PDE4B2 is Strongly Expressed in Blood Vessels and is Inhibited by Rolipram, Ameliorating Hepatic Ischemia Reperfusion Injury in a Rat Model

BACKGROUND Ischemia reperfusion (IR) injury is caused by the loss and subsequent return of blood flow. It is a limiting factor in the success of liver transplantation. Reperfusion initiates inflammation and recruitment of neutrophils, which are main effectors of liver damage in IR. The Phosphodiesterase‐4 (PDE4) inhibitor rolipram is anti‐inflammatory, since PDEs degrade cAMP, and higher cAMP levels are known to dampen inflammatory signaling. Isoform PDE4B2 is strongly expressed in the brain, in neutrophils and activated monocytes, and in Kupffer cells in the liver. Here we show for the first time that PDE4B2 antibody also stains blood vessel cells in the liver. HYPOTHESIS Rolipram is able to protect the liver during IR injury. METHODS Male Sprague‐Daley rats were randomized to six groups (n=6–8 per group): 1) sham, 2) sham + rolipram, 3) 40% IR, 4) 40% IR + rolipram, 5) 70% IR, and 6) 70% IR + rolipram. Two levels of severity were attained by clamping either 40% (left lobe) or 70% (left and median lobes) of the rat liver. 1 hour of occlusion was followed by 3 hours of reperfusion. I.V. rolipram (3 mg/kg) was given at the end of the ischemic period. After sacrifice, blood chemistry, histology, and RT PCR were performed. ANOVA was followed by Tukey’s or Bonferroni’s tests. P values < 0.05 were significant. Frozen immunofluorescent histology revealed colocalization of PDE4B2 (short isoform) with CD31, a marker for blood vessels. RESULTS Both 40% and 70% IR elevated serum ALT, an indicator of liver damage. Rolipram lowered ALT levels in 40% IR. Histology showed necrosis and increased neutrophil infiltration, which improved with rolipram, in both 40% and 70% IR. Expression of PDE4B2 was elevated in 70% IR, and lowered by rolipram. Correlation analysis revealed that PDE4B2 mRNA increased with increasing ALT, necrosis, and neutrophil count. Both neutrophils and injury markers correlated most strongly with expression of the transcription factors (TFs) activating transcription factor‐3 (ATF3) and interferon regulatory factor‐1 (IRF1). These two genes also correlated positively with PDE4B2 expression. PDE4B2 antibody strongly stained some endothelial and perivascular cells of the liver, indicating that these cells may be playing some kind of role in the protection of rolipram in IR. CONCLUSION These data suggest that inhibition of PDE4 reduces IR injury by decreasing neutrophil recruitment, and that IR injury increases with TFs IRF1 and ATF3. Further experiments are needed to determine conclusive effects of this change in TF expression. Histology showed strong staining of PDE4B2 in blood vessels. Mechanisms of PDE4B2 inhibition protection of the liver in blood vessels cells, however, remain to be elucidated. Ultimately, liver injury was decreased by rolipram; therefore, phosphodiesterase‐4 inhibition is a potential therapy for IR injury. Support or Funding Information NIH