Search for Active-Sterile Antineutrino Mixing Using Neutral-Current Interactions with the NOvA Experiment.

This Letter reports results from the first long-baseline search for sterile antineutrinos mixing in an accelerator-based antineutrino-dominated beam. The rate of neutral-current interactions in the two NOvA detectors, at distances of 1 and 810 km from the beam source, is analyzed using an exposure of 12.51×10^{20} protons-on-target from the NuMI beam at Fermilab running in antineutrino mode. A total of 121 of neutral-current candidates are observed at the far detector, compared to a prediction of 122±11(stat.)±15(syst.) assuming mixing only between three active flavors. No evidence for ν[over ¯]_{µ}→ν[over ¯]_{s} oscillation is observed. Interpreting this result within a 3+1 model, constraints are placed on the mixing angles θ_{24}<25° and θ_{34}<32° at the 90% C.L. for 0.05 eV^{2}≤Δm_{41}^{2}≤0.5 eV^{2}, the range of mass splittings that produces no significant oscillations at the near detector. These are the first 3+1 confidence limits set using long-baseline accelerator antineutrinos.

Dentists of tomorrow 2020: An analysis of the results of the 2020 ADEA Survey of U.S. Dental School Seniors.

PURPOSE/OBJECTIVES: This study examines the journey of U.S. dental schools' predoctoral senior class of 2020, from the influences on and their motivations to pursue careers in dentistry, aspects of their dental school experiences, to plans upon graduation and the investment in their careers. METHODS: The study is an analysis of the results of the ADEA Survey of Dental School Seniors, 2020 Graduating Class. Each year, ADEA surveys senior predoctoral students from the accredited U.S. dental schools. Whenever feasible, the answers of the survey respondents from the 2020 class were compared with their 2015 counterparts. RESULTS: The analysis revealed that 46% of the 2020 respondents decided to become a dentist before going to undergraduate college, more than the proportion of those deciding while in college (42%). When it comes to preparedness to practice dentistry, the responses indicated a high level of readiness to go into the profession. Seventy-seven percent of survey participants reported the COVID-19 pandemic did not affect their plans after graduation. Between 2015 and 2020, the share of survey respondents who planned to go into advanced dental education immediately after graduation increased from 35% to 40%. Almost a third of the 2020 respondents who planned to go into private practice immediately upon graduation intended to join a Dental Service Organization (DSO). Grants and scholarships represented a higher share of the average funding for dental education for the 2020 respondents than five years ago. The share of respondents expecting to graduate without any loans to finance their dental degrees and predental education (educational debt) increased significantly, from 12% in 2015 to 17% in 2020. CONCLUSION(S): This research shows that during these uncertain times, U.S. dental schools continued their mission to train and graduate oral health professionals fully prepared to go into the profession.

Massa mediastinal anterior em jovem: aneurisma de artéria descendente anterior como diagnóstico diferencial, um relato de caso / Anterior mediastinal mass in a young woman: anterior descending artery aneurysm as a differential diagnosis, a case report

