Synchrotron radiation-based Fourier transform infrared microspectroscopy investigation of WRL68 cells treated with doxorubicin

Abstract

Doxorubicin is an effective chemotherapeutic agent applied in a wide variety of cancers. Despite its potent anticancer activity towards cancer cells, doxorubicin is also toxic to noncancerous cells. Therefore, doxorubicin can cause serious side effects in various organs, especially when dose escalation is required for patients with advanced disease. The liver is the major detoxification organ that metabolizes drugs, and hepatotoxicity is one of the most common adverse effects of doxorubicin administration. However, the exact mechanisms of doxorubicin-induced hepatotoxicity have not been clearly identified, and how doxorubicin treatment affects the biomolecular contents of normal human hepatocytes has rarely been studied. Synchrotron radiation-based Fourier transform infrared (SR-FTIR) microspectroscopy is a state-of-the-art analytical technique for characterizing the biomolecules present in cells. In this research, the biomolecular alterations of doxorubicin-treated normal human hepatocytes compared to untreated control cells were investigated at the single-cell level by combining SR-FTIR microspectroscopy with the Cell Counting Kit-8 (CCK-8) assay and flow cytometry. WRL68 human normal embryonic liver cells, which have been shown to be very promising for assessing the cytotoxicity of toxic compounds and investigating hepato-toxicology, were used in this research. Principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA) were used to further analyse the biomolecular contents of WRL68 cells. The order of lipid acyl chains and protein α-helix structures in doxorubicin-treated WRL68 cells was found to be distinctly changed, while the nucleic acids were altered relatively less. No alteration in the carbohydrate content was distinguishable after doxorubicin treatment. These results provide more comprehensive information about the biomolecular changes in hepatocytes induced by doxorubicin treatment and help to elucidate the mechanism of doxorubicin-induced hepatotoxicity. This research also proves that SR-FTIR microspectroscopy, combined with PCA and OPLS-DA, is a promising approach for investigating drug-cell interaction systems.