Prospective memory: the combined impact of cognitive load and task focality.

Prospective Memory (PM) entails a set of executive processes primarily associated with the activation of frontal and parietal regions. Both the number of PM-targets to be monitored (i.e. task load) and the relationship between the type of PM-targets and the ongoing (ONG) task (i.e. task focality) can impact executive monitoring and PM performance. In the present imaging study, we manipulated load and focality of an event-based PM task to test the hypothesis that common resources engage in situations requiring high levels of cognitive control: that is, in high-load (i.e. monitor multiple PM-targets) and non-focal conditions (i.e. monitor at the same time letters' identity and color). We investigated monitoring-related and detection-related processes by assessing behavior and brain activity separately for ONG trials (monitoring) and PM-targets (detection). At the behavioral level, we found a significant interaction between load and focality during detection, with slowest reaction times for focal, high-load PM-targets. The imaging analyses of the detection phase revealed the activation of the left intraparietal sulcus in the high-load conditions. Both in the monitoring and the detection phases, we found overlapping effects of non-focality and low-load in the fusiform gyrus. Our results suggest that under low-load conditions, cognitive control operates via early selection mechanisms in the ventral occipito-temporal cortex. By contrast, high-load conditions entail control at later processing stages within the dorsal parietal cortex. We conclude that load and focality operate via different mechanisms, with the level of task load largely determining how cognitive control selects the most relevant information.

Speaking with mask in the COVID-19 era: Multiclass machine learning classification of acoustic and perceptual parameters.

The intensive use of personal protective equipment often requires increasing voice intensity, with possible development of voice disorders. This paper exploits machine learning approaches to investigate the impact of different types of masks on sustained vowels /a/, /i/, and /u/ and the sequence /a'jw/ inside a standardized sentence. Both objective acoustical parameters and subjective ratings were used for statistical analysis, multiple comparisons, and in multivariate machine learning classification experiments. Significant differences were found between mask+shield configuration and no-mask and between mask and mask+shield conditions. Power spectral density decreases with statistical significance above 1.5 kHz when wearing masks. Subjective ratings confirmed increasing discomfort from no-mask condition to protective masks and shield. Machine learning techniques proved that masks alter voice production: in a multiclass experiment, random forest (RF) models were able to distinguish amongst seven masks conditions with up to 94% validation accuracy, separating masked from unmasked conditions with up to 100% validation accuracy and detecting the shield presence with up to 86% validation accuracy. Moreover, an RF classifier allowed distinguishing male from female subject in masked conditions with 100% validation accuracy. Combining acoustic and perceptual analysis represents a robust approach to characterize masks configurations and quantify the corresponding level of discomfort.

Treatment outcomes in patients with laryngotracheal lesions and compromised airway during and prior to the coronovirus disease 2019 pandemic: a tertiary institution's experience.

OBJECTIVE: This study aimed to compare treatment outcomes in patients with laryngeal and tracheal stenosis treated during and prior to the coronavirus disease 2019 pandemic period. METHOD: Patients treated for laryngotracheal lesions with impending airway compromise during the active pandemic period were matched with those treated for similar lesions in the preceding years in a monocentric tertiary hospital setting. RESULTS: During the pandemic period of 55 days, 31 patients underwent 47 procedures. Seven patients (2 children, 5 adults) had open airway surgery, and one had an operation-specific complication. Twenty-four patients (10 children, 14 adults) underwent 40 endoscopic interventions without any complications. Operation specific results during and prior to the pandemic were comparable. CONCLUSION: The management strategy in patients with laryngotracheal lesions and impending airway compromise should not be altered during periods of risk from coronavirus disease 2019. Avoiding a tracheostomy by performing primary corrective surgery or proceeding with a definitive decannulation would be beneficial in these patients to reduce the risk of contagion.

Smartphones Offer New Opportunities in Clinical Voice Research.

Smartphone technology provides new opportunities for recording standardized voice samples of patients and sending the files by e-mail to the voice laboratory. This drastically improves the collection of baseline data, as used in research on efficiency of voice treatments. However, the basic requirement is the suitability of smartphones for recording and digitizing pathologic voices (mainly characterized by period perturbations and noise) without significant distortion. In this experiment, two smartphones (a very inexpensive one and a high-level one) were tested and compared with direct microphone recordings in a soundproof room. The voice stimuli consisted in synthesized deviant voice samples (median of fundamental frequency: 120 and 200 Hz) with three levels of jitter and three levels of added noise. All voice samples were analyzed using PRAAT software. The results show high correlations between jitter, shimmer, and noise-to-harmonics ratio measured on the recordings via both smartphones, the microphone, and measured directly on the sound files from the synthesizer. Smartphones thus appear adequate for reliable recording and digitizing of pathologic voices.

