Published online before print February 7, 2005, doi: 10.1161/01.HYP.0000154365.30593.d3
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(Hypertension. 2005;45:773.)
© 2005 American Heart Association, Inc.
Original Articles |
Aldosterone Activates Vascular p38MAP Kinase and NADPH Oxidase Via c-Src
From CIHR Multidisciplinary Research Group on Hypertension (G.E.C., R.M.T., Y.H., S.M., E.L.S.), Clinical Research Institute of Montreal, University of Montreal, Canada; and the Department of Pharmacology (R.C.T., A.Y.), Institute of Biomedical Sciences, University of Sao Paulo, Brazil.
Correspondence to Glaucia E. Callera, Clinical Research Institute of Montreal, 110 Pine Ave, West Montreal, Quebec H2W 1H7. E-mail callereg{at}ircm.qc.ca
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Abstract |
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Increasing evidence indicates that aldosterone elicits vascular effects through nongenomic signaling pathways. We tested the hypothesis that aldosterone induces activation of vascular mitogen-activated protein (MAP) kinases and NADPH oxidase via c-Src–dependent mechanisms in vascular smooth muscle cells (VSMCs). Aldosterone effects on activation of c-Src, p38MAP kinase, and NADPH oxidase, and incorporation of [3H]proline, an index of collagen synthesis, were assessed in cultured rat VSMCs. Studies were performed in the absence and presence of eplerenone, a selective mineralocorticoid receptor blocker, PP2, a selective Src inhibitor, and SB212190, a selective p38MAPK inhibitor. Phosphorylation of c-Src was dose-dependently increased by aldosterone, with maximal responses obtained at 10–7 mol/L. Aldosterone increased p38MAP kinase phosphorylation, NAD(P)H oxidase activation, and [3H]proline incorporation. These responses were abrogated by eplerenone and almost abolished by PP2. Aldosterone-stimulated incorporation of [3H]proline was significantly reduced by SB212190, indicating that p38MAP kinase plays a role in profibrotic actions of aldosterone. To unambiguously demonstrate the importance of aldosterone in c-Src signaling, VSMCs from c-Src+/+ and c-Src+/– mice were also studied. Aldosterone increased phosphorylation of c-Src, p38MAP kinase, and cortactin, a Src-specific substrate, in c-Src+/+ VSMCs, but not in c-Src-deficient cells. Taken together, our findings demonstrate that nongenomic signaling by aldosterone occurs through c-Src–dependent pathways. These processes may play an important role in profibrotic actions of aldosterone.
Key Words: aldosterone • mineralocorticoids • oxidative stress • vasculature • signal transduction • collagen
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Introduction |
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Aldosterone, a mineralocorticoid hormone that classically acts via intracellular mineralocorticoid receptors (MRs), is a key regulator of blood pressure and electrolytic balance.1,2 Aldosterone also plays an important pathophysiological role in hypertension and cardiovascular diseases by promoting changes in vascular reactivity and endothelial function, cardiovascular fibrosis, tissue remodeling, inflammation, and oxidative stress.3–7 Some of these events occur through angiotensin II (Ang II)–dependent pathways.8–10
Binding of aldosterone to intracellular MRs, which belong to the superfamily of ligand-regulated transcription factors, causes dissociation of the ligand-activated MR from a multiprotein complex containing molecular chaperones, translocation of the ligand–MR complex to the nucleus, and interactions with the regulatory region of target gene promoters. Aldosterone-induced genomic effects are characterized by a delay corresponding to a long series of subcellular events.1,2
Besides its well-known genomic actions, there is evidence of aldosterone-mediated short-term effects. Aldosterone induces rapid cellular responses by interfering with intracellular Ca2+ and cAMP levels, Na+/H+ exchanger activity, and phosphorylation of signaling molecules including protein kinase C, epidermal growth factor receptor, and mitogen-activated protein kinases (MAPKs), c-Jun NH2-terminal kinase (JNK) and extracellular signal-regulated kinases (ERKs) 1/2.11–18 Activation of these pathways are known to be critically involved in vascular smooth muscle cell (VSMC) processes associated with remodeling, inflammation, and altered tone in hypertension.19
