Short-term creep approach to redefining the role of 17-4PH stainless steel for high-temperature applications.

The creep response of the 17-4PH martensitic age-hardening steel in H1150 state was investigated at 427 and 482 °C. Hardness measurements of the heads of the creep samples demonstrated that the material underwent additional age hardening during the high-temperature exposure. Microstructural investigations confirmed that the additional precipitation of carbides and the G-phase occurred at the lowest temperature. A set of constitutive equations previously developed to describe the creep response of particle-strengthened alloys was successfully used to obtain a comprehensive description of the experimental data. The value of the particle strengthening term was obtained from the hardness measurements and corresponded to the Orowan stress. The model accurately described the observed minimum creep rate dependence on the applied stress and explained the occurrence of lower values of the minimum strain rate observed during variable-load experiments.

Real-life experience of edoxaban treatment for venous thromboembolism (VTE)/pulmonary embolism (PE) in patients with isolated positive Lupus Anticoagulant (LAC) during the COVID-19 pandemic lockdown in Italy.

OBJECTIVE: Direct-acting oral anticoagulants (DOACs) have established indications, according to recent guidelines for the treatment and prevention of venous thromboembolism (VTE), including pulmonary embolism (PE), with a safer profile compared to vitamin K antagonist (VKA) in terms of a lower risk for major bleeding and no need of blood coagulation tests. However, DOACs are not indicated in the treatment of patients with triple-positive antiphospholipid syndrome (APS). This limitation is often extended in clinical practice to patients with isolated positivity. The COVID-19 pandemic has sometimes made it difficult to maintain a safe VKA treatment, due to the practical difficulties of performing INR. PATIENTS AND METHODS: We evaluated 39 patients with a previous unprovoked VTE/PE, who were no longer eligible for VKA treatment due to the difficulty of performing INR during the COVID-19 pandemic lockdown, in Italy. All patients had a positive LAC and refused a long-term anticoagulation with low molecular weight heparin. They were shifted to edoxaban. RESULTS: Any recurrence of VTE/PE occurred during the observation period (up to eight months of treatment), while only one minor bleeding event was recorded (Hazard ratio=0.06, 95% confidence interval 0.03-0.11, p=0.094). No arterial events occurred during the observation period. Hemoglobin, platelets, and creatinine were unchanged during the observation period. CONCLUSIONS: Edoxaban treatment may be safe and effective in preventing the recurrence of VTE/PE in patients with isolated LAC positivity, without the occurrence of arterial events.

Fine tuning of the DNAM-1/TIGIT/ligand axis in mucosal T cells and its dysregulation in pediatric inflammatory bowel diseases (IBD).

De-regulated T-cell activation and functions are pivotal in the orchestration of immune-mediated tissue damage in IBD. We investigated the role of DNAM-1 (co-activating)/TIGIT (co-inhibitory)/ligand axis in the regulation of T-cell functions and its involvement in IBD pathogenesis. We show that DNAM-1 and TIGIT display a peculiar expression pattern on gut mucosa T-cell populations, in a microenvironment where their shared ligands (PVR and Nectin-2) are physiologically present. Moreover, DNAM-1 family receptor/ligand system is perturbed in IBD lesions, in a disease activity-dependent manner. The expression profile of CCR6 and CD103 mucosa addressins suggests that microenvironment-associated factors, rather than skewed recruitment of circulating T-cell populations, play a more relevant role in supporting the establishment of DNAM-1 and TIGIT expression pattern in mucosal T-cell populations, and may explain its alteration in IBD. Although both co-receptors mark functionally competent T cells, DNAM-1 and TIGIT segregate on T cells endowed with different proliferative potential. Moreover, their opposing role in regulating T-cell proliferation exquisitely depends on ligand availability. All together, our data propose a role for DNAM-1 and TIGIT in regulating mucosal T-cell activation and immune homeostasis, and highlight the involvement of an imbalance of this system in IBD.

Differential microRNA expression between decidual and peripheral blood natural killer cells in early pregnancy.

