Nuclear localization of mouse fibroblast growth factor 2 requires N-terminal and C-terminal sequences.

In vertebrates, different isoforms of fibroblast growth factor 2 (FGF2) exist, which differ by their N-terminal extension. They show different localization and expression levels and exert distinct biological effects. Nevertheless, genetic inactivation of all FGF2 isoforms in the mouse results in only mild phenotypes. Here, we analyzed mouse FGF2, and show that, as in the human, mouse FGF2 contains CTG-initiated high molecular-weight (HMW) isoforms, which contain a nuclear localization signal, and which mediate localization of this isoform to the nucleus. Using green fluorescent protein-FGF2 fusions, we furthermore observed, that C-terminal deletions disable nuclear localization of the short low-molecular-weight (LMW) 18-kDa isoform. This loss of specific localization is accompanied by a loss in heparin binding. We therefore suggest that, first, localization of mouse FGF2 is comparable to that in other vertebrates and, second, FGF2 contains at least two sequences important for nuclear localization, a nuclear localization sequence at the N terminus which is only contained in the HMW isoform, and another sequence at the C terminus, which is only required for localization of the LMW 18-kDa isoform.

Multi institutional phase II study of concomitant stereotactic reirradiation and cetuximab for recurrent head and neck cancer.

PURPOSE: Recurrent head and neck cancer is associated to a poor survival prognosis. A high toxicity rate is demonstrated when surgery and/or radiotherapy and/or chemotherapy are combined. Furthermore, the duration of treatment is often not ethically compatible with the expected survival (median survival<1year). Normal tissues tolerance limits the use of reirradiation and stereotactic body radiotherapy (SBRT) could offer precise irradiation while sparing healthy tissues. After completion of a feasibility study, results of a multicentric study (Lille, Nancy & Nice) using SBRT with cetuximab are reported. The aim of the study was to deliver non toxic short course SBRT (2weeks) in order to get the same local control as the one demonstrated with longer protocols. METHODS AND MATERIALS: Patients with inoperable recurrent, or new primary tumor in a previously irradiated area, were included (WHO<3). Reirradiation (RT) dose was 36Gy in six fractions of 6Gy to the 85% isodose line covering 95% of the PTV with 5 injections of concomitant cetuximab (CT). All patients had previous radiotherapy, 85% had previous surgery and 48% previous chemotherapy. RESULTS: Between 11/2007 and 08/2010, 60 were included (46 men and 14 women), 56 received CT+RT, 3 were not treated and 1 received only CT. Median age was 60 (42-87)) and all 56 patients had squamous carcinoma and received concomitant cetuximab. Mean time between previous radiotherapy and the start of SBRT was 38months. Cutaneous toxicity was observed for 41 patients. There was one toxic death from hemorrhage and denutrition. Median follow-up was 11.4months. At 3months, response rate was 58.4% (95% CI: 43.2-72.4%) and disease control rate was 91.7% (95% CI: 80.0-97.7%). The one-year OS rate was 47.5% (95% CI: 30.8-62.4). CONCLUSION: These results suggest that short SBRT with cetuximab is an effective salvage treatment with good response rate in this poor prognosis population with previously irradiated HNC. Treatment is feasible and, with appropriate care to limiting critical structure, acute toxicities are acceptable. This combination may be the reference treatment is this population.

Comparison of quality of life after stereotactic body radiotherapy and surgery for early-stage prostate cancer

Background: As the long-term efficacy of stereotactic body radiation therapy (SBRT) becomes established and other prostate cancer treatment approaches are refined and improved, examination of quality of life (QOL) following prostate cancer treatment is critical in driving both patient and clinical treatment decisions. We present the first study to compare QOL after SBRT and radical prostatectomy, with QOL assessed at approximately the same times pre- and post-treatment and using the same validated QOL instrument. Methods: Patients with clinically localized prostate cancer were treated with either radical prostatectomy (n = 123 Spanish patients) or SBRT (n = 216 American patients). QOL was assessed using the Expanded Prostate Cancer Index Composite (EPIC) grouped into urinary, sexual, and bowel domains. For comparison purposes, SBRT EPIC data at baseline, 3 weeks, 5, 11, 24, and 36 months were compared to surgery data at baseline, 1, 6, 12, 24,and 36 months. Differences in patient characteristics between the two groups were assessed using Chi-squared tests for categorical variables and t-tests for continuous variables. Generalized estimating equation (GEE) models were constructed for each EPIC scale to account for correlation among repeated measures and used to assess the effect of treatment on QOL. Results: The largest differences in QOL occurred in the first 16 months after treatment, with larger declines following surgery in urinary and sexual QOL as compared to SBRT, and a larger decline in bowel QOL following SBRT as compared to surgery. Long-term urinary and sexual QOL declines remained clinically significantly lower for surgery patients but not for SBRT patients. Conclusions: Overall, these results may have implications for patient and physician clinical decision making which are often influenced by QOL. These differences in sexual, urinary and bowel QOL should be closely considered in selecting the right treatment, especially in evaluating the value of non-invasive treatments, such as SBRT.

