PANDER-induced cell-death genetic networks in islets reveal central role for caspase-3 and cyclin-dependent kinase inhibitor 1A (p21).
PANcreatic DERived factor is an islet-specific cytokine that promotes apoptosis in primary islets and islet cell lines. To elucidate the genetic mechanisms of PANDER-induced cell death we performed expression profiling using the mouse PancChip version 5.0 in conjunction with Ingenuity Pathway Analysis. Murine islets were treated with PANDER and differentially expressed genes were identified at 48 and 72 h post-treatment. 64 genes were differentially expressed in response to PANDER treatment. 22 genes are associated with cell death. In addition, the genes with the highest fold change were linked with cell death or apoptosis. The most significantly affected gene at 48 h was the downregulated cyclin-dependent kinase inhibitor 1A (CDKN1A or p21). Approximately half of the genes impacted at 72 h were linked to cell death. Cell death differentially expressed genes were confirmed by quantitative RT-PCR. Further analysis identified cell death genetic networks at both time points with 21 of the 22 cell death genes related in various biological pathways. Caspase-3 (CASP3) was biologically linked to CDKN1A in several genetic networks and these two genes were further examined. Elevated cleaved CASP3 levels in PANDER-treated beta-TC3 insulinoma cells were found to abrogate CDKN1A expression. Levels of CDKN1A were not affected in the absence of cleaved CASP3. PANDER-induced downregulation of CDKN1A expression coupled with induced CASP3-activation may serve a central role in islet cell death and offers further insight into the mechanisms of cytokine-induced beta-cell apoptosis.
Burkhardt
Brant R
BR
Department of Pathology and Laboratory Medicine, 803D Abramson Research Center 3516 Civic Center Blvd., Children's Hospital of Philadelphia, Philadelphia, PA 19104-4318, United States. burkhardtb@email.chop.edu
Greene
Scott R
SR
White
Peter
P
Wong
Ryan K
RK
Brestelli
John E
JE
Yang
Jichun
J
Robert
Claudia E
CE
Brusko
Todd M
TM
Wasserfall
Clive H
CH
Wu
Jianmei
J
Atkinson
Mark A
MA
Gao
Zhiyong
Z
Kaestner
Klaus H
KH
Wolf
Bryan A
BA
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D045744:Cell Line, Tumor; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism*; D016207:Cytokines / Q000502:physiology*; D015536:Down-Regulation; D007515:Islets of Langerhans / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D020411:Oligonucleotide Array Sequence Analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction
C493957:Cdkn1a protein, mouse; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016207:Cytokines; C504749:PANDER protein, mouse; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases