Effects of oxysterols on cell viability, inflammatory cytokines, VEGF, and reactive oxygen species production on human retinal cells: cytoprotective effects and prevention of VEGF secretion by resveratrol.

BACKGROUND AND AIMS: Oxysterols are assumed to play important roles in age-related macular degeneration, a major cause of blindness. So we characterized the cytotoxic, oxidative, inflammatory, and angiogenic activities of oxysterols (7ß-hydroxycholesterol (7ß-OH), 7-ketocholesterol (7KC), 25-hydroxycholesterol (25-OH)) in human retinal ARPE-19 cells, and evaluated the protective effects of resveratrol (Rsv: 1 µM), a polyphenol from red wine. METHODS: ARPE-19 cells were treated with 7ß-OH, 7KC, or 25-OH (5-40 µg/mL; 24-48 h) without or with Rsv. Cell viability was determined using trypan blue and the MTT assay. Cell death was characterized by electron microscopy and in situ detection of activated caspases with fluorochrome-labeled inhibitors of caspases. Reactive oxygen species (ROS) production was measured with hydroethidine. ELISA methods and a cytometric bead assay were used to quantify cytokines involved in inflammation (IL-8, IL-1ß, IL-6, IL-10, IL-12p70, TNF-α, MCP-1) and VEGF. RESULTS: 7ß-OH and 7KC triggered a caspase-independent cell death process associated with the presence of multilamellar cytoplasmic structures evocating phospholipidosis, increased ROS production, and IL-8 secretion. 7ß-OH enhanced VEGF secretion. No cytotoxic effects were identified with 25-OH, which highly stimulated ROS production, MCP-1, and VEGF secretion. With oxysterols, no IL-10, TNF-α, and IL-12p70 secretion were detected. 25-OH induced IL-8 secretion through the MEK/ERK½ signaling pathway, and Rsv showed cytoprotective activities and inhibited VEGF secretion. CONCLUSION: 7ß-OH, 7KC, and 25-OH have cytotoxic, oxidative, inflammatory, and/or angiogenic activities on ARPE-19 cells. As Rsv has some protective effects against oxysterol-induced cell death and VEGF secretion it could be valuable in ARMD treatment.

Prevention of inflammation-mediated acquisition of metastatic properties of benign mouse fibrosarcoma cells by administration of an orally available superoxide dismutase.

Weakly tumorigenic and nonmetastatic QR-32 cells derived from a fibrosarcoma in C57BL6 mouse are converted to malignant cells once they have grown after being coimplanted with a gelatine sponge which induces inflammation. We administered a newly developed peroral superoxide dismutase (SOD), oxykine, and as control vehicle, gliadin and saline, starting 2 days before the coimplantation and continued daily throughout the experiment. In the oxykine group, tumour incidence was lower (41%) than in the gliadin or saline group (83 and 79%, respectively). The inhibitory effect of oxykine was lost when an individual component of oxykine was administered, that is, SOD alone and gliadin alone. The effect was also abolished when administered by intraperitoneal route. When perfused in situ with nitroblue tetrazolium, an indicator of superoxide formation, the tumour masses from gliadin and saline groups displayed intense formazan deposition, whereas, those from oxykine group had less deposition. Enzymatic activity of SOD was also increased in oxykine group. Arising tumour cells in gliadin and saline groups acquired metastatic phenotype, but those in oxykine group showed reduced metastatic ability. These results suggested that the orally active SOD derivative prevented tumour progression promoted by inflammation, which is thought to be through scavenging inflammatory cell-derived superoxide anion.

Contribution of nitric oxide to the apoptotic process in human B cell chronic lymphocytic leukaemia.

B cell chronic lymphocytic leukaemia (B-CLL) is characterised by defective apoptosis that cannot be explained solely on the basis of the known chromosomal abnormalities. We and other have now reported that the leukemic cells spontaneously display the inducible isoform of nitric oxide synthase, iNOS. Inhibition of the iNOS pathway leads to increased apoptosis of the tumoral cells in vitro, indicating that the endogenous release of NO contributes to their resistance to the normal apoptotic process. The factors that induce the expression of iNOS in vivo in the leukemic cells are not yet identified. Yet, as interaction of B-CLL leukemic cells with bone marrow stromal cells promotes their survival, the involvement of adhesion molecules and integrins may be suspected. The engagement of CD23 stimulates iNOS activation in the tumoral cells, suggesting that in vivo interaction of CD23 with one of its recognised ligands may contribute to iNOS induction. A role for CD40-CD40 ligand interaction may also be hypothesised. The mechanisms involved in the anti-apoptotic role of NO are not fully understood, but may implicate the inhibition of caspase activity, hence the impairment of the Fas pathway. In addition, the mitochondrial membrane potential disruption appears to be a NO-sensitive step in the apoptosis cascade. The presence of a NOS displaying anti-apoptotic properties has now been recognised in different cell types, including various leukaemia. A better knowledge of the mechanisms governing the ultimate fate of NO, anti- versus pro-apoptotic would allow the development of new therapeutic approaches for the treatment of these diseases.

Expression of a functional inducible nitric oxide synthase in hairy cell leukaemia and ESKOL cell line.

The expression of nitric oxide synthase (NOS) isoforms was investigated in the established ESKOL hairy cell line and in leukemic cells of patients with hairy cell leukemia (HCL). By reverse transcription-polymerase chain reaction (RT-PCR), these cells were found to spontaneously express inducible NOS (iNOS)-specific mRNA, but not endothelial constitutive NOS (ecNOS) mRNA. The iNOS protein was detected by immunofluorescence in the cytoplasm of permeabilized leukemic cells and ESKOL cells, using different anti-iNOS monoclonal antibodies. A protein of 135 kDa was identified by Western blotting in ESKOL and HCL lysates, confirming the presence of an iNOS in these cells. Cytosolic homogenates displayed NOS catalytic activity, as measured by the conversion of 14C-labelled L-arginine into 14C L-citrulline and by detection in situ using the DAF-2DA (diaminofluorescein diacetate) NO-sensitive fluorescent probe. Ligation of CD23 (low affinity IgE receptor) was found to increase iNOS expression in ESKOL and conversely to decrease the percentage of cells undergoing apoptosis, as measured by the percentage of cells expressing annexin V. These results indicate that, as in chronic B cell lymphocytic leukemia cells (B-CLL) a functional iNOS is expressed constitutively in hairy cells that contributes to protecting these tumoral cells from apoptosis.

Regulation of ICAM-1/CD54 expression on human endothelial cells by hydrogen peroxide involves inducible NO synthase.

Expression of the inducible isoform of nitric oxide synthase (iNOS) is stimulated by cytokines in human epithelial cells. This work indicates that incubation of human umbilical cord endothelial cells with combinations of interleukin-1beta, tumor necrosis factor alpha, and interferon-gamma stimulated the synthesis of iNOS mRNA, as detected by reverse transcriptase-polymerase chain reaction. It is important to note that 50, 100, and 200 microM hydrogen peroxide was able to stimulate iNOS directly. Furthermore, 100 microM H2O2 enhanced synthesis of the oxidation products, nitrite (NO2-) and nitrate (NO3-) at 12 and 36 h. iNOS protein, detected by Western blot analysis, as well as L-citrulline levels, were also increased. When endothelial cell monolayers were incubated for 1 h with 100 microM H2O2 and subsequently with cytokines, iNOS mRNA was further augmented. Under the same conditions, we regularly observed an inhibition (25%) of intercellular adhesion molecule-1 (ICAM-1/CD54) expression. The latter was reversed when the NOS inhibitor N(G)-monomethyl-L-arginine was added, as shown by flow cytometry. These data suggest a specific effect of endogenous hydroperoxides on the biosynthesis and processing of the human endothelial iNOS isoform. We propose that H2O2 induces a temporary NO-dependent modulation of adhesion molecule expression to limit the tissue destruction that accompanies the vascular recruitment of leukocytes.

Fc-receptor-mediated intracellular delivery of Cu/Zn-superoxide dismutase (SOD1) protects against redox-induced apoptosis through a nitric oxide dependent mechanism.

