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Biomedical subjects

Roland Baron

Publications and source records attributed to Roland Baron.

At least 37 records · Page 2Linked to original sources
PubMed2005

Subcutaneous injections of a TNF-alpha antagonistic peptide inhibit both inflammation and bone resorption in collagen-induced murine arthritis.

The cyclic peptide WP9QY (YCWSQYLCY), which was designed to mimic the most critical tumor necrosis factor (TNF) alpha recognition loop on type1 TNF receptor, antagonizes the effects of TNFalpha. In this study, we investigated the effects of WP9QY peptide on collagen-induced arthritis (CIA) mice to evaluate its effects on inflammatory bone destruction. DBA/1J mice were injected intradermally at the base of the tail with bovine type II collagen, emulsified in complete Freund's adjuvant on day 0 and 21. The three sets of WP9QY peptide injections (24 mg/kg x 8 times per day) were performed before the onset of paw swelling. Mice were sacrificed at day 38 and thereafter, the arthritis scores as well as radiographical and histological outcomes were assessed. WP9QY peptide inhibited CIA-induced increase in the arthritis score. Furthermore, histomorphometric analysis of the tibial epiphysis region revealed that WP9QY peptide inhibited the increase of synovial pannus infiltration and the decrease of bone volume, which were induced by the CIA. The WP9QY treatment prevented the inflammation as well as bone destruction of the joints in the CIA mice, suggesting that the administration of WP9QY peptide might be useful for developing a drug to prevent inflammatory bone destruction.

Animals↗
PubMed2004

Src kinase activity is essential for osteoclast function.

Deletion of the c-src gene impairs osteoclast bone resorbing activity, causing osteopetrosis. Although it has been concluded that restoring only the Src adaptor function at least partly rescues the cell attachment and skeletal phenotypes, the contribution of Src kinase activity remains controversial. Src forms a complex with Pyk2 and Cbl after adhesion-induced stimulation of alpha(V)beta(3) integrin. To demonstrate the importance of the Pyk2-Src association in osteoclasts and to distinguish the contributions of the Src adaptor and kinase activities in cytoskeletal organization and osteoclast function, we expressed mutants of Src and Pyk2 in osteoclasts using adenovirus vectors. Eliminating the Src-binding site on Pyk2 (Pyk2(Y402F)) markedly inhibited bone resorption by osteoclast-like cells, whereas kinase-dead Pyk2 had little effect. Kinase-dead Src, unlike kinase-dead Pyk2, markedly inhibited the bone-resorbing activity of wild type osteoclasts and failed to significantly restore bone-resorbing activity to Src(-/-) osteoclast-like cells. Activation of Src kinase by overexpressing kinase-dead Csk failed to reverse the inhibitory effect of Pyk2(Y402F), suggesting that osteoclastic bone resorption requires both c-Src kinase activity and the targeting of Src kinase by Pyk2. Src-catalyzed phosphorylation of Cbl on Tyr-731 is reported to induce the activation and recruitment of phosphatidylinositol 3-kinase to the cell membrane in a signaling pathway that is critical for osteoclast function. Expressing the Cbl(Y731F) mutant in osteoclasts markedly reduced their bone resorbing activity, suggesting that phosphorylation of Cbl(Y731) and the subsequent recruitment and activation of phosphatidylinositol 3-kinase may be critical signaling events downstream of Src in osteoclasts.

Adenoviridae↗
PubMed2004

Interaction of estrogen receptor alpha with protein kinase C alpha and c-Src in osteoblasts during differentiation.