Recent studies have demonstrated that aldosterone induces phosphorylation of c-Src in renal cortical and Chinese hamster ovary cells.20,21 The c-Src is abundantly expressed in VSMCs and rapidly activated by G-protein–coupled receptors. Furthermore, c-Src plays an important role in phospholipase C phosphorylation, inositol 1,4,5-trisphosphate formation, and Ca2+ mobilization. Src also induces activation of MAPKs (p38MAPK, JNK, and ERK1/2) associated with cell growth, apoptosis, and collagen deposition, as well as activation of other downstream proteins including focal adhesion kinase, Pyk2, and paxillin, involved in cell adhesion processes.19 In addition, c-Src is a critical proximal regulator of reduced nicotinamide adenine dinucleotide (phosphate) [NAD(P)H] oxidase-driven superoxide anion generation.22,23 Of clinical relevance, recent studies have highlighted the contribution of c-Src in molecular and cellular processes underlying vascular changes that occur in human and experimental hypertension.24,25
MRs do not have intrinsic kinase activity and molecular mechanisms underlying nongenomic actions of aldosterone in VSMCs are not fully understood. In the present study, we questioned whether c-Src plays a role in signaling pathways to mediate aldosterone-induced short-term vascular effects. Our findings demonstrate that aldosterone rapidly increases activation of p38MAP kinase and NAD(P)H oxidase through c-Src–dependent pathways. Profibrotic action of aldosterone, assessed by determining 3H-proline incorporation, marker of collagen synthesis, was also dependent on c-Src–regulated p38MAP kinase. These data provide novel insights into nongenomic signaling by aldosterone and highlight the importance of c-Src in these processes.
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Methods |
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Cell Culture
The study was approved by the Animal Ethics Committee of the Clinical Research Institute of Montreal and performed according to the recommendations of the Canadian Council for Animal Care. VSMCs from adult male Wistar-Kyoto rats (WKY), c-Src+/– mice, heterozygous for a disruption in the c-Src gene, and wild-type c-Src+/+ mice were studied. The c-Src+/– mice were generated from a c-Src+/– F1 cross26 and were genotyped by polymerase chain reaction. Rats (16 weeks old) and mice (8 to 10 weeks old) were euthanized by decapitation. VSMCs derived from mesenteric arteries were isolated and characterized as described in detail previously.27 Briefly, arteries were cleaned of adipose and connective tissue; VSMCs were dissociated by digestion of vascular arcades with enzymatic solution (collagenase, elastase, soybean trypsin inhibitor, and bovine serum albumin type I) during 30 to 60 minutes at 37°C; the tissue was filtered; and the cell suspension was centrifuged and resuspended in Dulbecco modified Eagle medium containing 10% fetal calf serum, 2 mmol/L glutamine, 20 mmol/L HEPES (pH 7.4), and antibiotics. At subconfluence, the culture medium was replaced with serum-free medium for 24 hours to render the cells quiescent. Low-passage cells (passages 1 to 7) were studied. The concentrations of pharmacological agents that we used were based on previous studies, which demonstrated that at these doses the signaling pathways were completely abolished.28–30
Western Blotting
VSMCs from WKY rats were stimulated with aldosterone (0.1 nmol/L to 1 µmol/L) for 60 minutes. The concentration of aldosterone that induced maximal c-Src phosphorylation (0.1 µmol/L) was used in further studies. VSMCs were stimulated with aldosterone (1 to 60 minutes), and in some experiments cells were pre-exposed for 30 minutes to 10 µmol/L eplerenone (selective aldosterone receptors antagonist) or 10 µmol/L PP2 (selective Src inhibitor). Proteins were extracted from VSMCs, separated by electrophoresis on a 10% polyacrylamide gel, and transferred onto a nitrocellulose membrane as previously described.25 Nonspecific binding sites were blocked with 5% skim milk in Tris-buffered saline solution with Tween for 1 hour at 24°C. Membranes were then incubated with phospho-specific antibodies (1:1000) overnight at 4°C. Antibodies were as follows: anti–c-Src (pY418) (Biosource), anti-p38MAPK (pY180/102) (Cell Signaling); and anti-cortactin (pY421) (Upstate). The respective nonphospho-antibodies were also used in the present study. After incubation with secondary antibodies, signals were revealed with chemiluminescence, visualized by autoradiography, and quantified densitometrically.