Sodium/hydrogen exchanger NHA2 in osteoclasts: subcellular localization and role in vitro and in vivo.

NHA2 was recently identified as a novel sodium/hydrogen exchanger which is strongly upregulated during RANKL-induced osteoclast differentiation. Previous in vitro studies suggested that NHA2 is a mitochondrial transporter required for osteoclast differentiation and bone resorption. Due to the lack of suitable antibodies, NHA2 was studied only on RNA level thus far. To define the protein's role in osteoclasts in vitro and in vivo, we generated NHA2-deficient mice and raised several specific NHA2 antibodies. By confocal microscopy and subcellular fractionation studies, NHA2 was found to co-localize with the late endosomal and lysosomal marker LAMP1 and the V-ATPase a3 subunit, but not with mitochondrial markers. Immunofluorescence studies and surface biotinylation experiments further revealed that NHA2 was highly enriched in the plasma membrane of osteoclasts, localizing to the basolateral membrane of polarized osteoclasts. Despite strong upregulation of NHA2 during RANKL-induced osteoclast differentiation, however, structural parameters of bone, quantified by high-resolution microcomputed tomography, were not different in NHA2-deficient mice compared to wild-type littermates. In addition, in vitro RANKL stimulation of bone marrow cells isolated from wild-type and NHA2-deficient mice yielded no differences in osteoclast development and activity. Taken together, we show that NHA2 is a RANKL-induced plasmalemmal sodium/hydrogen exchanger in osteoclasts. However, our data from NHA2-deficient mice suggest that NHA2 is dispensable for osteoclast differentiation and bone resorption both in vitro and in vivo.

GLUT8 subcellular localization and absence of translocation to the plasma membrane in PC12 cells and hippocampal neurons.

GLUT8 is a high-affinity glucose transporter present mostly in testes and a subset of brain neurons. At the cellular level, it is found in a poorly defined intracellular compartment in which it is retained by an N-terminal dileucine motif. Here we assessed GLUT8 colocalization with markers for different cellular compartments and searched for signals, which could trigger its cell surface expression. We showed that when expressed in PC12 cells, GLUT8 was located in a perinuclear compartment in which it showed partial colocalization with markers for the endoplasmic reticulum but not with markers for the trans-Golgi network, early endosomes, lysosomes, and synaptic-like vesicles. To evaluate its presence at the plasma membrane, we generated a recombinant adenovirus for the expression of GLUT8 containing an extracellular myc epitope. Cell surface expression was evaluated by immunofluorescence microscopy of transduced PC12 cells or primary hippocampal neurons exposed to different stimuli. Those included substances inducing depolarization, activation of protein kinase A and C, activation or inhibition of tyrosine kinase-linked signaling pathways, glucose deprivation, AMP-activated protein kinase stimulation, and osmotic shock. None of these stimuli-induced GLUT8 cell surface translocation. Furthermore, when GLUT8myc was cotransduced with a dominant-negative form of dynamin or GLUT8myc-expressing PC-12 cells or neurons were incubated with an anti-myc antibody, no evidence for constitutive recycling of the transporter through the cell surface could be obtained. Thus, in cells normally expressing it, GLUT8 was associated with a specific intracellular compartment in which it may play an as-yet-uncharacterized role.

Anthracene-tethered ruthenium(II) arene complexes as tools to visualize the cellular localization of putative organometallic anticancer compounds.

