title: Cryopreservation of monocytes or differentiated immature DCs leads to an altered cytokine response to TLR agonists and microbial stimulation.
abstract: Literature on the effects of cryopreservation and thawing of monocytes or monocyte-derived immature dendritic cells (iDCs) on the subsequent functional capacities of the DCs is limited to a few specific maturation stimuli and is focused on applications in clinical immunotherapy. Given the cardinal role of DCs in regulating tolerance and immunity at mucosal surfaces there is a growing interest in understanding the effect of stromal, microbial and probiotic signals on DC function. Therefore our aim was to investigate the effects of cryopreservation on the functional properties of DCs stimulated with bacteria or the bacterial components using a standardized method. Surface markers CD83 and CD86 were expressed at similar levels on iDCs generated from cryopreserved or freshly isolated monocytes. Cryopreservation of iDCs led to slightly decreased expression of CD86 and CD83 compared to freshly generated iDCs prepared from unfrozen cells but this did not affect the capacity of DCs to acquire fully mature characteristics after stimulation. In contrast the cytokine response to lipoteichoic acid and bacterial stimulation was altered by cryopreservation of monocytes or iDCs, particularly for IL-12p70 which was decreased up to 250 fold or not detected. Cryopreservation also decreased TNF-α and IL-1β production in stimulated iDCs but to a lesser extent than for IL-12p70, depending on the maturation factors used. The amounts of IL-10 produced by stimulated iDCs were increased up to 3.6 fold when iDCs were cryopreserved, but decreased up to 90 fold when generated from cryopreserved monocytes. Immature DCs are often used to investigate the immunomodulatory properties of probiotics and here we show for the first time that cryopreserved monocytes and cryopreserved iDCs have a skewed cytokine response to microbial stimulation. These findings have implications for the methods used in bacterial-DC immune assays and highlight the importance of comparing different cytokines and stimuli in immune cell cryopreservation protocols.
authors: Top Institute Food and Nutrition, Nieuwe Kanaal 9A, 6709 PA, Wageningen, The Netherlands.
authors: Marjolein
authors: M
authors: Meijerink
authors: Dulantha
authors: D
authors: Ulluwishewa
authors: Rachel C
authors: RC
authors: Anderson
authors: Jerry M
authors: JM
authors: Wells
mesh_terms: D015703:Antigens, CD / Q000276:immunology / Q000378:metabolism; D051940:B7-2 Antigen / Q000276:immunology / Q000378:metabolism; D002454:Cell Differentiation / Q000276:immunology; D015925:Cryopreservation / Q000379:methods*; D016207:Cytokines / Q000276:immunology* / Q000378:metabolism; D003713:Dendritic Cells / Q000187:drug effects / Q000276:immunology* / Q000382:microbiology; D005434:Flow Cytometry; D054884:Host-Pathogen Interactions; D006801:Humans; D007118:Immunoassay / Q000379:methods; D007136:Immunoglobulins / Q000276:immunology / Q000378:metabolism; D016753:Interleukin-10 / Q000276:immunology / Q000378:metabolism; D018664:Interleukin-12 / Q000276:immunology / Q000378:metabolism; D053583:Interleukin-1beta / Q000276:immunology / Q000378:metabolism; D007778:Lactobacillus / Q000502:physiology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism; D009000:Monocytes / Q000187:drug effects / Q000276:immunology* / Q000382:microbiology; D013682:Teichoic Acids / Q000494:pharmacology; D013997:Time Factors; D051193:Toll-Like Receptors / Q000819:agonists / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology / Q000378:metabolism; D000097802:CD83 Antigen
chemical_list: D015703:Antigens, CD; D051940:B7-2 Antigen; D016207:Cytokines; D007136:Immunoglobulins; D016753:Interleukin-10; D018664:Interleukin-12; D053583:Interleukin-1beta; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D013682:Teichoic Acids; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha; D000097802:CD83 Antigen; C009900:lipoteichoic acid