Ischemic tolerance modulates TRAIL expression and its receptors and generates a neuroprotected phenotype.

TNF-related apoptosis inducing ligand (TRAIL), a member of the TNF superfamily released by microglia, appears to be involved in the induction of apoptosis following focal brain ischemia. Indeed, brain ischemia is associated with progressive enlargement of damaged areas and prominent inflammation. As ischemic preconditioning reduces inflammatory response to brain ischemia and ameliorates brain damage, the purpose of the present study was to evaluate the role of TRAIL and its receptors in stroke and ischemic preconditioning and to propose, by modulating TRAIL pathway, a new therapeutic strategy in stroke. In order to achieve this aim a rat model of harmful focal ischemia, obtained by subjecting animals to 100 min of transient occlusion of middle cerebral artery followed by 24 h of reperfusion and a rat model of ischemic preconditioning in which the harmful ischemia was preceded by 30 mins of tMCAO, which represents the preconditioning protective stimulus, were used. Results show that the neuroprotection elicited by ischemic preconditioning occurs through both upregulation of TRAIL decoy receptors and downregulation of TRAIL itself and of its death receptors. As a counterproof, immunoneutralization of TRAIL in tMCAO animals resulted in significant restraint of tissue damage and in a marked functional recovery. Our data shed new light on the mechanisms that propagate ongoing neuronal damage after ischemia in the adult mammalian brain and provide new molecular targets for therapeutic intervention. Strategies aimed to repress the death-inducing ligands TRAIL, to antagonize the death receptors, or to activate the decoy receptors open new perspectives for the treatment of stroke.

Autologous fat injection to face and neck: from soft tissue augmentation to regenerative medicine.

Minimally-invasive autologous fat injection of the head and neck region can be considered a valid alternative to major invasive surgical procedures both for aesthetic and functional purposes. The favourable outcomes of autologous fat injection in otolaryngological practice are due to the filling of soft tissue and, mainly, to the potential regenerative effect of adipose-derived mesenchymal stem cells. Herewith, some important biological preliminary remarks are described underlying the potential of autologous fat injection in regenerative medicine, and personal experience in using it for both consolidated clinical applications, such as fat grafting to the face and vocal fold augmentation in the treatment of glottic incompetence, and more recent applications including the treatment of post-parotidectomy Frey syndrome and velopharyngeal insufficiency.

Outcomes of structural fat grafting for paralytic and non-paralytic dysphonia.

Aims of this prospective study were to evaluate the results of vocal fold structural fat grafting for glottic insufficiency and to compare the outcomes obtained in unilateral vocal fold paralysis (UVFP) and congenital or acquired soft tissue defects in vocal folds. Sixty-six consecutive patients with breathy dysphonia, in 43 cases (aged 16-79 years) related to UVFP and in 23 cases (aged 16-67 years) related to vocal fold iatrogenic scar or sulcus vocalis, underwent autologous structural fat grafting into vocal folds. Lipoaspirates were centrifuged at 1200 g for 3 min to separate and remove blood, cell debris and the oily layer. The refined fat was injected under direct microlaryngoscopy in a multilayered way. The main outcome measures were grade, roughness, breathiness, asthenicity and strain (GRBAS) perceptual evaluation, maximum phonation time (MPT), self-assessed Voice Handicap Index (VHI), and voice acoustic analysis, considered pre-operatively and at 3 and 6 months after fat grafting. After surgery, MPT, VHI, G and B improved in both groups (p < 0.05). In particular, G and VHI functional subscales showed a significantly greater decrease in patients with UVFP (p < 0.05). The acoustic variables improved significantly only in the UVFP group (p < 0.005). From 3 to 6 months postoperatively, most variables showed a trend with further improvement. Vocal fold structural fat grafting was significantly effective in treating glottic insufficiency due to UVFP or soft tissue defects. Perceptual, acoustic and subjective assessments confirmed that patients with UVFP had better outcomes than those with soft tissue defects.

Endothelial activation and injury by cigarette smoke exposure.

