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PubMed2006

Vascular endothelial growth factor-A induces plaque expansion in ApoE knock-out mice by promoting de novo leukocyte recruitment.

Vascular endothelial growth factor-A is widely used in clinical trials for the treatment of cardiac ischemia. VEGF-A was recently suggested to act in a proinflammatory manner, which could aggravate adjacent atherogenesis in VEGF-A-based therapy. To assess potential bystander effects, VEGF-A was focally overexpressed in advanced atherosclerotic plaques in ApoE-/- mice. Sheer-induced carotid artery plaques were transluminally incubated with Ad.hVEGF-A leading to neointimal overexpression of VEGF-A. Ad.hVEGF-A treatment of pre-existing lesions was seen to promote plaque expansion, with a concomitant increase in macrophage and lipid content, whereas it lowered collagen content. In general, Ad.hVEGF-A-treated plaques displayed a more vulnerable phenotype. VEGF-A overexpression was not accompanied by increased microvessel development in the neointima, suggesting that VEGF-A destabilizes atherosclerotic plaques through an angiogenesis-independent mechanism. Intravital microscopy confirmed that treatment with Ad.hVEGF-A led to an increased monocyte adhesion, which was mediated by a VCAM-1/PECAM-1-dependent pathway. VEGF-A indeed induced a differential expression of VCAM-1 and PECAM-1 in endothelial cells. Our data underline the importance of regular monitoring of stenotic vessels adjacent to the site of VEGF-A application. We propose that VCAM-1/PECAM-1-directed cotherapy may be an efficient strategy to prevent bystander effects of focal VEGF-A therapy in patients suffering from cardiovascular disease.

Adenoviridae↗
PubMed2002

VEGF regulates haematopoietic stem cell survival by an internal autocrine loop mechanism.

Vascular endothelial growth factor (VEGF) is a principal regulator of blood vessel formation and haematopoiesis, but the mechanisms by which VEGF differentially regulates these processes have been elusive. Here we describe a regulatory loop by which VEGF controls survival of haematopoietic stem cells (HSCs). We observed a reduction in survival, colony formation and in vivo repopulation rates of HSCs after ablation of the VEGF gene in mice. Intracellularly acting small-molecule inhibitors of VEGF receptor (VEGFR) tyrosine kinase dramatically reduced colony formation of HSCs, thus mimicking deletion of the VEGF gene. However, blocking VEGF by administering a soluble VEGFR-1, which acts extracellularly, induced only minor effects. These findings support the involvement in HSC survival of a VEGF-dependent internal autocrine loop mechanism (that is, the mechanism is resistant to inhibitors that fail to penetrate the intracellular compartment). Not only ligands selective for VEGF and VEGFR-2 but also VEGFR-1 agonists rescued survival and repopulation of VEGF-deficient HSCs, revealing a function for VEGFR-1 signalling during haematopoiesis.

Animals↗
PubMed1998

Sequential DNA damage-independent and -dependent activation of NF-kappaB by UV.

NF-kappaB activation in response to UV irradiation of HeLa cells or of primary human skin fibroblasts occurs with two overlapping kinetics but totally different mechanisms. Although both mechanisms involve induced dissociation of NF-kappaB from IkappaBalpha and degradation of IkappaBalpha, targeting for degradation and signaling are different. Early IkappaBalpha degradation at 30 min to approximately 6 h is not initiated by UV-induced DNA damage. It does not require IkappaB kinase (IKK), as shown by introduction of a dominant-negative kinase subunit, and does not depend on the presence of the phosphorylatable substrate, IkappaBalpha, carrying serines at positions 32 and 36. Induced IkappaBalpha degradation requires, however, intact N- (positions 1-36) and C-terminal (positions 277-287) sequences. IkappaB degradation and NF-kappaB activation at late time points, 15-20 h after UV irradiation, is mediated through DNA damage-induced cleavage of IL-1alpha precursor, release of IL-1alpha and autocrine/paracrine action of IL-1alpha. Late-induced IkappaBalpha requires the presence of Ser32 and Ser36. The late mechanism indicates the existence of signal transfer from photoproducts in the nucleus to the cytoplasm. The release of the 'alarmone' IL-1alpha may account for some of the systemic effects of sunlight exposure.

