cells Article Diclofenac Alters the Cell Cycle Progression of the Green Alga Chlamydomonas reinhardtii Darya Harshkova 1, * , Ivan Liakh 2 , Vitali Bialevich 3 , Kamila Ondrejmišková 3 , Anna Aksmann 1 and Kateˇ rina Bišová 3   Citation: Harshkova, D.; Liakh, I.; Bialevich, V.; Ondrejmišková, K.; Aksmann, A.; Bišová, K. Diclofenac Alters the Cell Cycle Progression of the Green Alga Chlamydomonas reinhardtii. Cells 2021, 10, 1936. https://doi.org/10.3390/cells10081936 Academic Editor: Suleyman Allakhverdiev Received: 29 June 2021 Accepted: 28 July 2021 Published: 30 July 2021 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). 1 Department of Plant Physiology and Biotechnology, Faculty of Biology, University of Gdansk, Wita Stwosza 59, 80-308 Gda ´ nsk, Poland; anna.aksmann@ug.edu.pl 2 Department of Toxicology, Medical University of Gda´ nsk, Al. Gen. J. Haller 107, 80-416 Gda ´ nsk, Poland; liakh_ivan@mail.ru 3 Laboratory of Cell Cycles of Algae, Centre Algatech, Institute of Microbiology, Czech Academy of Sciences, Novohradská 237, 379-01 Trebon, Czech Republic; bialevich@alga.cz (V.B.); ondrejmiskova@alga.cz (K.O.); bisova@alga.cz (K.B.) * Correspondence: darya.harshkova@phdstud.ug.edu.pl; Tel.: +48-690-379-071 Abstract: The aim of the study was to verify the hypothesis that a potential cause of the phytotoxicity of diclofenac (DCF, a non-steroidal anti-inflammatory drug) is an effect of cell cycle progression. This research was conducted using synchronous cultures of a model organism, green alga Chlamy- domonas reinhardtii. The project examined DCF effects on selected parameters that characterize cell cycle progression, such as cell size, attainment of commitment points, DNA replication, number of nuclei formed during cells division and morphology of cells in consecutive stages of the cell cycle, together with the physiological and biochemical parameters of algae cells at different stages. We demonstrated that individual cell growth remained unaffected, whereas cell division was delayed in the DCF-treated groups grown in continuous light conditions, and the number of daughter cells from a single cell decreased. Thus, the cell cycle progression is a target affected by DCF, which has a similar anti-proliferative effect on mammalian cells. Keywords: non-steroidal anti-inflammatory drug; diclofenac; cell cycle; Chlamydomonas reinhardtii 1. Introduction Algae are a basic element of the water environment, and thus any abnormality in their function may lead to a disruption in the stability of a particular ecosystem. Hence, it is important to understand how different toxic substances influence algal physiology and biochemistry. Diclofenac (DCF), belonging to the non-steroidal anti-inflammatory (NSAID) class of drugs, is one of the pharmaceuticals on the hazardous substances list [1] and one of the most common pollutants of water environments [2]. As with other pharmaceuticals, DCF is designed to influence specific metabolic pathways in animal and human cells. Thus, ecotoxicological research is mostly focused on the threat DCF poses to non-target animal organisms [3]. In this regard, some investigations into DCF phytotoxicity mainly describe general observations. For example, it was shown that DCF inhibits the growth of duck- weed Lemna minor (Magnoliophyta) as well as the reproduction of green algae Scenedesmus vacuolatus, Desmodesmus subspicatus and Dunaliella tertiolecta (Chlorophyta) [4–7]. Further- more, DCF was found to be phytotoxic in standard tests as defined by the OECD or ISO guidelines [8]. In our earlier studies, Chlamydomonas reinhardtii (C. reinhardtii) population growth was found to be suppressed by DCF [9], which could have resulted from either cell growth inhibition or cell division blockage. However, when using an asynchronous culture, it is impossible to separate the two; thus, the cause of the abovementioned effects at the biochemical and molecular level remained unresolved. Here, we attempt to fill this Cells 2021, 10, 1936. https://doi.org/10.3390/cells10081936 https://www.mdpi.com/journal/cells