(68)Ga-labeled superparamagnetic iron oxide nanoparticles (SPIONs) for multi-modality PET/MR/Cherenkov luminescence imaging of sentinel lymph nodes.

The aim of this study was to develop (68)Ga-SPIONs for use as a single contrast agent for dynamic, quantitative and high resolution PET/MR imaging of Sentinel Lymph Node (SLN). In addition (68)Ga enables Cherenkov light emission which can be used for optical guidance during resection of SLN. SPIONs were labeled with (68)Ga in ammonium acetate buffer, pH 5.5. The labeling yield and stability in human serum were determined using instant thin layer chromatography. An amount of 0.07-0.1 mL (~5-10 MBq, 0.13 mg Fe) of (68)Ga-SPIONs was subcutaneously injected in the hind paw of rats. The animals were imaged at 0-3 h and 25 h post injection with PET/CT, 9.4 T MR and CCDbased Cherenkov optical systems. A biodistribution study was performed by dissecting and measuring the radioactivity in lymph nodes, kidneys, spleen, liver and the injection site. The labeling yield was 97.3 ± 0.05% after 15 min and the (68)Ga-SPIONs were stable in human serum. PET, MR and Cherenkov luminescence imaging clearly visualized the SLN. Biodistribution confirmed a high uptake of the (68)Ga-SPIONs within the SLN. We conclude that generator produced (68)Ga can be labeled to SPIONs. Subcutaneously injected (68)Ga-SPIONs can enhance the identification of the SLNs by combining sensitive PET and high resolution MR imaging. Clinically, hybrid PET/MR cameras are already in use and (68)Ga-SPIONs have a great potential as a single-dose, tri-modality agent for diagnostic imaging and potential Cherenkov luminescent guided resection of SLN

Prevalence and predictors of HIV-related stigma among institutional- and community-based caregivers of orphans and vulnerable children living in five less-wealthy countries

<p>Abstract</p> <p>Background</p> <p>In the face of the HIV/AIDS epidemic that has contributed to the dramatic increase in orphans and abandoned children (OAC) worldwide, caregiver attitudes about HIV, and HIV-related stigma, are two attributes that may affect caregiving. Little research has considered the relationship between caregiver attributes and caregiver-reported HIV-related stigma. In light of the paucity of this literature, this paper will describe HIV-related stigma among caregivers of OAC in five less wealthy nations.</p> <p>Methods</p> <p>Baseline data were collected between May 2006 through February 2008. The sample included 1,480 community-based and 192 institution-based caregivers. Characteristics of the community-based and institution-based caregivers are described using means and standard deviations for continuous variables or counts and percentages for categorical variables. We fit logistic regression models, both for the full sample and separately for community-based and institution-based caregivers, to explore predictors of acceptance of HIV.</p> <p>Results</p> <p>Approximately 80% of both community-based and institution-based caregivers were female; and 84% of institution-based caregivers, compared to 66% of community-based caregivers, said that they would be willing to care for a relative with HIV. Similar proportions were reported when caregivers were asked if they were willing to let their child play with an HIV-infected child. In a multivariable model predicting willingness to care for an HIV-infected relative, adjusted for site fixed effects, being an institution-based caregiver was associated with greater willingness (less stigma) than community-based caregivers. Decreased willingness was reported by older respondents, while willingness increased with greater formal education. In the adjusted models predicting willingness to allow one's child to play with an HIV-infected child, female gender and older age was associated with less willingness. However, willingness was positively associated with years of formal education.</p> <p>Conclusions</p> <p>The caregiver-child relationship is central to a child's development. OAC already face stigma as a result of their orphaned or abandoned status; the addition of HIV-related stigma represents a double burden for these children. Further research on the prevalence of HIV-related acceptance and stigma among caregivers and implications of such stigma for child development will be critical as the policy community responds to the global HIV/AIDS orphan crisis.</p

