Exosomal ROR1 in peritoneal fluid identifies peritoneal disseminated PDAC and is associated with poor survival

BackgroundPancreatic ductal adenocarcinoma (PDAC) is one of the deadliest forms of cancer and peritoneal dissemination is one major cause for this poor prognosis. Exosomes have emerged as promising biomarkers for gastrointestinal cancers and can be found in all kinds of bodily fluids, also in peritoneal fluid (PF). This is a unique sample due to its closeness to gastrointestinal malignancies. The receptor tyrosine kinase-like orphan receptor 1 (ROR1) has been identified as a potential biomarker in human cancers and represents a promising target for an immunotherapy approach, which could be considered for future treatment strategies. Here we prospectively analyzed the exosomal surface protein ROR1 (exo-ROR1) in PF in localized PDAC patients (PER-) on the one hand and peritoneal disseminated tumor stages (PER+) on the other hand followed by the correlation of exo-ROR1 with clinical-pathological parameters.MethodsExosomes were isolated from PF and plasma samples of non-cancerous (NC) (n = 15), chronic pancreatitis (CP) (n = 4), localized PDAC (PER-) (n = 18) and peritoneal disseminated PDAC (PER+) (n = 9) patients and the surface protein ROR1 was detected via FACS analysis. Additionally, soluble ROR1 in PF was analyzed. ROR1 expression in tissue was investigated using western blots (WB), qPCR, and immunohistochemistry (IHC). Exosome isolation was proven by Nano Tracking Analysis (NTA), WB, Transmission electron microscopy (TEM), and BCA protein assay. The results were correlated with clinical data and survival analysis was performed.ResultsPDAC (PER+) patients have the highest exo-ROR1 values in PF and can be discriminated from NC (p <0.0001), PDAC (PER-) (p <0.0001), and CP (p = 0.0112). PDAC (PER-) can be discriminated from NC (p = 0.0003). In plasma, exo-ROR1 is not able to distinguish between the groups. While there is no expression of ROR1 in the exocrine pancreatic tissue, PDAC and peritoneal metastasis show expression of ROR1. High exo-ROR1 expression in PF is associated with lower overall survival (p = 0.0482).ConclusionWith exo-ROR1 in PF we found a promising diagnostic and prognostic biomarker possibly discriminating between NC, PDAC (PER-) and PDAC (PER+) and might shed light on future diagnostic and therapeutic concepts in PDAC

Minimal information for studies of extracellular vesicles 2018 (MISEV2018):a position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines

The last decade has seen a sharp increase in the number of scientific publications describing physiological and pathological functions of extracellular vesicles (EVs), a collective term covering various subtypes of cell-released, membranous structures, called exosomes, microvesicles, microparticles, ectosomes, oncosomes, apoptotic bodies, and many other names. However, specific issues arise when working with these entities, whose size and amount often make them difficult to obtain as relatively pure preparations, and to characterize properly. The International Society for Extracellular Vesicles (ISEV) proposed Minimal Information for Studies of Extracellular Vesicles (“MISEV”) guidelines for the field in 2014. We now update these “MISEV2014” guidelines based on evolution of the collective knowledge in the last four years. An important point to consider is that ascribing a specific function to EVs in general, or to subtypes of EVs, requires reporting of specific information beyond mere description of function in a crude, potentially contaminated, and heterogeneous preparation. For example, claims that exosomes are endowed with exquisite and specific activities remain difficult to support experimentally, given our still limited knowledge of their specific molecular machineries of biogenesis and release, as compared with other biophysically similar EVs. The MISEV2018 guidelines include tables and outlines of suggested protocols and steps to follow to document specific EV-associated functional activities. Finally, a checklist is provided with summaries of key points

Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches

Extracellular vesicles (EVs), through their complex cargo, can reflect the state of their cell of origin and change the functions and phenotypes of other cells. These features indicate strong biomarker and therapeutic potential and have generated broad interest, as evidenced by the steady year-on-year increase in the numbers of scientific publications about EVs. Important advances have been made in EV metrology and in understanding and applying EV biology. However, hurdles remain to realising the potential of EVs in domains ranging from basic biology to clinical applications due to challenges in EV nomenclature, separation from non-vesicular extracellular particles, characterisation and functional studies. To address the challenges and opportunities in this rapidly evolving field, the International Society for Extracellular Vesicles (ISEV) updates its 'Minimal Information for Studies of Extracellular Vesicles', which was first published in 2014 and then in 2018 as MISEV2014 and MISEV2018, respectively. The goal of the current document, MISEV2023, is to provide researchers with an updated snapshot of available approaches and their advantages and limitations for production, separation and characterisation of EVs from multiple sources, including cell culture, body fluids and solid tissues. In addition to presenting the latest state of the art in basic principles of EV research, this document also covers advanced techniques and approaches that are currently expanding the boundaries of the field. MISEV2023 also includes new sections on EV release and uptake and a brief discussion of in vivo approaches to study EVs. Compiling feedback from ISEV expert task forces and more than 1000 researchers, this document conveys the current state of EV research to facilitate robust scientific discoveries and move the field forward even more rapidly

Updating the approaches to define susceptibility and resistance to anti-tuberculosis agents: implications for diagnosis and treatment

11 páginas, 2 figuras, 1 tablaInappropriately high breakpoints have resulted in systematic false-susceptible AST results to anti-TB drugs. MIC, PK/PD and clinical outcome data should be combined when setting breakpoints to minimise the emergence and spread of antimicrobial resistance.I. Comas was supported by PID2019-104477RB-I00 from the Spanish Science Ministry and by ERC (CoG 101001038)Peer reviewe

Earlier Intervention Leads to Better Sound Localization in Children with Bilateral Cochlear Implants.