Endothelial activation/injury following exposure to cigarette smoke may explain incidence of atherosclerosis and cardiovascular disease in smokers. We investigated cigarette smoke extract (CSE) effects relative to activation, injury, and survival of human umbilical vein endothelial cells (HUVEC) and compared circulating levels of specific endothelial activation markers between smokers and healthy non-smokers before and after smoking cessation. Viability and toxicity of HUVEC were tested by MTT and LDH assay. Release (by endothelial cells) and circulating levels (in smokers) of von Willebrand Factor (vWF), thrombomodulin (TM), was evaluated by ELISA. Incubation with increasing concentrations of CSE reduced the percentage of viable cells, being 33.9%, 23.9% after CSE 4%, 6% respectively. Dose- and time-dependent release of LDH was observed after incubation with CSE. vWF, TM release were assayed after CSE 2% HUVEC stimulation. Significant 42%, 61%, 76% increase in vWF concentration was detected respectively at 30', 60', 120'. Reduction in circulating levels of vWF, from a median value of 144.0% to 123.7%, was observed in the quitters group after smoking cessation. Exposure to cigarette smoke is cytotoxic and induces activation/injury of endothelium in vitro and in vivo. These findings may provide pathogenetic basis by which smoking can predispose to development of atherothrombosis and cardiovascular disease.

Do mesenchymal stem cells play a role in vocal fold fat graft survival?

OBJECTIVES: Adipose tissue in vocal fold lipoinjection is currently used to treat patients affected by laryngeal hemiplegia or anatomical defects. The aim of this study has been to evaluate the efficacy of this clinical strategy, by long-term follow-up of the patients and to investigate whether the fat samples used to treat them contain a stem cell population with a wide differentiation potential. MATERIALS AND METHODS: Fat samples harvested from 12 patients affected by severe breathy dysphonia who had undergone vocal fold lipoinjection were analysed by immunocytochemistry, by flow cytometry and reverse transcription-polymerase chain reaction, and the isolated adipose derived mesenchymal stem cells (ADMSCs) were evaluated in order to define their ability to produce soluble factors possibly involved in tissue regeneration, and to differentiate towards different lineages. RESULTS: ADMSCs were efficiently and successfully isolated from all of the samples. They were positive for SSEA-4, an embryonic marker recently identified on bone marrow MSCs and which could explain their high differentiation plasticity. Molecular analysis showed that these cells also expressed Oct-4, Runx-1 and ABCG-2, which characterize the stem cell state, and a number of other specific lineage markers. Flow cytometry revealed mesenchymal markers expressed on ADMSCs and identified a subpopulation characterized by CD146(+)/34(-)/45(-) cells consistent with perivascular/pericyte-like cells. Osteogenic, adipogenic and endothelial tissue differentiation were obtained. CONCLUSIONS: Our results confirmed the therapeutic efficacy of this clinical approach and showed that adipose tissue, administered to patients in order to restore glottic competence, contains mesenchymal stem cells.

TRAIL inhibits angiogenesis stimulated by VEGF expression in human glioblastoma cells.

Tumour growth is tightly related to new blood vessel formation, tissue remodelling and invasiveness capacity. A number of tissular factors fuel the growth of glioblastoma multiforme, the most aggressive brain neoplasm. In fact, gene array analyses demonstrated that the proapoptotic cytokine tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) inhibited mRNA expression of VEGF, along with those of matrix metalloproteinase-2 (MMP-2), its inhibitor tissue inhibitor of matrix metalloproteinases-2 (TIMP-2), as well as the tumour invasiveness-related gene secreted protein acid rich in cysteine (SPARC) in different human glioblastoma cell lines. Particularly, VEGF mRNA and protein expression and release from glioblastoma cells were also inhibited by TRAIL. The latter also exerted antimitogenic effects on human umbilical vein endothelial cells (HUVECs). With the same cells, TRAIL inhibited new vessel formation in the in vitro matrigel model, as well as it exerted powerful inhibition of blood vessel formation induced by an angiogenic cocktail administered in subcutaneous pellets in vivo in the C57 mouse. Moreover, the expression of MMP-2, its inhibitor TIMP-2 and the tumour invasiveness-related protein SPARC were effectively inhibited by TRAIL in glioblastoma cell lines. In conclusion, our data indicate that TRAIL inhibits the orchestra of factors contributing to glioblastoma biological aggressiveness. Thus, the TRAIL system could be regarded as a molecular target to exploit for innovative therapy of this type of tumour.