A puzzle of life: crafting ribosomal subunits

The biogenesis of eukaryotic ribosomes is a complicated process during which the transcription, modification, folding, and processing of the rRNA is coupled with the ordered assembly of ∼80 ribosomal proteins (r-proteins). Ribosome synthesis is catalyzed and coordinated by more than 200 biogenesis factors as the preribosomal subunits acquire maturity on their path from the nucleolus to the cytoplasm. Several biogenesis factors also interconnect the progression of ribosome assembly with quality control of important domains, ensuring that only functional subunits engage in translation. With the recent visualization of several assembly intermediates by cryoelectron microscopy (cryo-EM), a structural view of ribosome assembly begins to emerge. In this review we integrate these first structural insights into an updated overview of the consecutive ribosome assembly steps

Structural and functional study of the Rpf2/Rrs1 complex in ribosome biogenesis

La biogenèse des ribosomes est un processus complexe qui implique la production et l'assemblage de 4 ARN et d'environ 80 protéines. Chez l'Homme, la production des deux sous-unités ribosomiques débute dans le nucléole par la synthèse par l'ARN polymérase I d'un long transcrit contenant les séquences des ARN ribosomiques 5.8S, 18S et 25S, qui s'associe de manière co-transcriptionnelle à des protéines ribosomiques et à des facteurs d'assemblage. Le quatrième ARN ribosomique, l'ARNr 5S est transcrit séparément par l'ARN polymérase III, et s'associe avec les protéines ribosomiques Rpl5 et Rpl11 en dehors du ribosome. Ce sous-complexe, appelé particule 5S, est ensuite intégré au sein de la grande sous-unité. La particule 5S est également impliquée dans le contrôle de la prolifération cellulaire. En effet, en cas de dé-régulation de la biogenèse du ribosome, la particule 5S s'accumule dans le nucléoplasme et interagit directement avec l'ubiquitine-ligase MDM2, provoquant la stabilisation du suppresseur de tumeur p53. L'objectif principal de ma thèse est d'étudier le rôle des facteurs d'assemblage Rpf2 et Rrs1 dans la biogenèse du ribosome. Ces protéines assurent deux fonctions distinctes : elles sont requises pour l'association de la particule 5S avec la sous-unité pré-60S, et stimulent la transcription des ARNr par l'ARN polymérase I. Elles sont donc impliquées dans deux événements fondamentaux qui conditionnent les capacités de prolifération cellulaire. La combinaison d'études structurales par cristallographie aux rayons X, et d'études d'interactions protéine-ARN in vitro et in vivo, m'ont permis de mieux appréhender le rôle du complexe Rpf2/Rrs1 dans l'intégration de la particule 5S et dans la maturation de la grande sous-unité. J'ai également étudié le rôle du complexe Rpf2/Rrs1 dans la régulation de la transcription des ARNr, en caractérisant ses interactions avec la polymérase I.Ribosome Biogenesis is a complex process that requires the production and the correct assembly of the 4 rRNA with more than 80 proteins. Ribosome biogenesis starts by the transcription of a pre-RNA precursor in the nucleolus. Three of the four ribosomal RNAs, the 5.8S, 18S, and 25S rRNAs, are cotranscribed as a single 35S precursor by polymerase I. This precursor is cotranscriptionally modified, folded, cleaved, and assembled with both ribosomal proteins and non-ribosomal factors to generate the mature ribosomes. The fourth rRNA, the 5S rRNA, is transcribed by RNA polymerase III and is assembled into the 5S particle, containing ribosomal proteins Rpl5 and Rpl11, prior to its incorporation into preribosomes. In mammals, the 5S RNP is also a central regulator of the homeostasis of the tumor suppressor p53 The main objective of my thesis was to understand the precise roles of the two assembly factors Rpf2 and Rrs1 in ribosome biogenesis. These proteins have two distinctive functions : Rpf2 and Rrs1 are required for the 5S particle incorporation into the large subunit, and stimulate the rRNA transciption by polymerase I. Using a combination of structural studies by X-Ray crystallography and biochemical approaches as in vitro and in vivo methods to study proteins-RNA interactions, I was able to uncover the function of the Rpf2/Rrs1 dimer in the maturation of the large subunit through the recruitment of the 5S particle. I also studied the function of Rpf2 and Rrs1 in the rRNA transcription regulation, by characterizing physical connection with polymerase I subunits

