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Impact of plastic adhesion in vitro on analysis of Th1 and Th2 cytokines and immune cell distribution from mice with multiple low-dose streptozotocin-induced diabetes.
J Immunol Methods. 2005 Dec 20; 307(1-2):73-81.JI

Abstract

Cytokines produced by Th1 or Th2 cells have been postulated to be important in the development of type 1 diabetes in humans and animal models, such as murine multiple low-dose streptozotocin (MLDSTZ)-induced diabetes. The aim of this study was to investigate cytokine production with or without in vitro depletion of plastic adherent cells from spleens isolated after MLDSTZ treatment. Spleen cells were prepared on day 14 from MLDSTZ- and saline-treated mice and divided into two fractions. One cell fraction was depleted of adherent cells by plastic adherence and the other was not. Both cell fractions were analysed by FACS for the distribution of immune cells. In other experiments, the cells were cultured for 48 h with concanavalin A stimulation. Supernatant samples were analysed by ELISA for TNFalpha, IFNgamma and IL-10 production. Either before or after the 48-h culture cytokine mRNA expression was determined by RT-PCR. Plastic adhesion decreased the macrophage numbers by approximately 30% and CD4(+)CD25(+) cells by about 60%. This was accompanied by increased medium levels of TNFalpha, IFNgamma and IL-10, which suggest that either CD4(+)CD25(+) cells, macrophages, or both, down-regulate production of both Th1 and certain Th2 cytokines. Depletion of adherent cells also decreased IL-4 mRNA amounts. MLDSTZ treatment increased the production of Th1 cytokines mainly at the protein level, and IL-10 mainly at the mRNA level. This indicates a sustained increase in Th1 production after MLDSTZ treatment and an increase in IL-10 that might reflect an attempt to counteract the MLDSTZ-induced immune damage. Plastic adhesion during cell preparation may affect the relative distribution of certain immune cells.

Authors+Show Affiliations

Department of Medical Cell Biology, Biomedicum, Uppsala, Sweden. Lina.Thorvaldson@medcellbiol.uu.seNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

16263129

Citation

Thorvaldson, Lina, et al. "Impact of Plastic Adhesion in Vitro On Analysis of Th1 and Th2 Cytokines and Immune Cell Distribution From Mice With Multiple Low-dose Streptozotocin-induced Diabetes." Journal of Immunological Methods, vol. 307, no. 1-2, 2005, pp. 73-81.
Thorvaldson L, Johansson SE, Höglund P, et al. Impact of plastic adhesion in vitro on analysis of Th1 and Th2 cytokines and immune cell distribution from mice with multiple low-dose streptozotocin-induced diabetes. J Immunol Methods. 2005;307(1-2):73-81.
Thorvaldson, L., Johansson, S. E., Höglund, P., & Sandler, S. (2005). Impact of plastic adhesion in vitro on analysis of Th1 and Th2 cytokines and immune cell distribution from mice with multiple low-dose streptozotocin-induced diabetes. Journal of Immunological Methods, 307(1-2), 73-81.
Thorvaldson L, et al. Impact of Plastic Adhesion in Vitro On Analysis of Th1 and Th2 Cytokines and Immune Cell Distribution From Mice With Multiple Low-dose Streptozotocin-induced Diabetes. J Immunol Methods. 2005 Dec 20;307(1-2):73-81. PubMed PMID: 16263129.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Impact of plastic adhesion in vitro on analysis of Th1 and Th2 cytokines and immune cell distribution from mice with multiple low-dose streptozotocin-induced diabetes. AU - Thorvaldson,Lina, AU - Johansson,Sofia E, AU - Höglund,Petter, AU - Sandler,Stellan, Y1 - 2005/10/17/ PY - 2005/06/30/received PY - 2005/08/22/revised PY - 2005/09/08/accepted PY - 2005/11/3/pubmed PY - 2006/2/28/medline PY - 2005/11/3/entrez SP - 73 EP - 81 JF - Journal of immunological methods JO - J Immunol Methods VL - 307 IS - 1-2 N2 - Cytokines produced by Th1 or Th2 cells have been postulated to be important in the development of type 1 diabetes in humans and animal models, such as murine multiple low-dose streptozotocin (MLDSTZ)-induced diabetes. The aim of this study was to investigate cytokine production with or without in vitro depletion of plastic adherent cells from spleens isolated after MLDSTZ treatment. Spleen cells were prepared on day 14 from MLDSTZ- and saline-treated mice and divided into two fractions. One cell fraction was depleted of adherent cells by plastic adherence and the other was not. Both cell fractions were analysed by FACS for the distribution of immune cells. In other experiments, the cells were cultured for 48 h with concanavalin A stimulation. Supernatant samples were analysed by ELISA for TNFalpha, IFNgamma and IL-10 production. Either before or after the 48-h culture cytokine mRNA expression was determined by RT-PCR. Plastic adhesion decreased the macrophage numbers by approximately 30% and CD4(+)CD25(+) cells by about 60%. This was accompanied by increased medium levels of TNFalpha, IFNgamma and IL-10, which suggest that either CD4(+)CD25(+) cells, macrophages, or both, down-regulate production of both Th1 and certain Th2 cytokines. Depletion of adherent cells also decreased IL-4 mRNA amounts. MLDSTZ treatment increased the production of Th1 cytokines mainly at the protein level, and IL-10 mainly at the mRNA level. This indicates a sustained increase in Th1 production after MLDSTZ treatment and an increase in IL-10 that might reflect an attempt to counteract the MLDSTZ-induced immune damage. Plastic adhesion during cell preparation may affect the relative distribution of certain immune cells. SN - 0022-1759 UR - https://www.unboundmedicine.com/medline/citation/16263129/Impact_of_plastic_adhesion_in_vitro_on_analysis_of_Th1_and_Th2_cytokines_and_immune_cell_distribution_from_mice_with_multiple_low_dose_streptozotocin_induced_diabetes_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0022-1759(05)00313-3 DB - PRIME DP - Unbound Medicine ER -