Evaluation and diagnostic potential of circulating extracellular vesicle-associated microRNAs in adrenocortical tumors

There is no available blood marker for the preoperative diagnosis of adrenocortical malignancy. The objective of this study was to investigate the expression of extracellular vesicle-associated microRNAs and their diagnostic potential in plasma samples of patients suffering from adrenocortical tumors. Extracellular vesicles were isolated either by using Total Exosome Isolation Kit or by differential centrifugation/ultracentrifugation. Preoperative plasma extracellular vesicle samples of 6 adrenocortical adenomas (ACA) and 6 histologically verified adrenocortical cancer (ACC) were first screened by Taqman Human Microarray A-cards. Based on the results of screening, two miRNAs were selected and validated by targeted quantitative real-time PCR. The validation cohort included 18 ACAs and 16 ACCs. Beside RNA analysis, extracellular vesicle preparations were also assessed by transmission electron microscopy, flow cytometry and dynamic light scattering. Significant overexpression of hsa-miR-101 and hsa-miR-483-5p in ACC relative to ACA samples has been validated. Receiver operator characteristics of data revealed dCT hsa-miR-483-5p normalized to cel-miR-39 to have the highest diagnostic accuracy (area under curve 0.965), the sensitivity and the specifity were 87.5 and 94.44, respectively. Extracellular vesicle-associated hsa-miR-483-5p thus appears to be a promising minimally invasive biomarker in the preoperative diagnosis of ACC but needs further validation in larger cohorts of patients

Regulation of zebrafish hatching by tetraspanin cd63

Tetraspanins cause the clustering of membrane proteins into a level of organisation essential for cellular function. Given the importance and complicated nature of this mechanism, we attempted a novel approach to identify the function of a single component in a biologically relevant context. A morpholino knockdown strategy was used to investigate the role of cd63, a membrane protein associated with intracellular transport and a melanoma marker, in embryonic zebrafish. By using three separate morpholinos targeting cd63, we were able to identify a specific phenotype. Strikingly, morphant fish failed to hatch due to the lack of secreted proteolytic enzymes required for chorion-softening. The morphology of the hatching gland at both the cellular and intracellular levels was disorganised, suggesting a role for cd63 in the functioning of this organ. This work identifies a specific role for cd63 in the zebrafish embryo and provides evidence for the suitability of zebrafish as a model system for the investigation of tetraspanin enriched microdomains

EGF stimulates annexin 1-dependent inward vesiculation in a multivesicular endosome subpopulation

Here we show that EGF and EGF receptor (EGFR) are trafficked through a subpopulation of multivesicular endosomes/bodies (MVBs) that are distinct from morphologically identical vacuoles that label for the late endosomal marker lyso-bisphosphatidic acid (LBPA). EGF stimulation increases both MVB biogenesis and inward vesiculation within EGFR-containing MVBs. Deletion of annexin 1, a substrate of EGFR tyrosine kinase, abolishes the effect of EGF stimulation on inward vesiculation. This phenotype is reversible by transfection with wild-type but not Y21F phosphorylation mutant annexin 1. Deletion of annexin 1 has no effect on EGF-stimulated MVB biogenesis, suggesting that MVB biogenesis and inward vesiculation within MVB are mediated by separate mechanisms. Loss or depletion of annexin 1 has no effect on EGF degradation and causes only a small delay in EGFR degradation, indicating that annexin 1 operates downstream of Hrs- and ESCRT-mediated sorting and is required solely for EGF-stimulated inward vesiculation. Annexin 1 accumulates on internal vesicles of MVB after EGF-stimulated inward vesiculation, suggesting that it may be required for a late stage in inward vesiculation

The biogenesis of multivesicular endosomes

Recent progress has been made in our understanding of the molecular mechanisms that regulate the biogenesis of multivesicular transport intermediates in the degradation pathway that leads to lysosomes. Here, we discuss recent work that uncovers some of the mechanisms that cause both membrane invagination within these newly forming intermediates and the detachment of these intermediates from early endosomal membranes

At the Feet of the Fortress: Analysis of Inka Period (ca. AD 1430-1536) Archaeofaunal Assemblages from Residential Unit 1 (RU1), Pucara de Tilcara (Jujuy, Argentina).

This paper reports the results of a zooarchaeological analysis conducted on the occupation layer of a compound structure (Residential Unit 1) of the Pucara de Tilcara archaeological site (Jujuy Province, northwestern Argentina). Its occupation span extends between the 13th and 15th centuries AD, but evidence diagnostic of the Inka Period (AD 1430-1536) is predominant. Residential Unit 1 was a house-workshop that hosted specialized crafts like metallurgy and lapidary during the Inka Period. It was proposed in previous works that artisans living at Pucara de Tilcara were provisioned with agropastoral products by the Inka administration. This paper aims to test that hypothesis against the zooarchaeological evidence of Residential Unit 1. Three variables were used as proxies for state-sponsored distribution: taxonomic diversity (family and species ranks), and skeletal and age profiles of the predominant zoological family (Camelidae) in the assemblage. The results show a high degree of continuity with the regional record, characterized by a herding-hunting strategy focused on domestic and wild species of Camelidae and a mixed mortality pattern. The skeletal profile shows a strong and negative correlation with the desiccation potential of elements, which could be indicative of local production of chalona. Overall, faunal evidence does not show any sign of centralized distribution