are able to abolish the vasoactive properties of cholesterol. MK-886, an inhibitor of 5-lipoxygenase is inefficient at blocking the vasoactive properties of cholesterol whereas NS-398, a selective inhibitor of cyclooxygenase-2 (COX-2) completely abolishes cholesterol-induced vasoconstriction. In intact rat aortae, cholesterol stimulates prostaglandin E(2) and prostaglandin F(2 alpha) production, an effect that can be completely prevented by inhibiting p38 MAPK, or COX-2. In vitro, cholesterol appears to stimulate a similar pro-inflammatory pathway in human cerebrovascular smooth muscle cells. Disruption of the MAPK/COX-2 pathway may represent a valuable therapy to block the hypertension associated with HC, as well as the development of atherosclerosis.     

Correlation between apolipoprotein B and endothelin-1-induced vasoconstriction in humans

Low-density lipoprotein (LDL) concentrations have been linked to altered responses to endogenous vasodilators and vasoconstrictors. We evaluated retrospectively the relationship between LDL and vasoconstrictor (endothelin-1, phenylephrine) responsiveness of the forearm vasculature in 15 elderly healthy volunteers with apolipoprotein B and LDL levels in the normal range. In contrast to phenylephrine, changes in forearm vascular resistance induced by endothelin-1 were correlated with apolipoprotein B, LDL, and total cholesterol concentrations in women but not in men. These findings might suggest that lipids may increase vascular tone through both impaired endothelial vasodilation and increased vasoconstriction to endothelin-1 at least in women.     

Adiponectin opposes endothelin-1-mediated vasoconstriction in the perfused rat hindlimb

Recent studies have shown that adiponectin is able to increase nitric oxide (NO) production by the endothelium and relax preconstricted isolated aortic rings, suggesting that adiponectin may act as a vasodilator. Endothelin-1 (ET-1) is a potent vasoconstrictor, elevated levels of which are associated with obesity, type 2 diabetes, hypertension, and cardiovascular disease. We hypothesized that adiponectin has NO-dependent vascular actions opposing the vasoconstrictor actions of ET-1. We studied the vascular and metabolic effects of a physiological concentration of adiponectin (6.5 μg/ml) on hooded Wistar rats in the constant-flow pump-perfused rat hindlimb. Adiponectin alone had no observable vascular activity; however, adiponectin pretreatment and coinfusion inhibited the increase in perfusion pressure and associated metabolic stimulation caused by low-dose (1 nM) ET-1. Adiponectin was not able to oppose vasoconstriction when infusion was commenced after ET-1. This is in contrast to the NO donor sodium nitroprusside, which significantly reduced the pressure due to established ET-1 vasoconstriction, suggesting dissociation of the actions of adiponectin and NO. In addition, adiponectin had no effect on vasoconstriction caused by either high-dose (20 nM) ET-1 or low-dose (50 nM) norepinephrine. Our findings suggest that adiponectin has specific, apparently NO-independent, vascular activity to oppose the vasoconstrictor effects of ET-1. The hemodynamic actions of adiponectin may be an important aspect of its insulin-sensitizing ability by regulating access of insulin and glucose to myocytes. Imbalance in the relationship between adiponectin and ET-1 in obesity may contribute to the development of insulin resistance and cardiovascular disease.     

Caveolin-1蛋白在171β-雌二醇抑制内皮素-1诱导血管平滑肌细胞增殖中的作用

本工作旨在研究Caveolin-1在17β-雌二醇（17β-estradiol,E2)抑制内皮素-1（endothelin-1,ET-1)诱导血管平滑肌细胞（vascular smooth muscle cells,VSMCs前后ET-1对DNA合成和caveolin-1蛋白表达的影响。结果显示,ET-1可以刺激VSMCs增殖。E2作用24h后,可明显抑制ET-1的上述作用。免疫荧光证实,在VSMCs上有cavellin-1分布,ET-1刺激VSMCs增殖的过程中,VSMCs上caveolin-1蛋白荧光强度下,而事称给予E2可逆转这种下降。Western bolt证实,ET-1可抑制caveolin-1蛋白的表达而E2则可增加caveolin-1蛋白的表达。以上结果表明E2抑制ET-1诱导的VSMCs增殖可能与其增加caveolin-1蛋白表达有关。     

Survivin expression induced by endothelin-1 promotes myofibroblast resistance to apoptosis

