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. 2019 Feb 22;9:2567. doi: 10.1038/s41598-018-37495-6

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Cocaine treatment induces Ser/Thr phosphorylation of prolidase. HBMECs were treated with cocaine overnight and the cellular lysates were used to measure mRNA levels of prolidase, and serine/threonine phosphorylated prolidase. (A) Quantification of prolidase mRNA levels. Total RNA from untreated and cocaine-treated cells was subjected to qPCR and prolidase mRNA levels were normalized to the levels of GAPDH mRNA. Prolidase mRNA in cocaine-treated cells were expressed as fold changes over the untreated control cells. (B) Immunoblot analysis of phospho-prolidase expression. To determine levels of Ser/Thr phosphorylation, cellular lysates were immunoprecipitated with anti-prolidase antibody and analyzed by immunoblot using anti-phospho Ser/Thr antibody or with anti-prolidase antibody. (C) Densitometry analysis of phospho-prolidase levels relative to total prolidase from three independent experiments. Data represents the mean ± SEM of at least three determinations. *P < 0.05 represents statistical comparison of untreated and cocaine-treated samples.