STUDY QUESTION: Have decidual natural killer (dNK) cells a different microRNA (miRNA or miR) expression pattern compared to NK cells circulating in the peripheral blood (pb) of healthy pregnant women in the first trimester of gestation? SUMMARY ANSWER: dNK cells have a unique miRNA profile, showing exclusive expression of a set of miRNAs and significant up- or down-regulation of most of the miRNAs shared with pbNK cells. WHAT IS KNOWN ALREADY: dNK cells differ from pbNK cells both phenotypically and functionally, and their origin is still debated. Many studies have indicated that miRNAs regulate several important aspects of NK cell biology, such as development, activation and effector functions. STUDY DESIGN, SIZE, DURATION: Decidua basalis and peripheral blood specimens were collected from women (n = 7) undergoing voluntary termination of gestation in the first trimester of pregnancy. dNK and pbNK cells were then highly purified by cell sorting. PARTICIPANTS/MATERIALS, SETTING, METHODS: miRNAs expression was analysed by quantitative RT-PCR (qRT-PCR)-based arrays using RNA purified from freshly isolated and highly purified pbNK and dNK cells. Results from arrays were validated by qRT-PCR assays. The bioinformatics tool ingenuity pathway analysis (IPA) was applied to determine the cellular network targeted by validated miRNAs and the correlated biological functions. MAIN RESULTS AND THE ROLE OF CHANCE: Herein, we identified the most differentially expressed miRNAs in NK cells isolated from peripheral blood and uterine decidua of pregnant women. We found that 36 miRNAs were expressed only in dNK cells and two miRNAs only in pbNK cells. Moreover, 48 miRNAs were commonly expressed by both NK cell preparations although at different levels: 28 were upregulated in dNK cells, while 15 were downregulated compared to pbNK cells. Validation of a selected set (n = 11) of these miRNAs confirmed the differential expression of nine miRNAs: miR-10b and miR-214 expressed only in dNK cells and miR-200a-3p expressed only in pbNK cells; miR-130b-3p, miR-125a-5p, miR-212-3p and miR-454 were upregulated while miR-210-3p and miR-132 were downregulated in dNK cells compared to pbNK cells. IPA network analysis identified a single network connecting all the miRNAs as well as their significant involvement in several classes of functions: 'Organismal injury, Reproductive system disease, Inflammatory disease' and 'Cellular development'. These miRNAs target molecules such as argonaute 2, tumour protein p53, insulin and other genes that belong to the same network and significantly influence cell differentiation and pregnancy. LIMITATIONS, REASONS FOR CAUTION: In the present study, the cellular network and biological functions modulated by miRNAs differentially expressed in dNK and pbNK cells were identified by IPA considering only molecules and relationships that were with confidence 'experimentally observed' in leucocytes. The decidual and pbNK cells that were analysed here are a heterogeneous population and further study will help to disentangle whether there are differences in miRNA production by the different subsets of NK cells. WIDER IMPLICATIONS OF THE FINDINGS: This is the first study describing a different miRNA expression profile in dNK cells compared to matched pbNK cells during the first trimester of pregnancy. Our findings improved the body of knowledge on dNK cell biology and strongly suggest further investigation into the roles of miRNAs that are differentially expressed in human dNK compared to pbNK cells. Our results suggest that specific miRNAs can modulate dNK cell origin and functions, highlighting a potential role of this miRNA signature in human development and diseases. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by grants from the Istituto Pasteur, Fondazione Cenci Bolognetti, the European NoE EMBIC within FP6 (Contract number LSHN-CT-2004-512040), Istituto Italiano di Tecnologia, and Ministero dell'Istruzione, dell'Università e della Ricerca (Ricerche Universitarie), and from Università Politecnica delle Marche. There are no conflicts of interest to declare.

Utility of a stool antigen test to detect the incidence of helicobacter pylori infection and familial and community enviromental risk factors for this infection in pediatric age.