In cultured osteoblasts, protein kinase C (PKC) activity increases and estrogen receptor alpha (ERalpha) binding capacity decreases upon confluence. We investigated potential interactions between ERalpha and PKC isoforms and their confluence-induced modulations in clonal ROS.SMER#14 cells and primary osteoblasts. In sub-confluent ROS.SMER#14 cells, which express an exogenous plus small amounts of the endogenous ERalpha gene, the receptor appeared as two main bands of approximately 66 and approximately 46 kDa. In over-confluent, more differentiated cells, the cytosolic approximately 66 kDa ERalpha appeared decreased and the approximately 46 kDa variant increased. Enhanced expression and/or membrane translocation of PKCalpha and PKCepsilon, but not PKCzeta, was evidenced at over-confluence, along with transient increases in expression and kinase activity of c-Src, accompanied by membrane translocation of the kinase-activated enzyme. In contrast, negligible membrane translocation of PKCalpha and/or activated c-Src was observed in parental ROS 17/2.8 cells, which express low levels of full-length ERalpha. PKCalpha from over-confluent cells phosphorylated p60c-Src in vitro, suggesting functional interaction between the two kinases. ERalpha co-immunoprecipitated c-Src and PKCalpha, mostly in its cleaved form (PKMalpha). An analogous interaction was observed in primary osteoblasts. However, in these cells, much more PKCalpha/PKMalpha was ERalpha-co-immunoprecipitated at over-confluence, a condition in which the shorter, approximately 46 kDa ERalpha variant is increased. This interaction was enhanced by estradiol treatment or PKC down-regulation, but was unaffected by c-Src inhibition. These data highlight direct PKCalpha-c-Src-ERalpha interactions, which may be crucial in the modulation of estrogen responsiveness and the differentiation process in osteoblasts.

Animals↗
PubMed2004

Essential role for ADAM19 in cardiovascular morphogenesis.

Congenital heart disease is the most common form of human birth defects, yet much remains to be learned about its underlying causes. Here we report that mice lacking functional ADAM19 (mnemonic for a disintegrin and metalloprotease 19) exhibit severe defects in cardiac morphogenesis, including a ventricular septal defect (VSD), abnormal formation of the aortic and pulmonic valves, leading to valvular stenosis, and abnormalities of the cardiac vasculature. During mouse development, ADAM19 is highly expressed in the conotruncus and the endocardial cushion, structures that give rise to the affected heart valves and the membranous ventricular septum. ADAM19 is also highly expressed in osteoblast-like cells in the bone, yet it does not appear to be essential for bone growth and skeletal development. Most adam19(-/-) animals die perinatally, likely as a result of their cardiac defects. These findings raise the possibility that mutations in ADAM19 may contribute to human congenital heart valve and septal defects.

ADAM Proteins↗
PubMed2004

DeltaFosB induces osteosclerosis and decreases adipogenesis by two independent cell-autonomous mechanisms.

Osteoblasts and adipocytes may develop from common bone marrow mesenchymal precursors. Transgenic mice overexpressing DeltaFosB, an AP-1 transcription factor, under the control of the neuron-specific enolase (NSE) promoter show both markedly increased bone formation and decreased adipogenesis. To determine whether the two phenotypes were linked, we targeted overexpression of DeltaFosB in mice to the osteoblast by using the osteocalcin (OG2) promoter. OG2-DeltaFosB mice demonstrated increased osteoblast numbers and an osteosclerotic phenotype but normal adipocyte differentiation. This result firmly establishes that the skeletal phenotype is cell autonomous to the osteoblast lineage and independent of adipocyte formation. It also strongly suggests that the decreased fat phenotype of NSE-DeltaFosB mice is independent of the changes in the osteoblast lineage. In vitro, overexpression of DeltaFosB in the preadipocytic 3T3-L1 cell line had little effect on adipocyte differentiation, whereas it prevented the induction of adipogenic transcription factors in the multipotential stromal cell line ST2. Also, DeltaFosB isoforms bound to and altered the DNA-binding capacity of C/EBPbeta. Thus, the inhibitory effect of DeltaFosB on adipocyte differentiation appears to occur at early stages of stem cell commitment, affecting C/EBPbeta functions. It is concluded that the changes in osteoblast and adipocyte differentiation in DeltaFosB transgenic mice result from independent cell-autonomous mechanisms.

Adipocytes↗
PubMed2003

Regulation of osteoclast apoptosis by ubiquitylation of proapoptotic BH3-only Bcl-2 family member Bim.

Osteoclasts (OCs) undergo rapid apoptosis without trophic factors, such as macrophage colony-stimulating factor (M-CSF). Their apoptosis was associated with a rapid and sustained increase in the pro-apoptotic BH3-only Bcl-2 family member Bim. This was caused by the reduced ubiquitylation and proteasomal degradation of Bim that is mediated by c-Cbl. Although the number of OCs was increased in the skeletal tissues of bim-/- mice, the mice exhibited mild osteosclerosis due to reduced bone resorption. OCs differentiated from bone marrow cells of bim-/- animals showed a marked prolongation of survival in the absence of M-CSF, compared with bim+/+ OCs, but the bone-resorbing activity of bim-/- OCs was significantly reduced. Overexpression of a degradation-resistant lysine-free Bim mutant in bim-/- cells abrogated the anti-apoptotic effect of M-CSF, while wild-type Bim did not. These results demonstrate that ubiquitylation-dependent regulation of Bim levels is critical for controlling apoptosis and activation of OCs.