Autocrine Communication↗
PubMed2006

Calcitonin gene-related peptide inhibits interleukin-1beta-induced endogenous monocyte chemoattractant protein-1 secretion in type II alveolar epithelial cells.

As important multifunctional cells in the lung, alveolar epithelial type II (AEII) cells secrete numerous chemokines on various stimuli. Our previous data showed that AEII cells also express the neuropeptide calcitonin gene-related peptide (CGRP) and the proinflammatory factor interleukin (IL)-1beta induces CGRP secretion in the A549 human AEII cell line. In the present study, the CGRP-1 receptor antagonist human (h)CGRP(8-37) (0.1-1 nM) greatly amplified the production of IL-1beta-induced monocyte chemoattractant protein (MCP)-1. The inhibition of CGRP expression by small interfering RNA significantly increased MCP-1 secretion on IL-1beta stimulation. However, exogenous hCGRP (10-100 nM) suppressed IL-1beta-evoked MCP-1 secretion in MCP-1 promoter activity, and CGRP gene stably transfected cell clones significantly inhibited both the mRNA and protein levels of MCP-1 induced by IL-1beta. These data imply that AEII-derived CGRP suppressed IL-1beta-induced MCP-1 secretion in an autocrine/paracrine mode. Subsequent investigation revealed that CGRP inhibited IL-1beta-evoked NF-kappaB activity by suppressing IkappaBalpha phosphorylation and degradation. Moreover, CGRP attenuated IL-1beta-induced reactive oxygen species (ROS) formation, the early event in proinflammatory factor signaling. We previously showed that the CGRP inhibitory effect was mediated by elevated intracellular cAMP and show here that analogs of cAMP, 8-bromoadenosine 3',5'-cyclic monophosphothioate and the Sp isomer of adenosine 3',5'-cyclic monophosphothioate, mimicked the CGRP suppressive effect on IL-1beta-induced ROS formation, NF-kappaB activation, and MCP-1 secretion. Thus increased endogenous CGRP secretion in lung inflammatory disease might eliminate the excessive response by elevating the cAMP level through inhibiting the ROS-NF-kappaB-MCP-1 pathway.

Autocrine Communication↗
PubMed2003

Epiregulin as a major autocrine/paracrine factor released from ERK- and p38MAPK-activated vascular smooth muscle cells.

BACKGROUND: The coordinated activation of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (p38MAPK) is critical for the induction of vascular and visceral smooth muscle cell (SMC) dedifferentiation. We previously reported that on the forced activation of both MAPKs, visceral SMCs secrete a non-heparin-binding protein factor(s) that is involved in the dedifferentiation of neighboring SMCs. In this study, we sought to identify the dedifferentiation factor(s) derived from vascular SMCs (VSMCs). METHODS AND RESULTS: We fractionated the VSMC dedifferentiation factor(s) in the conditioned medium obtained from differentiated VSMCs in which both ERK and p38MAPK were forcedly activated and identified epiregulin as a major autocrine/paracrine factor for VSMC dedifferentiation. The epiregulin-induced VSMC dedifferentiation was mediated through the coordinated activation of ERK and p38MAPK. Unsaturated lysophosphatidic acid and platelet-derived growth factor-BB, which are potent VSMC dedifferentiation factors, rapidly upregulated epiregulin mRNA expression in an ERK- and p38MAPK-dependent manner. Reverse transcriptase-polymerase chain reaction and/or immunohistological analyses revealed the restricted expression of epiregulin in human atherosclerotic and balloon-injured rat arteries, in which the phenotypic modulation of medial VSMCs occurred in vivo. CONCLUSIONS: Epiregulin is released from VSMCs primed by atherogenic factors and acts as a major autocrine/paracrine factor for VSMC dedifferentiation. It may be involved in the progression of vascular remodeling such as atherosclerosis.

Animals↗
PubMed1998

The survival-promoting effect of glial cell line-derived neurotrophic factor on axotomized corticospinal neurons in vivo is mediated by an endogenous brain-derived neurotrophic factor mechanism.

