Modulation of Apoptosis Pathways by Oxidative Stress and Autophagy in β Cells

Table 2

Mean fold (>2.0) changes in apoptosis pathway gene expression in islets after exposure to oxidative stress (H₂O₂ and peroxynitrite), hypoxia, and cytokines.

Genes	Role in apoptosis	Peroxynitrite	H2O2	Hypoxia	Cytokines
Bcl2A1	Anti-apoptotic; Bcl2 family	3.2	2.1	2.4	11.4
Bid	Proapoptotic; BH3 only	3.7	3.5	2.5	3.4
Fas	Extrinsic; receptor	4.3	3.1	3.4	7.4
Fas ligand	Extrinsic; receptor ligand	2.4	2.1	NS	3.4
TRAIL	Extrinsic; receptor ligand	4.6	3.6	NS	4.2
A20	Extrinsic; anti-apoptotic	2.3	NS	2.5	7.4
c-Flip	Extrinsic; anti-apoptotic	2.6	NS	NS	4.2
Caspase-3	Marker for apoptosis	4.5	3.2	3.2	2.3
BIRC3	Caspase inhibitor	3.2	2.4	NS	8.7
CARD8	Caspase-9 inhibitor	−2.3	−2.4	−2.1	−2.3
BRAF	Signaling kinase	−3.7	−3.5	−2.5	NS
BFAR	Anti-apoptotic; links both pathways	−4.8	−3.5	−4.2	NS
CIDE-A	Causes DNA fragmentation	3.2	2.4	3.8	NS
BNIP3	Autophagy Inducer	2.6	NS	5.6	NS

Human islets (2000 IEQ) were exposed to 200 μM of peroxynitrite, 200 μM of H₂O₂ or a mixture of cytokines (2 ng/mL of IL-1β, 10 ng/mL of TNF-α and 10 ng/mL of IFN-γ) for 24 h or cultured under hypoxic conditions (1% oxygen) for 8 h. The cDNA synthesized from isolated RNA was mixed with Master Mix containing SYBR Green and distributed into 96 wells containing primers for the 84 genes associated with the apoptotic pathway. Five housekeeping control genes and three RNA and PCR quality controls were also included. PCR analysis was carried out and the fold changes between control and treated were calculated based on Δ𝐶_𝑡 and corrected for GAPDH expression. Results are the mean obtained from four different batches of human islets. NS: not significant.