BACKGROUND: Using specific antibodies against bovine Cu/Zn-superoxide dismutase (EC 1.15.1.1, SOD1) we demonstrated that anti-SOD antibodies (IgG1) are able to promote the intracellular translocation of the antioxidant enzyme. The transduction signalling mediated by IgG1 immune complexes are known to promote a concomitant production of superoxide and nitric oxide leading to the production of peroxynitrites and cell death by apoptosis. The Fc-mediated intracellular delivery of SOD1 thus limited the endogenous production of superoxide. It was thus of interest to confirm that in the absence of superoxide anion, the production of nitric oxide protected cells against apoptosis. Study in greater detail clearly stated that under superoxide anion-free conditions, nitric oxide promoted the cell antioxidant armature and thus protected cells against redox-induced apoptosis. MATERIALS AND METHODS: The murine macrophage cell-lines J774 A1 were preactivated or not with interferon-gamma and were then stimulated by IgG1 immune complexes (IC), free SOD1 or SOD1 IC and superoxide anion, nitric oxide, peroxynitrite, and tumor necrosis factor-alpha (TNF-alpha) production was evaluated. The redox consequences of these activation processes were also evaluated on mitochondrial respiration and apoptosis as well as on the controlled expression of the cellular antioxidant armature. RESULTS: We demonstrated that SOD1 IC induced a Fcgamma receptor (FcgammaR)-dependent intracellular delivery of the antioxidant enzyme in IFN-gamma activated murine macrophages (the J774 AI cell line). The concomitant stimulation of the FcyR and the translocation of the SOD1 in the cytoplasm of IFN-gamma-activated macrophages not only reduced the production of superoxide anion but also induced the expression of the inducible form of nitric oxide synthase (iNOS) and the related NO production. This inducing effect in the absence of superoxide anion production reduced mitochondrial damages and cell death by apoptosis and promoted the intracellular antioxidant armature. CONCLUSIONS: To define the pharmacologic mechanism of action of bovine SOD1, we attempted to identify the second messengers that are induced by SOD1 IC. In this work, we propose that Fc-mediated intracellular delivery of the SOD1 that reduced the production of superoxide anion and of peroxynitrite, promoted a NO-induced protective effect in inducing the antioxidant armature of the cells. Taken together, these data suggested that specific immune responses against antigenic SOD1 could promote the pharmacological properties of the antioxidant enzyme likely via a NO-dependent mechanism.

FcepsilonRII/CD23 is expressed in Parkinson's disease and induces, in vitro, production of nitric oxide and tumor necrosis factor-alpha in glial cells.

Oxidative stress is thought to be involved in the mechanism of nerve cell death in Parkinson's disease (PD). Among several toxic oxidative species, nitric oxide (NO) has been proposed as a key element on the basis of the increased density of glial cells expressing inducible nitric oxide synthase (iNOS) in the substantia nigra (SN) of patients with PD. However, the mechanism of iNOS induction in the CNS is poorly understood, especially under pathological conditions. Because cytokines and FcepsilonRII/CD23 antigen have been implicated in the induction of iNOS in the immune system, we investigated their role in glial cells in vitro and in the SN of patients with PD and matched control subjects. We show that, in vitro, interferon-gamma (IFN-gamma) together with interleukin-1beta (Il-1beta) and tumor necrosis factor-alpha (TNF-alpha) can induce the expression of CD23 in glial cells. Ligation of CD23 with specific antibodies resulted in the induction of iNOS and the subsequent release of NO. The activation of CD23 also led to an upregulation of TNF-alpha production, which was dependent on NO release. In the SN of PD patients, a significant increase in the density of glial cells expressing TNF-alpha, Il-1beta, and IFN-gamma was observed. Furthermore, although CD23 was not detectable in the SN of control subjects, it was found in both astroglial and microglial cells in parkinsonian patients. Altogether, these data demonstrate the existence of a cytokine/CD23-dependent activation pathway of iNOS and of proinflammatory mediators in glial cells and their involvement in the pathophysiology of PD.

The good old days: a look back at cosmetic surgery.

During the 25-year, history of the American Society of Plastic and Reconstructive Surgical Nurses (ASPRSN), cosmetic surgery procedures and nursing care have undergone constant change. Lessons learned over the past quarter-century will be discussed as we live and learn from our past experiences.

Regulation by endogenous INTERLEUKIN-10 of the expression of nitric oxide synthase induced after ligation of CD23 in human macrophages.