Anthracene derivatives of ruthenium(II) arene compounds with 1,3,5-triaza-7-phosphatricyclo[3.3.1.1]decane (pta) or a sugar phosphite ligand, viz., 3,5,6-bicyclophosphite-1,2-O-isopropylidene-α-d-glucofuranoside, were prepared in order to evaluate their anticancer properties compared to the parent compounds and to use them as models for intracellular visualization by fluorescence microscopy. Similar IC(50) values were obtained in cell proliferation assays, and similar levels of uptake and accumulation were also established. The X-ray structure of [{Ru(η(6)-C(6)H(5)CH(2)NHCO-anthracene)Cl(2)(pta)] is also reported.

Tumor localization of radiolabeled antibodies against carcinoembryonic antigen in patients with carcinoma: a critical evaluation.

Purified, [131I]-labeled goat antibodies against carcinoembryonic antigen, which have been shown to localize in human carcinoma in nude mice, were injected into 27 patients with carcinoma. Patients were scanned with a scintillation camera at various intervals. In 11 patients, radioactivity was detectable in the tumor 48 hours after injection. Computerized subtraction of blood-pool radioactivity provided clearer pictures in positive cases, but in 16 patients the scans remained doubtful or negative. To study the specificity of [131I]-antibody localization, we gave some patients simultaneous injections of [125I]-labeled normal IgG. Both isotopes were measured by means of scintillation counting in tumors and normal tissues recovered after surgery. The results demonstrated that only the anti-CEA antibodies localized in tumors. However, the total antibody-derived radioactivity in the tumor was only about 0.001 of the injected dose. We conclude that, despite the present demonstration of specificity, this method of tumor detection is not yet clinically useful.

Coarse Global Visual Localization of a Mobile Robot

Localization, which is the ability of a mobile robot to estimate its position within its environment, is a key capability for autonomous operation of any mobile robot. This thesis presents a system for indoor coarse and global localization of a mobile robot based on visual information. The system is based on image matching and uses SIFT features as natural landmarks. Features extracted from training images arestored in a database for use in localization later. During localization an image of the scene is captured using the on-board camera of the robot, features are extracted from the image and the best match is searched from the database. Feature matching is done using the k-d tree algorithm. Experimental results showed that localization accuracy increases with the number of training features used in the training database, while, on the other hand, increasing number of features tended to have a negative impact on the computational time. For some parts of the environment the error rate was relatively high due to a strong correlation of features taken from those places across the environment.

Electrical source imaging for presurgical focus localization in epilepsy patients with normal MRI.

PURPOSE: Patients with magnetic resonance (MR)-negative focal epilepsy (MRN-E) have less favorable surgical outcomes (between 40% and 70%) compared to those in whom an MRI lesion guides the site of surgical intervention (60-90%). Patients with extratemporal MRN-E have the worst outcome (around 50% chance of seizure freedom). We studied whether electroencephalography (EEG) source imaging (ESI) of interictal epileptic activity can contribute to the identification of the epileptic focus in patients with normal MRI. METHODS: We carried out ESI in 10 operated patients with nonlesional MRI and a postsurgical follow-up of at least 1 year. Five of the 10 patients had extratemporal lobe epilepsy. Evaluation comprised surface and intracranial EEG monitoring of ictal and interictal events, structural MRI, [(18)F]fluorodeoxyglucose positron emission tomography (FDG-PET), ictal and interictal perfusion single photon emission computed tomography (SPECT) scans. Eight of the 10 patients also underwent intracranial monitoring. RESULTS: ESI correctly localized the epileptic focus within the resection margins in 8 of 10 patients, 9 of whom experienced favorable postsurgical outcomes. DISCUSSION: The results highlight the diagnostic value of ESI and encourage broadening its application to patients with MRN-E. If the surface EEG contains fairly localized spikes, ESI contributes to the presurgical decision process.

Phosphorylation-dependent dimerization and subcellular localization of islet-brain 1/c-Jun N-terminal kinase-interacting protein 1.

