Dicle ÇAYAN, Erdoğan UNUR, MEHTAP NİSARİ, Dilara PATAT, Ertuğrul DAĞLI, Hilal AKALIN

The Effect of Triclosan on In vitro Embryonic Development in Rat

Triclosan (TCS) is a broad spectrum antimicrobial agent showing its effect by deactivating the fatty acid synthesis of bacteria. The aim of this study was to investigate the effects of TCS on in vitro embryonic development in rats and to determine the levels of caspases 2, 7, 8, and 9 inducing cells to apoptosis through gene expression. According to the TCS dose added to the culture whole rat serum, 3 experimental groups and a control group were formed with each including 10 embryos. After 48 h culturing period, embryos were subjected to morphological scoring for developmental evaluation. The levels of caspases 2, 7, 8, and 9 were measured by performing gene expression on 40 embryos. Significant decreases were obtained in all parameters of morphological scoring in the experimental groups as the dose of TCS increased. While the caspase-2 level showed a significant decrease among the groups and was inversely proportional to the level of TCS, the caspase-9 level showed a significant increase among the groups and was directly proportional to the level of TCS. In conclusion, TCS was determined to cause apoptosis in the cells via the intrinsic pathway during pregnancy period and lead to embryonic growth retardation, which increased with increased dose of TCS.

Triklosanın In vitro Embriyonik Rat Gelişimi Üzerine Etkisi

Triklosan, bakterilerin yağ asidi sentezini bozarak etkisini gösteren geniş spektrumlu bir antimikrobiyal ajandır. Bu çalışmanın amacı, triklosanın ratlarda in vitro embriyonik gelişim üzerindeki etkisini araştırmak ve gen ekspresyon yöntemi ile hücreleri apoptoza sürükleyen kaspaz 2, 7, 8 ve 9 değerlerini belirlemektir. Kültür şişesine eklenen triklosan dozuna göre, kontrol ve 3 deney grubu oluşturuldu. 48 saatlik kültür periyodundan sonra embriyolar gelişimsel değerlendirme amacıyla morfolojik skorlamaya tabi tutuldu. Ardından 40 embriyoya gen ekspresyonu yapılarak kaspaz-2, kaspaz-7, kaspaz-8 ve kaspaz-9 değerleri ölçüldü. Morfolojik skorlamaya ait tüm parametrelerde istatistiksel olarak anlamlı bir gerileme tespit edildi. Kaspaz-2 değerlerinin triklosan miktarı ile ters orantılı bir şekilde, gruplar arasında istatistiksel olarak anlamlı bir azalma gösterdiği, kaspaz-9 değerlerinin ise triklosan miktarıyla orantılı bir biçimde istatistiksel olarak anlamlı bir artış gösterdiği tespit edildi. Bu verilere göre, gebelik döneminde triklosan kullanımının, hücrelerde intrinsik yol aracılığıyla apoptoza yol açarak, embriyonik gelişme geriliğine neden olduğu, bu durumun doza bağlı olarak arttığı belirlendi.

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1. Glaser A: The Ubiquitous TCS. Pest You J, 24 (3): 12-17, 2004.

2. Szychowski KA, Wnuk A, Rzemieniec J, Kajta M, Leszczyńska T, Wójtowicz AK: Triclosan-evoked neurotoxicity involves NMDAR subunits with the specific role of glun2a in caspase-3-dependent apoptosis. Mol Neurobiol, 56, 1‐12, 2019. DOI: 10.1007/s12035-018-1083-z

3. Cheng W, Yang S, Liang F, Wang W, Zhou R, Li Y, Feng Y, Wang Y: Low-dose exposure to TCS disrupted osteogenic differentiation of mouse embryonic stem cells via BMP/ERK/Smad/Runx-2 signalling pathway. Food Chem Toxicol, 127, 1‐10, 2019. DOI: 10.1016/j.fct.2019.02.038

4. Oliveira R, Domingues I, Koppe Grisolia C, Soares AMVM: Effects of triclosan on zebrafish early-life stages and adults. Environ Sci Pollut Res, 16, 679‐688, 2009. DOI: 10.1007/s11356-009-0119-3

5. Weatherly LM, Gosse JA: Triclosan exposure, transformation, and human health effects. J Toxicol Environ Health B Crit Rev, 20 (8): 447‐469, 2017. DOI: 10.1080/10937404.2017.1399306

6. Dhillon GS, Kaur S, Pulicharla R, Brar SK, Cledon M, Verma M, Surampalli RY: Triclosan: Current status, occurrence, environmental risks and bioaccumulation potential. Int J Environ Res Public Health, 12 (5): 5657‐5684, 2015. DOI: 10.3390/ijerph120505657