Animals↗
PubMed2003

Murine Frizzled-1 behaves as an antagonist of the canonical Wnt/beta-catenin signaling.

Activation of the Wnt signaling cascade provides key signals during development and in disease. Wnt signals are transduced by seven-transmembrane Frizzleds (Fzs) and the single transmembrane low density lipoprotein receptor-related proteins 5 or 6. In the course of the analysis of genes regulated by bone morphogenetic protein 2 in mesenchymal cells we found a significant induction of murine Frizzled-1 (mFz1) gene expression. Unexpectedly overexpression of mFz1 dramatically repressed the induction of alkaline phosphatase mediated by either bone morphogenetic protein 2 or Wnt3a in these cells. Moreover mFz1 overexpression significantly repressed both beta-catenin translocation into the nucleus and T cell factor signaling mediated by Wnt3a. Importantly microinjection of mFz1 transcript in Xenopus embryo inhibited the ability of Wnt1 to induce the expression of the Wnt/beta-catenin target gene Siamois in animal cap assay and secondary axis formation in whole embryo. By using chimeric constructs in which N- and C-terminal segments of mFz1 were replaced by the corresponding parts of Xfz3 we demonstrated that the antagonistic activity resides in the cysteine-rich domain of the N-terminal part. The antagonist activity of mFz1 could be prevented by overexpression of Galphaq-(305-359), which specifically uncouples Gq-coupled receptors, suggesting that Galphaq signaling contributes to the inhibition of Wnt/beta-catenin pathway by mFz1. This is the first time that a Frizzled receptor has been reported to antagonize Wnt/beta-catenin.

Active Transport, Cell Nucleus↗
PubMed2003

Gp130-mediated signaling is necessary for normal osteoblastic function in vivo and in vitro.

Previous studies have shown that mice missing gp130, the common receptor subunit for many cytokines, die at or before birth with multiple skeletal abnormalities. Furthermore, interactions between PTH and gp130 signaling have suggested that gp130 signaling might influence calcium homeostasis. We, therefore, examined the function of osteoblasts, osteoclasts, and calcium homeostasis in gp130(-/-) mice, both in vivo and in vitro. Osteoblasts from these mice exhibit widespread abnormalities, including decreased alkaline phosphatase mRNA and protein, both in vivo and in osteoblast cultures. Although osteoclast number is increased in gp130(-/-) fetuses, these osteoclasts exhibit abnormalities in the resorptive organelle and the ruffled border, and the mice are mildly hypocalcemic. Although the hypocalcemia is associated with secondary hyperparathyroidism, the increase in PTH does not explain the increase in osteoclast number because removal of the PTH gene in gp130(-/-) fetuses does not importantly change osteoclast number. Calvarial bone resorption in response to PTH is defective, as is the ability of osteoblastic cells from gp130(-/-) mice to stimulate osteoclastogenesis from normal precursors in vitro or to increase receptor activator of nuclear factor-kappa B ligand mRNA levels after exposure to PTH. These studies demonstrate the importance of gp130 signaling for osteoblast function and calcium homeostasis.

Animals↗
PubMed2003

Tyrosine phosphatase epsilon is a positive regulator of osteoclast function in vitro and in vivo.