Protective effects of estradiol on TRAIL-induced apoptosis in a human oligodendrocytic cell line: evidence for multiple sites of interactions.

Demyelinating diseases are high impact neurological disorders. Steroids are regarded as protective molecules in the susceptibility to these diseases. Here, we studied the interactions between tumour necrosis factor-related apoptosis-inducing ligand (TRAIL), a potent proapoptotic molecule toxic to oligodendrocytes, and 17-beta-estradiol (E-17-beta), in human oligodendrocytic MO3.13 cells. Exposure of cells to TRAIL resulted in the upregulation of both death receptors DR4 and DR5 and apoptosis, as well as the activation of caspase-8 and -3, increased phosphorylation of Jun-N-terminal kinase and p38 kinase, and the reduction of bcl-2 and bcl-xL proteins. TRAIL-mediated MO3.13 cell apoptosis was abrogated by the dominant-negative form of the adaptor protein FADD and by caspase inhibitors. Preincubation with E-17-beta completely prevented both TRAIL-induced DR4 and DR5 upregulation and apoptosis. Estrogen-induced cytoprotection was time and concentration dependent and reverted by antiestrogens. Estrogen treatment per se reduced kinase phosphorylation, and upregulated bcl-2 and bcl-xL proteins. In conclusion, our data show that the detrimental role of TRAIL on oligodendrocytes can be effectively counteracted by estrogens, thus suggesting that the underlying molecular interactions can be of potential relevance in characterizing novel targets for therapy of demyelinating disorders.

Neutralization of TRAIL death pathway protects human neuronal cell line from beta-amyloid toxicity.

Here we report that a novel member of the TNF-alpha family, TNF-related apoptosis-inducing ligand (TRAIL), contributes substantially to amyloid-induced neurotoxicity in human SH-SY5Y neuronal cell line. Involvement of TRAIL in the amyloid-induced cell death is supported by cDNA array, Northern blot, and Western blot data, demonstrating increased TRAIL expression after treatment of the cells with a neurotoxic fragment of amyloid protein (betaAP). TRAIL was also found to be released in the culture media after betaAP treatment with a time-course overlapping to contents of the intracellular protein. Contribution of TRAIL to betaAP neurotoxicity is demonstrated by data showing that TRAIL-neutralizing monoclonal antibody protects neuronal SH-SY5Y cells from betaAP neurotoxicity. Moreover, exposure of neuronal SH-SY5Y cells to TRAIL leads to cell death, indicating that this substance per se is endowed with neurotoxic properties. We also found that, similarly to betaAP and TRAIL, activation of the death-domain adaptor protein FADD results in neuronal cell death. Lack of FADD function, by overexpression of its dominant negative, rescued cells from either TRAIL- or betaAP-induced neurotoxicity, supporting the hypothesis that these three molecules share common intracellular pathways. Finally, we found that betaAP strongly activated caspase-8, and the cell-permeable, selective caspase-8 inhibitor z-IETD-FMK prevents both betaAP- and TRAIL-induced neurotoxicity. In view of TRAIL's potency in inducing neuronal death, and its role as mediator of betaAP, it is plausible to hypothesize that TRAIL can be regarded as a molecule that provides substantial contribution to betaAP-dependent cell death, which takes part in the progression of the neurodegenerative process and related chronic inflammatory response.

Divergent effects of corticotropin releasing hormone on endothelial cell nitric oxide synthase are associated with different expression of CRH type 1 and 2 receptors.

1. Endothelium is a target for an array of factors involved in inflammation. Endothelial cells express receptors for CRH, a neuropeptide produced during inflammation. We report both the concentration-dependent inhibitory effect of CRH upon cytokine-stimulated nitrite release by H5V murine endothelioma cells, and its stimulatory one in HUVEC cells. 2. Western blot analysis showed that CRH inhibits cytokine-stimulated iNOS protein in H5V cells, and, instead, potentiated it in HUVEC cells. 3. H5V cells expressed both CRH receptors (CRH-R1 and R2) mRNAs, whereas HUVEC cells expressed the CRH-R2 mRNA solely. 4. CRH increased medium nitrites and iNOS protein expression in H5V cells pretreated with the selective CRH-R1 antagonist CP 154,526. However, the selective CRH-R2 antagonist anti-Svg-30 failed to produce similar effects. In fact, anti-Svg-30 inhibited CRH-induced increase of nitrite release and iNOS expression in HUVEC cells. 5. Our results confirm the activating role of CRH on endothelial cells, although it suggests its possible inhibitory role in the late phase of the inflammatory response. NO-mediated effects of CRH on endothelial cells could be exploited in therapeutic strategies related to inflammatory and/or degenerative diseases.

