The Cat As A Naturally Occurring Model Of Renal Interstitial Fibrosis: Characterisation Of Primary Feline Proximal Tubular Epithelial Cells And Comparative Pro-Fibrotic Effects Of Tgf-Β1

Chronic kidney disease (CKD) is common in both geriatric cats and aging humans, and is pathologically characterised by chronic tubulointerstitial inflammation and fibrosis in both species. Cats with CKD may represent a spontaneously occurring, non-rodent animal model of human disease, however little is known of feline renal cell biology. In other species, TGF-β1 signalling in the proximal tubular epithelium is thought to play a key role in the initiation and progression of renal fibrosis. In this study, we first aimed to isolate and characterise feline proximal tubular epithelial cells (FPTEC), comparing them to human primary renal epithelial cells (HREC) and the human proximal tubular cell line HK-2. Secondly, we aimed to examine and compare the effect of human recombinant TGF-β1 on cell proliferation, pro-apoptotic signalling and genes associated with epithelial-to-mesenchymal transition (EMT) in feline and human renal epithelial cells. FPTEC were successfully isolated from cadaverous feline renal tissue, and demonstrated a marker protein expression profile identical to that of HREC and HK-2. Exposure to TGF-β1 (0-10 ng/ml) induced a concentration-dependent loss of epithelial morphology and alterations in gene expression consistent with the occurrence of partial EMT in all cell types. This was associated with transcription of downstream pro-fibrotic mediators, growth arrest in FPTEC and HREC (but not HK-2), and increased apoptotic signalling at high concentrations of TGF- β1. These effects were inhibited by the ALK5 (TGF-β1RI) antagonist SB431542 (5 µM), suggesting they are mediated via the ALK5/TGF-β1RII receptor complex. Taken together, these results suggest that TGF-β1 may be involved in epithelial cell dedifferentiation, growth arrest and apoptosis in feline CKD as in human disease, and that cats may be a useful, naturally occurring model of human CKD.慢性肾脏病（CKD）在老年猫与衰老人类中均十分常见，二者的病理特征均表现为慢性肾小管间质性炎症与纤维化。罹患CKD的猫可作为人类疾病的自发非啮齿类动物模型，但目前对猫肾脏细胞生物学的了解仍十分有限。在其他物种中，近端小管上皮细胞的转化生长因子-β1（TGF-β1）信号通路被认为在肾纤维化的起始与进展过程中发挥关键作用。本研究首先旨在分离并鉴定猫近端小管上皮细胞（FPTEC），并将其与人原代肾上皮细胞（HREC）及人近端肾小管上皮细胞系HK-2进行对比；其次，本研究旨在探究并比较人重组转化生长因子-β1对猫源及人源肾上皮细胞的细胞增殖、促凋亡信号通路以及与上皮间质转化（EMT）相关基因的影响。 本研究成功从尸体猫肾脏组织中分离出FPTEC，其标志物蛋白表达谱与HREC及HK-2完全一致。用0~10 ng/ml的TGF-β1处理后，所有细胞类型均出现浓度依赖性的上皮形态丢失与基因表达改变，符合部分上皮间质转化的发生特征。该变化与下游促纤维化介质的转录、FPTEC及HREC的生长停滞（但HK-2未出现此现象）相关，且在高浓度TGF-β1处理时凋亡信号通路活性升高。上述效应可被ALK5（转化生长因子-β1I型受体）拮抗剂SB431542（5 µM）抑制，表明其通过ALK5/转化生长因子-β1II型受体复合物介导。综上，本研究结果提示，与人类疾病中一致，TGF-β1可能参与猫CKD中的上皮细胞去分化、生长停滞与凋亡过程，且猫或可成为人类CKD的实用自然发生模型。