Antiapoptotic effect of tacrolimus on cytokine-challenged human islets.
Our goal was to investigate whether previously related antiapoptotic and anti-inflammatory effects of tacrolimus could be useful in protecting human islets cultured in the presence of several proinflammatory mediators. Human islets obtained from cadaveric donors after intraductal infusion with collagenase, mechanical digestion, and continuous Ficoll gradient purification were cultured in RPMI-1640 medium for 24 h. Escherichia coli lipopolysaccharide (10 microg/ml) or interleukin-1 (50 UI/ml) + gamma-IF (1000 UI/ml) and low-dose tacroliumus (5 ng/ml) were added. Homogenized samples (300 IE) from five different donors where assigned to four different experimental groups (control, treatment, cytokines, and cytokines + treatment). To evaluate islet damage and apoptotic response, nucleosome content, Bcl-2 protein levels, caspase-3, -8, and -9 levels, and insulin concentration were measured. Also, TNF-alpha and IL-6 levels where assessed as indicators of the inflammatory response. All proapoptotic markers, TNF-alpha, and IL-6 levels were augmented after both LPS and cytokine stimulation. Tacrolimus reduced significantly all of them and restored baseline values of nucleosome and caspase-9 in both experiments and Bcl-2 and caspase-3 when IL-1 + gamma-IF was added. Twenty-four-hour insulin concentration diminished when LPS or IL-1 + gamma-IF were present. Tacrolimus treatment restored insulin levels in both experiments. These results suggest that in vitro apoptotic events and media insulin concentration decrease after proinflammatory stimulation can be reverted using low-dose tacrolimus.
Del Castillo
José M Balibrea
JM
Department of Surgery, Universidad Complutense, Madrid, Spain. balibrea@gmail.com
García-Martín
M Cruz
MC
Arias-Díaz
Javier
J
Giné
Elena
E
Vara
Elena
E
Cantero
José L Balibrea
JL
D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000378:metabolism; D053181:Caspase 8 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D002478:Cells, Cultured; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D007328:Insulin / Q000378:metabolism; D007371:Interferon-gamma / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D015850:Interleukin-6 / Q000378:metabolism; D007515:Islets of Langerhans / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D016559:Tacrolimus / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D007166:Immunosuppressive Agents; D007328:Insulin; D007375:Interleukin-1; D015850:Interleukin-6; D008070:Lipopolysaccharides; D019253:Proto-Oncogene Proteins c-bcl-2; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D016559:Tacrolimus