Autocrine trophic functions of brain-derived neurotrophic factor (BDNF) have been proposed for many central neurons because this neurotrophin displays striking colocalization with its receptor trkB within the CNS. In the cortex, the distribution patterns of BDNF and trkB expression are almost identical. Corticospinal neurons (CSNs) are a major cortical long-distance projecting system. They are localized in layer V of the somatosensory cortex, and their axons project into the spinal cord where they contribute to the innervation of spinal motoneurons. We have shown recently that adult CSNs express trkB mRNA and are rescued from axotomy-induced death by BDNF treatment. Half of the axotomized CSNs survived without BDNF infusions. These findings raise the possibility that endogenous cortical BDNF is involved in the trophic support of this neuronal population. To test the hypothesis that endogenous cortical BDNF promotes survival of adult CSNs, we infused the BDNF-neutralizing affinity-purified antibody RAB to axotomized and unlesioned CSNs for 7 d. This treatment resulted in increased death of axotomized CSNs. Survival of unlesioned CSNs was not affected by RAB treatment. In situ hybridizations for BDNF and trkB mRNA revealed that virtually all CSNs express trkB, whereas only half of them express BDNF. Thus, autocrine/paracrine mechanisms are likely to contribute to the endogenous BDNF protection of axotomized CSNs. We have demonstrated previously that, in addition to BDNF, glial cell line-derived neurotrophic factor (GDNF) and neurotrophin 3 (NT-3) also rescue CSNs from axotomy-induced death. We now show that the rescuing by GDNF requires the presence of endogenous cortical BDNF, implicating a central role of this neurotrophin in the trophic support of axotomized CSNs and a trophic cross-talk between BDNF and GDNF regarding the maintenance of lesioned CSNs. In contrast, NT-3 promotes survival of axotomized CSNs even when endogenous cortical BDNF is neutralized by RAB, indicating a potential of compensatory mechanisms for the trophic support of CSNs.

Animals↗
PubMed2003

Expression of NOS1 and soluble guanylyl cyclase by human kidney epithelial cells: morphological evidence for an autocrine/paracrine action of nitric oxide.

BACKGROUND: Nitric oxide plays an important role in the kidney through effects on both renal hemodynamics and tubular functions. Tubular epithelial cells are thus a target for nitric oxide. However, as to whether tubular epithelial cells endogeneously produce nitric oxide under physiologic conditions in human kidney is currently unknown. The aim of the present study was to characterize and localize in situ the nitric oxide synthase (NOS) isoforms (NOS1, NOS2, and NOS3) expressed in human normal kidney, and soluble guanylyl cyclase, the well-known target for nitric oxide. METHODS: Five complementary experimental approaches were used: (1) detection of NOS reductase activity by nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry, (2) immunolocalization of the NOS isoforms (NOS1, NOS2, NOS3), (3) immunoblot analysis, (4) quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis of NOS mRNA, and (5) measurement of NOS activity as the conversion rate of l-[14C]-arginine to l-[14C]-citrulline. In addition, in situ detection of soluble guanylyl cyclase was assessed by immunohistochemistry. RESULTS: All these techniques led to consistent results showing that epithelial cells of most tubules along the human nephron exhibit functional NOS1, with a corticomedullary gradient observed both at the protein and mRNA levels. Moreover, epithelial cells expressing NOS1 also express soluble guanylyl cyclase, indicating that these cells possess the machinery for autocrine/paracrine effect of nitric oxide. CONCLUSION: The present study demonstrates that NOS1 is strongly expressed in most tubules of the human nephron and therefore invites to consider epithelial cells as one of the major source of nitric oxide in the human kidney under physiologic conditions.

Autocrine Communication↗
PubMed2000

Progesterone is an autocrine/paracrine regulator of human granulosa cell survival in vitro.

Ovarian follicles are composed of granulosa cells (GC), which undergo apoptosis within 24 hours of culture in serum-free medium. The present study was designed to assess the role of progesterone in regulating human GC survival. Human GC were isolated from follicular aspirates of women undergoing in vitro fertilization. GC were then cultured for 24 hours in serum-free media supplemented with progesterone and/or the progesterone antagonist RU486 and dexamethasone. Cells were then fixed and assessed for apoptosis by in situ end labeling of DNA fragments, cell cycle analysis of DNA content, and electron microscopy. When compared with controls, progesterone reduced and RU486 increased the percentage of apoptotic GC (p < 0.05), whereas dexamethasone had no effect. In addition, RU486 inhibited the protective effect of progesterone on GC survival (p < 0.05). Taken together, these data indicate that progesterone inhibits human GC apoptosis, and this effect is mediated through the progesterone receptor.