Fibrosis of the lungs and other organs is characterized by the accumulation of myofibroblasts, effectors of wound-repair that are responsible for the deposition and organization of new extracellular matrix (ECM) in response to tissue injury. During the resolution phase of normal wound repair, myofibroblast apoptosis limits the continued deposition of ECM. Mounting evidence suggests that myofibroblasts from fibrotic wounds acquire resistance to apoptosis, but the mechanisms regulating this resistance have not been fully elucidated. Endothelin-1 (ET-1), a soluble peptide strongly associated with fibrogenesis, decreases myofibroblast susceptibility to apoptosis through activation of phosphatidylinositol 3′-OH kinase (PI3K)/AKT. Focal adhesion kinase (FAK) also promotes myofibroblast resistance to apoptosis through PI3K/AKT-dependent and -independent mechanisms, although the role of FAK in ET-1 mediated resistance to apoptosis has not been explored. The goal of this study was to investigate whether FAK contributes to ET-1 mediated myofibroblast resistance to apoptosis and to examine potential mechanisms downstream of FAK and PI3K/AKT by which ET-1 regulates myofibroblast survival. Here, we show that ET-1 regulates myofibroblast survival by Rho/ROCK-dependent activation of FAK. The anti-apoptotic actions of FAK are, in turn, dependent on activation of PI3K/AKT and the subsequent increased expression of Survivin, a member of the inhibitor of apoptosis protein (IAP) family. Collectively, these studies define a novel mechanism by which ET-1 promotes myofibroblast resistance to apoptosis through upregulation of Survivin.     

Angiotensin II enhances endothelin-1-induced vasoconstriction through upregulating endothelin type A receptor

Endothelin-1 (ET-1) is the most potent vasoconstrictor by binding to endothelin receptors (ET_AR) in vascular smooth muscle cells (VSMCs). The complex of angiotensin II (Ang II) and Ang II type one receptor (AT₁R) acts as a transient constrictor of VSMCs. The synergistic effect of ET-1 and Ang II on blood pressure has been observed in rats; however, the underlying mechanism remains unclear. We hypothesize that Ang II leads to enhancing ET-1-mediated vasoconstriction through the activation of endothelin receptor in VSMCs. The ET-1-induced vasoconstriction, ET-1 binding, and endothelin receptor expression were explored in the isolated endothelium-denuded aortae and A-10 VSMCs. Ang II pretreatment enhanced ET-1-induced vasoconstriction and ET-1 binding to the aorta. Ang II enhanced ET_AR expression, but not ET_BR, in aorta and increased ET-1 binding, mainly to ET_AR in A-10 VSMCs. Moreover, Ang II-enhanced ET_AR expression was blunted and ET-1 binding was reduced by AT₁R antagonism or by inhibitors of PKC or ERK individually. In conclusion, Ang II enhances ET-1-induced vasoconstriction by upregulating ET_AR expression and ET-1/ET_AR binding, which may be because of the AngII/Ang II receptor pathways and the activation of PKC or ERK. These findings suggest the synergistic effect of Ang II and ET-1 on the pathogenic development of hypertension.     

Rat gastric relaxation induced by stimulation of endothelin-1 selective receptors.

We have investigated the smooth muscle activity of ET-1 and ET-3 on rat fundus strips in vitro as well as the effects of the peptides on gastric motility in vivo. In the isolated tissue with no precontraction ET-1 and ET-3 were potent spasmogens which produced half maximal contractions at concentrations 4.5 and 8.0 nM, respectively. In contrast, under conditions where the isolated tissue was precontracted to approx. 50% of maximum by prostaglandin E2, ET-1 dose-dependently (5 x 10(-10) - 10(-8) M) and temporarily relaxed the fundus strip, whereas ET-3 further increased the contraction. The relaxing capacity of ET-1 was absent when the tissue was precontracted by potassium yet was resistant to pretreatments with tetrodotoxin, capsaicin, propranolol, indomethacin, NG-methyl-L-arginine or glibenclamide. In addition in vivo ET-1 and ET-3 (less than 1 nmol/kg) showed opposite effects on gastric motility as the former reduced basal tonus and spontaneous activity, whereas the latter increased the motor activity of the gastric ventricle. The results support the notion that ET-1 may induce gastric relaxation by stimulation of selective receptors whereas stimulation of nonselective receptors may promote gastric smooth muscle contraction.     