Spindle cell lipoma of the hypopharynx.

Larynx and hypopharynx lipomas are reported to account for approximately 0.6% of benign laryngeal neoplasms. Spindle cell lipoma is a histologically distinct variant characterized by mature adipocytes mixed with collagen-forming spindle cells; only one case of spindle cell lipoma of the larynx has been previously reported. We here describe a new case of spindle cell lipoma of the pyriform sinus successfully treated by means of endoscopic surgical excision. A 77-year-old woman with a 40-year history of dysphagia reported that the condition had markedly worsened over the three years before she came to us. She had difficulty swallowing even semisolid food and she experienced occasional nasal regurgitation of liquid or solid food. Flexible videolaryngoscopy showed a very large mass, covered by normal mucosa that almost totally occupied the right pyriform sinus and was apparently attached to the right arytenoid. Functional endoscopic study and videofluoroscopy of swallowing showed that the bolus progressed exclusively in the left pyriform sinus, with postdeglutitory pooling in the right pyriform sinus and a reflux toward the valleculae during consecutive deglutitions. Computed tomography demonstrated that the hypopharyngeal mass had low attenuation values and negative densitometry. The entire mass was surgically removed during suspension microlaryngoscopy. The histological sections showed mature adipocytes mixed with small and slender spindle cells. Postoperative endoscopic and videofluorosocpic deglutition studies revealed the recovery of normal swallowing. This case indicates that hypopharyngeal lipomas should be included in the differential diagnosis of slowly occurring swallowing impairments.

Tamoxifen inhibits nerve growth factor-induced proliferation of the human breast cancerous cell line MCF-7.

An array of polypeptide growth factors contribute to the development of breast cancer, the most common tumor-related cause of death in women of Western countries. Therefore, breast cancer therapy should be aimed at inhibition of growth factor-dependent breast cancerous cell proliferation. However, the relative contribution of each individual factor in the development and maintenance of the transformed phenotype is largely unknown. Here we report for the first time that the proliferative effects of nerve growth factor, (NGF) a typical neurotrophin, are similar to those of epidermal growth factor (EGF) and insulin-like growth factor II, and are enhanced by 17beta-estradiol in the human breast cancer cell line MCF-7. The effect of NGF appeared to be mediated by its trkA receptors (trkA(NGFR)), as suggested by the potent inhibition of both MCF-7 cell proliferation and trkA(NGFR) phosphorylation occurring upon treatment of cultures with the selective trkA(NGFR) inhibitor K252a. Surprisingly, the antiestrogen drug tamoxifen (TAM) inhibited NGF-induced MCF-7 cell proliferation and trkA(NGFR) phosphorylation in a concentration-related fashion. The effect of TAM seemed to be estrogen receptor-independent, because the pure estrogen receptor antagonist ICI 182.780 was unable to block NGF-induced trkA(NGFR) phosphorylation. Our data underline the new emerging role of trkA(NGFR) in breast tumor growth, and suggest a related novel therapeutic use of TAM in breast cancer.

Increased ACTH and cortisol secretion after interleukin-alpha injection in the common marmoset (Callithrix jacchus jacchus).

We have studied the effect of intravenous injection of interleukin-1 (dose range: from 0.25 to 4.5 microg/kg of body weight) on plasma ACTH and cortisol levels in the marmoset, a primate paradygm of peripheral glucocorticoid resistance. Blood sampling were collected and body temperature recorded 0, 15, 30, 60, 120, 180, 240 and 300 min after injection. Interleukin-1 stimulated secretion of ACTH in a dose-dependent fashion. Maximal secretion occurred 120 min after injection, and lasted up to 240 min. Plasma ACTH levels returned to baseline 300 min after interleukin-1 injection. Plasma cortisol levels were related to ACTH levels. Body temperature elevation, which occurred 10-15 min after injection was dose-dependent, and lasted 3 h. Results suggest that the pyrogenic effect of interleukin is associated, in the marmoset, with integrated activation of the hypothalamic-pituitary-adrenal axis. In light of the proneness of marmosets to hyperimmune disorders, our data are consistent with the hypothesized central biological role of IL-1, as well as the pathophysiological relevance of the neuro-endocrine-immune cross-talk during the acute phase response.