Determination of Proline Incorporation
[3H]proline incorporation was considered a marker of collagen deposition and was measured according to the protocol of Dubey et al.31 Quiescent cells were stimulated for 24 hours with aldosterone (0.01 nmol/L to 1 0.1 µmol/L). In some experiments, cells were pre-exposed to eplerenone, PP2, or SB212190 (a selective p38MAPK inhibitor) (10 µmol/L) and stimulated with aldosterone 0.1 µmol/L.
Measurement of NAD(P)H Oxidase Activity
VSMCs were stimulated with 0.1 µmol/L aldosterone (1 to 60 minutes). In some experiments, cells were pre-exposed for 30 minutes to eplerenone, and PP2 (10 µmol/L) and stimulated with aldosterone for 60 minutes, when activity of the oxidase was maximal. Cells were washed in ice-cold phosphate-buffered saline, scraped off in lysis buffer (20 mmol/L KH2PO4, 1 mmol/L EGTA, and protease inhibitors, pH 7.4), transferred to Eppendorf tubes, and sonicated for 3 seconds. The lucigenin-derived chemiluminescence assay was used to determine NAD(P)H oxidase activity in total protein cell homogenates. The reaction was started by the addition of NAD(P)H (0.1 mmol/L) to the suspension (250 µL final volume) containing sample (50 µL), lucigenin (5 µmol/L), and assay phosphate buffer (50 mmol/L KH2PO4, 1 mmol/L EGTA, 150 mmol/L sucrose, pH 7.4). Luminescence was measured every 1.8 seconds for 3 minutes in a luminometer (Lumistar Galaxy; BMG Labtechnologies). Buffer blank was subtracted from each reading. Activity was expressed as arbitrary units/mg protein.
Data Analysis
Aldosterone-stimulated effects were determined as the percent increase over control, with the control normalized to 100%. Results are presented as mean±SEM and compared by ANOVA or by the Student t test when appropriate. Values of P<0.05 were considered to be significant.
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Results |
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Aldosterone Effects on c-Src and p38MAPK Phosphorylation in VSMCs From WKY
Figure 1 demonstrates that aldosterone induces concentration-dependent phosphorylation of c-Src in VSMCs from WKY. Maximal c-Src phosphorylation was observed with 0.1 µmol/L of aldosterone, and this concentration was used in further studies. As shown in Figure 2, c-Src phosphorylation was time-dependently increased by aldosterone, with a small response occurring within 1 to 5 minutes, followed by a maximal response at 30 minutes. Aldosterone receptor antagonism with eplerenone, as well as inhibition of c-Src with PP2, reduced aldosterone-induced c-Src activation. Aldosterone induced a biphasic increase in p38MAPK activation with a peak response obtained within 1 minute, followed by a second response within 30 minutes (Figure 3). PP2 pretreatment inhibited aldosterone-induced p38MAPK activation. Total protein expression of c-Src and p38MAPK was unaltered by aldosterone stimulation.
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Aldosterone Effects on Phosphorylation of cSrc and p38MAPK in VSMCs From c-Src–Deficient Mice
As shown in Figure 4A, aldosterone-induced maximal c-Src phosphorylation was observed at 1 minute and was sustained for up to 30 in VSMCs from wild-type c-Src+/+ mice. The c-Src phosphorylation was not significantly altered by aldosterone in VSMCs from c-Src+/– mice. Aldosterone also stimulated phosphorylation of cortactin in VSMCs from wild type c-Src+/+ mice (Figure 4B), but not in c-Src–deficient mice. Aldosterone significantly increased phosphorylation of p38MAPK in c-Src+/+ VSMCs, but not in c-Strc+/– cells (Figure 5).
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Aldosterone Effects on NAD(P)H Oxidase Activity
In VSMCs cells from WKY, aldosterone induced a time-dependent increase in NAD(P)H-oxidase activity with maximal effect at 60 minutes, as assessed by low-dose lucigenin chemiluminescence (Figure 6A). Aldosterone also increased NADPH-oxidase activity in VSMCs from wild-type c-Src+/+ mice (234.6±4 arbitrary units/mg protein), whereas in VSMCs from c-Src+/– the response was significantly lower (100.5±1 arbitrary units/mg protein). Figure 6B shows that pretreatment with eplerenone or PP2 significantly reduced aldosterone-mediated activation of NAD(P)H oxidase in VSMCs cells from WKY. Neither PP2 nor eplerenone influenced the basal state of NAD(P)H oxidase activity. In the absence of NAD(P)H, lucigenin-derived chemiluminescence was almost undetectable (data not shown). A possible explanation for this finding is that basal superoxide anion in VSMCs from WKY rats is very low.
