RESUMEN
Cancer progression and physiological changes within the cells are accompanied by alterations in the biophysical properties. Therefore, the cell biophysical properties can serve as promising markers for cancer detection and physiological activities. To aid in the investigation of the biophysical markers of cells, a microfluidic chip has been developed which consists of a constriction channel and embedded microelectrodes. Single-cell impedance magnitudes at four frequencies and entry and travel times are measured simultaneously during their transit through the constriction channel. This microchip provides a high-throughput, label-free, automated assay to identify biophysical signatures of malignant cells and monitor the therapeutic efficacy of drugs. Here, we monitored the dynamic cellular biophysical properties in response to sphingosine kinase inhibitors (SphKIs), and compared the effectiveness of drug delivery using poly lactic-co-glycolic acid (PLGA) nanoparticles (NPs) loaded with SphKIs versus conventional delivery. Cells treated with SphKIs showed significantly higher impedance magnitudes at all four frequencies. The bioelectrical parameters extracted using a model also revealed that the highly aggressive breast cells treated with SphKIs shifted electrically towards that of a less malignant phenotype; SphKI-treated cells exhibited an increase in cell-channel interface resistance and a significant decrease in specific membrane capacitance. Furthermore, SphKI-treated cells became slightly more deformable as measured by a decrease in their channel entry and travel times. We observed no significant difference in the bioelectrical changes produced by SphKI delivered conventionally or with NPs. However, NPs-packaged delivery of SphKI decreased the cell deformability. In summary, this study showed that while the bioelectrical properties of the cells were dominantly affected by SphKIs, the biomechanical properties were mainly changed by the NPs.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Capacidad Eléctrica , Nanopartículas/química , Fosfotransferasas (Aceptor de Grupo Alcohol)/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Análisis de la Célula Individual/métodos , Antineoplásicos/química , Antineoplásicos/farmacología , Fenómenos Biomecánicos/efectos de los fármacos , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Progresión de la Enfermedad , Sistemas de Liberación de Medicamentos , Femenino , Humanos , Ácido Láctico/química , Técnicas Analíticas Microfluídicas/instrumentación , Estructura Molecular , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Inhibidores de Proteínas Quinasas/química , Relación Estructura-ActividadRESUMEN
Triple negative breast cancer (TNBC) is highly aggressive and has a poor prognosis when compared to other molecular subtypes. In particular, the claudin-low subtype of TNBC exhibits tumor-initiating/cancer stem cell like properties. Here, we seek to find new biomarkers to discriminate different forms of TNBC by characterizing their bioimpedance. A customized bioimpedance sensor with four identical branched microelectrodes with branch widths adjusted to accommodate spreading of individual cells was fabricated on silicon and pyrex/glass substrates. Cell analyses were performed on the silicon devices which showed somewhat improved inter-electrode and intra-device reliability. We performed detailed analysis of the bioimpedance spectra of four TNBC cell lines, comparing the peak magnitude, peak frequency and peak phase angle between claudin-low TNBC subtype represented by MDA-MB-231 and Hs578T with that of two basal cells types, the TNBC MDA-MB-468, and an immortalized non-malignant basal breast cell line, MCF-10A. The claudin-low TNBC cell lines showed significantly higher peak frequencies and peak phase angles than the properties might be useful in distinguishing the clinically significant claudin-low subtype of TNBC.
Asunto(s)
Microelectrodos , Neoplasias de la Mama Triple Negativas/diagnóstico , Línea Celular Tumoral , Supervivencia Celular , Claudinas/genética , Claudinas/metabolismo , Impedancia Eléctrica , Femenino , Humanos , Microscopía Confocal , Microscopía Electrónica de Rastreo , Modelos Biológicos , Células Madre Neoplásicas/metabolismo , Reproducibilidad de los Resultados , Silicio/químicaRESUMEN
The relative sensitivity of standard gold microelectrodes for electric cell-substrate impedance sensing was compared with that of gold microelectrodes coated with gold nanoparticles, carbon nanotubes, or electroplated gold to introduce nano-scale roughness on the surface of the electrodes. For biological solutions, the electroplated gold coated electrodes had significantly higher sensitivity to changes in conductivity than electrodes with other coatings. In contrast, the carbon nanotube coated electrodes displayed the highest sensitivity to MDA-MB-231 metastatic breast cancer cells. There was also a significant shift in the peak frequency of the cancer cell bioimpedance signal based on the type of electrode coating. The results indicate that nano-scale coatings which introduce varying degrees of surface roughness can be used to modulate the frequency dependent sensitivity of the electrodes and optimize electrode sensitivity for different bioimpedance sensing applications.
Asunto(s)
Neoplasias de la Mama/metabolismo , Materiales Biocompatibles Revestidos/química , Oro/química , Nanotubos de Carbono/química , Neoplasias de la Mama/patología , Línea Celular Tumoral , Impedancia Eléctrica , Electrodos , Femenino , HumanosRESUMEN
Detection of a few cancer cells within a complex cellular mixture is a key challenge presented by clinical human biopsy samples. We have designed and tested a microfabricated bioimpedance device that can detect a few human MDA-MB-231 breast cancer cells in a mixed cell culture model of a breast tissue sample. The normal tissue components were modelled using non-cancerous MCF10A human breast epithelial cells and normal human HS68 fibroblasts. The sensor is a silicon chip 0.5 cm in diameter that contains one counter electrode and four 40 µm-wide multi-branched sensing electrodes. The cells' bioimpedances were characterized in pure monocultures and in mixed cell cultures following a brief cultivation on the sensor. After cell seeding, a stable bioimpedance signal was achieved indicative of cell attachment. A cancer-selective bioimpedance signal was elicited by addition of suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor with selective actions on the cytoskeleton in breast cancer cells. SAHA elicited a 50% rise in peak bioimpedance in MDA-MB-231 breast cancer cells by 15 h. In mixed cultures of MDA-MB-231, MCF10A, and HS68 cells, the contribution of cancer cells present in the mixture dominated impedance response to SAHA. A single adherent cancer cell on any one of four electrodes in a background of â¼100 normal cells resulted in ≥5% increase in bioimpedance. The estimated sensitivity of this device is therefore one cancer cell among a background of 400 normal cells or the equivalent of 25 cancer cells in a biopsy sample of 10 000 cells.