BACKGROUND: Helicobacter pylori (Hp) infection is mainly acquired during childhood; it is recognised as a cause of gastritis and peptic ulcer and it has been classified as a group A carcinogen by World Health Organization. The exact mode of transmission is as yet, not known. Aim of our study has been to identify risk factors associated with Helicobacter pylori infection in a preschool and school population and to confirm if Hp antigen in faeces is useful as screening in epidemiological studies. METHODS: We interviewed, with questionnaire, 400 children (203 male; age range 3-10 years; mean age 6 years) of 3 different schools and stool samples were collected of all children too. 35 of 400 (8%) children underwent to upper gastrointestinal endoscopy because of a suspect of upper gastrointestinal disease. RESULTS: stool were collected from 400 school children and 35 of them shown positivity of Hp antigen test. A questionnaire about presence of nausea, vomit, recurrent abdominal pain, family size, parent's occupations and education, use of antibiotics, country of birth of child and parents, personal hygiene, breast feeding, presence of the animals was completed. 35 children with positive Hp stool antigen test and a suspicious of upper gastrointestinal disease (recurrent abdominal pain, diurnal or nocturnal abdominal pain, nausea, vomiting, iron deficiency) underwent to esophagogastroduodenoscopy (EGDS) that demonstrated antral gastritis and positive histology and urease rapid test. CONCLUSIONS: the results of this study suggest that risk factors for Hp infection are low socioeconomics factors, hygiene and living conditions and that Hp antigen in faeces is useful as screening test.

Size and dynamics of mucosal and peripheral IL-17A+ T-cell pools in pediatric age, and their disturbance in celiac disease.

Mucosal interleukin (IL)-17A-producing T cells contribute to protective antimicrobial responses and to epithelial barrier integrity; their role in celiac disease (CD) is debated. We analyzed the frequency and developmental dynamics of mucosal (intraepithelial lymphocytes (IEL)) and circulating (peripheral blood (PB)) IL-17A (T17) and/or interferon (IFN)-γ-producing (T1, T1/T17) T-cell populations in 86 pediatric controls and 116 age-matched CD patients upon phorbol myristate acetate/ionomycin or CD3/CD28 stimulation. T17 and T1/17 are physiologically present among IEL and PB populations, and their frequency is selectively and significantly reduced in CD IEL. The physiological age-dependent increase of Th17 IEL is also absent in CD, while IFN-γ-producing PB-T cells significantly accumulate with patient's age. Finally, the amplitude of IL-17A+ and IFN-γ+ T-cell pools are significantly correlated in different individuals; this relationship only applies to CD4+ T cells in controls, while it involves also the CD4- counterpart in CD patients. In conclusion, both size and dynamics of mucosa-associated and circulating IL-17A+ T-cell pools are finely regulated in human pediatric subjects, and severely disturbed in CD. The impaired IL-17A+ IEL-T pool may negatively impact on epithelial barrier efficiency, and contribute to CD mucosa damage; the disturbed dynamics of circulating IL-17A+ and IFN-γ+ T-cell pools may be involved in the extraintestinal autoimmune manifestations associated with CD.

A comprehensive overview of grain development in Brachypodium distachyon variety Bd21.

A detailed and comprehensive understanding of seed reserve accumulation is of great importance for agriculture and crop improvement strategies. This work is part of a research programme aimed at using Brachypodium distachyon as a model plant for cereal grain development and filling. The focus was on the Bd21-3 accession, gathering morphological, cytological, and biochemical data, including protein, lipid, sugars, starch, and cell-wall analyses during grain development. This study highlighted the existence of three main developmental phases in Brachypodium caryopsis and provided an extensive description of Brachypodium grain development. In the first phase, namely morphogenesis, the embryo developed rapidly reaching its final morphology about 18 d after fertilization (DAF). Over the same period the endosperm enlarged, finally to occupy 80% of the grain volume. During the maturation phase, carbohydrates were continuously stored, mainly in the endosperm, switching from sucrose to starch accumulation. Large quantities of ß-glucans accumulated in the endosperm with local variations in the deposition pattern. Interestingly, new ß-glucans were found in Brachypodium compared with other cereals. Proteins (i.e. globulins and prolamins) were found in large quantities from 15 DAF onwards. These proteins were stored in two different sub-cellular structures which are also found in rice, but are unusual for the Pooideae. During the late stage of development, the grain desiccated while the dry matter remained fairly constant. Brachypodium exhibits some significant differences with domesticated cereals. Beta-glucan accumulates during grain development and this cell wall polysaccharide is the main storage carbohydrate at the expense of starch.