Protein tyrosine phosphorylation is a major regulator of bone metabolism. Tyrosine phosphatases participate in regulating phosphorylation, but roles of specific phosphatases in bone metabolism are largely unknown. We demonstrate that young (<12 weeks) female mice lacking tyrosine phosphatase epsilon (PTPepsilon) exhibit increased trabecular bone mass due to cell-specific defects in osteoclast function. These defects are manifested in vivo as reduced association of osteoclasts with bone and as reduced serum concentration of C-terminal collagen telopeptides, specific products of osteoclast-mediated bone degradation. Osteoclast-like cells are generated readily from PTPepsilon-deficient bone-marrow precursors. However, cultures of these cells contain few mature, polarized cells and perform poorly in bone resorption assays in vitro. Podosomes, structures by which osteoclasts adhere to matrix, are disorganized and tend to form large clusters in these cells, suggesting that lack of PTPepsilon adversely affects podosomal arrangement in the final stages of osteoclast polarization. The gender and age specificities of the bone phenotype suggest that it is modulated by hormonal status, despite normal serum levels of estrogen and progesterone in affected mice. Stimulation of bone resorption by RANKL and, surprisingly, Src activity and Pyk2 phosphorylation are normal in PTPepsilon-deficient osteoclasts, indicating that loss of PTPepsilon does not cause widespread disruption of these signaling pathways. These results establish PTPepsilon as a phosphatase required for optimal structure, subcellular organization, and function of osteoclasts in vivo and in vitro.

Animals↗
PubMed2003

Deletion of the gene encoding c-Cbl alters the ability of osteoclasts to migrate, delaying resorption and ossification of cartilage during the development of long bones.

During development of the skeleton, osteoclast (OC) recruitment and migration are required for the vascular invasion of the cartilaginous anlage and the ossification of long bones. c-Cbl lies downstream of the vitronectin receptor and forms a complex with c-Src and Pyk2 in a signaling pathway that is required for normal osteoclast motility. To determine whether the decreased motility we observed in vitro in c-Cbl(-/-) OCs translated into decreased cell migration in vivo, we analyzed the long bones of c-Cbl(-/-) mice during development. Initiation of vascularization and replacement of cartilage by bone were delayed in c-Cbl(-/-) mice, due to decreased osteoclast invasion of the hypertrophic cartilage through the bone collar. Furthermore, c-Cbl(-/-) mice show a delay in the formation of secondary centers of ossification, a thicker hypertrophic zone of the growth plate, and a prolonged presence of cartilaginous remnants in the spongiosa, confirming a decrease in resorption of the calcified cartilage. Thus, the decrease in motility of c-Cbl(-/-) osteoclasts observed in vitro results in a decreased ability of osteoclasts to invade and resorb bone and mineralized cartilage in vivo. These results confirm that c-Cbl plays an important role in osteoclast motility and resorbing activity.

Animals↗
PubMed2003

The alternatively spliced deltae13 transcript of the rabbit calcitonin receptor dimerizes with the C1a isoform and inhibits its surface expression.

Numerous alternatively spliced transcripts are generated from the gene for the G protein-coupled calcitonin receptor, and some of the splice variants show differences in receptor-mediated signaling events. This study showed that the deltae13 splice variant of the rabbit calcitonin receptor is expressed together with the more common C1a in osteoclast-like cells. Since other G protein-coupled receptors form homo- or heterodimers, we examined whether heterodimerization of the calcitonin receptor splice variants occurs and, if so, whether it affects the function of the receptor. Homodimers of both isoforms and deltae13/C1a heterodimers were detected by co-immunoprecipitation and fluorescence resonance energy transfer analysis. In contrast to the C1a isoform, the deltae13 isoform was not efficiently transported to the cell surface. When co-expressed with the C1a splice variant, the deltae13 isoform colocalized with the C1a isoform within the cell but not at the cell surface. Furthermore, the overexpression of the deltae13 variant led to a significant reduction of the C1a surface expression and consequently a reduction of the cAMP response and Erk phosphorylation after ligand stimulation. We therefore suggest that the deltae13 variant of the rabbit calcitonin receptor acts to regulate the surface expression of the C1a isoform.

Alternative Splicing↗
PubMed2003

Regulation of cytochrome c oxidase activity by c-Src in osteoclasts.

The function of the nonreceptor tyrosine kinase c-Src as a plasma membrane-associated molecular effector of a variety of extracellular stimuli is well known. Here, we show that c-Src is also present within mitochondria, where it phosphorylates cytochrome c oxidase (Cox). Deleting the c-src gene reduces Cox activity, and this inhibitory effect is restored by expressing exogenous c-Src. Furthermore, reducing endogenous Src kinase activity down-regulates Cox activity, whereas activating Src has the opposite effect. Src-induced Cox activity is required for normal function of cells that require high levels of ATP, such as mitochondria-rich osteoclasts. The peptide hormone calcitonin, which inhibits osteoclast function, also down-regulates Cox activity. Increasing Src kinase activity prevented the inhibitory effect of calcitonin on Cox activity and osteoclast function. These results suggest that c-Src plays a previously unrecognized role in maintaining cellular energy stores by activating Cox in mitochondria.