The possible role of interleukin 10 (IL-10) as an endogenous inhibitor of CD23-driven inducible nitric oxide synthase (iNOS) expression in human macrophages was investigated. Cross-linking of CD23 by a monoclonal antibody induced iNOS mRNA, as detected by RT-PCR, and the production of NO measured as the stable derivative, nitrite. A linear correlation was observed between CD23 expression and iNOS activity or NO2- production. The iNOS activity reached a maximum 48 h after ligation of CD23, then declined rapidly until 72 h. In parallel, nitrite production was detected after 24 h and reached a maximum after 48 h. In addition, ligation of the CD23 molecule induced, in a time-dependent manner, the production of IL-10. As this cytokine is known to regulate iNOS induction and activity, we evaluated the effect of a neutralizing mAb to IL-10 on CD23-induced iNOS activity and nitrite production by CD23-bearing macrophages and found that both were significantly enhanced. Furthermore, the addition of exogenous IL-10 suppressed CD23-driven iNOS mRNA expression, iNOS activity and production of nitrite. These data suggest that, after CD23-ligation at the cell surface of human phagocytes, the secretion of IL-10 downregulates the CD23-induced NO production at the transcriptional level, thus providing an efficient feed-back mechanism.

The induction of nitric oxide by interleukin-12 and tumor necrosis factor-alpha in human natural killer cells: relationship with the regulation of lytic activity.

We have investigated the interleukin-12 (IL-12) and tumor necrosis factor-alpha (TNFalpha)-induced regulation of human natural killer (NK) cell function and their relationship with nitric oxide (NO) generation. We demonstrate that both cytokines were efficient to trigger the transcription of the inducible nitric oxide synthase (iNOS) mRNA, as detected by reverse transcriptase-polymerase chain reaction (RT-PCR). Western blot analysis and intracytoplasmic fluorescence showed that iNOS protein was also induced by both cytokines. However, our data indicate that NO does not play a significant role in the effector phase of the cytotoxic activity mediated by NK-stimulated cells, inasmuch as the lytic activity was not affected in the presence of specific NO synthase inhibitors. When aminoguanidine (AMG), an inhibitor of iNOS, was added during the afferent phase of NK stimulation with IL-12 and TNFalpha, a subsequent increase in the lytic potential of the effector cells towards the NK-sensitive target cells (K562) and lymphokine-activated killer (LAK) target cells (Daudi) was observed. Conversely, the addition of chemical NO donors during the afferent step resulted in a dose-dependent inhibition of the NK and LAK cytotoxicity. Our data suggest that the enhancement of NK-cell cytotoxic activity resulting from iNOS inhibition may be correlated, at least in part, to an increase in interferon-gamma production and granzyme B expression.

B-cell chronic lymphocytic leukemia cells express a functional inducible nitric oxide synthase displaying anti-apoptotic activity.

The expression of different isoforms of nitric oxide synthase (NOS) was investigated in B-cell chronic lymphocytic leukemia (B-CLL) to delineate a possible role for nitric oxide (NO) in the control of apoptosis of the tumoral cells. By reverse transcription-polymerase chain reaction (RT-PCR), all B-CLL cells were found to express spontaneously inducible NOS (iNOS) mRNA, whereas endothelial constitutive NOS (ecNOS) mRNA was undetectable. The iNOS protein was detected by immunofluorescence in the cytoplasm of permeabilized leukemic cells and identified by Western blotting, using different anti-iNOS antibodies, as a protein of 135 kD in B-CLL cytoplasmic extracts. B-CLL cell lysates also displayed basal NOS enzymatic activity, as measured by the conversion of 14C-labeled L-arginine into 14C-L-citrulline. Ligation of CD23, expressed on the vast majority of B-CLL cells, resulted in increased iNOS expression and activity. The NO released exerted an anti-apoptotic effect on B-CLL cells that was counteracted by NOS inhibitors and engagement of the APO-1/Fas pathway. Therefore, the existence of a functional iNOS in B-CLL cells will provide further insights into the mechanisms that control proliferation and apoptosis in these tumor cells.

Nitric oxide production in human macrophagic cells phagocytizing opsonized zymosan: direct characterization by measurement of the luminol dependent chemiluminescence.