INTRODUÇÃO: As massas mediastinais anteriores (MMA) são as mais comuns do grupo de tumores do mediastino. O timoma é a forma mais comum, já os tumores de células germinativas mediastinais, os quais caracterizam um grupo heterogêneo de neoplasias, correspondem a cerca de 15%, dentre eles o teratoma. O presente caso, no entanto, elucida a importância do diagnóstico diferencial para essas MMAs. MÉTODOS: Relato de caso com análise de prontuário. RESULTADOS: Masculino, 14 anos, antecedentes patológicos de pseudoaneurisma de artéria mamária interna esquerda embolizado aos 8 anos e aftas orais recorrentes. Relatou dor anginosa há 5 meses de média intensidade sem irradiação, com melhora à analgesia simples e repouso. Procurou atendimento médico devido à piora dos sintomas há um mês, acompanhados de tosse seca, dispneia e febre. Admitido com eletrocardiograma normal e hipótese diagnóstica de pneumonia complicada com derrame pleural a esquerda, o paciente realizou ecocardiograma transtorácico (ECOTT) com evidência de derrame pericárdico leve. Em uso de antibioticoterapia com pesquisa infecciosa negativa realizou angiotomografia de tórax, a qual evidenciou tromboembolismo pulmonar à direita e massa na região apical do ventrículo direito (VD) de 3,4cm. Encaminhado a um hospital terciário para investigação realizou novo ECOTT: imagem heterogênea na porção médio-apical do VD, medindo 5,1 cm x 4,2 cm, podendo corresponder a trombo ou massa. Seguiu investigação com ressonância nuclear magnética cardíaca que mostrou massa intramiocárdica, sendo aventado hipótese de teratoma. Heart Team optou por prosseguir investigação através de angiotomografia de coronárias, a qual mostrou em artéria descendente anterior (ADA) aneurisma sacular em terço distal medindo 63x53mm. A cinecoronarioangiografia subsequente evidenciou ADA ocluída no terço médio com aspecto de compressão extrínseca. Equipe manteve tratamento conservador e investigação diagnóstica etiológica em conjunto com reumatologia, cuja suspeita firmou-se em Doença de Behçet (DB), desordem inflamatóri multisistêmica. CONCLUSÃO: O caso em questão partiu de investigação de MMAs com análise de imagens criteriosa, a qual evidenciou aneurisma de ADA, com hipótese no contexto de DB, cujo acometimento de coronárias é raro. Dessa maneira, houve impacto profundo na opção terapêutica. Dessa forma demostramos a importância do estudo de imagem detalhado para diagnóstico de massas cardíacas. O paciente do caso recebeu pulsoterapia com corticoesteróide e seguiu em acompanhamento com reumatologia e cardiologia de hospital terciário.

Role of selenium addition to CdZnTe matrix for room-temperature radiation detector applications.

Because of its ideal band gap, high density and high electron mobility-lifetime product, cadmium zinc telluride (CdZnTe or CZT) is currently the best room-temperature compound-semiconductor X- and gamma-ray detector material. However, because of its innate poor thermo-physical properties and above unity segregation coefficient for Zn, the wide spread deployment of this material in large-volume CZT detectors is still limited by the high production cost. The underlying reason for the low yield of high-quality material is that CZT suffers from three major detrimental defects: compositional inhomogeneity, high concentrations of dislocation walls/sub-grain boundary networks and high concentrations of Te inclusions/precipitates. To mitigate all these disadvantages, we report for the first time the effects of the addition of selenium to the CZT matrix. The addition of Se was found to be very effective in arresting the formation of sub-grain boundaries and its networks, significantly reducing Zn segregation, improving compositional homogeneity and resulting in much lower concentrations of Te inclusions/precipitates. Growth of the new quaternary crystal Cd1-xZnxTe1-ySey (CZTS) by the Traveling Heater Method (THM) is reported in this paper. We have demonstrated the production of much higher yield according to its compositional homogeneity, with substantially lower sub-grain boundaries and their network, and a lower concentration of Te inclusions/precipitates.

Morphological and functional imaging of neck paragangliomas.

OBJECTIVE: To review the optimal techniques for localization and characterization of neck paragangliomas (PGL). MATERIAL AND METHODS: Systematic review of the literature from the PubMed/Medline database. RESULTS: Neck PGL are hypervascular tumours essentially arising from paraganglionic tissue situated at the carotid bifurcation (carotid body) and along the vagus nerve. Morphological and functional imaging are indicated to confirm the diagnosis, identify multifocal disease and for local and regional staging. MR angiography is the noninvasive technique of choice. CT scan and especially CT angiography are excellent alternatives for diagnosis and staging. Conventional arteriography remains useful preoperatively for embolization and occlusion tests. Functional imaging allows localization and characterization of PGLs. Somatostatin receptor scintigraphy (SRS) was the reference imaging technique for staging of sporadic PGLs. The indications for PET imaging have been extended over recent years in parallel with the development of new tracers such as [18F]-FDOPA PET or 68Gallium-labelled DOTA peptides. 68Gallium-labelled DOTA peptides has become the first-line imaging modality in the evaluation of cervical PGLs, regardless of the genetic background. CONCLUSION: Morphological and functional imaging is essential for the staging of neck PGL.

