Impact of chitosan membrane culture on the expression of pro and antiinflammatory cytokines in mesenchymal stem cells

Osteoarthritis (OA) is a chronic inflammatory joint condition caused by various inflammatory cytokines. The proinflammatory cytokines controlling OA include interleukin (IL)1β, tumor necrosis factor (TNF)α, IL6 and IL18. The antiinflammatory cytokines include IL4, IL10, IL13, leukemia inhibitory factor (LIF), glycoprotein 130 (IL6ST), TNFαstimulated gene 6 and transforming growth factor (TGF)β1. Mesenchymal stem cells (MSCs) serve an antiinflammatory role in the treatment of OA by secreting various cytokines. Previous studies demonstrated that the antiinflammatory ability of MSCs decreased rapidly in a traditional plate culture. Maintaining the antiinflammatory properties of MSCs in vitro remains challenging. Therefore, it is necessary to develop a more stable and efficient method to culture MSCs in vitro. Chitosan is a deacetylated derivative of chitin and is the second most abundant natural polysaccharide worldwide. The present study demonstrated that that MSCs cultured on chitosan membranes (CM) spontaneously formed multicellular spheroids. Compared with the control group without CM, the formation of multicellular spheres in the CM enhanced the antiinflammatory properties of MSCs. Expression levels of pro and antiinflammatory genes mRNA and their proteins in MSCs were detected by reverse transcriptionquantitative PCR, western blot analysis and immunofluorescence assay. Protein and mRNA expression levels of proinflammatory cytokines IL1β, TNFα, IL6 and IL18 were significantly decreased in CMcultured MSCs compared with the control group (P<0.05). Furthermore, mRNA and protein expression levels of antiinflammatory cytokines TGFβ1 in CMcultured MSCs were significantly increased compared with the control group (P<0.01). These results indicated that the formation of multicellular spheroids by CMcultured MSCs aided in maintaining antiinflammatory effects.