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Aldosterone Effects on [3H]Proline Incorporation
Aldosterone induced a concentration-dependent increase in [3H]proline incorporation, a marker of collagen synthesis, in VSMCs from WKY rats (Figure 7A). Eplerenone did not change the basal levels of [3H]proline incorporation and inhibited aldosterone effects in VSMCs from WKY rats (Figure 7B). PP2 and SB212190 decreased basal levels of [3H]proline incorporation. In the presence of these inhibitors, aldosterone stimulation did not change [3H]proline incorporation (Figure 7B). [3H]proline incorporation increased in a dose-dependent manner in response to aldosterone in VSMCs from c-Src+/+ (aldosterone 0.1 µmol/L: 140.7±12% of basal), whereas in cells from c-Src+/– aldosterone did not induce [3H]proline incorporation. PP2 and eplerenone inhibited aldosterone-induced [3H]proline incorporation in c-Src+/+ VSMCs (data not shown).
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Discussion |
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Aldosterone can elicit rapid nongenomic effects, both in vivo and in vitro. Major findings of the present study demonstrate that aldosterone induces rapid activation of p38MAPK and NAD(P)H oxidase through c-Src–dependent pathways in VSMCs. These phenomena are associated with profibrotic processes, as evidenced by increased3[H]proline incorporation in response to aldosterone.
Multiple signaling pathways are activated by aldosterone, of which protein kinase C, epidermal growth factor receptor, ERK 1/2, and JNK cascades are best characterized.11–14 Src kinases are a family of nonreceptor tyrosine kinases, of which the prototype, c-Src, is the major isoform in the vasculature.19 By using a phospho-specific antibody that recognizes the activated site of c-Src, we demonstrated that aldosterone increases c-Src phosphorylation in rat mesenteric VSMCs. The ability of aldosterone to induce c-Src phosphorylation was lost in the presence of PP2, a selective c-Src inhibitor. To unambiguously demonstrate the importance of c-Src in aldosterone-mediated events, we also examined VSMCs from c-Src–deficient mice. Aldosterone increased phosphorylation of c-Src in c-Src+/+ VSMCs, with blunted effects in c-Src+/– cells. To further support a role of c-Src in aldosterone signaling and to evaluate the functional significance of c-Src activation, we assessed aldosterone actions on phosphorylation of cortactin. Cortactin, an actin-binding protein, is a Src-specific downstream target of c-Src tyrosine kinase signaling and plays an important role in regulating actin assembly and organization.32 Aldosterone stimulated cortactin phosphorylation in c-Src+/+ VSMCs, but not in Src+/– cells.
VSMCs express multiple MAPKs, including ERK1/2, a major growth-signaling kinase, JNK, and p38MAPK, involved in cell survival, apoptosis, differentiation, inflammation, and collagen deposition.19 Because MAPKs are downstream targets involved in c-Src signaling, we questioned whether aldosterone induces c-Src–dependent MAPK activation, specifically p38MAPK, in VSMCs. We observed a biphasic increase in p38MAP kinase activation by aldosterone in rat VSMCs, with an acute response within 1 to 3 minutes, followed by a sustained effect at 30 minutes. PP2 inhibited aldosterone-induced p38MAP kinase phosphorylation, indicating that c-Src is upstream of p38MAP kinase. Maximal activation of c-Src was temporally dissociated from the acute p38MAP kinase response, suggesting that even a modest activation of c-Src by aldosterone is sufficient to induce p38MAP kinase activation. Confirming the pharmacological data, aldosterone was not able to induce p38MAPK phosphorylation in cells from c-Src–deficient mice. Previous studies demonstrated that p38MAPK phosphorylation is important in collagen production.5–7 Here we show that aldosterone stimulates [3H]proline incorporation through a c-Src p38MAP kinase-regulated pathway in VSMCs, as evidenced by findings using PP2 and SB212190. The c-Src and p38MAPK activity are also important for basal protein synthesis, particularly collagen, because PP2 and SB212190 decreased basal [3H]proline incorporation.
