Abstract
This study used nanofluidic protein posttranslational modification (PTM) profiling to measure the effects of six cannabidiol (CBD) oils and isolated CBD on the signaling pathways of a cultured SH-SY5Y neuronal cell line. Chemical composition analysis revealed that all CBD oils met the label claims and legal regulatory limit regarding the CBD and tetrahydrocannabinol (THC) contents, respectively. Isolated CBD was cytotoxic, with an effective concentration (EC50) of 40 µM. In contrast, the CBD oils had no effect on cell viability at CBD concentrations exceeding 1.2 mM. Interestingly, only an unadulterated CBD oil had strong and statistically significant suppressive effects on the pI3K/Akt/mTOR signaling pathway with an EC50 value of 143 µM and a slow-acting timescale requiring hours. Systematic profiling of twenty-six proteins, which served as biomarkers for nine signaling pathways, revealed that the unadulterated CBD oil downregulated seven signaling pathways but had no measurable effect on the other two signaling pathways. The remaining CBD oils, which were adulterated, and isolated CBD had weak, variable, or undetectable effects on neuronal signaling pathways. Our data clearly showed that adulteration diminished the biological activities of CBD oils. In addition, nanofluidic protein PTM profiling provided a robust means for potency assessment of CBD oils.
Highlights
The consumer demand for hemp-derived products is growing rapidly in the United States following a recent change in federal legislation and the approval of Epidiolex, an oral cannabidiol (CBD) solution, for the treatment of rare and severe forms of epilepsy by the Food and Drug Administration (FDA) [1].The 2018 Farm Bill legalized hemp and removed its designation as a Schedule I controlled substance.Hemp is any part of the Cannabis sativa plant that contains less than 0.3% tetrahydrocannabidiol (THC)by weight, which includes the concentrated liquid extract known as CBD oil
Multiplexed nanofluidic protein posttranslational modification (PTM) profiling assays were used to measure the activities of nine signaling pathways in cultured SH-SY5Y neuronal cells following treatment with CBD oils
The cannabinoid and terpene profiles were analyzed with high-performance lipid chromatography (HPLC) and gas chromatography coupled with mass spectrometry (GC-MS) methods, respectively
Summary
The consumer demand for hemp-derived products is growing rapidly in the United States following a recent change in federal legislation and the approval of Epidiolex, an oral cannabidiol (CBD) solution, for the treatment of rare and severe forms of epilepsy by the Food and Drug Administration (FDA) [1]. In addition to determining the chemical profiles of cannabinoids and terpenes, which are the chemical compounds that affect neurotransmission and give cannabis its distinctive smell, respectively, CBD oil potency was further evaluated by their effects on selected signaling pathways of an SH-SY5Y cell line. Due to the ease of their maintenance, SH-SY5Y cells can serve as a robust model to evaluate the biological effects of CBD oils on neuronal signaling pathways. Measuring the effects of CBD oils on cell signaling pathways provides a robust means to assess their biological activities and potency. Multiplexed nanofluidic protein posttranslational modification (PTM) profiling assays were used to measure the activities of nine signaling pathways in cultured SH-SY5Y neuronal cells following treatment with CBD oils. Multiplexed nanofluidic protein PTM profiling was deployed to assess the potency of CBD oils by measuring their effects on neuronal signaling pathways
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