Publicação
Assessment of the viability of circulating tumor cells in microfluidics
| Resumo: | Cancer remains the leading cause of morbidity and premature mortality in developed countries. Despite numerous scientific breakthroughs and emerging therapeutic approaches, the challenges associated with cancer metastasis remain an obstacle to achieving substantial improvements in long-term patient survival. Circulating tumor cells (CTCs) are cells released from the primary tumor that invade the endothelium and circulate in the bloodstream. They thus provide a non-invasive opportunity to gain insight into one of the main drivers of metastasis. This work aimed to study the formation of cancer spheroids originating from single breast cancer cells. A microfluidic cell sorter, coined the RUBYChip™, was used to separate breast cancer cells (SKBR-3 and MDA-MB-231) spiked in whole blood samples obtained from healthy individuals. The cell capture efficiency was determined at 54.5 ± 5.1% and 24.2 ± 6.1% for SKBR-3 and MDA-MB-231 respectively, contrasting with the 53% for MCF-7 determined in earlier studies. Single MCF-7 cells were cultured in decellularized extracellular matrix derived from human adipose tissue (adECM) and their growth tracked over time. The cells were able to proliferate and form multicellular spheroids with sizes averaging 130.4 ± 46.2 µm over the course of 14 days. MCF-7 spheroids were then grown in the presence of increasing concentrations of gold nanostars (GNSs), envisaging their use as plasmonic nanoparticles in surface-enhanced Raman spectroscopy measurements tracking the secretomics of single CTCderived spheroids. Data from live/dead, immunocytochemistry, metabolic activity and DNA quantification assays showed that concentrations up to 2mM did not affect cell viability and proliferation. This work reiterated that cancer cells can be efficiently separated from whole blood samples using a microfluidic-based filter; and has defined the protocols and minimum conditions to obtain single cell-derived cancer spheroids in minute volumes, thus holding promise for the study of CTC-derived spheroids aiming to unravel new insights on key factors driving cancer metastasis. |
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| Autores principais: | Esteves, Ana Margarida Freitas |
| Assunto: | Breast cancer CTCs Spheroid GNSs adECM |
| Ano: | 2024 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso embargado |
| Instituição associada: | Universidade de Trás-os-Montes e Alto Douro |
| Idioma: | inglês |
| Origem: | Repositório da UTAD |
| Resumo: | Cancer remains the leading cause of morbidity and premature mortality in developed countries. Despite numerous scientific breakthroughs and emerging therapeutic approaches, the challenges associated with cancer metastasis remain an obstacle to achieving substantial improvements in long-term patient survival. Circulating tumor cells (CTCs) are cells released from the primary tumor that invade the endothelium and circulate in the bloodstream. They thus provide a non-invasive opportunity to gain insight into one of the main drivers of metastasis. This work aimed to study the formation of cancer spheroids originating from single breast cancer cells. A microfluidic cell sorter, coined the RUBYChip™, was used to separate breast cancer cells (SKBR-3 and MDA-MB-231) spiked in whole blood samples obtained from healthy individuals. The cell capture efficiency was determined at 54.5 ± 5.1% and 24.2 ± 6.1% for SKBR-3 and MDA-MB-231 respectively, contrasting with the 53% for MCF-7 determined in earlier studies. Single MCF-7 cells were cultured in decellularized extracellular matrix derived from human adipose tissue (adECM) and their growth tracked over time. The cells were able to proliferate and form multicellular spheroids with sizes averaging 130.4 ± 46.2 µm over the course of 14 days. MCF-7 spheroids were then grown in the presence of increasing concentrations of gold nanostars (GNSs), envisaging their use as plasmonic nanoparticles in surface-enhanced Raman spectroscopy measurements tracking the secretomics of single CTCderived spheroids. Data from live/dead, immunocytochemistry, metabolic activity and DNA quantification assays showed that concentrations up to 2mM did not affect cell viability and proliferation. This work reiterated that cancer cells can be efficiently separated from whole blood samples using a microfluidic-based filter; and has defined the protocols and minimum conditions to obtain single cell-derived cancer spheroids in minute volumes, thus holding promise for the study of CTC-derived spheroids aiming to unravel new insights on key factors driving cancer metastasis. |
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