Étude structurale et fonctionnelle du complexe Rpf2/Rrs1 impliqué dans la biogenèse du ribosome

Ribosome Biogenesis is a complex process that requires the production and the correct assembly of the 4 rRNA with more than 80 proteins. Ribosome biogenesis starts by the transcription of a pre-RNA precursor in the nucleolus. Three of the four ribosomal RNAs, the 5.8S, 18S, and 25S rRNAs, are cotranscribed as a single 35S precursor by polymerase I. This precursor is cotranscriptionally modified, folded, cleaved, and assembled with both ribosomal proteins and non-ribosomal factors to generate the mature ribosomes. The fourth rRNA, the 5S rRNA, is transcribed by RNA polymerase III and is assembled into the 5S particle, containing ribosomal proteins Rpl5 and Rpl11, prior to its incorporation into preribosomes. In mammals, the 5S RNP is also a central regulator of the homeostasis of the tumor suppressor p53 The main objective of my thesis was to understand the precise roles of the two assembly factors Rpf2 and Rrs1 in ribosome biogenesis. These proteins have two distinctive functions : Rpf2 and Rrs1 are required for the 5S particle incorporation into the large subunit, and stimulate the rRNA transciption by polymerase I. Using a combination of structural studies by X-Ray crystallography and biochemical approaches as in vitro and in vivo methods to study proteins-RNA interactions, I was able to uncover the function of the Rpf2/Rrs1 dimer in the maturation of the large subunit through the recruitment of the 5S particle. I also studied the function of Rpf2 and Rrs1 in the rRNA transcription regulation, by characterizing physical connection with polymerase I subunits.La biogenèse des ribosomes est un processus complexe qui implique la production et l'assemblage de 4 ARN et d'environ 80 protéines. Chez l'Homme, la production des deux sous-unités ribosomiques débute dans le nucléole par la synthèse par l'ARN polymérase I d'un long transcrit contenant les séquences des ARN ribosomiques 5.8S, 18S et 25S, qui s'associe de manière co-transcriptionnelle à des protéines ribosomiques et à des facteurs d'assemblage. Le quatrième ARN ribosomique, l'ARNr 5S est transcrit séparément par l'ARN polymérase III, et s'associe avec les protéines ribosomiques Rpl5 et Rpl11 en dehors du ribosome. Ce sous-complexe, appelé particule 5S, est ensuite intégré au sein de la grande sous-unité. La particule 5S est également impliquée dans le contrôle de la prolifération cellulaire. En effet, en cas de dé-régulation de la biogenèse du ribosome, la particule 5S s'accumule dans le nucléoplasme et interagit directement avec l'ubiquitine-ligase MDM2, provoquant la stabilisation du suppresseur de tumeur p53. L'objectif principal de ma thèse est d'étudier le rôle des facteurs d'assemblage Rpf2 et Rrs1 dans la biogenèse du ribosome. Ces protéines assurent deux fonctions distinctes : elles sont requises pour l'association de la particule 5S avec la sous-unité pré-60S, et stimulent la transcription des ARNr par l'ARN polymérase I. Elles sont donc impliquées dans deux événements fondamentaux qui conditionnent les capacités de prolifération cellulaire. La combinaison d'études structurales par cristallographie aux rayons X, et d'études d'interactions protéine-ARN in vitro et in vivo, m'ont permis de mieux appréhender le rôle du complexe Rpf2/Rrs1 dans l'intégration de la particule 5S et dans la maturation de la grande sous-unité. J'ai également étudié le rôle du complexe Rpf2/Rrs1 dans la régulation de la transcription des ARNr, en caractérisant ses interactions avec la polymérase I