Isolation, culture and identification of goat alveolar macrophages / 中国比较医学杂志

Objective In order to study the biological characteristics of macrophages and provide the materials to study the survival mechanism of intracellular parasites, we conducted this study to establish a high-purity alveolar macrophage isolation and culture method.Methods Goat lungs were lavaged with normal saline in sterile environment several times, and cells were collected and then goat alveolar macrophages were purified by density gradient centrifugation using peripheral blood mononuclear cells (PBMC) solution.The isolated goat alveolar macrophages were cultured in cell culture medium containing 10% fetal bovine serum and cell morphology was observed under an inverted microscope every day,and the phagocytic activity of the cells was detected by chicken red blood cell phagocytosis test.Flow cytometry was used to detect CD14, a characteristic monocyte-macrophage surface marker.Results The adherent cells were characterized by typical macrophage morphology, pseudopodia and protrusions, showing round and irregular shape, rich cytoplasm, and large cell body.Of the cultured macrophages, 54.5% could phagocytize chicken erythrocytes and showed good phagocytic activity.After one month of in vitro culture, 93.7% of the cells were able to express CD14 antigen, which had a macrophage-specific immunophenotype.Conclusions The alveolar macrophages obtained in this study have high purity and good bioactivity, thus provide a cell model for studying the immune mechanism of intracellular parasites.

Contribution of IL-1β to migration of lung cancer H460 cells with acquired resistance to tumor necrosis factor-related apoptosis-inducing ligand / 实用医学杂志

Objective To determine the association of high IL-1β levels with the migration of lung cancer H460 cells with acquired resistance to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Methods The resistant cells were referred to as H460-TR in this study. IL-1β levels in H460-TR cells and the parent H460 (H460-WT) cells were measured through RT-PCR, Western blot, and enzyme-linked immunosorbent assay. The migration capacity of the cells was determined using the migration transwell assay. The extent of migration and the activation of phosphatidyl-inositol 3-kinase/serine-threonine kinase (PI3K/Akt) were detected in H460 cells treated with or without human recombinant IL-1 or IL-1R antagonist. Results Migration capacity of H460-TR cells in the conditioned medium and its IL-1β level were higher than those of H460-WT cells . The migration capacity and Akt activation were consistent with the IL-1β level in lung cancer H460 cells. Conclusions Significantly elevated IL-1β expression in cancer cells is associated with the high migration capacity of H460-TR cells, and Akt activation. Akt signaling as the downstream pathway of IL-1β and IL-1βmay function as a therapeutic target for metastatic lung cancer.