Animals↗
PubMed2003

Binding of filamin to the C-terminal tail of the calcitonin receptor controls recycling.

Many G protein-coupled receptors undergo endocytosis, but the mechanisms involved in endocytic sorting and recycling remain to be fully elucidated. We found that the G protein-coupled calcitonin receptor (CTR) undergoes tonic internalization and accumulates within the cell. Using a fluorescence loss in photobleaching assay, we classified these vesicles functionally as recycling vesicles. In a two-hybrid screening, we found that the actin-binding protein filamin interacted with the C-terminal tail of the CTR. The degradation of the receptor was profoundly increased in the absence of filamin or the CTR-filamin interaction. The absence of filamin was also associated with a marked decrease in recycling of the receptor from the endosomes to the cell surface. In contrast, calcitonin-induced inhibition of spontaneous filamin proteolysis was associated with increased recycling of the receptor to the cell surface and decreased degradation of the CTR, suggesting an important role for filamin in the endocytic sorting and recycling of the internalized CTR.

Binding Sites↗
PubMed2003

Potential role for ADAM15 in pathological neovascularization in mice.

ADAM15 (named for a disintegrin and metalloprotease 15, metargidin) is a membrane-anchored glycoprotein that has been implicated in cell-cell or cell-matrix interactions and in the proteolysis of molecules on the cell surface or extracellular matrix. To characterize the potential roles of ADAM15 during development and in adult mice, we analyzed its expression pattern by mRNA in situ hybridization and generated mice carrying a targeted deletion of ADAM15 (adam15(-/-) mice). A high level of expression of ADAM15 was found in vascular cells, the endocardium, hypertrophic cells in developing bone, and specific areas of the hippocampus and cerebellum. However, despite the pronounced expression of ADAM15 in these tissues, no major developmental defects or pathological phenotypes were evident in adam15(-/-) mice. The elevated levels of ADAM15 in endothelial cells prompted an evaluation of its role in neovascularization. In a mouse model for retinopathy of prematurity, adam15(-/-) mice had a major reduction in neovascularization compared to wild-type controls. Furthermore, the size of tumors resulting from implanted B16F0 mouse melanoma cells was significantly smaller in adam15(-/-) mice than in wild-type controls. Since ADAM15 does not appear to be required for developmental angiogenesis or for adult homeostasis, it may represent a novel target for the design of inhibitors of pathological neovascularization.

ADAM Proteins↗
PubMed2003

A functional androgen receptor is not sufficient to allow estradiol to protect bone after gonadectomy in estradiol receptor-deficient mice.

Although the role of estradiol in maintaining bone mass is well established, the relative contributions of the estradiol receptors ERalpha and ERbeta and of the androgen receptor (AR) remain controversial. To determine the role of ERalpha-mediated, ERbeta-mediated, and non-ER-mediated mechanisms in maintaining bone mass, gonadectomy and estradiol treatment were studied in ER-knockout mice. Estradiol treatment of ovariectomized ERalphabeta(-/-) mice failed to prevent bone loss, precluding significant effects of estradiol on bone through non-ER-signaling pathways. In contrast, estradiol prevented ovariectomy-induced bone loss in ERbeta(-/-) mice, as in WT males and females, indicating that ERalpha is the major mediator of estradiol effects in bone. No response of bone to estradiol was detected in orchidectomized ERalpha(-/-) mice, suggesting estradiol cannot protect bone mass via the AR in vivo. In contrast to female ERalphabeta(-/-) and male ERalpha(-/-) mice, female ERalpha(-/-) mice were partially protected against ovariectomy-induced bone loss by estradiol, confirming that ERbeta mediates estradiol effects in bone, but only in females and with a lower efficacy than ERalpha. We conclude that ERalpha is the main effector of estradiol's protective function in bone in both male and female mice, and that, in its absence, AR is not sufficient to mediate this response.

Androgen Antagonists↗
PubMed2003

BMP-2 controls alkaline phosphatase expression and osteoblast mineralization by a Wnt autocrine loop.