Adult↗
PubMed2000

Insulin-like growth factor (IGF)-binding protein-related protein-1: an autocrine/paracrine factor that inhibits skeletal myoblast differentiation but permits proliferation in response to IGF.

Skeletal myogenic cells respond to the insulin-like growth factors (IGF-I and IGF-II) by differentiating or proliferating, which are mutually exclusive pathways. What determines which of these responses to IGF skeletal myoblast undergo is unclear. IGF-binding protein-related protein 1 (IGFBP-rP1) is a secreted protein with close homology to the IGF-binding proteins (IGFBPs) in the N-terminal region. IGFBP-rP1, previously called mac25 and IGFBP-7, is highly expressed in C2 skeletal myoblasts during the proliferative phase, but is down-regulated during myoblast differentiation. To determine the role of IGFBP-rP1 in myogenesis, IGFBP-rP1 was overexpressed in C2 myoblasts using a retroviral vector. Western blots indicated that the resulting C2-rP1 myoblasts secreted approximately 27-fold higher levels of IGFBP-rP1 than control C2-LX myoblasts that were transduced with a control vector (LXSN). Compared with C2-LX myoblasts, the differentiation responses of C2-rP1 myoblasts to IGF-I, IGF-II, insulin, and des(1-3)IGF-I were significantly reduced (P < 0.05). However, proliferation responses of C2-rP1 and C2-LX myoblasts to these same factors were not significantly different. Exposure of control C2-LX myoblasts to factors secreted by C2-rP1 myoblasts using a transwell coculture system reduced C2-LX myoblast differentiation significantly (P < 0.05). Experiments with the mitogen-activated protein kinase (MAPK) kinase inhibitor PD098059 suggested that IGFBP-rP1 inhibits a MAPK-dependent differentiation pathway. In confirmation of this idea, levels of phosphorylated extracellular signal-regulated kinase-2 (a MAPK) were reduced in C2-rP1 myoblasts compared with those in C2-LX myoblasts. These findings indicate that IGFBP-rP1 may function as an autocrine/paracrine factor that specifies the proliferative response to the IGFs in myogenesis.

Autocrine Communication↗
PubMed2005

Activation of nuclear factor-kappaB via endogenous tumor necrosis factor alpha regulates survival of axotomized adult sensory neurons.

Embryonic dorsal root ganglion (DRG) neurons die after axonal damage in vivo, and cultured embryonic DRG neurons require exogenous neurotrophic factors that activate the neuroprotective transcription factor nuclear factor-kappaB (NF-kappaB) for survival. In contrast, adult DRG neurons survive permanent axotomy in vivo and in defined culture media devoid of exogenous neurotrophic factors in vitro. Peripheral axotomy in adult rats induces local accumulation of the cytokine tumor necrosis factor alpha (TNFalpha), a potent activator of NF-kappaB activity. We tested the hypothesis that activation of NF-kappaB stimulated by endogenous TNFalpha was required for survival of axotomized adult sensory neurons. Peripheral axotomy of lumbar DRG neurons by sciatic nerve crush induced a very rapid (within 2 h) and significant elevation in NF-kappaB-binding activity. This phenomenon was mimicked in cultured neurons in which there was substantial NF-kappaB nuclear translocation and a significant rise in NF-kappaB DNA-binding activity after plating. Inhibitors of NF-kappaB (SN50 or NF-kappaB decoy DNA) resulted in necrotic cell death of medium to large neurons (> or =40 microm) within 24 h (60 and 75%, respectively), whereas inhibition of p38 and mitogen-activated protein/extracellular signal-regulated kinase did not effect survival. ELISA revealed that these cultures contained TNFalpha, and exposure to an anti-TNFalpha antibody inhibited NF-kappaB DNA-binding activity by approximately 35% and killed approximately 40% of medium to large neurons within 24 h. The results show for the first time that cytokine-mediated activation of NF-kappaB is a component of the signaling pathway responsible for maintenance of adult sensory neuron survival after axon damage.

Animals↗
PubMed2003

Adrenomedullin is an autocrine/paracrine growth factor for rat vascular smooth muscle cells.

