The Influence of Culture Time and Passage Number on the Morphological and Physiological Development of Caco-2 Cells

Abstract

The Caco-2 cell line is used by many investigators as a model of the intestinal epithelium to study nutrient uptake and transport. Our goal was to create an awareness of inherent variabilities in the Caco-2 cell line which may influence their suitability as a model or their application to specific problems. To study the influence of passage on the model, cultures were monitored from passage 20 to 109. Transepithelial electrical resistance (TEER) and sucrase activity (measured in 21-day-old cultures) increased through about passage 36. TEER values declined after about passage 60; sucrase remained elevated but variable. Cells at passage 22, 33, and 72 were grown simultaneously for 24 days. Older-passaged cells reached plateau phase sooner. Before Day 15, passage 72 cells had higher TEER and lower permeability to 14C-mannitol than passages 22 and 33; however, after Day 15 all passages showed similar permeability. On Day 21, passage 72 cells had significantly lower alkaline phosphatase activity than did the other passages. Electron microscopy did not reveal any major morphological differences between the passages; however, it did show that some areas of cells grown on membranes were not monolayers but were several cells thick with varied morphology. Investigation of the formation of these multilayered areas showed them to be an inherent part of cell growth under the conditions used. These results emphasize the inherent variability in Caco-2 cell models and emphasize the need to monitor closely the culture characteristics during growth and differentiation under specific experimental conditions.

Bioactivation of zearalenone by porcine hepatic biotransformation

“…To this end, a series of initial experiments was conducted with 17β-oestradiol showing that in 96 well plates MCF-7 cells reach the stationary phase of growth after 144 h, when seeded at a density of 10 4 cells per well. Although our results are in accordance with previously published data [1,3,11], the exact assay conditions have to be established prior to the testing of new chemicals, since Table II the number of cell passages prior to the actual assay might influence cell proliferation rates as seen in SaOS cells [14], in MC3T3-E1 pre-neoblastic cells [18], in C-6 glioma cells [17], in LNCap prostatic adenocarcinoma cells [12], in Caco-2 cells [6], and in rat mesangial cells [16].…”

Caco-2 Cell Line

“…Thus, the expression of differentiation markers typical of enterocytes, change with increasing numbers of passages (Artursson et al 2001 ). Also, parameters like transepithelial electric resistance (TEER) and proliferation rate have been reported to increase with passage number (Briske Andersson et al 1997 ). It has also been documented that late passage cells may start growing in multilayers, a phenomenon that will affect TEER measurements, and make comparisons with results based on early passage cells diffi cult.…”

Tumour and normal tissue radiobiology in mouse models: how close are mice to mini-humans?

“…However, when preclinical tumours are established from cultured cell lines, the heterogeneity of human cancers is lost to a great extent. 13 Tumour cell lines have been grown in vitro for many years and hence may have altered characteristics compared with de novo tumours. In vitro culturing of cancer cells may introduce additional stress that lead to changes in the genotype and the phenotype of tumour cells.…”