Because aldosterone has also been linked to oxidative stress,33,34 it is of functional relevance to evaluate whether aldosterone short-term effects are associated with activation of redox-sensitive pathways. Our data demonstrate that aldosterone induces c-Src–dependent NAD(P)H oxidase activation. Because c-Src appears to be both upstream and downstream of NAD(P)H oxidase,19,22,23 activation of c-Src by aldosterone could result in amplification of NAD(P)H oxidase-mediated generation of reactive oxygen species and, consequently, oxidative stress-induced vascular damage. Moreover, redox-sensitive pathways are important in vascular remodeling through activation of MAPK events that underlie vascular functional, structural, and mechanical effects of aldosterone.19
The identity of the primary effector on aldosterone short-term effects—an unidentified membrane-bound receptor, the classical MR, or a related protein—is undergoing debate. Specific high-affinity binding sites for aldosterone have been characterized in membranes from different cells.35 Moreover, it was reported that aldosterone increased intracellular Ca2+ and cAMP levels in cultured skin cells from MR knockout mice, indicating that these rapid actions still occur in the absence of classical MRs.36 Conflicting reports have been demonstrated that nongenomic effects of aldosterone are not blocked by the classical MR antagonist, spironolactone. As examples, MR antagonists do not block aldosterone-induced changes in intracellular cAMP in VSMCs or ERK activation in cortical collecting duct cell.18,37 Conversely, others have demonstrated that MR antagonism inhibited signaling cascades activated by aldosterone.10,12,21 In agreement with these studies, we found that eplerenone, a selective MR antagonist, inhibited phosphorylation of c-Src and p38MAPK, as well as activation of NAD(P)H oxidase activity and [3H]proline incorporation induced by aldosterone. Taken together, these observations indicate that at least some of the rapid nongenomic effects of aldosterone may be mediated by activation of the classical MRs.
In conclusion, c-Src is an important signaling molecule in aldosterone-induced rapid effects in VSMCs. Aldosterone regulates MAP kinases and NAD(P)H-inducible generation of superoxide anion through c-Src–dependent mechanisms. Functional effectors of these processes may be associated with increased collagen production as evidenced by altered incorporation of [3H]-proline.
Perspectives
The present study demonstrates that c-Src plays an important role in aldosterone-mediated actions in VSMCs. We identify a novel nongenomic signaling pathway for aldosterone, involving c-Src–regulated activation of p38MAP kinase and NAD(P)H oxidase. Activation of this pathway may be important in the profibrotic actions of aldosterone. Although the exact mechanisms linking aldosterone to c-Src and the receptors through which aldosterone induces rapid nongenomic vascular actions remain unclear, findings from our studies highlight the functional importance of c-Src/p38MAP kinase/NAD(P)H oxidase and could help to identify molecular mechanisms contributing to aldosterone-mediated nongenomic effects in (patho)physiology. By elucidating such mechanisms, a better understanding to how drugs, such as eplerenone, may have therapeutic potential in hypertension.
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Acknowledgments |
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This study was supported by grants 44018 (to R.M.T.), 37917 (to E.L.S.), and a Group Grant to the Multidisciplinary Research Group on Hypertension, all from the Canadian Institutes for Health Research and an educational grant from Pharmacia/Pfizer. R.M.T. and G.C. received scholarships from the Fonds de la Recherché en Sante du Quebec and the Quebec Hypertension Society, respectively.
Received October 11, 2004; first decision November 4, 2004; accepted December 15, 2004.