Adrenomedullin is a potent vasodilator peptide secreted by vascular endothelial and smooth muscle cells. Adrenomedullin stimulates the proliferation of quiescent rat vascular smooth muscle cells (VSMCs) via p42/p44 ERK/MAP kinase activation. Recently, receptor-activity-modifying proteins (RAMPs) have been shown to transport calcitonin-receptor-like-receptor (CRLR) to the cell surface to present either as CGRP receptor or adrenomedullin receptor. We investigated whether adrenomedullin acts as an autocrine/paracrine growth factor for cultured rat VSMCs and whether coexpressions of RAMP isoform and CRLR may mediate p42/p44 ERK/MAP kinase activation by adrenomedullin. Adrenomedullin dose-dependently stimulated the proliferation of quiescent rat VSMCs, and this effect was inhibited by an adrenomedullin receptor antagonist, a MAP kinase kinase inhibitor and phosphatidylinositol 3-kinase inhibitors. Addition of either CGRP(8-37) or anti-adrenomedullin antibody to exponentially growing rat VSMCs inhibited the serum-induced cell proliferation, suggesting its role as an autocrine/paracrine growth factor. Cotransfection of RAMP2 or RAMP3 with CRLR into rat VSMCs potentiated activation of cAMP activity, but not of p42/p44 ERK/MAP kinase activity in response to adrenomedullin. Our results suggest that adrenomedullin is an autocrine/paracrine growth factor for rat VSMCs via p42/p44 ERK/MAP kinase and phosphatidylinositol 3-kinase pathways and that it is not mediated by human RAMP-CRLR receptors.

Adrenomedullin↗
PubMed2005

Autocrine/paracrine involvement of insulin-like growth factor-I and its receptor in chronic lymphocytic leukaemia.

Chronic lymphocytic leukaemia (CLL) is characterized by the accumulation of long-lived B lymphocytes blocked in G(0/1) by impaired apoptosis. As insulin-like growth factor-I (IGF-I) is known for its antiapoptotic effects in different cell types, we investigated whether IGF-I and its receptor (IGF-IR) participate in autocrine/paracrine loops affecting the survival of CLL cells. IGF-IR protein and mRNA was present in CLL cells in 44% and 59% of patients respectively. IGF-IR expression in CLL patients was positively correlated with the expression of the antiapoptotic protein Bcl-2 and was involved in CLL cell survival in vitro. Serum IGF-I was elevated in CLL patients, but growth hormone (GH) was normal. CLL cells expressed IGF-I mRNA and secreted the growth factor in vitro. Therefore, local production of IGF-I can account for the increased levels of serum IGF-I, independently of GH, and may be related to autocrine/paracrine control of lymphocyte survival acting at IGF-IR. This is the first demonstration of IGF-IR expression in a subgroup of CLL patients and of its antiapoptotic effects in vitro, highlighting the importance of this growth factor receptor as a possible therapeutic target in CLL.

Aged↗
PubMed2000

Interleukin-1alpha-dependent regulation of matrix metalloproteinase-9(MMP-9) secretion and activation in the epithelial cells of odontogenic jaw cysts.

Interleukin-1alpha (IL-1alpha) and matrix metalloproteinase-9 (MMP-9) are thought to be involved in odontogenic cyst expansion. In this study, we investigated the effects of IL-1alpha on the secretion and activation of MMP-9 in odontogenic jaw cysts. An active form of MMP-9 was present in odontogenic keratocyst (6 of 8 cases) fluids more frequently than dentigerous cyst (3 of 10 cases) and radicular cyst (3 of 10 cases) fluids, although proMMP-9 was present in all cyst fluids. Odontogenic keratocyst fragments in explant culture secreted a larger amount of IL-1alpha than dentigerous cyst and radicular cyst fragments in explant culture, and spontaneously secreted both proMMP-9 and an active form of MMP-9. The fragments of dentigerous cysts and radicular cysts secreted a small amount of proMMP-9, but no active form of MMP-9. Exogenously added recombinant human IL-1alpha (rhlL-1alpha) increased the secretion and activation of proMMP-9 in the fragments of dentigerous cysts and radicular cysts. The epithelial cells isolated from odontogenic keratocysts secreted IL-1alpha and proMMP-9 without stimulation. Under the cultivation on a fibronectin-coated dish, rhIL-1alpha increased the secretion of proMMP-9 from the epithelial cells in a dose-dependent manner. Moreover, rhIL-1alpha induced the secretion of proMMP-3 and plasminogen activator urokinase (u-PA) from the epithelial cells, and converted the secreted proMMP-3 to the active form in the presence of plasminogen. The secreted proMMP-9 was also activated in the presence of rhIL-1alpha and plasminogen. Hence, our results suggest that IL-1alpha may up-regulate not only proMMP-9 secretion but also proMMP-9 activation by inducing proMMP-3 and u-PA production in the cyst epithelial cells by autocrine/paracrine regulatory mechanisms.

