Miller J H, Heath L N
Animal Physiology Research Unit, School of Biological Sciences, Victoria University, Wellington, New Zealand.
J Cell Physiol. 1989 Jun;139(3):538-49. doi: 10.1002/jcp.1041390313.
Three clones of the pig kidney cell line LLC-PK1 were isolated and characterized with regard to morphology, growth, proximal tubule enzyme activity, sugar uptake capacity, and hormone and drug responsiveness in a defined medium. Clone N4 was similar in morphology to the wild type (WT), whereas clone F8 showed loose attachment to the substrate, formed large, sweeping domes, and had an elongated desmosome junction between cells. The third clone, F2, did not form domes and showed a marked reduction in growth rate. Cultures of WT, N4, and F8 had higher specific activities of the enzyme alkaline phosphatase and gamma-glutamyl transpeptidase at confluence relative to growing cells; however, there was no evidence of an increase in activity of either enzyme at confluence in F2. Phlorizin-sensitive alpha-methyl-D-glucoside uptake and cytochalasin B-sensitive 2-deoxy-D-glucose uptake were measured in confluent cultures grown on porous filter supports. None of the clones lacked either of the hexose transport systems, although quantitative differences were evident. N4 cells grown in a defined medium in 96-well culture plates were tested in situ for their enzyme responses to differentiation inducers, tumor promoters, and hormones. Alkaline phosphatase activity was significantly increased at confluence by serum, parathyroid hormone (PTH), and vasopressin (AVP), and was decreased by tetradecanoylphorbol acetate (TPA) and epinephrine (EPI). Glutamyl transpeptidase activity was decreased at confluence by serum, TPA, and EPI. Similar tests on alpha-methyl-D-glucoside uptake showed that serum, TPA, PTH, and AVP had no significant effect on phlorizin-sensitive uptake; however, calcitonin increased uptake by 84% (n = 18). It was concluded that LLC-PK1 clones maintained in a defined medium are useful models for studying renal cell function.
分离出猪肾细胞系LLC-PK1的三个克隆，并在限定培养基中对其形态、生长、近端小管酶活性、糖摄取能力以及激素和药物反应性进行了表征。克隆N4在形态上与野生型（WT）相似，而克隆F8与底物的附着松散，形成大的、呈扫状的穹顶，并且细胞间有伸长的桥粒连接。第三个克隆F2不形成穹顶，且生长速率显著降低。与生长中的细胞相比，WT、N4和F8的培养物在汇合时碱性磷酸酶和γ-谷氨酰转肽酶的比活性更高；然而，没有证据表明F2在汇合时这两种酶的活性增加。在多孔滤膜支持物上生长的汇合培养物中测量了根皮素敏感的α-甲基-D-葡萄糖苷摄取和细胞松弛素B敏感的2-脱氧-D-葡萄糖摄取。尽管存在定量差异，但没有一个克隆缺乏任何一种己糖转运系统。在96孔培养板中限定培养基中生长的N4细胞原位测试了它们对分化诱导剂、肿瘤启动子和激素的酶反应。汇合时，血清、甲状旁腺激素（PTH）和血管加压素（AVP）显著增加碱性磷酸酶活性，而十四酰佛波醇乙酸酯（TPA）和肾上腺素（EPI）则降低其活性。汇合时，血清、TPA和EPI降低谷氨酰转肽酶活性。对α-甲基-D-葡萄糖苷摄取的类似测试表明，血清、TPA、PTH和AVP对根皮素敏感摄取没有显著影响；然而，降钙素使摄取增加了84%（n = 18）。得出的结论是，在限定培养基中维持的LLC-PK1克隆是研究肾细胞功能的有用模型。
