Stem Cells International / 2019 / Article / Fig 4

Research Article

Kupffer Cells Promote the Differentiation of Adult Liver Hematopoietic Stem and Progenitor Cells into Lymphocytes via ICAM-1 and LFA-1 Interaction

Figure 4

Kupffer cells secrete hematopoiesis-promoting cytokines and promote the differentiation of liver HSPCs. (a) Collagenase IV was used to digest the liver tissue, and kupffer cells were isolated by density gradient centrifugation. The cells were then inoculated into 24-well plates ( cells/well). Kupffer cells were further purified by cell adhesion selection. After culturing for 30 min, the supernatant was absorbed and washed three times with 1× PBS, then replaced with fresh DMEM medium (500 μL). Cell culture supernatants were collected at 24 h and 48 h. An ELISA was used to detect the levels of SCF, IL-6, and IL-3 in the kupffer cell culture supernatants. The bars represent the of three independent experiments. (b) The sorted liver LSK cells were seeded onto kupffer cell monolayers in the presence of SCF (50 ng/mL), and fresh kupffer cells were replaced every three days. The cocultured cells were collected and analyzed by flow cytometry on days 7 and 14. The proportion of lymphocytes (CD3+ T, CD19+ B, and NK1.1+ NK cells) and myeloid cells (CD11b+ cells) gated from total cells in the coculture system was detected by flow cytometry on day 14. (c) Statistical chart of the proportion of CD3+, CD19+, NK1.1+, and CD11b+ cells among the total coculture cells in the kupffer+LSK+SCF and kupffer+LSK+SCF+LPS coculture groups. (d) The level of proinflammatory factor IL-6, TNF-α, and IL-1β in the coculture supernatants from kupffer+LSK+SCF, kupffer+LSK+SCF+LPS, kupffer+CTRL, and kupffer+LPS groups was detected by ELISA. The data are represented as the . ns: not significantly different. ; .
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