UNLABELLED: Wnt/beta-catenin signaling has recently been suggested to be involved in bone biology. The precise role of this cascade in osteoblast differentiation was examined. We show that a Wnt autocrine loop mediates the induction of alkaline phosphatase and mineralization by BMP-2 in pre-osteoblastic cells. INTRODUCTION: Loss of function of LRP5 leads to osteoporosis (OPPG syndrome), and a specific point mutation in this same receptor results in high bone mass (HBM). Because LRP5 acts as a coreceptor for Wnt proteins, these findings suggest a crucial role for Wnt signaling in bone biology. MATERIALS AND METHODS: We have investigated the involvement of the Wnt/LRP5 cascade in osteoblast function by using the pluripotent mesenchymal cell lines C3H10T1/2, C2C12, and ST2 and the osteoblast cell line MC3T3-E1. Transfection experiments were carried out with a number of elements of the Wnt/LRP5 pathway. Measuring osteoblast and adipocyte differentiation markers addressed the effect of this cascade on osteoblast differentiation. RESULTS: In mesenchymal cells, only Wnt's capable of stabilizing beta-catenin induced the expression of alkaline phosphatase (ALP). Wnt3a-mediated ALP induction was inhibited by overexpression of either Xddl, dickkopf 1 (dkk1), or LRP5deltaC, indicating that canonical beta-catenin signaling is responsible for this activity. The use of Noggin, a bone morphogenic protein (BMP) inhibitor, or cyclopamine, a Hedgehog inhibitor, revealed that the induction of ALP by Wnt is independent of these morphogenetic proteins and does not require de novo protein synthesis. In contrast, blocking Wnt/LRP5 signaling or protein synthesis inhibited the ability of both BMP-2 and Shh to induce ALP in mesenchymal cells. Moreover, BMP-2 enhanced Wntl and Wnt3a expression in our cells. In MC3T3-E1 cells, where endogenous ALP levels are maximal, antagonizing the Wnt/LRP5 pathway led to a decrease of ALP activity. In addition, overexpression of dkkl reduced extracellular matrix mineralization in a BMP-2-dependent assay. CONCLUSIONS: Our data strongly suggest that the capacity of BMP-2 and Shh to induce ALP relies on Wnt expression and the Wnt/LRP5 signaling cascade. Moreover the effects of BMP-2 on extracellular matrix mineralization by osteoblasts are mediated, at least in part, by the induction of a Wnt autocrine/paracrine loop. These results may help to explain the phenotype of OPPG patients and HBM.

Adipocytes↗
PubMed2002

Src kinase mediates phosphatidylinositol 3-kinase/Akt-dependent rapid endothelial nitric-oxide synthase activation by estrogen.

17beta-Estradiol activates endothelial nitric oxide synthase (eNOS), enhancing nitric oxide (NO) release from endothelial cells via the phosphatidylinositol 3-kinase (PI3-kinase)/Akt pathway. The upstream regulators of this pathway are unknown. We now demonstrate that 17beta-estradiol rapidly activates eNOS through Src kinase in human endothelial cells. The Src family kinase specific-inhibitor 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) abrogates 17beta-estradiol- but not ionomycin-stimulated NO release. Consistent with these results, PP2 blocked 17beta-estradiol-induced Akt phosphorylation but did not inhibit NO release from cells transduced with a constitutively active Akt. PP2 abrogated 17beta-estradiol-induced activation of PI3-kinase, indicating that the PP2-inhibitable kinase is upstream of PI3-kinase and Akt. A 17beta-estradiol-induced estrogen receptor/c-Src association correlated with rapid c-Src phosphorylation. Moreover, transfection of kinase-dead c-Src inhibited 17beta-estradiol-induced Akt phosphorylation, whereas constitutively active c-Src increased basal Akt phosphorylation. Estrogen stimulation of murine embryonic fibroblasts with homozygous deletions of the c-src, fyn, and yes genes failed to induce Akt phosphorylation, whereas cells maintaining c-Src expression demonstrated estrogen-induced Akt activation. Estrogen rapidly activated c-Src inducing an estrogen receptor, c-Src, and P85 (regulatory subunit of PI3-kinase) complex formation. This complex formation results in the successive activation of PI3-kinase, Akt, and eNOS with consequent enhanced NO release, implicating c-Src as a critical upstream regulator of the estrogen-stimulated PI3-kinase/Akt/eNOS pathway.

Adenoviridae↗