Bilateral laryngeal pseudoparalysis in xanthoma disseminatum treated by endoscopic laser medial arytenoidectomy.

Xanthoma disseminatum is a rare non-Langerhans' cell histiocytosis, characterized by papular cutaneous eruption, possible mucosal involvement, and frequent association with vasopressin-sensitive diabetes insipidus. Herein we report a case of xanthoma disseminatum with pharyngolaryngeal involvement. In this patient, mucosal xanthomas involving the arytenoid cartilages and the interarytenoid area resulted in laryngeal stenosis and severe impairment of both cricoarytenoid joints' motility. Endoscopic CO2 laser medial arytenoidectomy, according to the technique described by Crumley (1993), and vaporization of interarytenoid xanthomas were successfully performed, thus reestablishing bilateral cordal motility and the laryngeal airway. Four years later, a CO2 laser revision was necessary because of recurrence of xanthomas in the posterior larynx. Two years after the latter operation, the patient has no signs of laryngeal obstruction and has a normal voice quality. This case report suggests that endoscopic medial arytenoidectomy may be successfully used in the treatment of bilateral laryngeal pseudoparalysis secondary to xanthoma disseminatum.

Increased frequency dependence of specific airway resistance in patients with laryngeal hemiplegia.

The aim of this study was to investigate whether patients with laryngeal hemiplegia (LH) show a frequency-dependent increase in specific airway resistance (sRaw), measured by body plethysmography. In addition to the flow-volume loop, usually considered in the functional evaluation of upper airway obstructions, variations in sRaw at respiratory frequencies of 30+/-5 (=0.5 Hz), 60+/-5 (=1 Hz) and 90+5 breaths x min(-1) (=1.5 Hz) in 21 never-smoking patients (LH group, mean age+/-SD 55+/-12.09 yrs; 17 females) whose unilateral vocal-cord paralysis was documented by laryngoscopy and who had no signs or symptoms of other respiratory diseases studied. They were compared to 21 healthy control subjects (C group: 50.1+/-15.44 yrs; 10 females). The sRaw values at 30+/-5 breaths min(-1) were similar in the two groups (5.54+/-1.88 versus 5.68+/-1.06 cmH2O x s(-1); p=NS), but at increasing frequencies (30+/-5, 60+/-5 and 90+/-5 breaths min(-1)), they progressively and significantly increased in the LH patients (from 5.54+/-1.88 to 6.63+/-1.96 and 8.05+/-2.6 mH2O x s(-1); p<0.0005), and not significantly in controls (5.68+/-1.06, 5.85+/-0.95 and 5.9+/-1.12 cmH2O x s(-1); p=NS). Linear discriminant analysis using AsRaw (sRaw at 1.5 Hz-sRaw at 0.5 Hz) and forced inspiratory flow at 50% of the vital capacity made it possible to correctly classify all of the controls and 19 of the 21 patients. In conclusion, the multiple, rapid and noninvasive plethysmographical testing of frequency-dependent increase in specific airway resistance with the flow-volume loop, allows the sufficiently satisfactory discrimination of laryngeal hemiplegia patients from controls.

Recruitment of the IKK signalosome to the p55 TNF receptor: RIP and A20 bind to NEMO (IKKgamma) upon receptor stimulation.

The adapter protein RIP plays a crucial role in NF-kappaB activation by TNF. Here we show that triggering of the p55 TNF receptor induces binding of RIP to NEMO (IKKgamma), a component of the I-kappa-B-kinase (IKK) "signalosome" complex, as well as recruitment of RIP to the receptor together with the three major signalosome components, NEMO, IKK1 and IKK2, and some kind of covalent modification of the recruited RIP molecules. It also induces binding of NEMO to the signaling inhibitor A20, and recruitment of A20 to the receptor. Enforced expression of NEMO in cells revealed that NEMO can both promote and block NF-kappaB activation and dramatically augments the phosphorylation of c-Jun. The findings suggest that the signaling activities of the IKK signalosome are regulated through binding of NEMO to RIP and A20 within the p55 TNF receptor complex.