Endothelium-independent and -dependent contractions induced by endothelin-1 in canine basilar arteries

Endothelium-dependence of contractile responses to endothelin-1 was examined in isolated canine basilar arteries. Within 2 hrs after mounting tissue preparations, endothelin-1 (10(-9) M) caused a monophasic tonic contraction that developed very slowly and was sustained in intact and endothelium-removed arteries. More than 5 hrs after tissue mounting, endothelin-1 (10(-9) M) caused a biphasic contraction consisting of phasic and tonic components in intact arteries, and caused a monophasic tonic contraction in endothelium-removed arteries. This phasic component was significantly decreased by aspirin (5 x 10(-5) M,), OKY-046 (10(-5) M) (a TXA2 synthetase inhibitor) and ONO-3708 (10(-8) M) (a TXA2 antagonist). The present experiments demonstrate that endothelin-1 causes an endothelium-independent tonic contraction and an endothelium-dependent phasic contraction in canine basilar arteries, and suggest that TXA2 plays a role as an endothelium-derived contracting factor.     

Long-lasting vasoconstriction and efficient regional extraction of endothelin-1 in human splanchnic and renal tissues

Six healthy subjects were given endothelin-1, intravenously in a dose of 4 pmol.kg-1.min-1 for 20 min. Blood samples were drawn from arterial, hepatic and renal vein catheters for determinations of splanchnic and renal blood flows and the extraction of endothelin-1 in these vascular beds. Intravenous infusion of endothelin-1 increased the mean arterial blood pressure by 6.8 +/- 2.0 mm Hg (p less than 0.05) and reduced splanchnic and renal blood flows by 34% (p less than 0.005) and 26% (p less than 0.001) respectively. Return to basal flow values occurred after about 1 hr for the splanchnic and 3 hrs for the renal blood flow. The fractional extractions of endothelin-1-like immunoreactivity corresponded to 75 +/- 2% and 60 +/- 2% in the splanchnic and renal vascular beds, respectively. The disappearance curve in plasma and two half-lives of 1.4 +/- 0.1 min and 35 +/- 2.8 min respectively.     

17beta-Estradiol inhibits osteoprotegerin production by the estrogen receptor-alpha-positive human breast cancer cell line MCF-7

Estrogen regulates various cytokines and growth factors in estrogen receptor (ER)-positive human breast cancer. Receptor activator of NF-κB ligand (RANKL) is an essential cytokine for osteoclasts, whereas osteoprotegerin (OPG) is a soluble inhibitor for RANKL. We analyzed the regulation of the RANKL/OPG system by estrogens and androgens in the ER-positive breast cancer cell line MCF-7 and the ER-negative breast cancer cell line MDA-MB-231. In MCF-7 cells, which predominantly express ER-α, 17β-estradiol and testosterone dose-dependently decreased OPG mRNA levels and protein secretion by 70 and 65%, respectively (p < 0.0001 by ANOVA). The inhibition of OPG production by 17β-estradiol and testosterone was specifically prevented by the pure anti-estrogen ICI 182,780, and the testosterone effect was prevented by an aromatase inhibitor. In conclusion, 17β-estradiol suppressed OPG production by human breast cancer cell lines in a dose-dependent and specific manner, indicating that the RANKL/OPG cytokine system is an estrogen-responsive target in breast cancer.     

Mechanism of the salutary effects of 17beta-estradiol following trauma-hemorrhage: direct downregulation of Kupffer cell proinflammatory cytokine production

Kupffer cells have been reported as a major source of proinflammatory cytokines (i.e. IL-6, TNF-alpha), which have been implicated in the pathogenesis of trauma-hemorrhage. Previous studies have shown a protective effect of 17beta-estradiol on immune function and physiological responses following trauma-hemorrhage. In this study, we investigated whether 17beta-estradiol has a direct effect on Kupffer cell cytokine production following trauma-hemorrhage. Male Sprague-Dawley rats were subjected to trauma (midline laparotomy) and hemorrhage (35-40 mmHg for 90 min followed by fluid resuscitation) or sham operation. Two hours later, Kupffer cells were isolated and cultured with 17beta-estradiol in the presence and absence of lipopolysaccharide stimulation. Kupffer cell IL-6 and TNF-alpha production increased following trauma-hemorrhage. Incubation with 17beta-estradiol attenuated the production of IL-6 by cells from both sham and trauma-hemorrhage animals in a dose-dependent manner. The suppression of IL-6 production by 17beta-estradiol was paralleled by a decrease in mRNA levels. In contrast to IL-6, the effects of 17beta-estradiol on TNF-alpha production were minimal. In conclusion, these results indicate the direct downregulation of Kupffer cell IL-6 production by 17beta-estradiol at a molecular level, which might explain in part the previously observed salutary effects of estradiol treatment following trauma-hemorrhage.     