DNA microarray analysis and functional profile of pituitary transcriptome under core-clock protein BMAL1 control.

Although it is known to contain five cell types that synthesize and release hormones with a circadian pattern, the pituitary gland is poorly characterized as a circadian oscillator. By a differential microarray analysis, 252 genes were found to be differentially expressed in pituitaries from Bmal1(-/-) knockout versus wild-type mice. By integrative analyses of the data set with the Annotation, Visualization, and Integrated Discovery (DAVID) Bioinformatics Resources annotation analysis system, pituitary genes with altered expression in Bmal1(-/-) mice were dispatched among functional categories. Clusters of genes related to signaling and rhythmic processes as well as transcription regulators, in general, were found enriched in the data set, as were pathways such as circadian rhythm, transforming growth factor ß (TGFß) signaling, valine, leucine, and isoleucine degradation, and peroxisome proliferator-activated receptor (PPAR) signaling pathways. Gene Ontology term overrepresentation analyses revealed significant enrichment for genes involved in 10 key biological processes. To determine whether genes with altered expression in Bmal1(-/-) mice were actually circadian genes, we further characterized in the mouse pituitary gland the daily pattern of some of these genes, including core-clock genes. Core-clock genes and genes selected from three identified overrepresented biological processes, namely, hormone metabolic process, regulation of transcription from RNA polymerase II promoter, and cell adhesion, displayed a rhythmic pattern. Given the enrichment in genes dedicated to cell adhesion and their daily changes in the pituitary, it is hypothesized that cell-cell interactions could be involved in the transmission of information between endocrine cells, allowing rhythmic hormone outputs to be controlled in a temporally precise manner.

[Complications from sinonasal surgery]. / Complications de la chirurgie rhinosinusienne.

Endoscopic sinonasal surgery is the main procedure in sinonasal pathology. Complications are rare but potentially severe given the close relationship between the nasal cavities and sinuses and the orbit, skull base and carotid arteries. The different types of surgeries along with the mechanisms of injury, presenting signs and symptoms, and imaging features of the different surgical complications will be reviewed. We will also review the anatomical variants increasing the surgical risk that radiologists should describe on preoperative imaging studies.

Increase in polysialyltransferase gene expression following LTP in adult rat dentate gyrus.

Neural cell adhesion molecule (NCAM) is frequently associated with polysialic acid (PSA), and its function is highly dependent on this polysialylation. PSA-NCAM plays an important role in synaptic plasticity in the hippocampus. STX and PST are the enzymes responsible for NCAM polysialylation. We investigated whether unilateral long-term potentiation (LTP) induction in vivo, in adult rat dentate gyrus (DG), triggered NCAM polysialylation by STX and PST produced in the hippocampus. We found that levels of STX and PST mRNA increased strongly since the early stage of hippocampal LTP and remained high during the maintenance of DG-LTP for 4 h. This rapid increase in polysialyltransferase gene expression occurred in both the hippocampi, probably resulting from bilateral LTP induction by strong unilateral HFS. Thus, LTP triggers interhemispheric molecular changes in the hippocampal network. This study is the first to describe the effects of LTP induction and maintenance on polysialyl-transferases in vivo. Our findings suggest that hippocampal synaptic remodeling requires NCAM polysialylation.

[Vertebroplasty: 10 years clinical and radiological follow-up]. / Vertébroplastie: évaluation clinique et radiologique à plus de dix ans.

OBJECTIVES: Ten years follow-up of the first patients treated with percutaneous vertebroplasty. PATIENT AND METHODS: Eighteen patients were retrospectively reviewed having undergone vertebroplasty in our centre between 1989 and 1998. Eight were treated for angioma, eight for osteoporotic compression and two followed for myeloma. They all underwent clinical and radiological evaluation in 2007 (standard X-rays, CT scan and MRI). These examinations were compared to prior baseline pre- and post-therapeutic images. RESULTS: Radiological characteristic of cement remained unchanged in the long term and there was no modification of anatomical structures in contact with it. Even if the distribution of cement was asymmetrical there was no fracture of the treated vertebras at distance. Degenerative changes of discs facing the vertebroplasty were not more pronounced than for distant discs. We found no significant signal or density anomaly of disc in contact direct with cement. 38.8 % of the patients presented new fractures (n=30). Seventy percent of the fractures were multiple and contiguous. In the long term, all patients reported improvement of pain after the procedure. CONCLUSION: In our series, we found a good stability of treatment over time. This study shows the long-term safety of percutaneous acrylic vertebroplasty, in particular harmlessness of cement for bone and discs in contact.

