Xela DS2 and Xela VS2: two novel skin epithelial-like cell lines from adult African clawed frog (Xenopus laevis) and their response to an extracellular viral dsRNA analogue

Abstract

18The skin epithelial layer acts as an important immunological barrier against pathogens 19 and is capable of recognizing and responding to pathogen associated molecular patterns 20 (PAMPs) in human and mouse models. Although presumed, it is unknown whether amphibian 21 skin epithelial cells exhibit the ability to respond to PAMPs such as viral double-stranded RNA 22(dsRNA). To address this, two cell lines from the dorsal skin (Xela DS2) and ventral skin (Xela 23 VS2) of the African clawed frog (Xenopus laevis) were established. Xela DS2 and Xela VS2 24 cells have an epithelial-like morphology, express genes associated with epithelial cells, and lack 25 senescence-associated beta-galactosidase activity. Cells grow optimally in 70% Leibovitz's L-15 26 medium supplemented with 15% fetal bovine serum at 26°C. Upon treatment with poly(I:C), a 27 synthetic viral dsRNA analogue and known type I interferon inducer, Xela DS2 and Xela VS2 28 exhibit marked upregulation of key pro-inflammatory and antiviral transcripts suggesting frog 29 epithelial cells participate in the recognition of extracellular viral dsRNA and production of local 30 inflammatory signals; similar to human and mouse models. Currently, these are the only known 31Xenopus laevis skin epithelial-like cell lines and will be important for future research in 32 amphibian epithelial cell biology, initial host-pathogen interactions, and rapid screening of the 33 effects of environmental stressors, including contaminants, on frog skin epithelial cells. 34 35

Discovery of frog virus 3 microRNAs and their roles in evasion of host antiviral responses

“…Xela complete medium was prepared as previously described (Bui-Marinos et al, 2020) and is composed of amphibian-adjusted Leibovitz’s L-15 (AL-15; Wisent, Mont-Saint-Hilaire, Canada) medium supplemented with 15% fetal bovine serum (FBS; lot # 234K18; VWR, Radnor, United States). Xela low serum medium is composed of AL-15 medium supplemented with 2% FBS.…”

“…The Xela DS2 dorsal skin epithelial-like cell line was maintained in Xela complete medium as previously described (Bui-Marinos et al, 2020). Briefly, Xela DS2 cells were cultured at 26 °C in 75 cm 2 plug-seal tissue-culture treated flasks (BioLite; Thermo Fisher Scientific, Waltham, United States) and confluent (~90%) cultures were split 1:4, every 3-4 days.…”