When differentiated into mature macrophages by the combination of all-trans retinoic acid and 1,25-dihydroxyvitamin D3, the human promonocytic cell lines U937 and THP-1 expressed inducible nitric oxide synthase (iNOS) transcripts. During their differentiation, the cells acquired the capacity to produce not only superoxide anion (O2.-) but also nitric oxide (.NO) in response to IgG (or IgE)-opsonized zymosan. The inhibitors of the iNOS pathway, aminoguanidine and NG-monomethyl-L-arginine (L-NMMA), suppressed the production of .NO and enhanced the steady-state concentration of O2.- determined. Conversely, superoxide dismutase (SOD) scavenged the O2.- released and increased the .NO-derived nitrite concentration detected. These data suggested a possible interaction between O2.- and .NO. In differentiated U937 (or THP-1) cells, IgG or IgE-opsonized zymosan induced a strong time-dependent luminol-dependent chemiluminescence (LDCL), which was abrogated by SOD and partially inhibited by aminoguanidine or L-NMMA. Since the iNOS inhibitors did not directly scavenge O2.-, LDCL determination in the presence or absence of SOD and/or iNOS inhibitors demonstrated a concomitant production of O2.- and .NO. These radicals induced the formation of a .NO-derived product(s), probably peroxynitrite (ONOO-), which was required to elicit maximal LDCL. Finally, LDCL measurement provided a convenient tool to characterize iNOS triggering and demonstrated an interaction between NADPH oxidase and iNOS products in human macrophagic cells phagocytizing opsonized-zymosan. These findings show that in activated macrophages, iNOS activity can be involved in LDCL and support the debated hypothesis of iNOS participation to the microbicidal activity of human macrophages.

Characterization of a constitutive type III nitric oxide synthase in human U937 monocytic cells: stimulation by soluble CD23.

The soluble cleavage fragment of the low-affinity immunoglobulin E (IgE) receptor/CD23 (sCD23 25000 MW) and antibodies directed against their receptors on monocytes, CD11b and CD11c, stimulate the production of nitric oxide (NO) by these cells and we have suggested that the enzyme involved could be related to the endothelial constitutive type III nitric oxide synthase (ecNOS). In the present work, we have analysed the characteristic properties of this NOS isoform in the model of the human promonocytic cells U937 By reverse-transcription polymerase chain reaction (RT-PCR), the presence of an mRNA coding for type III NOS was found in U937 cells and the corresponding protein was detected by immunofluorescence in permeabilized cells with a specific anti-ecNOS monoclonal antibody (mAb). Membrane extracts displayed a NOS activity dependent on the presence of calcium and calmodulin in the reaction medium and that was abrogated in the presence of EGTA. Recombinant soluble CD23 (25000 MW) was found to trigger an NO-dependent cGMP accumulation in these cells, which was abrogated by calcium chelators and inhibitors of the calcium/calmodulin complex. Moreover, sCD23 elicited a transient augmentation of intracytoplasmic free calcium concentration [Ca2+]i that was dependent on the presence of calcium in the external buffer and was prevented in the presence of EGTA, indicating that it was due to a calcium influx. In conclusion, human promonocytic cells such as U937 exhibit a functional type III NOS that can be stimulated by calcium-raising agents, such as sCD23.

Role of nitric oxide in the anti-tumoral effect of retinoic acid and 1,25-dihydroxyvitamin D3 on human promonocytic leukemic cells.

All trans retinoic acid and vitamin D3 derivatives are well known for their antileukemic activity, while the precise mechanism of this effect remains to be clarified. Using human leukemic U937 and THP-1 promonocytic cell lines, we analyzed the effect of all-trans retinoic acid (RA) and/or 1,25-dihydroxyvitamin D3 (VD) on the generation of nitric oxide (NO), a potent antitumoral mediator. U937 cell differentiation with VD or with both RA and VD (RA/VD) correlated with gene transcription and functional expression of inducible nitric oxide synthase (iNOS). Nitrites and L-citrulline were also detected in U937 cell supernatants as soon as 24 hours following cell incubation with VD or RA/VD, but not in cells treated with RA alone. Inhibition of iNOS activity by NG-monomethyl-L-arginine (LNMMA) significantly decreased in vitro U937 cell differentiation with VD and RA/VD as shown by the expression of cell differentiation markers (CD14 and CD68) and by the capacity of these cells to undergo a luminol-dependent chemiluminescence in response to opsonized zymosan. Similar results were obtained using the THP-1 cell line strengthening the role of NO in the VD- and RA/VD-induced growth arrest and terminal differentiation of promonocytic leukemia cells.

Triggering of CD23b antigen by anti-CD23 monoclonal antibodies induces interleukin-10 production by human macrophages.