[Arterial spin labeling: state of the art]. / Etat de l'art sur le Marquage de Spin Artériel ou ASL.

UNLABELLED: Arterial spin labeling (ASL) perfusion MR imaging is a technique by which water from circulating arterial blood is magnetically labeled and acts as a diffusible tracer allowing non-invasive measurement of cerebral blood flow. In this paper, the technique and current applications in neuroimaging will be reviewed. CURRENT STATUS: First, the technical principles of ASL will be reviewed and both available techniques (continuous and pulsed ASL) explained. A review of the literature will demonstrate advances with the techniques of ASL and its clinical impact. Clinical research involves normal volunteers and patients with ischemic and tumoral pathologies. CONCLUSION: Recent technical advances have improved the sensitivity of ASL perfusion MR imaging. The routine clinical use of ASL at 3.0 Tesla should increase over the next few years.

Masking technique for coating thickness control on large and strongly curved aspherical optics.

We discuss a method to control the coating thickness deposited onto large and strongly curved optics by ion beam sputtering. The technique uses an original design of the mask used to screen part of the sputtered materials. A first multielement mask is calculated from the measured two-dimensional coating thickness distribution. Then, by means of an iterative process, the final mask is designed. By using such a technique, it has been possible to deposit layers of tantalum pentoxide having a high thickness gradient onto a curved substrate 500 mm in diameter. Residual errors in the coating thickness profile are below 0.7%.

[Bithalamic infarct: is there an evocative aspect? Radioclinical study]. / Accident vasculaire ischémique bithalamique: existe-t-il un tableau évocateur ? Etude radioclinique.

INTRODUCTION: Bithalamic paramedian infarcts are uncommon. This stroke results in a complex clinical syndrome. CASE REPORT: We report four cases of bithalamic paramedian infarcts with a presumed mechanism of occlusion of a single thalamic paramedian artery. DISCUSSION: This normal anatomic variant corresponds to an asymmetrical common trunk for the two thalamosubthalamic paramedian arteries arising from a P1 segment (type IIb in the G. Percheron classification dating from 1977). A literature analysis (from 1985 to 2006) allowed us to identify the most widely reported clinical signs. Four main clinical findings are described: vertical gaze palsy (65%), memory impairment (58%), confusion (53%) and coma (42%). We also found these symptoms in our patients but rarely associated; however, all four patients had exhibited episodes of drowsiness. In this article, we discuss the anatomy-function correlation responsible for such clinical variability. CONCLUSION: Clinicians should be aware of this diagnosis to better understand the imaging results which provide confirmation. Although the literature describes frequently severe consciousness disorders such as coma, this diagnosis must also be considered in patients presenting a simple fluctuation of consciousness, e.g. hypersomnia.

Efficient and tolerant resonant grating coupler for multimode optical interconnections.

More than 60% overall coupling efficiency is achieved in the demonstrator of an optical interconnect comprising an input grating coupler, a multimode slab waveguide section and an output grating coupler. The grating coupling strength is enhanced by means of a leaky mode resonance. The efficiency of the resonant grating coupler compares favourably with the performancs reported on mirror inserts.

Pituitary transcription factors: from congenital deficiencies to gene therapy.

