Patrizia Mancini

Cochlear Implants in Special Cases: Deafness in the Presence of Disabilities and/or Associated Problems

The aim of the present study is to construct a reference model with the indication for the attitude, the requirements and the resources needed in order to be able to deal with deafness in the presence of disabilities or associated problems. The study group consisted of 13 adults and 18 children affected by profound deafness, with associated problems and disabilities, who were implanted with Clarion® and Med-El® devices. Selection criteria for candidacy to cochlear implantation and counselling, hospitalization, fitting and speech therapy/rehabilitation are described. Findings were assessed evaluating: (i) use of acoustic feedback, on the ground of Erbers model; (ii) self-sufficiency: assessed by a questionnaire; and (iii) social and family relationships: qualitative judgment based on direct observation, analysis of drawings and structured interviews with family teachers and therapists. The whole group showed benefit from cochlear implantation, with particular satisfaction for post-lingual deaf-blind adults, as well as for subjects with associated psychopathologies and mental retardation. In conclusion, cochlear implants can improve life quality in profoundly deaf subjects with associated disabilities, increasing both listening and communication skills as well as self-sufficiency while family and social relationships tend to remain stable.

Sym1, the yeast ortholog of the MPV17 human disease protein, is a stress-induced bioenergetic and morphogenetic mitochondrial modulator

A peculiar form of hepatocerebral mtDNA depletion syndrome is caused by mutations in the MPV17 gene, which encodes a small hydrophobic protein of unknown function located in the mitochondrial inner membrane. In order to define the molecular basis of MPV17 variants associated with the human disorder, we have previously taken advantage of S. cerevisiae as a model system thanks to the presence of an MPV17 ortholog gene, SYM1. We demonstrate here that the SYM1 gene product is essential to maintain OXPHOS, glycogen storage, mitochondrial morphology and mtDNA stability in stressing conditions such as high temperature and ethanol-dependent growth. To gain insight into the molecular basis of the Sym1-less phenotype, we identified and characterized multicopy suppressor genes and metabolic suppressor compounds. Our results suggest that (i) metabolic impairment and mtDNA instability occur independently from each other as a consequence of SYM1 ablation; (ii) ablation of Sym1 causes depletion of glycogen storage, possibly due to defective anaplerotic flux of tricarboxylic acid (TCA) cycle intermediates to the cytosol; (iii) flattening of mitochondrial cristae in Sym1-defective organelles suggests a role for Sym1 in the structural preservation of the inner mitochondrial membrane, which could in turn control mtDNA maintenance and stability.

Positional vertigo and cochlear implantation.

Objective: To identify patients developing positional vertigo after cochlear implantation. Study Design: Prospective study on a cohort of patients undergoing cochlear implantation. Setting: Academic tertiary referral center. Patients: The study included 70 consecutive patients who underwent vestibular evaluation before and after cochlear implantation. Intervention: Medical record review. Main Outcome Measure: Recorded vestibular symptoms after cochlear implantation. Patients with positional vertigo were considered case subjects, whereas those without vestibular symptoms were considered case controls. Results: Benign paroxysmal positional vertigo (BPPV) occurred in 8 patients (on the cochlear implant [CI] side in 7 patients, and in the other ear in 1). One patient had BPPV of the lateral semicircular canal on the implanted side, and 7 patients had BPPV of the posterior semicircular canal (on the same CI side in 6 patients, and on the opposite side in 1), which were detected and presented during the last examination. In 5 patients, the onset of symptoms varied from 7 to 130 days after implant activation; in 2 patients, the onset occurred before activation. Conclusion: Three different mechanisms are proposed for the occurrence of BPPV in patients with CI. The first focuses on the fall of bone dust particles into the cochlea during cochleostomy. In the second, the vibration caused by drilling the cochlea would be sufficient to dislodge otoconia into the labyrinth. The third hypothesis suggests dislodging of an otolith because of the electric stimulation. In our patients, conservative approaches have been used with a minimal invasive cochleostomy and without perilymph suction. Thus, the vibratory trauma affecting the cochlea during cochleostomy seems to play a fundamental role in the development of paroxysmal vertigo in patients with implant.

Intratympanic steroid therapy in moderate sudden hearing loss: a randomized, triple-blind, placebo-controlled trial.

To investigate the efficacy of an intratympanic steroid as a first‐line therapy in patients affected by moderate idiopathic sudden sensorineural hearing loss (ISSNHL).