We present sound localization results from 30 children with bilateral cochlear implants. All children received their implants sequentially, at ages from 6 months to 9 years for the first implant and 1.5-12 years for the second implant, with delays of 10 months to 9 years. Localization was measured in the sound field, with a broadband bell-ring presented from 1 of 9 loudspeakers positioned in the frontal horizontal plane. The majority of the children (63%) were able to localize this signal significantly better than chance level. Mean absolute error scores varied from 9 to 51 degrees (root mean square error scores from 13 to 63 degrees ). The best scores were obtained by children who received their first implant before the age of 2 years and by children who used hearing aids prior to implantation for a period of 18 months or longer. Age at second implantation was important in the group of children who did not use a contralateral hearing aid during the unilateral implant period. Additionally, children who attended a mainstream school had significantly better localization scores than children who attended a school for the deaf. No other child or implantation variables were related to localization performance. Data of parent questionnaires derived from the Speech, Spatial and Qualities of Hearing Scale were significantly correlated with localization performance. This study shows that the sound localization ability of children with bilateral cochlear implants varies across subjects, from near-normal to chance performance, and that stimulation early in life, acoustically or electrically, is important for the development of this capacity

Earlier intervention leads to better sound localization in children with bilateral cochlear implants

We present sound localization results from 30 children with bilateral cochlear implants. All children received their implants sequentially, at ages from 6 months to 9 years for the first implant and 1.5–12 years for the second implant, with delays of 10 months to 9 years. Localization was measured in the sound field, with a broadband bell-ring presented from 1 of 9 loudspeakers positioned in the frontal horizontal plane. The majority of the children (63%) were able to localize this signal significantly better than chance level. Mean absolute error scores varied from 9 to 51° (root mean square error scores from 13 to 63°). The best scores were obtained by children who received their first implant before the age of 2 years and by children who used hearing aids prior to implantation for a period of 18 months or longer. Age at second implantation was important in the group of children who did not use a contralateral hearing aid during the unilateral implant period. Additionally, children who attended a mainstream school had significantly better localization scores than children who attended a school for the deaf. No other child or implantation variables were related to localization performance. Data of parent questionnaires derived from the Speech, Spatial and Qualities of Hearing Scale were significantly correlated with localization performance. This study shows that the sound localization ability of children with bilateral cochlear implants varies across subjects, from near-normal to chance performance, and that stimulation early in life, acoustically or electrically, is important for the development of this capacity.status: publishe

Sound localization, sound lateralization and binaural masking level differences in young children with normal hearing

OBJECTIVES: In this study, procedures for measuring sound localization, sound lateralization, and binaural masking level differences (BMLDs) in young children were developed. Sensitivity for these tasks was assessed in large groups of children between 4 and 9 yr of age to investigate potential developmental trends. DESIGN: Sound localization was measured in the sound field, with a broadband bell-ring presented from one of nine loudspeakers positioned in the frontal horizontal field. A group of 33 children between 4 and 6 yr of age and 5 adults took part in this experiment. Sound lateralization based on interaural time differences was measured with headphones in 49 children between 4 and 9 yr of age and 10 adults. A low-frequency stimulus containing harmonics 2 to 5 from a click train with a rate of 160 Hz was used. In the BMLD test, the same filtered click train was presented diotically or dichotically (phase reversed or time delayed) in a broadband (200 to 1000 Hz) frozen noise to 23 children between 4 and 6 yr of age and 10 adults. For comparison with literature, additional measurements with a 500-Hz sinusoid were administered to adults. All tasks were adapted to the interest and attention span of young children. RESULTS: Children of 5 yr of age did not perform significantly different from adults on the sound localization task, but mean absolute errors were larger for the 4-yr-olds. Also on the BMLD task, 5-yr-old children performed at the adult level, whereas the 4-yr-old children obtained significantly less binaural unmasking compared with the adults. Concerning sound lateralization, a small but significant difference between adults and children existed, but no age effects were apparent in the 4- to 9-yr-old group. Overall, the variation was relatively large in the 4-yr-old group, with some of the children performing at adult level, in all three tasks. CONCLUSIONS: The results of this study show that the modified procedures are suitable for testing children from the age of 4 to 5 yr. Furthermore, it seems that binaural hearing capacities of the 5-yr-olds are similar to those of adults. Several observations led to the hypothesis that the observed age differences between 4-yr-olds and older subjects on localization and BMLD or between those 4- to 9-yr old and adults on lateralization, were attributable to both a development in binaural hearing and to nonauditory factors, such as task comprehension, attention, and testing conditions. It is possible that the developmental process is more obvious and prolonged in other aspects of binaural hearing, which require more dynamic or more central processing.status: publishe