Despite the existence of interspecies phenotypic variability, animal models have yielded valuable insights into human pituitary diseases. Studies on Snell and Jackson mice known to have growth hormone, prolactin and thyroid-stimulating hormone deficiencies involving the hypoplastic pituitary gland have led to identifying alterations of the pituitary specific POU homeodomain Pit-1 transcription factor gene. The human phenotype associated with rare mutations in this gene was found to be similar to that of these mice mutants. Terminal differentiation of lactotroph cells and direct regulation of the prolactin gene both require interactions between Pit-1 and cell type specific partners, including panpituitary transcriptional regulators such as Pitx1 and Pitx2. Synergistic activation of the prolactin promoter by Pitx factors and Pit-1 is involved not only in basal condition, but also in responsiveness to forskolin, thyrotrophin-releasing-hormone and epidermal growth factor. In corticotroph cells, Pitx1 interacts with Tpit. Tpit mutations have turned out to be the main molecular cause of neonatal isolated adrenocorticotrophin deficiency. This finding supports the idea that Tpit plays an essential role in the differentiation of the pro-opiomelanocortin pituitary lineage. The effects of Pit-1 are not restricted to hormone gene regulation because this factor also contributes to cell division and protects the cell from programmed cell death. Lentiviral vectors expressing a Pit-1 dominant negative mutant induced time- and dose-dependent cell death in somatotroph and lactotroph adenomas in vitro. Gene transfer by lentiviral vectors should provide a promising step towards developing an efficient specific therapeutic approach by which a gene therapy programme for treating human pituitary adenomas could be based.

Identification of thyroglobulin domain(s) involved in cell-surface binding and endocytosis.

Thyroglobulin (Tg) binds to cell surfaces through various binding sites of high, moderate and low affinity. We have previously shown that binding with low to moderate affinity is pH dependent, selective, but not tissue specific. To identify the regions of Tg involved in this cell surface binding, we studied the binding of (125)I-labeled cyanogen bromide peptides from human Tg to cell surfaces of thyroid cells (inside-out follicles) and of CHO cells. Electrophoretic analysis of cell homogenates after binding of native or of reduced and alkylated (125)I-labeled peptides showed that three peptides, P1, P2 and P3, were always associated with the cells. Sequence analysis allowed the identification of P1 (Ser-2445 to Met-2596 or Met-2610) and P2 (Phe-2156 to Met-2306). P3 proved to be a mixture of several peptides among which two were identified: P3-1 (Cys-1306 to Met-1640) and P3-2 (Cys-2035 to Met-2413) which includes P2. P1, P2 and P3-2 are entirely (P1) or partly (P2 and P3-2) located in the C-terminal domain of Tg homologous with acetylcholinesterase. The smallest peptides, P1 and P2, were purified by preparative electrophoresis. They both displayed strong binding properties towards cell surfaces. Inhibition experiments of (125)I-labeled Tg binding by P1 or P2 indicated that they were involved in Tg binding to cell surfaces. All the other peptides tested for their binding abilities were either not or only poorly involved in Tg binding to cell surfaces, which suggested that P1 and P2 are major Tg sites of binding to cell surfaces. These two peptides are not involved in the binding of Tg to the known Tg 'receptors' described in the literature, to which recycling, transcytosis and regulation functions have been ascribed. Thus they are potential tools to identify cell surface components involved in the process of Tg endocytosis leading to lysosomal degradation.

Alternatively spliced form of human thyroperoxidase, TPOzanelli: activity, intracellular trafficking, and role in hormonogenesis.

Thyroperoxidase (TPO), a type I transmembrane heme containing glycoprotein, catalyzes iodide organification and thyroid hormone synthesis. One of the two main alternatively spliced forms of this enzyme, TPOzanelli, which is present in Graves's disease thyroid tissue, has a cytoplasmic domain completely modified. In the first stage of this study, the results of RT-PCR experiments showed that the TPOzanelli mRNA is present in normal thyroid tissue. We then generated CHO cell lines expressing the wild-type TPO (TPO1) and the alternatively spliced form TPOzanelli. Upon investigating a panel of 12 mAbs directed against the extracellular domain of TPO1 and sera from patients with a high titer of TPO autoantibodies, we observed that (i) the three-dimensional structure of this domain is similar in both isoforms; (ii) the autoantibodies recognize TPOzanelli as well as TPO1. The results of pulse chase and cell surface biotinylation experiments showed that the TPOzanelli has a shorter half-life (7 versus 11 h) and is expressed at the cell surface in lesser amounts than TPO1 (7 versus 15%). The total enzymatic activity and cell surface activity were determined in CHO cells expressing TPO1 and TPOzanelli, and TPO1 and TPOzanelli were found to have similar levels of activity. It was established that approximately 20% of the TPO purified from a Graves' disease thyroid gland was precipitated by polyclonal antibodies directed against a specific part of the cytoplasmic tail of TPOzanelli. This confirmed that the protein corresponding to the mRNA is present in the thyroid tissue. All in all, these results indicate that TPOzanelli can be expected to play a role in thyroid hormone synthesis and in thyroid autoimmunity.