Loss of pericentromeric DNA methylation pattern in human glioblastoma is associated with altered DNA methyltransferases expression and involves the stem cell compartment.

Cancer is generally characterized by loss of CG dinucleotides methylation resulting in a global hypomethylation and the consequent genomic instability. The major contribution to the general decreased methylation levels seems to be due to demethylation of heterochromatin repetitive DNA sequences. In human immunodeficiency, centromeric instability and facial anomalies syndrome, demethylation of pericentromeric satellite 2 DNA sequences has been correlated to functional mutations of the de novo DNA methyltransferase 3b (DNMT3b), but the mechanism responsible for the hypomethylated status in tumors is poorly known. Here, we report that human glioblastoma is affected by strong hypomethylation of satellite 2 pericentromeric sequences that involves the stem cell compartment. Concomitantly with the integrity of the DNMTs coding sequences, we report aberrations in DNA methyltrasferases expression showing upregulation of the DNA methyltransferase 1 (DNMT1) and downregulation of the de novo DNA methyltransferase 3a (DNMT3a). Moreover, we show that DNMT3a is the major de novo methyltransferase expressed in normal neural progenitor cells (NPCs) and its forced re-expression is sufficient to partially recover the methylation levels of satellite 2 repeats in glioblastoma cell lines. Thus, we speculate that DNMT3a decreased expression may be involved in the early post-natal inheritance of an epigenetically altered NPC population that could be responsible for glioblastoma development later in adult life.

Signaling pathways activated by chemokine receptor CXCR2 and AMPA-type glutamate receptors and involvement in granule cells survival.

We show that treatment of cerebellar granules with interleukin-8 (IL-8), growth-related gene product beta (GRObeta) or AMPA induced activation of PI3-K/Akt and of ERK pathways, the latter being independent of PI3-K and dependent on PTX-sensitive G proteins. We also show that AMPA-mediated neuron survival was abolished both by ERK kinase inhibitor PD98059 and AMPA-Rs blocker CNQX, and that chemokine-mediated survival was blocked by the PI3-K inhibitors LY294002 and wortmannin. We conclude that the neurotrophic effects of AMPA need the contemporary activation of ERKs and stimulation of AMPA-Rs, and that PI3-K/Akt activation is a determinant pathway for the IL-8/GRObeta anti-apoptotic activity.

Ubiquitination and degradation of Syk and ZAP-70 protein tyrosine kinases in human NK cells upon CD16 engagement.

Syk and ZAP-70 nonreceptor protein tyrosine kinases (PTKs) are essential elements in several cascades coupling immune receptors to intracellular responses. The critical role of these kinases in promoting the propagation of intracellular signaling requires a tight regulation of their activity, thus the existence of a negative feedback loop regulating their expression can be hypothesized. Herein, we have investigated whether ubiquitin-dependent proteolysis could be a mechanism responsible for controlling the fate of Syk and ZAP-70 after their immunoreceptor-induced activation. We found that both Syk and ZAP-70 become ubiquitinated in response to aggregation of the low affinity Fc receptor for IgG (CD16) on human natural killer cells. We confirmed the identity of the major in vivo ubiquitinated kinase species by performing an in vitro ubiquitination assay. In addition, we found that after CD16 stimulation, ubiquitinated forms of Syk and ZAP-70 associate with the receptor complex. After CD16 engagement, we also observed a decrease in the stability of Syk and ZAP-70 PTKs that is counteracted by pretreatment with either proteasome or lysosomal inhibitors. Moreover, in the presence of the proteasome inhibitor, epoxomicin, we observed an accumulation of ubiquitinated forms of both kinases. Our findings provide evidence of ligand-induced ubiquitination of nonreceptor PTKs belonging to the Syk family and propose the ubiquitin-dependent proteasome-mediated degradation pathway as a mechanism for attenuating the propagation of intracellular signaling initiated by immune receptor engagement.