Islet-brain 1 [IB1; also termed c-Jun N-terminal kinase (JNK)-interacting protein 1 (JIP-1] is involved in the apoptotic signaling cascade of JNK and functions as a scaffold protein. It organizes several MAP kinases and the microtubule-transport motor protein kinesin and relates to other signal-transducing molecules such as the amyloid precursor protein. Here we have identified IB1/JIP-1 using different antibodies that reacted with either a monomeric or a dimeric form of IB1/JIP-1. By immunoelectron microscopy, differences in the subcellular localization were observed. The monomeric form was found in the cytoplasmic compartment and is associated with the cytoskeleton and with membranes, whereas the dimeric form was found in addition in nuclei. After treatment of mouse brain homogenates with alkaline phosphatase, the dimeric form disappeared and the monomeric form decreased its molecular weight, suggesting that an IB1/JIP-1 dimerization is phosphorylation dependent and that IB1 exists in several phospho- forms. N-methyl-D-aspartate receptor activation induced a dephosphorylation of IB1/JIP-1 in primary cultures of cortical neurons and reduced homodimerization. In conclusion, these data suggest that IB1/JIP-1 monomers and dimers may differ in compartmental localization and thus function as a scaffold protein of the JNK signaling cascade in the cytoplasm or as a transcription factor in nuclei.

Mobile robot localization using sonar ranging and wlan intensity maps

 Main goal of this thesis was to implement a localization system which uses sonars and WLAN intensity maps to localize an indoor mobile robot. A probabilistic localization method, Monte Carlo Localization is used in localization. Also the theory behind probabilistic localization is explained. Two main problems in mobile robotics, path tracking and global localization, are solved in this thesis. Implemented system can achieve acceptable performance in path tracking. Global localization using WLAN received signal strength information is shown to provide good results, which can be used to localize the robot accurately, but also some bad results, which are no use when trying to localize the robot to the correct place. Main goal of solving ambiguity in office like environment is achieved in many test cases.

Localization and fine structure of a vaccinia virus gene encoding an envelope antigen.

The major antigen on the envelope of extracellular vaccinia virus particles is a polypeptide with an apparent molecular weight of 37,000 (p37K; G. Hiller and K. Weber, J. Virol. 55:651-659, 1985). The gene encoding p37K was mapped in the vaccinia virus genome by hybrid selection of RNA followed by in vitro translation. p37K was then identified among the in vitro translation products by immunoprecipitation with a monoclonal antibody. The gene is located close to the right-hand end of the HindIII F fragment. The corresponding region of the DNA was sequenced, and an open reading frame encoding a polypeptide of 41,748 daltons was observed. The 5' end of the mRNA, as defined by nuclease S1 analysis, maps within only a few nucleotides of the translation initiation codon. Examination of the DNA sequence around the putative initiation site of transcription revealed a characteristic sequence, TAAATG, which includes the ATG translation initiation codon and which is conserved in all but one late gene so far analyzed. It is therefore likely that this sequence is an important regulatory signal for late gene expression in vaccinia virus.

Stereotactic body radiotherapy with helical TomoTherapy for medically inoperable early stage primary and second-primary non-small-cell lung neoplasm: 1-year outcome and toxicity analysis.

OBJECTIVE: This study investigated the effectiveness of stereotactic body radiotherapy with helical TomoTherapy (T-SBRT) for treating medically inoperable primary and second-primary early stage non-small-cell lung neoplasm (SPLN) and evaluated whether the movement of organizing pneumonia (OP) within the irradiation field (IF) can be detected via analysis of radiological changes. METHODS: Patients (n = 16) treated for 1 year (2011-12) at our hospital by T-SBRT at a total dose of 60 Gy in five fractions were examined retrospectively. Outcome and toxicity were recorded and were separately described for SPLN. CT scans were reviewed by a single radiologist. RESULTS: Of the 16 patients, 5 (31.3%) had primary lung malignancies, 10 (62.5%) had SPLN, and 1 case (6.3%) had isolated mediastinal metastasis of lung neoplasm. Pathological evidence was obtained for 72.2% of all lesions. The median radiological follow-up was 11 months (10.5 months for SPLN). For all cases, the 6- and 12-month survival rates were 100% and 77.7% (100% and 71.4%, respectively, for SPLN), and the 6- and 12-month locoregional control rates were 100% in all cases. 2 (12.5%) of 16 patients developed grade 3 late transient radiation pneumonitis following steroid therapy and 1 (6.3%) presented asymptomatic infiltrates comparable to OP opacities. CONCLUSION: T-SBRT seems to be safe and effective. ADVANCES IN KNOWLEDGE: Mild OP is likely associated with radiation-induced anomalies in the IF, identification of migrating opacities can help discern relapse of radiation-induced opacities.

Dosimetric comparison of different treatment modalities for stereotactic radiosurgery of meningioma.

