Department of Biology, Faculty of Science, Silpakorn University, Nakhon Pathom 73000, Thailand.
Cells. 2023 May 19;12(10):1427. doi: 10.3390/cells12101427.
Previously established immune-responsive co-culture models with macrophages have limitations due to the dedifferentiation of macrophages in long-term cultures. This study is the first report of a long-term (21-day) triple co-culture of THP-1 macrophages (THP-1m) with Caco-2 intestinal epithelial cells and HT-29-methotrexate (MTX) goblet cells. We demonstrated that high-density seeded THP-1 cells treated with 100 ng/mL phorbol 12-myristate 13-acetate for 48 h differentiated stably and could be cultured for up to 21 days. THP-1m were identified by their adherent morphology and lysosome expansion. In the triple co-culture immune-responsive model, cytokine secretions during lipopolysaccharide-induced inflammation were confirmed. Tumor necrosis factor-alpha and interleukin 6 levels were elevated in the inflamed state, reaching 824.7 ± 130.0 pg/mL and 609.7 ± 139.5 pg/mL, respectively. Intestinal membrane integrity was maintained with a transepithelial electrical resistance value of 336.4 ± 18.0 Ω·cm. Overall, our findings suggest that THP-1m can be effectively employed in models of long-term immune responses in both normal and chronic inflammatory states of the intestinal epithelium, making them a valuable tool for future research on the association between the immune system and gut health.
先前建立的具有巨噬细胞的免疫反应共培养模型由于巨噬细胞在长期培养中的去分化而存在局限性。本研究首次报道了一种长期（21 天）的三重共培养体系，即 THP-1 巨噬细胞（THP-1m）与 Caco-2 肠上皮细胞和 HT-29-甲氨蝶呤（MTX）杯状细胞的共培养。我们证明，高密度接种的 THP-1 细胞经 100ng/mL 佛波醇 12-肉豆蔻酸 13-乙酸处理 48 小时后可稳定分化，并可培养长达 21 天。THP-1m 通过其贴壁形态和溶酶体扩张来识别。在三重共培养免疫反应模型中，证实了脂多糖诱导炎症期间细胞因子的分泌。肿瘤坏死因子-α和白细胞介素 6 水平在炎症状态下升高，分别达到 824.7±130.0 pg/mL 和 609.7±139.5 pg/mL。肠上皮的跨上皮电阻值为 336.4±18.0 Ω·cm，保持了肠道膜的完整性。总的来说，我们的发现表明，THP-1m 可有效地用于正常和慢性肠上皮炎症状态下的长期免疫反应模型，是未来研究免疫系统与肠道健康之间关系的有价值工具。
