Project description:Exosomes were isolated from plasma and saliva of healthy individuals and head and neck cancer (HNSCC) patients. miRNA profiling of plasma- and saliva-derived exosomes was performed using nCounter SPRINT system. Diagnostic panels were selected from the exosomal miRNA profile.
Project description:To investigate the contribution of fibroblast-derived extracellular matrices (ECMs) to the resistance to targeted therapies in BRAF-mutated melanoma cells, we generated native-like 3D ECMs from human primary fibroblasts obtained from healthy individuals or melanoma patients. Cell-derived matrices from human dermal fibroblasts (HDF), skin melanoma associated fibroblasts (MAF) and two different lymph node fibroblast reticular cells (FRC) were denuded of cells and their composition was analyzed by mass spectrometry.
Project description:Extracellular vesicles (EVs) are important players in melanoma progression, but their use as clinical biomarkers has been limited by the difficulty of profiling blood-derived EV proteins with high depth of coverage, the requirement for large input amounts, and complex protocols. Here, we provide a streamlined and reproducible experimental workflow to identify plasma- and serum- derived EV proteins of healthy donors and melanoma patients using minimal amounts of sample input. SEC-DIA-MS couples size-exclusion chromatography to EV concentration and deep-proteomic profiling using data-independent acquisition. From as little as 200 µl of plasma per patient in a cohort of three healthy donors and six melanoma patients, we identified and quantified 2’896 EV-associated proteins, achieving a 3.5-fold increase in depth compared to previously published melanoma studies. To compare the EV-proteome to unenriched blood, we employed an automated workflow to deplete the 14 most abundant proteins from plasma and serum and thereby approximately doubled protein group identifications versus native blood. The EV proteome diverged from corresponding unenriched plasma and serum, and unlike the latter, separated healthy donor and melanoma patient samples. Furthermore, known melanoma markers such as MCAM, TNC, and TGFBI were upregulated in melanoma EVs but not in depleted melanoma plasma, highlighting the specific information contained in EVs. Overall, EVs were significantly enriched in intact membrane proteins and proteins related to SNARE protein interactions and T cell biology. Taken together, we demonstrated the increased sensitivity of an EV-based proteomic workflow that can be easily applied to larger melanoma cohorts and other indications.
Project description:Exosomes, extracellular vesicles of 30-150 nm, are released by normal and tumor cells. These nanovesicles play a major role in cell communication and can be found in biological fluids as carriers of biomarkers. Tumor exosomes can inhibit immune responses, mediate drug resistance and transform mesenchymal stem cells. In contrast to healthy donors, cancer patients’ plasma contain higher levels of exosomes. Cutaneous melanoma is a very aggressive cancer whose incidence has rapidly increased worldwide and the prognosis is generally poor, given the propensity of melanoma cells to spread to distant sites while evading immune system control.We investigated potential differences between plasma exosomes derived from healthy donors (HD) and melanoma patients at 0-I, II, and III-IV stages of disease.
Project description:We sequenced 8 colorectal cancer patients' PBMC samples, and 6 healthy donors' PBMC samples. These individuals' plasma RNA have been profiled.
Project description:TaqMan low density array (TLDA) was carried out to screen of the profiles of circulating miRNAs in pooled plasma samples from healthy controls and pre-operative osteosarcoma patients. The expression changes of circulating miRNAs in osteosarcoma patients were identified. To select candidate plasma miRNAs for osteosarcoma detection and monitoring, we employed TLDA technique to screen expression levels of 739 miRNAs in pooled plasma samples from healthy controls and pre-operative osteosarcoma patients (each pooled from 10 individuals).
Project description:Background: Familial melanoma accounts for 10% of cases, withCDKN2A being the main high-risk gene. However, the mechanisms underlying melanomagenesis in individuals at high risk of developing melanoma remain poorly understood. Objective: To analyze the transcriptome of melanocyte/keratinocyte co-cultures derived from healthy skin from familial melanoma patients vs. controls, to comprehend melanoma development. Methods: Primary melanocyte-keratinocyte co-cultures were established from healthy skin biopsies from 16 unrelated familial melanoma patients (8 CDKN2A mutant, 8 CDKN2A wild-type) and from 7 healthy controls. Whole transcriptome was captured using the SurePrint G3 Human Microarray. Transcriptome analyses included: differential gene expression, functional enrichment, and protein-protein interaction (PPI) networks. Results: We identified a gene profile associated with familial melanoma independently of CDKN2A germline status. Functional enrichment analysis of this profile showed a downregulation of pathways related to DNA repair and immune response in familial melanoma (P < 0.05). In addition, the PPI network analysis revealed a network that consisted of double stranded DNA repair genes (including BRCA1, BRCA2, BRIP1, and FANCA), immune response genes and regulation of chromosome segregation. The hub gene was BRCA1. Conclusion: The constitutive deregulation of BRCA1 pathway genes and immune response in healthy skin could be a mechanism related with melanoma risk.
Project description:This study examined the miRNA expression level in exosomal derived from the plasma of first episode schizophrenia (FOS) patients and Healthy controls (HC), and explored the the potential of exosomes as biomarkers for schizophrenia. This study examined the lncRNA expression level in exosomal derived from the plasma of first episode schizophrenia (FOS) patients and Healthy controls (HC), and explored the the potential of exosomes as biomarkers for schizophrenia. This study examined the mRNA expression level in exosomal derived from the plasma of first episode schizophrenia (FOS) patients and Healthy controls (HC), and explored the the potential of exosomes as biomarkers for schizophrenia.
Project description:We report the sequencing of small RNAs present in the plasma of three normal subjects. In addition to microRNAs we identified abundant non-human small RNA sequences. The organisms from which these were derived were identified by BLAST searches with contigs assembled from the short sequences. The taxonomic profiles were very consistent between individuals, including plants and microbes reported previously in the microbiome, but in proportions distinct from other sites. The majority of bacterial reads were from the phylum Proteobacteria, whilst for 5 of 6 individuals over 90% of the more abundant fungal reads were from the phylum Ascomycota; of these over 90% were from the order Hypocreales. Most reads mapped to rRNA sequences and the presence of specific common sequences was confirmed by RT-PCR. In addition, extremely abundant small RNAs derived from human Y RNAs were detected. We conclude that a characteristic profile of a subset of the human microbiome can be obtained by sequencing small RNAs present in the blood. The origin and potential function of these molecules remains to be determined, but the specific profile is likely to reflect health status. This facile test has immense potential to provide biomarkers for the diagnosis and prognosis of human disease. The profile of small RNAs present in the plasma of three normal subjects was determined