BACKGROUND: The objective of this study was to compare the three most prominent systems for stereotactic radiosurgery in terms of dosimetric characteristics: the Cyberknife system, the Gamma Knife Perfexion and the Novalis system. METHODS: Ten patients treated for recurrent grade I meningioma after surgery using the Cyberknife system were identified; the Cyberknife contours were exported and comparative treatment plans were generated for the Novalis system and Gamma Knife Perfexion. Dosimetric values were compared with respect to coverage, conformity index (CI), gradient index (GI) and beam-on time (BOT). RESULTS: All three systems showed comparable results in terms of coverage. The Gamma Knife and the Cyberknife system showed significantly higher levels of conformity than the Novalis system (Cyberknife vs Novalis, p = 0.002; Gamma Knife vs Novalis, p = 0.002). The Gamma Knife showed significantly steeper gradients compared with the Novalis and the Cyberknife system (Gamma Knife vs Novalis, p = 0.014; Gamma Knife vs Cyberknife, p = 0.002) and significantly longer beam-on times than the other two systems (BOT = 66 ± 21.3 min, Gamma Knife vs Novalis, p = 0.002; Gamma Knife vs Cyberknife, p = 0.002). CONCLUSIONS: The multiple focal entry systems (Gamma Knife and Cyberknife) achieve higher conformity than the Novalis system. The Gamma Knife delivers the steepest dose gradient of all examined systems. However, the Gamma Knife is known to require long beam-on times, and despite worse dose gradients, LINAC-based systems (Novalis and Cyberknife) offer image verification at the time of treatment delivery.

Reconceptualizing Chinese state-owned entreprise in Africa : double embeddedness and localization in Ghana

Challenging the view of asymmetrical power relations between China and Africa, this thesis questions the "Chinese comparative advantages" (monolithic state power and economic advantages) of Chinese state-owned enterprises (SOEs) in Africa. It argues that the power dynamics between Chinese and African actors are dialectical and pluralistic, with localized social capital representing the true Chinese competitive advantage in Africa. Based on ethnographical fieldwork conducted in Ghana, this thesis shows that Chinese SOEs pursue their globalization in a double context - that of the deliberate "retreat" of the Chinese state, and more importantly, that of Ghanaian governance and society (characterized by political party patronage, extraversion dynamics, and worker agency). The trajectories of Chinese expatriates' expatriation/ social promotion and their SOEs' globalization/ localization are mutually influenced and reinforced. By cultivating local relationships and knowledge, a provincial Chinese SOE in Ghana can outperform a large Chinese central SOE, even if the latter has more support from the Chinese state. Moreover, the recent effort to build a "socially acceptable Chinese community" in Ghana has renewed the power dynamics between the Chinese state and the SOEs. All these observations provide for constructing a new perspective of Chinese SOEs in Africa - a "second-class" Chinese globalization - the SOEs may begin with few privileges, but promotion over time is possible. -- A contre pied des approches postulant des relations de pouvoir asymétriques entre la Chine et l'Afrique, cette thèse interroge les « avantages comparatifs chinois » (pouvoir de l'État monolithique et avantages économiques) des entreprises publiques chinoises (EPC) en Afrique. Elle soutient l'idée selon laquelle les dynamiques de pouvoir entre les acteurs chinois et africains est dialectique et pluraliste, et le capital social localisé étant le véritable avantage compétitif chinois en Afrique. S'appuyant sur un travail de terrain ethnographique au Ghana, cette thèse montre que les EPC poursuivent leur mondialisation dans un double contexte - celui de la «retraite» délibérée de l'État chinois, et, de façon plus importante, celui de la gouvernance et de la société ghanéennes (caractérisées par un clientélisme des partis politiques, une dynamique d'extraversion et le pouvoir de négociation des travailleurs). Les trajectoires d'expatriation / de promotion sociale des expatriés chinois et la mondialisation / localisation de leurs EPC s'influencent et se renforcent mutuellement. En cultivant des relations et des connaissances locales, une EPC provinciale au Ghana peut surpasser une grande EPC centrale, même si cette dernière reçoit plus de soutien de l'État chinois. En outre, les efforts récents visant à construire une «communauté chinoise socialement acceptable» au Ghana ont renouvelé la dynamique du pouvoir entre l'État chinois et les EPC. Ces observations permettent de construire une nouvelle perspective des EPC en Afrique - la globalisation chinoise de « deuxième classe » - les EPC peuvent débuter avec peu de privilèges, mais leur promotion reste possible avec le temps.