Autocrine Communication↗
PubMed2003

Autocrine/paracrine activation of the GABA(A) receptor inhibits the proliferation of neurogenic polysialylated neural cell adhesion molecule-positive (PSA-NCAM+) precursor cells from postnatal striatum.

GABA and its type A receptor (GABA(A)R) are present in the immature CNS and may function as growth-regulatory signals during the development of embryonic neural precursor cells. In the present study, on the basis of their isopycnic properties in a buoyant density gradient, we developed an isolation procedure that allowed us to purify proliferative neural precursor cells from early postnatal rat striatum, which expressed the polysialylated form of the neural cell adhesion molecule (PSA-NCAM). These postnatal striatal PSA-NCAM+ cells were shown to proliferate in the presence of epidermal growth factor (EGF) and formed spheres that preferentially generated neurons in vitro. We demonstrated that PSA-NCAM+ neuronal precursors from postnatal striatum expressed GABA(A)R subunits in vitro and in situ. GABA elicited chloride currents in PSA-NCAM+ cells by activation of functional GABA(A)R that displayed a typical pharmacological profile. GABA(A)R activation in PSA-NCAM+ cells triggered a complex intracellular signaling combining a tonic inhibition of the mitogen-activated protein kinase cascade and an increase of intracellular calcium concentration by opening of voltage-gated calcium channels. We observed that the activation of GABA(A)R in PSA-NCAM+ neuronal precursors from postnatal striatum inhibited cell cycle progression both in neurospheres and in organotypic slices. Furthermore, postnatal PSA-NCAM+ striatal cells synthesized and released GABA, thus creating an autocrine/paracrine mechanism that controls their proliferation. We showed that EGF modulated this autocrine/paracrine loop by decreasing GABA production in PSA-NCAM+ cells. This demonstration of GABA synthesis and GABA(A)R function in striatal PSA-NCAM+ cells may shed new light on the understanding of key extrinsic cues that regulate the developmental potential of postnatal neuronal precursors in the CNS.

Animals↗
PubMed2003

Perturbation of autocrine/paracrine loops of burst-forming units of erythroid-derived cells in rHuEPO-hyporesponsive hemodialysis patients.

BACKGROUND: Quantitative or qualitative abnormalities of erythroid progenitors in patients with chronic renal failure (CRF) could be the major factor for recombinant human erythropoietin (rHuEPO) hyporesponsiveness and severe anemia in hemodialysis (HD) patients receiving rHuEPO therapy. METHODS: Purified 1 x 10(4) circulating CD34+ cells isolated from rHuEPO-hyporesponsive HD patients (EPO-H; n = 10), rHuEPO-responsive non-HD patients with CRF (EPO-R; n = 8), nonanemic HD patients without rHuEPO therapy (EPO-W/O; n = 10), and healthy volunteer controls (CON; n = 10) were subjected to a methylcellulose culture system supplemented with rHuEPO, recombinant human interleukin-3 (IL-3), recombinant human stem cell factor (SCF), and recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) for 14 days. RESULTS: The average number of burst-forming units of erythroids (BFU-Es) was significantly less in the EPO-H group compared with the CON and EPO-W/O groups. Furthermore, colony size also was significantly smaller in the EPO-H group. Total RNAs were extracted from approximately 100 colonies/patient and subjected to complementary DNA expression array studies of 268 growth factors, cytokines, chemokines, and their receptors. A characteristic cluster upregulated in the EPO-R and EPO-W/O groups and downregulated in the EPO-H group was identified that contained various cytokines and growth factors, including IL-6, GM-CSF, vascular endothelial growth factor B, IL-9, IL-3, leukemia inhibitory factor, and interferon alpha-2, and such receptors as thrombopoietin receptor, IL-9 receptor, and colony-stimulating factor 1 receptor. CONCLUSION: These data suggest that the cross-talk network or autocrine/paracrine regulatory loop is critically impaired in BFU-E-derived cells in EPO-H patients, and investigation of these cluster genes would facilitate the development of novel therapeutic strategies for such patients.

