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. 2021 Jul;85(3):236-240.

Establishment of a canine lens epithelial cell line

Akira Matsuda  1 , Yuki Shimizu  1 , Teppei Kanda  1 , Akihiro Ohnishi  1 , Noritaka Maeta  1 , Masahiro Miyabe  1 , Kaori Saeki  1 , Yoshiki Itoh  1
Affiliations

Establishment of a canine lens epithelial cell line

Akira Matsuda et al. Can J Vet Res. 2021 Jul.

Abstract
in English, French

Some immortalized lens epithelial cell lines have been established and are useful for molecular analysis. The establishment of additional cell lines must, however, enable a variety of in-vitro examinations. The objective of this study was to establish a new canine lens epithelial cell line by isolating CLC-1 cells from the lens tissue of a dog with cataracts. In CLC-1 cells, transforming growth factor beta (TGF-β) treatment significantly decreased gene expression of an epithelial marker and elevated that of mesenchymal markers; these characteristics are similar to those of a human lens epithelial cell line. Interestingly, CLC-1 cells exhibited lower expression of an epithelial marker and higher expression of mesenchymal markers than an anterior lens capsule. These results suggest that CLC-1 cells were derived from a cell population that was committed to epithelial-mesenchymal transition in cataract lens tissue. In conclusion, CLC-1 cells could be useful for analyzing molecular pathogenesis in canine cataracts.

Certaines lignées de cellules épithéliales du cristallin immortalisées ont été établies et sont utiles pour analyse moléculaire. L’établissement de lignées cellulaires supplémentaires doit cependant permettre une variété d’examens in vitro. L’objectif de cette étude était d’établir une nouvelle lignée cellulaire épithéliale du cristallin canin en isolant les cellules CLC-1 du tissu du cristallin d’un chien atteint de cataracte. Dans les cellules CLC-1, le traitement par le facteur de croissance transformant bêta (TGF-β) a significativement diminué l’expression génique d’un marqueur épithélial et élevé celle des marqueurs mésenchymateux; ces caractéristiques sont similaires à celles d’une lignée cellulaire épithéliale du cristallin humain. Fait intéressant, les cellules CLC-1 présentaient une expression inférieure d’un marqueur épithélial et une expression plus élevée de marqueurs mésenchymateux qu’une capsule antérieure du cristallin. Ces résultats suggèrent que les cellules CLC-1 étaient dérivées d’une population cellulaire qui était impliquée dans la transition épithéliale-mésenchymateuse dans le tissu du cristallin de la cataracte. En conclusion, les cellules CLC-1 pourraient être utiles pour analyser la pathogenèse moléculaire dans les cataractes canines.(Traduit par Docteur Serge Messier).

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Figures

Figure 1
Figure 1
Characteristics of CLC-1 cells. CLC-1 cells have a polygonal shape (A) and proliferate linearly (B). Proliferation activity was evaluated by a Trypan blue dye exclusion test. Cells were suspended in culture medium at 1 × 104 cells/mL, then the number of cells was counted every 24 h. The expression of CRYαB protein was detected by western blotting (C) and immunocytochemistry (D). Rabbit anti-CRYαB antibody and Alexa Fluor 594-conjugated secondary antibody were used for the detection. 6-Diamidino-2-phenylindole (DAPI) was used for nuclear counterstaining. Scale bar = 10 μm.
Figure 2
Figure 2
Relative expression levels of epithelial or mesenchymal markers. Real-time polymerase chain reaction (RT-PCR) was carried out to determine the expression levels of CDH, αSMA, and ITGAV in ALC, pLEC, and CLA-1 cells (A). The values of target genes were normalized to that of GAPDH. Real-time PCR was carried out to detect the changes in gene expression after TGF-β treatment (10 ng/mL) in CLC-1 cells (B). Three independent experiments were conducted. The values of target genes were normalized to that of GAPDH and then standardized to that of medium alone. * P < 0.05 versus medium alone. CDH — E-cadherin; αSMA — alpha smooth muscle action; ITGAV — integrin subunit alpha V; ALC — anterior lens capsule; pLEC — primary lens epithelial cells; GAPDH — glyceraldehyde-3-phosphate dehydrogenase; TGF-β — transforming growth factor beta; p3 — 3rd passage; p14 — 14th passage; p45 — 45th passage.
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