Biochemical properties of encapsulated high-density 3-D HepG2 aggregates formed in an ultrasound trap for application in hepatotoxicity studies

Abstract

This paper describes the alginate encapsulation of preformed high-density 3-D HepG2 cell aggregates that guarantees good maintenance of liver-specific biomarker expression. The process involves forming a high-density (> or =7 x 10(4) cells/aggregate) discoid 3-D cell aggregate in an ultrasound trap, which is subsequently recovered and encapsulated in alginate/CaCl(2) hydrogel. Glucose secretion/consumption, lactate release, detoxifying enzyme capacity, cytokeratin-18 expression as well as hypoxia were characterized in encapsulated 3-D HepG2 aggregates over 10 days in culture. Encapsulated 3-D HepG2 aggregates released glucose into the media, although this ability was exhibited only after 1 day in culture and was subsequently lost over the ensuing 9 days. In contrast, lactate was constantly released into the media. Significantly more lactate was secreted after 3 days in culture indicating a more hypoxic environment and hence a higher rate of anaerobic glycolysis. Aggregates consistently expressed cytokeratin-18. Cytochrome P450-1A1 activity reached a maximum on day 1 of culture followed by a progressive reduction to basal levels, while P450-3A4 activity was up-regulated in a time-dependent manner reaching a peak on day 7 in culture. Glutathione-S-transferase activity, on the other hand, was at more physiological levels and remained constant over the 10-day culture period. The ultrasound trap allowed the rapid (within 5 min) generation of uniformly shaped and sized aggregates. The results reported here suggest that ultrasound-formed 3-D HepG2 aggregates can serve as alternative in vitro models providing a quick outlook on toxicity, in a tissue-mimetic manner, thus offering the future option of a cost-effective screening platform for pharmaceutical development.

Culture phases, cytotoxicity and protein expressions of agarose hydrogel induced Sp2/0, A549, MCF-7 cell line 3D cultures

“…Many studies have highlighted the importance of the 3D cell culture systems in inducing a differential gene and protein expression for several cell lines (Zschenker et al 2012). This if of significance is in utilizing this differential expression for cancer research and biomarker discovery (Bazou 2010;Lai et al 2011). …”

Therapy with Cell Encapsulation for Substitution of Organ Function and Tumor Treatment

“…[49] Further, parathyroid tissue has been encapsulated as well as hepatocyte aggregates. [50] In all three instances, the argument has been that the desired function is better preserved in a three-dimensional tissue context with all the various cell types present.…”

Ultrasound-Induced Cell–Cell Interaction Studies in a Multi-Well Microplate

“…For example, Augustsson et al [39] and Burguillos et al [40] used the XTT assay for measuring the mitochondrial dehydrogenase activity present in cells from prostate cancer cells and BV2 microglial cells, respectively. In another study, Bazou et al [41] performed different biochemical assays on HEP-G2 liver cell aggregates (e.g., detection of hypoxia, cytokeratin-18, glucose and lactate). In the following section (Section 4), we investigate the functions of natural killer (NK) cells interacting with different target cells during long-term ultrasound exposure.…”