Loratadine and terfenadine interaction with nefazodone: Both antihistamines are associated with QTc prolongation.

BACKGROUND AND OBJECTIVE: Nefazodone inhibits CYP3A; therefore coadministration with CYP3A substrates such as terfenadine or loratadine may result in increased exposure to these drugs. A potential pharmacodynamic consequence is electrocardiographic QTc prolongation, which has been associated with torsade de pointes cardiac arrhythmia. Therefore a clinical pharmacokinetic-pharmacodynamic evaluation of this potential interaction was conducted. METHODS: A randomized, double-blind, double-dummy, parallel group, multiple-dose design was used. Healthy men and women who were given doses of 60 mg of terfenadine every 12 hours, 20 mg of loratadine once daily, and 300 mg of nefazodone every 12 hours were studied. Descriptive pharmacokinetics (time to maximum concentration, maximum concentration, and area under the plasma concentration-time curve) were used for the examination of interactions among the respective parent drugs and metabolites. QTc prolongation (mean value over the dosing interval) was the pharmacodynamic parameter measured. Kinetic and dynamic analysis was used for the examination of pooled concentration and QTc data with the use of a linear model. RESULTS: Concomitant nefazodone treatment markedly increased the dose interval area under the plasma concentration-time curve of both terfenadine (mean value, 17.3 +/- 8.5 ng. mL/h versus 97.4 +/- 48.9 ng. mL/h; P <.001) and carboxyterfenadine (mean value, 1.69 +/- 0.48 microg. h/mL versus 2.88 +/- 0.53 microg. h/mL; P <.001) and moderately increased the dose interval area under the plasma concentration-time curve of both loratadine (mean value, 31.5 +/- 27.9 ng. h/mL versus 43.7 +/- 25.9 ng. h/mL; P <.014) and descarboethoxyloratadine (mean value, 73.4 +/- 54.9 ng. h/mL versus 81.9 +/- 26.2 ng. h/mL; P <.002). The mean QTc was unchanged with terfenadine alone; however, it was markedly prolonged with concomitant nefazodone and terfenadine (mean [90% confidence interval] prolongation 42.4 ms [34.2, 50.6 ms]; P <.05). Similarly, the mean QTc was unchanged with loratadine alone; however, it was prolonged with concomitant nefazodone and loratadine (21.6 ms [13.7, 29.4 ms]; P <.05). Nefazodone alone did not change mean QTc. QTc was positively correlated with terfenadine plasma concentration (r (2) = 0.21; P =.0001). Similarly, QTc was positively correlated with loratadine plasma concentration (r (2) = 0.056; P =.0008) but with a flatter slope. There was no relationship between QTc and nefazodone plasma concentration during treatment with nefazodone alone (r (2) = 0.002, not significant). CONCLUSIONS: In healthy men and women, concomitant nefazodone treatment at a therapeutic dose increases exposure to both terfenadine and carboxyterfenadine. This increased exposure is associated with marked QTc prolongation, which is correlated with terfenadine plasma concentration. A similar interaction occurs with loratadine, although it is of lesser magnitude. Concomitant administration of nefazodone with terfenadine may have predisposed individuals to the arrhythmia associated with QTc prolongation, torsade de pointes, when terfenadine was available for clinical use. However, a new finding is that in the context of higher than clinically recommended daily doses (20 mg) of loratadine concomitant administration with a metabolic inhibitor such as nefazodone can also result in QTc prolongation.

Role of complex asparagine-linked oligosaccharides in the expression of a functional thyrotropin receptor.