Aged↗
PubMed2000

Voltage-dependent, pertussis toxin insensitive inhibition of calcium currents by histamine in bovine adrenal chromaffin cells.

Histamine is a known secretagogue in adrenal chromaffin cells. Activation of G-protein linked H(1) receptors stimulates phospholipase C, which generates inositol trisphosphate leading to release of intracellular calcium stores and stimulation of calcium influx through store operated and other channels. This calcium leads to the release of catecholamines. In chromaffin cells, the main physiological trigger for catecholamine release is calcium influx through voltage-gated calcium channels (I(Ca)). Therefore, these channels are important targets for the regulation of secretion. In particular N- and P/Q-type I(Ca) are subject to inhibition by transmitter/hormone receptor activation of heterotrimeric G-proteins. However, the direct effect of histamine on I(Ca) in chromaffin cells is unknown. This paper reports that histamine inhibited I(Ca) in cultured bovine adrenal chromaffin cells and this response was blocked by the H(1) antagonist mepyramine. With high levels of calcium buffering in the patch pipette solution (10 mM EGTA), histamine slowed the activation kinetics and inhibited the amplitude of I(Ca). A conditioning prepulse to +100 mV reversed the kinetic slowing and partially relieved the inhibition. These features are characteristic of a membrane delimited, voltage-dependent pathway which is thought to involve direct binding of G-protein betagamma subunits to the Ca channels. However, unlike virtually every other example of this type of inhibition, the response to histamine was not blocked by pretreating the cells with pertussis toxin (PTX). The voltage-dependent, PTX insensitive inhibition produced by histamine was modest compared with the PTX sensitive inhibition produced by ATP (28% vs. 53%). When histamine and ATP were applied concomitantly there was no additivity of the inhibition beyond that produced by ATP alone (even though the agonists appear to activate distinct G-proteins) suggesting that the inhibition produced by ATP is maximal. When experiments were carried out under conditions of low levels of calcium buffering in the patch pipette solution (0.1 mM EGTA), histamine inhibited I(Ca) in some cells using an entirely voltage insensitive pathway. We demonstrate that activation of PTX insensitive G-proteins (most likely Gq) by H(1) receptors inhibits I(Ca). This may represent a mechanism by which histamine exerts inhibitory (in addition to previously identified stimulatory) effects on catecholamine release.

Adenosine Triphosphate↗
PubMed2002

Protein calorie restriction affects nonhepatic IGF-I production and the lymphoid system: studies using the liver-specific IGF-I gene-deleted mouse model.

Nutritional status is a critical factor that modulates the responsiveness of the liver to GH and the resulting production of endocrine (mostly liver-derived) IGF-I. Using a conditional Cre/lox P system, we have established a liver-specific IGF-I-deficient mouse model. Despite the reduction in the circulating IGF-I (75%), the growth parameters are normal, except for the reduced spleen size, providing a unique model to study the effect of protein restriction on the autocrine/paracrine GH/IGF-I axis. To determine the effects of protein calorie malnutrition on the spleen, liver-specific IGF-I-deficient mice were assigned to one of four isocaloric diets, differing in the protein content (20, 12, 4, and 0%), for a period of 10 d. A low protein intake decreased the nonhepatic IGF-I secretion into the circulation, whereas it caused an increase in the level of circulating GH. This supports the view that nonhepatic IGF-I production contributes to circulating IGF-I levels. The lack of dietary protein led to an up-regulation of GH and IGF-I receptors expression in the spleen, whereas the IGF-I mRNA remained unchanged, as was demonstrated by flow cytometry and ribonuclease protection assay. B lymphocytes seem to be responsible for the up-regulated GH/IGF-I receptor expression. Northern blot analysis showed an up-regulation of IGF-binding protein-3 mRNA levels, which suggests that the protein deprivation may lead to an increased sequestration of circulating or locally synthesized IGF-I. These results support the hypothesis that the splenic GH/IGF-I axis responds to the nutritional stress caused by a low protein intake, to maintain the tissue homeostasis.

Animals↗