Natural killer cells and nitric oxide.

Natural killer (NK) cells and nitric oxide (NO) are both important components of the natural or innate immune response. NK cells are large granular lymphocytes capable of destroying cells infected by virus or bacteria and susceptible tumor cells without prior sensitization and restriction by MHC antigens. They are abundant in blood, spleen, liver and lungs and are distinct from both T and B lymphocytes in their circulation patterns, profile of surface antigens, receptor repertoire and the way in which they discriminate between self and non-self. Uniquely, NK cells express receptors that can recognize and discriminate between normal and altered MHC class I determinants. NK cell cytotoxic activity is strongly induced by cytokines such as IL-2 and IL-12, and this activation is associated with synthesis of NO. Inhibitors of NO synthesis impair NK cell-mediated target cell killing, demonstrating a role for NO in NK cell function. Furthermore, NO itself can regulate NK cell activation. In this article, evidence that NO is a mediator of NK cell-mediated target cell killing, and that NO is a regulator of NK cell activation will be reviewed. Results of NO synthase gene deletion studies will be discussed, and rodent and human NK cells will be compared.

The adaptor protein shc is involved in the negative regulation of NK cell-mediated cytotoxicity.

The activation of protein tyrosine kinase(s) (PTK) is a critical event required for the development of NK cell-mediated cytotoxicity. Here we demonstrate that the adaptor protein shc undergoes tyrosine phosphorylation during the generation of antibody-dependent cellular cytotoxicity (ADCC) and natural killing. In addition, we report that, upon direct or antibody-dependent target cell interaction, shc coprecipitates with the Src homology 2 (SH2)-containing inositol phosphatase, SHIP. To gain information on the functional role of shc in NK cytotoxicity, we overexpressed wild-type or dominant negative shc constructs in the human NKL cell line. Our findings show a consistent shc-mediated down-regulation of ADCC and natural killing. Such functional effect correlates with a perturbation of the phosphoinositide (PI) metabolism by means of a shc-mediated negative regulation of inositol 1,4,5 triphosphate (IP3) generation and intracellular calcium flux upon CD16 ligation. Furthermore, our data show that dominant-negative shc-mediated perturbation of shc/SHIP interaction leads to inhibition of ligand-dependent SHIP recruitment to CD16 zeta chain. We suggest that shc plays a role of negative adaptor by modulating SHIP recruitment to activation receptors involved in the generation of NK cytotoxic function.

Polyclonal antibodies against gp185HER2 peptides: their putative role in the identification of a particular HER2 status in patients with breast cancer.

The HER2 oncogene and its relative oncoprotein, gp185HER2, a transmembrane glycoprotein belonging to the epidermal growth factor receptor family, are overexpressed in a wide range of solid tumors including breast and ovarian cancer. In patients with breast cancer, both humoral and cell-mediated HER2 immune responses have been found as well as in some patients with gp185HER2 nonoverexpressing tumors. To establish whether peptide sequences identified as HLA-A2-restricted T-cell epitopes are expressed in breast tumor cell lines and tissues, we produced and characterized by different methodologic approaches polyclonal antibodies raised against four gp185HER2 peptides. Two of the antibodies recognized peptides eluted from the HLA-A2 groove of the mDAmB231 breast cancer cell line expressing a basal level of gp185HER2. Paraffin-embedded primary and metastatic breast tumors were specifically immunostained by all four reagents, thereby showing an overlapping reactivity. When this immunoreactivity was compared with that obtained using two different monoclonal antibodies, in 105 breast primary tumors and 36 corresponding lymph node metastases, we identified a subset of tumors that were negative with anti-gp185HER2 monoclonal antibodies and positive with the four antipeptide antibodies. Our novel observations provide in vivo evidence of the complexity involved in evaluating HER2 expression, and open a new path for understanding the biologic significance of HER2 status in breast tumors.

I-309 binds to and activates endothelial cell functions and acts as an angiogenic molecule in vivo.