7. Sandborgh-Englund G, Adolfsson-Erici M, Odham G, Ekstrand J: Pharmacokinetics of Triclosan following oral ingestion in humans. J Toxicol Environ Health A, 69 (20): 1861-1873, 2006. DOI: 10.1080/ 15287390600631706

8. Allmyr M, Adolfsson-Erici M, McLachlan MS, Sandborgh-Englund G: Triclosan in plasma and milk from Swedish nursing mothers and their exposure via personal care products. Sci Total Environ, 372, 87-93, 2006. DOI: 10.1016/j.scitotenv.2006.08.007

9. Fang JL, Stingley RL, Beland FA, Harrouk W, Lumpkins DL, Howard P: Occurrence, efficacy, metabolism, and toxicity of Triclosan. J Environ Sci Health C Environ Carcinog Ecotoxicol Rev, 28, 147-171, 2010. DOI: 10.1080/10590501.2010.504978

10. Wu Y, Chitranshi P, Loukotková L, Gamboa da Costa G, Beland FA, Zhang J, Fang JL: Cytochrome P450-mediated metabolism of Triclosan attenuates its cytotoxicity in hepatic cells. Arch Toxicol, 91 (6): 2405‐2423, 2017. DOI: 10.1007/s00204-016-1893-6

11. Chaudhari U, Nemade H, Sureshkumar P, Vinken M, Ates G, Rogiers V, Hescheler J, Hengstler JG, Sachinidis A: Functional cardiotoxicity assessment of cosmetic compounds using human-induced pluripotent stem cell-derived cardiomyocytes. Arch Toxicol, 92 (1): 371‐381, 2018. DOI: 10.1007/s00204-017-2065-z

12. Park BK, Gonzales ELT, Yang SM, Bang M, Choi CS, Shin CY: Effects of triclosan on neural stem cell viability and survival. Biomol Ther (Seoul), 24 (1): 99‐107, 2016. DOI: 10.4062/biomolther.2015.164

13. Chen X, Xu B, Han X, Mao Z, Chen M, Du G, Talbot P, Wang X, Xia Y: The effects of Triclosan on pluripotency factors and development of mouse embryonic stem cells and zebrafish. Arch Toxicol, 89, 635-646, 2015. DOI: 10.1007/s00204-014-1270-2

14. Van Maele-Fabry G, Delhaise F, Picard JJ: Morphogenesis and quantification of the development of post-implantation mouse embryos. Toxicol In Vitro, 4 (2): 149‐156, 1990. DOI: 10.1016/0887- 2333(90)90037-t

15. Augustine-Rauch K, Zhang CX, Panzica-Kelly JM: In vitro developmental toxicology assays: A review of the state of the science of rodent and zebrafish whole embryo culture and embryonic stem cell assays. Birth Defects Res C Embryo Today, 90 (2): 87-98, 2010. DOI: 10.1002/ bdrc.20175

16. Unur E, Ülger H, Ekinci N, Hacıalioğulları M, Ertekin T, Kılıç E: Effect of anti-basic fibroblast growth factor (anti-bFGF) on in vitro embryonic development in rat. Anat Histol Embryol, 38, 241-245, 2009. DOI: 10.1111/j.1439-0264.2009.00927.x

17. Tekinarslan İİ, Unur E, Ülger H, Ekinci N, Ertekin T, Hacıalioğulları M, Arslan S: The effects of FGF-9 on in vitro embryonic development. Balkan Med J, 28, 18-22, 2011. DOI: 10.5174/tutfd.2009.02019.2

18. Ertekin T, Ülger H, Nisari M, Karaca Ö, Unur E, Şahin U, Elmalı F: Effects of angiostatin on in vitro embryonic rat development. Kafkas Univ Vet Fak Derg, 17 (5): 843-847, 2011. DOI: 10.9775/kvfd.2011.4637

19. Nisari M, Ulger H, Unur E, Karaca O, Ertekin T: Effect of interleukin 12 (IL-12) on embryonic development and yolk sac vascularisation. Bratisl Lek Listy, 115 (9): 532-537, 2014. DOI: 10.4149/bll_2014_103

20. Toder V, Carp H, Fein A, Torchinsky A: The role of pro- and antiapoptotic molecular interactions in embryonic maldevelopment. Am J Reprod Immunol, 48, 235-244, 2002. DOI: 10.1034/j.1600-0897.2002.01130.x

21. McArthur K, Kile BT: Apoptotic caspases: Multiple or mistaken identities? Trends Cell Biol, 28 (6): 475‐493, 2018. DOI: 10.1016/j. tcb.2018.02.003

22. Aşan E, Dağdeviren A: Hücre. In, Aşan E, Dağdeviren A (Eds): Moleküler Histoloji. 145-168, Atlas Kitapçılık, Ankara, 2012.