Mechanisms mediating canine renal vasoconstriction induced by nicotine infusion

We investigated the respective contributions of the renin-angiotensin and alpha-adrenergic systems to nicotine-induced, canine, renal vasoconstriction by using saralasin (4 micrograms/kg/min) and phentolamine (25 micrograms/kg/min) blockade respectively. Nicotine infusion (0.024 mg/kg/min) increased mean arterial blood pressure (MABP) (114 +/- 3.0 to 219 +/- 8.0 mmHg) and decreased total renal blood flow (TRBF) (3.12 +/- 0.34 to 1.60 +/- 0.37 ml/min/g). Nicotine infusion produced a significantly lesser blood flow in outer cortex (OC), inner cortex (IC), and outer medulla (OM) compared to control dogs. The intrarenal-artery infusion of saralasin or phentolamine had no effect on the nicotine-induced MABP changes. Phentolamine infusion prior to nicotine resulted in a significantly greater TRBF (P less than 0.01), OC (p less than 0.001), IC (p less than 0.001) and OM (p less than 0.01) flow than in the group that received nicotine only. Saralasin pretreatment prior to nicotine resulted only in a significantly (p less than 0.01) greater OC flow than nicotine only. Our data suggest that while angiotensin II mediates a portion of the action of nicotine on the OC renal vasculature, the alpha adrenergic system predominates as the mediator of nicotine-induced renal vasoconstriction in the first 7 minutes of nicotine infusion.     

Mechanism of estrogen-mediated intestinal protection following trauma-hemorrhage: p38 MAPK-dependent upregulation of HO-1

p38 MAPK has been reported to regulate the inflammatory response in various cell types via extracellular stimuli. p38 MAPK activation also results in the induction of heme oxygenase (HO)-1, which exerts potent anti-inflammatory effects. Although studies have shown that 17beta-estradiol (E(2)) prevented organ dysfunction following trauma-hemorrhage, it remains unknown whether p38 MAPK/HO-1 plays any role in E(2)-mediated attenuation of intestinal injury under those conditions. To study this, male rats underwent trauma-hemorrhage (mean blood pressure approximately 40 mmHg for 90 min) followed by fluid resuscitation. At the onset of resuscitation, rats were treated with vehicle, E(2) (1 mg/kg body wt), the p38 MAPK inhibitor SB-203580 (2 mg/kg body wt) or E(2) plus SB-203580. Two hours thereafter, intestinal myeloperoxidase (MPO) activity and lactate, TNF-alpha, IL-6, ICAM-1, cytokine-induced neutrophil chemoattractant (CINC)-1, and macrophage inflammatory protein (MIP)-2 levels were measured. Intestinal p38 MAPK and HO-1 protein levels were also determined. Trauma-hemorrhage led to an increase in intestinal MPO activity and lactate, TNF-alpha, IL-6, ICAM-1, CINC-1, and MIP-2 levels. This was accompanied with a decrease in intestinal p38 MAPK activity and increase in HO-1 expression. Administration of E(2) normalized all the above parameters except HO-1, which was further increased following trauma-hemorrhage. Administration of SB-203580 with E(2) abolished the E(2)-mediated restoration of the above parameters as well as the increase in intestinal HO-1 expression following trauma-hemorrhage. These results suggest that the p38 MAPK/HO-1 pathway plays a critical role in mediating the salutary effects of E(2) on shock-induced intestinal injury.     

BACE1 modulates filopodia-like protrusions induced by sodium channel beta4 subunit

Processing of APP by BACE1 plays a crucial role in the pathogenesis of Alzheimer disease (AD). Recently, the voltage-gated sodium channel (Na_v) β4 subunit (β4), an auxiliary subunit of Na_v that is supposed to serve as a cell adhesion molecule, has been identified as a substrate for BACE1. However, the biological consequence of BACE1 processing of β4 remains illusive. Here, we report the biological effects of β4 processing by BACE1. Overexpression of β4 in Neuro2a cells promoted neurite extension and increased the number of F-actin rich filopodia-like protrusions. While coexpression of BACE1 together with β4 further accelerated neurite extension, the number of filopodia-like protrusions was reduced. Overexpression of C-terminal fragment of β4 that was generated by BACE1 (β4-CTF) partially recapitulated the results obtained with BACE1 overexpression. These results suggest that the processing of β4 by BACE1 regulates neurite length and filopodia-like protrusion density in neurons.