Several chemokines have been shown to act as angiogenic molecules or to modulate the activity of growth factors such as fibroblast growth factor 2 (FGF-2) and vascular endothelial growth factor (VEGF). The detection of the CC chemokine receptor (CCR) 8 message in human umbilical vein endothelial cells (HUVECs) by reverse transcription- polymerase chain reaction (RT-PCR) and RNase protection assay (RPA), prompted us to investigate the potential role exerted by the CC chemokine I-309, a known ligand of such receptor, in both in vitro and in vivo angiogenesis assays. We show here that I-309 binds to endothelial cells, stimulates chemotaxis and invasion of these cells, and enhances HUVEC differentiation into capillary-like structures in an in vitro Matrigel assay. Furthermore, I-309 is an inducer of angiogenesis in vivo in both the rabbit cornea and the chick chorioallantoic membrane assay (CAM).

SDF-1alpha-mediated modulation of synaptic transmission in rat cerebellum.

The functional expression of the seven-transmembrane domain G protein-coupled chemokine receptor CXCR-4/fusin in rat nerve cell was demonstrated by staining with a polyclonal anti-CXCR-4 Ab, and by evaluating the calcium responses to the physiological agonist stromal-derived cell factor-1alpha (SDF-1alpha) in both cerebellar granule cells in culture and Purkinje neurons (PNs) in cerebellar slices. Cerebellar glial, granule and Purkinje cells showed a pronounced staining for CXCR-4. Furthermore, cultured granule cells exhibited Ca2+ transients elicited by the application of SDF-1alpha, both in cell bodies and in neuronal processes. Whole-cell patch-clamped PNs in cerebellar slices responded to SDF-1alpha application by a slow inward current followed by an increase of both intracellular Ca2+ level and spontaneous synaptic activity. In particular, the SDF-1alpha-induced slow inward current was considerably reduced by ionotropic glutamate receptor blockers, but developed fully in a medium in which synaptic transmission was inhibited, indicating that this current might be, at least in part, mediated by extrasynaptic glutamate, possibly released from the surrounding glial and/or nerve cells. Taken together, these findings indicate a functional involvement of CXCR-4 in the modulation of synaptic transmission, adding another member to the repertoire of the chemokine receptors exerting a neuromodulatory role in the cerebellum.

The chemokine growth-related gene product beta protects rat cerebellar granule cells from apoptotic cell death through alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptors.

Cultured cerebellar granule neurons are widely used as a cellular model to study mechanisms of neuronal cell death because they undergo programmed cell death when switched from a culture medium containing 25 mM to one containing 5 mM K(+). We have found that the growth-related gene product beta (GRObeta) partially prevents the K(+)-depletion-induced cell death, and that the neuroprotective action of GRObeta on granule cells is mediated through the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) type of ionotropic glutamate receptors. GRObeta-induced survival was suppressed by 6-cyano-7-nitroquinoxaline-2,3-dione, which is a specific antagonist of AMPA/kainate receptors; it was not affected by the inhibitor of N-methyl-D-aspartate receptors, 2-amino-5-phosphonopentanoic acid, and was comparable to the survival of granule cells induced by AMPA (10 microM) treatment. Moreover, GRObeta-induced neuroprotection was abolished when granule cells were treated with antisense oligonucleotides specific for the AMPA receptor subunits, which significantly reduced receptor expression, as verified by Western blot analysis with subunit-specific antibodies and by granule cell electrophysiological sensitivity to AMPA. Our data demonstrate that GRObeta is neurotrophic for cerebellar granule cells, and that this activity depends on AMPA receptors.

bcl-2 over-expression enhances NF-kappaB activity and induces mmp-9 transcription in human MCF7(ADR) breast-cancer cells.