23. New DAT: Whole-embryo culture and the study of mammalian embryos during organogenesis. Biol Rev Camb Philos Soc, 53, 81-122, 1978.

24. Horie Y, Yamagishi T, Takahashi H, Iguchi T, Tatarazako N: Effects of Triclosan on Japanese medaka (Oryzias latipes) during embryo development, early life stage and reproduction. J Appl Toxicol, 38 (4): 544‐551, 2018. DOI: 10.1002/jat.3561

25. Ho JCH, Hsiao CD, Kawakami K, Tse WKF: Triclosan (TCS) exposure impairs lipid metabolism in zebrafish embryos. Aquat Toxicol, 173, 29‐35, 2016. DOI: 10.1016/j.aquatox.2016.01.001

26. Guo J, Ito S, Nguyen HT, Yamamoto K, Tanoue R, Kunisue T, Iwata H: Effects of prenatal exposure to triclosan on the liver transcriptome in chicken embryos. Toxicol Appl Pharmacol, 347, 23‐32, 2018. DOI: 10.1016/j. taap.2018.03.026

27. Haggard DE, Noyes PD, Waters KM, Tanguay RL: Phenotypically anchored transcriptome profiling of developmental exposure to the antimicrobial agent, triclosan, reveals hepatotoxicity in embryonic zebrafish. Toxicol Appl Pharmacol, 308, 32‐45, 2016. DOI: 10.1016/j. taap.2016.08.013

28. Szychowski KA, Sitarz AM, Wojtowicz AK: Triclosan induces Fas receptor-dependent apoptosis in mouse neocortical neurons in vitro. Neuroscience, 284, 192-201, 2015. DOI: 10.1016/j.neuroscience.2014.10.001

29. Dubey D, Srivastav AK, Singh J, Chopra D, Qureshi S, Kushwaha HN, Singh N, Ray RS: Photoexcited Triclosan induced DNA damage and oxidative stress via p38 MAP kinase signaling involving type I radicals under sunlight/UVB exposure. Ecotoxicol Environ Saf, 174, 270‐282, 2019. DOI: 10.1016/j.ecoenv.2019.02.065

30. Lee GA, Choi KC, Hwang KA: Treatment with phytoestrogens reversed triclosan and bisphenol A-induced anti-apoptosis in breast cancer cells. Biomol Ther (Seoul), 26 (5): 503‐511, 2018. DOI: 10.4062/ biomolther.2017.160

31. O’Byrne KJ, Richard DJ: Nucleolar caspase-2: Protecting us from DNA damage. J Cell Biol, 216 (6): 1521‐1523, 2017. DOI: 10.1083/jcb.201704114

32. Ando K, Parsons MJ, Shah RB, Charendoff CI, Paris SL, Liu PH, Fassio SR, Rohrman BA, Thompson R, Oberst A, Sidi S, Bouchier- Hayes L: NPM1 directs PIDDosome-dependent caspase-2 activation in the nucleolus. J Cell Biol, 216 (6): 1795‐1810, 2017. DOI: 10.1083/ jcb.201608095

33. Lamkanfi M, Kanneganti TD: Caspase-7: A protease involved in apoptosis and inflammation. Int J Biochem Cell Biol, 42, 21-24, 2010. DOI: 10.1016/j.biocel.2009.09.013

34. Brentnall M, Rodriguez-Menocal L, De Guevara RL, Cepero E, Boise LH: Caspase-9, caspase-3 and caspase-7 have distinct roles during intrinsic apoptosis. BMC Cell Biol, 14:32, 2013. DOI: 10.1186/1471- 2121-14-32

35. Galluzzi L, López-Soto A, Kumar S, Kroemer G: Caspases connect cell-death signaling to organismal homeostasis. Immunity, 44 (2): 221- 231, 2016. DOI: 10.1016/j.immuni.2016.01.020

36. Li X, An J, Li H, Qiu X, Wei Y, Shang Y: The methyl-triclosan induced caspase-dependent mitochondrial apoptosis in HepG2 cells mediated through oxidative stress. Ecotoxicol Environ Saf, 182:109391, 2019. DOI: 10.1016/j.ecoenv.2019.109391 The Effect of Triclosan on ...