The aim of this study was to evaluate the capacity of human macrophages to produce interleukin (IL)-10 upon stimulation of membrane CD23. An anti-CD23 monoclonal antibody (mAb) was found to elicit the expression of the specific mRNA for IL-10 in CD23-bearing macrophages, and to induce a time-dependent production of this cytokine with a maximal effect reached after 12 h. Inasmuch as we previously reported that CD23 ligation evoked the generation of nitric oxide and of cAMP, the effect of the Rp diastereoisomer of adenosine 3', 5'-cyclic phosphorothioate (Rp-cAMP, an inhibitor of the cAMP pathway) and of NG-monomethyl-L-arginine (L-NMMA, an inhibitor of the nitric oxide pathway) were evaluated on CD23-induced IL-10 production. In the presence of Rp-cAMP, the CD23-induced production of IL-10 and of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) was totally abrogated, whereas, in the presence of L-NMMA, IL-10 production was enhanced and TNF-alpha production was suppressed. In addition, neutralization of IL-10 with an anti-IL-10 mAb increased both the magnitude and duration of CD23-driven TNF-alpha production. Such an inducing effect was observed with different anti-CD23 mAb (clone 135, MHM6 and 25), indicating that the triggering of the CD23 molecule at the surface of human macrophages induced the generation of IL-10 through a cAMP-dependent mechanism. Concomitantly this generation of IL-10 was down-regulated by nitric oxide, which was also produced after triggering of the CD23 antigen. Taken together these data indicated that human macrophages produced IL-10 after triggering of the CD23 molecule and that this production could regulate the inflammatory state of these cells.

Canine visceral leishmaniasis: successful chemotherapy induces macrophage antileishmanial activity via the L-arginine nitric oxide pathway.

Following successful chemotherapy in canine visceral leishmaniasis, monocyte-derived macrophages can induce antileishmanial activity via a gamma interferon-dependent mechanism in the presence of autologous lymphocytes. The killing of leishmania correlated with the induction of the NO synthase pathway, because it correlated with the generation of nitrogen derivative production and was abrogated in the presence of NG-monomethyl-L-arginine, a competitive inhibitor of the NO synthase pathway. The level of L-citrulline in serum, which was produced after activation of the NO synthase pathway, was markedly enhanced in dogs receiving successful chemotherapy. Taken together, these data indicate that following successful chemotherapy of visceral leishmaniasis, leishmania parasites are killed by macrophages activated by gamma interferon-producing lymphocytes via an NO-dependent mechanism.

Role of IgE immune complexes in the regulation of HIV-1 replication and increased cell death of infected U1 monocytes: involvement of CD23/Fc epsilon RII-mediated nitric oxide and cyclic AMP pathways.

BACKGROUND: IgE/anti-IgE immune complexes (IgE-IC) induce the release of multiple mediators from monocytes/macrophages and the monocytic cell line U937 following the ligation of the low-affinity Fc epsilon receptors (Fc epsilon RII/CD23). These effects are mediated through an accumulation of cAMP and the generation of L-arginine-dependent nitric oxide (NO). Since high IgE levels predict more rapid progression to acquired immunodeficiency syndrome, we attempted to define the effects of IgE-IC on human immunodeficiency virus (HIV) production in monocytes. MATERIALS AND METHODS: Two variants of HIV-1 chronically infected monocytic U1 cells were stimulated with IgE-IC and virus replication was quantified. NO and cAMP involvement was tested through the use of agonistic and antagonistic chemicals of these two pathways. RESULTS: IgE-IC induced p24 production by U1 cells with low-level constitutive expression of HIV-1 mRNAs and extracellular HIV capsid protein p24 levels (U1low), upon their pretreatment with interleukin 4 (IL-4) or IL-13. This effect was due to the crosslinking of CD23, as it was reversed by blocking the IgE binding site on CD23. The IgE-IC effect could also be mimicked by crosslinking of CD23 by a specific monoclonal antibody. p24 induction by IgE-IC was then shown to be due to CD23-mediated stimulation of cAMP, NO, and tumor necrosis factor alpha (TNF alpha) generation. In another variant of U1 cells with > 1 log higher constitutive production of p24 levels (U1high), IgE-IC addition dramatically decreased all cell functions tested and accelerated cell death. This phenomenon was reversed by blocking the nitric oxide generation. CONCLUSIONS: These data point out a regulatory role of IgE-IC on HIV-1 production in monocytic cells, through CD23-mediated stimulation of cAMP and NO pathways. IgE-IC can also stimulate increased cell death in high HIV producing cells through the NO pathway.