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1.
Clin Exp Med ; 24(1): 67, 2024 Apr 03.
Article in English | MEDLINE | ID: mdl-38568288

ABSTRACT

Colorectal cancer (CRC) is the second most prevalent cancer type worldwide, which highlights the urgent need for non-invasive biomarkers for its early detection and improved prognosis. We aimed to investigate the patterns of long non-coding RNAs (lncRNAs) in small extracellular vesicles (sEVs) collected from low-volume blood serum specimens of CRC patients, focusing on their potential as diagnostic biomarkers. Our research comprised two phases: an initial exploratory phase involving RNA sequencing of sEVs from 76 CRC patients and 29 healthy controls, and a subsequent validation phase with a larger cohort of 159 CRC patients and 138 healthy controls. Techniques such as dynamic light scattering, transmission electron microscopy, and Western blotting were utilized for sEV characterization. Optimized protocol for sEV purification, RNA isolation and preamplification was applied to successfully sequence the RNA content of sEVs and validate the results by RT-qPCR. We successfully isolated sEVs from blood serum and prepared sequencing libraries from a low amount of RNA. High-throughput sequencing identified differential levels of 460 transcripts between CRC patients and healthy controls, including mRNAs, lncRNAs, and pseudogenes, with approximately 20% being lncRNAs, highlighting several tumor-specific lncRNAs that have not been associated with CRC development and progression. The validation phase confirmed the upregulation of three lncRNAs (NALT1, AL096828, and LINC01637) in blood serum of CRC patients. This study not only identified lncRNA profiles in a population of sEVs from low-volume blood serum specimens of CRC patients but also highlights the value of innovative techniques in biomolecular research, particularly for the detection and analysis of low-abundance biomolecules in clinical samples. The identification of specific lncRNAs associated with CRC provides a foundation for future research into their functional roles in cancer development and potential clinical applications.


Subject(s)
Colorectal Neoplasms , Extracellular Vesicles , Neoplasms, Second Primary , RNA, Long Noncoding , Humans , RNA, Long Noncoding/genetics , Serum , Extracellular Vesicles/genetics , Biomarkers , Colorectal Neoplasms/diagnosis , Colorectal Neoplasms/genetics
2.
Cas Lek Cesk ; 161(5): 207-211, 2022.
Article in English | MEDLINE | ID: mdl-36443101

ABSTRACT

Standard autopsy does not always detect a cause of individuals death. It occurs often in cases of sudden death. The reason for decease, at least in a part of unsolved cases, can be revealed using methods of molecular biology and genetics. This approach is called molecular autopsy. First application dates to the end of 20th century when cause of sudden unexplained death of a young woman was provided only after execution of molecular autopsy. Molecular autopsy (also known as post-mortem genetic testing) finds its application particularly in cases of sudden death of young people or infants as their decease is more frequently associated with hereditary diseases linked for example to heart or metabolic conditions. In terms of methodical development, the form of molecular testing has been improved until now. Originally, targeted analysis of small number of genes was used. Nowadays, whole-exome and whole-genome sequencing slowly becomes a new standard for molecular autopsy. Although molecular autopsy has a potential to be integrated into an autopsy as a standard part of it, for now it has not become a standardised routine part of forensic autopsy.


Subject(s)
Death, Sudden , Forensic Medicine , Infant , Female , Humans , Adolescent , Autopsy , Exome , Genetic Testing
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