To evaluate the functional role of complex asparagine-linked oligosaccharides of the human thyrotropin receptor (TSHR), a Chinese hamster ovary cell line (JP09) and a K562 cell line (K562-TSHR) expressing this receptor were treated with deoxymannojirimycin (dMM), a mannosidase I inhibitor. dMM blocks the formation of complex-type structures and leads to the formation of high-mannose-type structures. Treatment of cells with dMM led to a decrease in the number of thyrotropin (TSH)-binding sites at the cell surface. Detection of the TSHR at the cell surface using a monoclonal antibody directed against the A subunit showed that this decrease was not due to a decrease in the number of TSHRs expressed at the cell surface. However the recognition of TSHR by a monoclonal antibody directed against the C peptide was greatly decreased. On immunoblotting, after deglycosylation using peptide N-glycanase F, the A subunit was visualized as a doublet (36 and 41 kDa). In control cells the species of higher molecular mass was more abundant whereas after dMM treatment the species of lower molecular mass became more abundant. This difference in molecular mass between the two peptides is compatible with the removal of the C peptide. In conclusion, the results show that inhibition of complex-type structure formation leads to (i) an incapacity for TSHR to bind TSH, without affecting its intracellular transport and (ii) an increase of TSHR susceptibility to proteases that remove the C peptide. We then hypothesized that removal of the C peptide could contribute to the formation of a non-functional TSHR.

Degradation of human thyroperoxidase in the endoplasmic reticulum involves two different pathways depending on the folding state of the protein.

Human thyroperoxidase (hTPO), a type I transmembrane glycoprotein, plays a key role in thyroid hormone synthesis. In a previous paper (Fayadat, L., Niccoli, P., Lanet, J., and Franc, J. L. (1998) Endocrinology 139, 4277-4285) we established that after the synthesis, only 15-20% of the hTPO molecules were recognized by a monoclonal antibody (mAb15) directed against a conformational structure and that only 2% were able to reach the cell surface. In the present study using pulse-chase experiments in the presence or absence of protease inhibitors followed by immunoprecipitation procedures with monoclonal antibodies recognizing unfolded or partially folded hTPO forms we show that: (i) unfolded hTPO forms are degraded by the proteasome and (ii) partially folded hTPO forms are degraded by other proteases. It was also established upon incubating endoplasmic reticulum (ER) membranes in vitro that the degradation of the partially folded hTPO was carried out by serine and cysteine integral ER membrane proteases. These data provide valuable insights into the quality control mechanisms whereby the cells get rid of misfolded or unfolded proteins. Moreover, this is the first study describing a protein degradation process involving two distinct degradation pathways (proteasome and ER cysteine/serine proteases) at the ER level, depending on the folding state of the protein.

Calnexin and calreticulin binding to human thyroperoxidase is required for its first folding step(s) but is not sufficient to promote efficient cell surface expression.

Human thyroperoxidase (hTPO) is a type I transmembrane-bound heme-containing glycoprotein that catalyzes the synthesis of thyroid hormones. In a previous study we stably expressed hTPO in Chinese hamster ovary cells and observed that after the synthesis, only 20% of the hTPO molecules were recognized by a monoclonal antibody (mAb 15) directed against a conformational structure, and that only 2% were able to reach the cell surface. In the present study it was proposed to determine how calnexin (CNX) and calreticulin (CRT) contribute to the folding of hTPO. Sequential immunoprecipitation was performed using anti-CNX or anti-CRT followed by anti-hTPO antibodies, and the results showed that CNX and CRT were associated with hTPO. Inhibiting the interactions between CNX or CRT and hTPO using castanospermine greatly reduced the first step(s) in the hTPO folding process. Under these conditions, the half-life of this enzyme was greatly reduced (2.5 vs. 17 h in the control experiments), and hTPO was degraded via the proteasome pathway. This reduced the rate of hTPO transport to the cell surface. Overexpression of CNX or CRT into the hTPO-CHO cells was found to enhance the first hTPO folding step(s) by 20-60%, but did not increase the level of hTPO present at the cell surface. All in all, these findings provide evidence that CNX and CRT are crucial to the first step(s) in hTPO folding, but that interactions with other molecular chaperones are required for the last folding steps to take place.