APY-1, a Novel Caenorhabditis elegans Apyrase Involved in Unfolded Protein Response Signalling and Stress Responses

Protein glycosylation modulates a wide variety of intracellular events and dysfunction of the glycosylation pathway has been reported in a variety of human pathologies. Endo-apyrases have been suggested to have critical roles in protein glycosylation and sugar metabolism. However, deciphering the physiological relevance of Endo-apyrases activity has actually proved difficult, owing to their complexity and the functional redundancy within the family. We report here that a UDP/GDPase, homologous to the human apyrase Scan-1, is present in the membranes of Caenorhabditis elegans, encoded by the ORF F08C6.6 and hereinafter-named APY-1. We showed that ER stress induced by tunicamycin or high temperature resulted in increased transcription of apy-1. This increase was not observed in C. elegans mutants defective in ire-1 or atf-6, demonstrating the requirement of both ER stress sensors for up-regulation of apy-1. Depletion of APY-1 resulted in constitutively activated unfolded protein response. Defects in the pharynx and impaired organization of thin fibers in muscle cells were observed in adult worms depleted of APY-1. Some of the apy-1(RNAi) phenotypes are suggestive of premature aging, because these animals also showed accumulation of lipofuscin and reduced lifespan that was not dependent on the functioning of DAF-2, the receptor of the insulin/IGF-1 signaling pathway.

Cochlear Implant Programming: A Global Survey on the State of the Art

The programming of CIs is essential for good performance. However, no Good Clinical Practice guidelines exist. This paper reports on the results of an inventory of the current practice worldwide. A questionnaire was distributed to 47 CI centers. They follow 47600 recipients in 17 countries and 5 continents. The results were discussed during a debate. Sixty-two percent of the results were verified through individual interviews during the following months. Most centers (72%) participated in a cross-sectional study logging 5 consecutive fitting sessions in 5 different recipients. Data indicate that general practice starts with a single switch-on session, followed by three monthly sessions, three quarterly sessions, and then annual sessions, all containing one hour of programming and testing. The main focus lies on setting maximum and, to a lesser extent, minimum current levels per electrode. These levels are often determined on a few electrodes and then extrapolated. They are mainly based on subjective loudness perception by the CI user and, to a lesser extent, on pure tone and speech audiometry. Objective measures play a small role as indication of the global MAP profile. Other MAP parameters are rarely modified. Measurable targets are only defined for pure tone audiometry. Huge variation exists between centers on all aspects of the fitting practice.

Distinct involvement of Cdc42 and RhoA GTPases in actin organization and cell shape in untransformed and Dbl oncogene transformed NIH3T3 cells

The Dbl oncogene is a putative exchange factor for the small GTPases RhoA and Cdc42, which are involved in actin polymerization into stress fibers and filopodia, respectively. We report here that, upon adhesion to fibronectin, Dbl-transformed NIH3T3 cells display a contracted, polygonal shape with a high number of short stress fibers. In contrast, untransformed NIH3T3 cells acquire the characteristic fibroblast morphology and organize a regular mesh of long stress fibers. We show that in Dbl-transformed and in untransformed NIH3T3 cells the different shape and actin cytoskeleton organization observed in the early steps of adhesion involves activation of distinct GTPases. Upon adhesion to fibronectin, cell morphology of Dbl-transformed NIH3T3 cells depends on activation of RhoA and not of Cdc42. In contrast Cdc42 activation is necessary to untransfected NIH3T3 cells to acquire their fibroblast shape. In both Dbl-transformed and in untransformed NIH3T3 cells a basal Rac activation is necessary to support stress fiber organization, while constitutive Rac activation promotes ruffles and lamellipodia formation. As a consequence of RhoA activation, Dbl-transformed cells show high activity of ROCK-α and CRIK kinases, two known RhoA effectors. In addition Dbl-transformed and NIH3T3 cells expressing the constitutive active form of RhoA are less motile on fibronectin than cells expressing constitutive active Cdc42. We conclude that in NIH3T3 cells in response to fibronectin the expression of the Dbl oncogene leads to a predominant activation of RhoA which both supports the peculiar cell shape and actin cytoskeleton organization in stress fibers and regulates cell motility.