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E. L. Schiffrin New Twist to the Role of the Renin-Angiotensin System in Heart Failure: Aldosterone Upregulates Renin-Angiotensin System Components in the Brain Hypertension, March 1, 2008; 51(3): 622 - 623. [Full Text] [PDF]
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 Z.-H. Zhang, Y. Yu, Y.-M. Kang, S.-G. Wei, and R. B. Felder Aldosterone acts centrally to increase brain renin-angiotensin system activity and oxidative stress in normal rats Am J Physiol Heart Circ Physiol, February 1, 2008; 294(2): H1067 - H1074. [Abstract] [Full Text] [PDF]
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N. J. Brown Aldosterone and Vascular Inflammation Hypertension, February 1, 2008; 51(2): 161 - 167. [Full Text] [PDF]
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H. Hitomi, H. Kiyomoto, A. Nishiyama, T. Hara, K. Moriwaki, K. Kaifu, G. Ihara, Y. Fujita, T. Ugawa, and M. Kohno Aldosterone Suppresses Insulin Signaling Via the Downregulation of Insulin Receptor Substrate-1 in Vascular Smooth Muscle Cells Hypertension, October 1, 2007; 50(4): 750 - 755. [Abstract] [Full Text] [PDF]
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A. Yogi, G.E. Callera, A.C.I. Montezano, A.B. Aranha, R.C. Tostes, E.L. Schiffrin, and R.M. Touyz Endothelin-1, but not Ang II, Activates MAP Kinases Through c-Src-Independent Ras-Raf-Dependent Pathways in Vascular Smooth Muscle Cells Arterioscler. Thromb. Vasc. Biol., September 1, 2007; 27(9): 1960 - 1967. [Abstract] [Full Text] [PDF]
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F. Nietlispach, B. Julius, R. Schindler, A. Bernheim, C. Binkert, W. Kiowski, and H. P. Brunner-La Rocca Influence of Acute and Chronic Mineralocorticoid Excess on Endothelial Function in Healthy Men Hypertension, July 1, 2007; 50(1): 82 - 88. [Abstract] [Full Text] [PDF]
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 H. Otani, F. Otsuka, K. Inagaki, M. Takeda, T. Miyoshi, J. Suzuki, T. Mukai, T. Ogura, and H. Makino Antagonistic effects of bone morphogenetic protein-4 and -7 on renal mesangial cell proliferation induced by aldosterone through MAPK activation Am J Physiol Renal Physiol, May 1, 2007; 292(5): F1513 - F1525. [Abstract] [Full Text] [PDF]
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Y. Hirono, T. Yoshimoto, N. Suzuki, T. Sugiyama, M. Sakurada, S. Takai, N. Kobayashi, M. Shichiri, and Y. Hirata Angiotensin II Receptor Type 1-Mediated Vascular Oxidative Stress and Proinflammatory Gene Expression in Aldosterone-Induced Hypertension: The Possible Role of Local Renin-Angiotensin System Endocrinology, April 1, 2007; 148(4): 1688 - 1696. [Abstract] [Full Text] [PDF]
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O. Jung, S. L. Marklund, N. Xia, R. Busse, and R. P. Brandes Inactivation of Extracellular Superoxide Dismutase Contributes to the Development of High-Volume Hypertension Arterioscler. Thromb. Vasc. Biol., March 1, 2007; 27(3): 470 - 477. [Abstract] [Full Text] [PDF]
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J.-K. Park, R. Fischer, R. Dechend, E. Shagdarsuren, A. Gapeljuk, M. Wellner, S. Meiners, P. Gratze, N. Al-Saadi, S. Feldt, et al. p38 Mitogen-Activated Protein Kinase Inhibition Ameliorates Angiotensin II-Induced Target Organ Damage Hypertension, March 1, 2007; 49(3): 481 - 489. [Abstract] [Full Text] [PDF]
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Y. Yu, Y.-M. Kang, Z.-H. Zhang, S.-G. Wei, Y. Chu, R. M. Weiss, and R. B. Felder Increased Cyclooxygenase-2 Expression in Hypothalamic Paraventricular Nucleus in Rats With Heart Failure: Role of Nuclear Factor {kappa}B Hypertension, March 1, 2007; 49(3): 511 - 518. [Abstract] [Full Text] [PDF]
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 R. Gros, Q. Ding, S. Armstrong, C. O'Neil, J. G. Pickering, and R. D. Feldman Rapid effects of aldosterone on clonal human vascular smooth muscle cells Am J Physiol Cell Physiol, February 1, 2007; 292(2): C788 - C794. [Abstract] [Full Text] [PDF]
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S. Shibata, M. Nagase, S. Yoshida, H. Kawachi, and T. Fujita Podocyte as the Target for Aldosterone: Roles of Oxidative Stress and Sgk1 Hypertension, February 1, 2007; 49(2): 355 - 364. [Abstract] [Full Text] [PDF]
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 K. Bedard and K.-H. Krause The NOX Family of ROS-Generating NADPH Oxidases: Physiology and Pathophysiology Physiol Rev, January 1, 2007; 87(1): 245 - 313. [Abstract] [Full Text] [PDF]
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B. M. Thobe, M. Frink, M. A. Choudhry, M. G. Schwacha, K. I. Bland, and I. H. Chaudry Src family kinases regulate p38 MAPK-mediated IL-6 production in Kupffer cells following hypoxia Am J Physiol Cell Physiol, September 1, 2006; 291(3): C476 - C482. [Abstract] [Full Text] |