bcl-2 expression is often associated with poor prognosis in several types of tumors; however, the role of this molecule in breast cancer is still controversial. We found earlier that over-expression of bcl-2 in a human breast-cancer cell line (MCF7(ADR)) enhances its tumorigenicity and metastatic potential by inducing metastasis-associated properties such as increased secretion of the matrix metalloproteinase-9 (mmp-9). In the present study, we investigated the effect of bcl-2 over-expression on the activity of the transcription factor NF-kappaB, an important regulator of genes involved in tumor progression and invasion. Transient transfection experiments indicate that over-expression of bcl-2 in the MCF7(ADR) cell line, enhances NF-kappaB-dependent transcriptional activity. Mobility-shift analysis revealed an increase of NF-kappaB DNA-binding in bcl-2-over-expressing clones that correlated with lower levels of the NF-kappaB cytoplasmic inhibitor IkappaBalpha. Moreover, point mutations of 2 highly conserved residues within the BH1 and BH2 domains that abrogate the interaction of bcl-2 with bax, or deletion of the N-terminal BH4 domain, completely eliminate the ability of this molecule to up-regulate NF-kappaB-dependent transactivation. Since mmp-9 is a NF-kappaB-regulated gene, we also investigated whether bcl-2 over-expression up-regulated mmp-9 transcription. We found that induction of mmp-9 mRNA correlates with the activation of an mmp-9-promoter-reporter-gene construct in transient transfection assay, and a mutation of the (-600)mmp-9-NF-kappaB binding element abolishes this effect. The overall data indicate that bcl-2-mediated regulation of NF-kappaB-transcription-factor activity may represent an important mechanism for the promotion of malignant behavior in MCF-7(ADR) cells.

Dichotomic effects of IFN-gamma on the development of systemic lupus erythematosus-like syndrome in MRL-lpr / lpr mice.

Systemic lupus erythematosus (SLE)-prone female MRL-lpr / lpr (MRL-lpr) mice were treated with mouse or rat IFN-gamma under different experimental conditions, both prophylactically in 6- to 8 week-old animals and therapeutically in 12- to 18-week-old SLE-affected mice. It was found that IFN-gamma heterogeneously modulated the course of the disease in MRL-lpr mice. When administered prophylactically, IFN-gamma favorably modulated the histological, serological and clinical signs of the disease. Relative to untreated or PBS-treated control animals, the MRL-lpr mice which received IFN gamma were virtually free of inflammatory infiltration of the kidneys and the lungs, had lower levels of azotemia with reduction of both circulating IgG1, IgG2a and IgG3 and anti-double strand (ds) and single strand (ss) DNA antibodies, milder skin vasculitis, significantly reduced enlargement of their lymph nodes and lower weight of the spleens. IFN-gamma also lowered the rate of mortality of MRL-lpr mice. In contrast to these findings, therapeutically administered IFN-gamma worsened the course of the disease in MRL-lpr mice, which exhibited increased proteinuria, higher levels of IgG2a and IgG3 and anti-ds and -ss DNA antibodies, more aggressive nephritis and died at an earlier age than PBS-treated control mice. The dichotomic effect of IFN-gamma on disease manifestation in MRL-lpr mice offers new insights into the complex role of this cytokine in the regulation of systemic autoimmunity such as SLE.

CD69-triggered ERK activation and functions are negatively regulated by CD94 / NKG2-A inhibitory receptor.

CD69 represents a functional triggering molecule on activated NK and T cells, capable of inducing cytotoxic activity and costimulating cytokine production. It belongs to the C-lectin type superfamily, and its gene maps in the NK gene complex, close to other genes coding for NK receptors. CD94 / NKG2-A complex is the inhibitory receptor for the non classical MHC class I molecule HLA-E on human NK cells. To investigate CD69-initiated signal transduction pathways, and to evaluate CD94 / NKG2-A interference on CD69 triggering ability, we have generated transfectants expressing both receptors in the RBL cell line. Here we report that CD69 engagement leads to the activation of extracellular signal-regulated kinase (ERK) enzymes belonging to the MAPK family, and that this event is required for CD69-mediated cell degranulation. Moreover, we show that the co-engagement of CD94 / NKG2-A inhibitory receptor effectively suppresses both CD69-triggered cell degranulation in RBL transfectants, through the inhibition of ERK activation, and CD69-induced cytotoxicity in human NK cells. Thus, here we provide new information on the molecular mechanisms initiated by CD69 activation receptor, and show that CD69-initiated signaling pathways and functional activity are negatively regulated by CD94 / NKG2-A inhibitory complex.