Involvement of the Src-cortactin pathway in migration induced by IGF-1 and EGF in human breast cancer cells

Cancer cells need to become motile in order to escape the primary tumor and move to distant areas to form metastasis. They move as single cells or as a group, following different stimuli, including growth factors. Among them, insulin-like growth factor‑1 (IGF-1) and epidermal growth factor (EGF) and their receptors have been implicated in the development and progression of human breast carcinoma. In this report, we provide evidence that the tyrosine kinase Src is responsible for migration promoted by both IGF-1 and EGF in MDA-MB-231 and MCF7 cells, although with a different effect. Moreover, both IGF-1 and EGF induce reorganization of actin cytoskeleton in lamellipodia and membrane ruffles in a time- and Src-dependent manner. Furthermore, we analyzed the tyrosine phosphorylation status of the actin-binding protein cortactin upon growth factor stimulation, showing that even the activation of cortactin is time- and Src-dependent. In addition, immunofluorescence analysis with anti-paxillin antibody reveals that, after treatment with growth factors, tyrosine phosphorylated cortactin is localized on the plasma membrane in correspondence of focal adhesions. Collectively, our findings suggest a crucial role for Src-mediated activation of cortactin in cell migration, reorganization of actin cytoskeleton and phosphotyrosine cortactin localization to the focal adhesions in human breast cancer cell lines upon both IGF-1 and EGF stimulation.

Phosphorylation-independent membrane relocalization of ezrin following association with Dbl in vivo

Ezrin, a widespread protein involved in cell migration, morphogenesis and cell adhesion, belongs to a large family of proteins known as ERM (ezrin, radixin, moesin). These three closely related proteins are thought to function as linkers between plasma membrane and actin cytoskeleton and their function is regulated by the small GTP-binding protein Rho. It has been previously shown that the active form of radixin can bind in vitro to Dbl, a Rho-specific guanine nucleotide exchange factor, although an in vivo interaction has not yet been demonstrated. In this paper, we attempted to investigate whether ezrin can also associate with Dbl. We show here that Dbl protein can effectively bind both in vitro and in vivo to the N-terminal region (amino acids 1–531) of a constitutively active mutant of ezrin and with the full-length molecule. We found that this binding is mediated by the Dbl pleckstrin homology domain, responsible for the proper subcellular localization of the Dbl protein. Moreover, we show that Dbl induces localization to the plasma membrane of both the active deletion mutant and the full-length ezrin proteins. Finally, we show that the relocalization of ezrin is independent of Dbl GEF activity. These results indicate that Dbl could induce translocation of ezrin to the plasma membrane through a mechanism that does not require ezrin C-terminus phosphorylation by Rho-associated kinases.

Polarized Endocytosis of the Keratinocyte Growth Factor Receptor in Migrating Cells: Role of Src-Signaling and Cortactin

Cell migration is a physiological process that requires endocytic trafficking and polarization of adhesion molecules and receptor tyrosine kinases (RTKs) to the leading edge. Many growth factors are able to induce motility by binding to specific RTK on target cells. Among them, keratinocyte growth factor (KGF or FGF7) and fibroblast growth factor 10 (FGF10), members of the FGF family, are motogenic for keratinocytes, and exert their action by binding to the keratinocyte growth factor receptor (KGFR), a splicing variant of FGFR2, exclusively expressed on epithelial cells. Here we analyzed the possible role of cortactin, an F-actin binding protein which is tyrosine phosphorylated by Src and is involved in KGFR-mediated cell migration, in the KGFR endocytosis and polarization to the leading edge of migrating cells upon ligand-induced stimulation. Biochemical phosphorylation study revealed that both KGF and FGF10 were able to induce tyrosine phosphorylation of Src and in turn of cortactin, as demonstrated by using the specific pharmacological Src-inhibitor SU6656, although FGF10 effect was delayed with respect to that promoted by KGF. Immunofluorescence analysis demonstrated the polarized localization of KGFR upon ligand stimulation to the leading edge of migrating keratinocytes, process that was regulated by Src. Moreover, we showed that the colocalization of cortactin with KGFR at the plasma membrane protrusions and on early endosomes after KGF and FGF10 treatment was Src-dependent. Further, by using a RNA interference approach through microinjection, we showed that cortactin is required for KGFR endocytosis and that the clathrin-dependent internalization of the receptor is a critical event for its polarization. Finally, KGFR expression and polarization enhanced cell migration in a scratch assay. Our results indicate that both Src and cortactin play a key role in the KGFR endocytosis and polarization at the leading edge of migrating keratinocytes, supporting the crucial involvement of RTK trafficking in cell motility.