Effect of granulocyte-colony stimulating factor on expression of selected proteins involved in regulation of apoptosis in the brain of newborn piglets after cardiopulmonary bypass and deep hypothermic circulatory arrest

Abstract

Objective: The study objective was to investigate the effect of granulocyte-colony stimulating factor on the expression of proteins that regulate apoptosis in newborn piglet brain after cardiopulmonary bypass and deep hypothermic circulatory arrest.

Methods: The newborn piglets were assigned to 3 groups: (1) deep hypothermic circulatory arrest (30 minutes of deep hypothermic circulatory arrest, 1 hour of low-flow cardiopulmonary bypass); (2) deep hypothermic circulatory arrest with prior injection of granulocyte-colony stimulating factor (17 μg/kg 2 hours before cardiopulmonary bypass); and (3) sham-operated. After 2 hours of post-bypass recovery, the frontal cortex, striatum, and hippocampus were dissected. The expression of proteins was measured by gel electrophoresis or protein arrays. Data are presented in arbitrary units. Statistical analysis was performed using 1-way analysis of variance.

Results: In the frontal cortex, only Fas ligand expression was significantly lower in the granulocyte-colony stimulating factor group when compared with the deep hypothermic circulatory arrest group. In the hippocampus, granulocyte-colony stimulating factor increased Bcl-2 (54.3 ± 6.4 vs 32.3 ± 2.2, P = .001) and serine/threonine-specific protein kinase (141.4 ± 19 vs 95.9 ± 21.1, P = .047) when compared with deep hypothermic circulatory arrest group. Caspase-3, Bax, Fas, Fas ligand, death receptor 6, and Janus protein tyrosine kinase 2 levels were unchanged. The Bcl-2/Bax ratio was 0.33 for deep hypothermic circulatory arrest group and 0.93 for the granulocyte-colony stimulating factor group (P = .02). In the striatum, when compared with the deep hypothermic circulatory arrest group, the granulocyte-colony stimulating factor group had higher levels of Bcl-2 (50.3 ± 7.4 vs 31.8 ± 3.8, P = .01), serine/threonine-specific protein kinase (132.7 ± 12.3 vs 14 ± 1.34, P = 2.3 × 10(6)), and Janus protein tyrosine kinase 2 (126 ± 17.4 vs 77.9 ± 13.6, P = .011), and lower levels of caspase-3 (12.8 ± 5.0 vs 32.2 ± 11.5, P = .033), Fas (390 ± 31 vs 581 ± 74, P = .038), Fas ligand (20.5 ± 11.5 vs 57.8 ± 15.6, P = .04), and death receptor 6 (57.4 ± 4.4 vs 108.8 ± 13.4, P = .007). The Bcl-2/Bax ratio was 0.25 for deep hypothermic circulatory arrest and 0.44 for the granulocyte-colony stimulating factor groups (P = .046).

Conclusions: In the piglet model of hypoxic brain injury, granulocyte-colony stimulating factor decreases proapoptotic signaling, particularly in the striatum.

Figure 1

Effect of DHCA and G-CSF on Bax, Fas-L and DR6 immunoreactivities in the frontal cortex of newborn brain. Bars represent the means ± SEM for the density of the bands for six independent experiments.

Figure 2

Effect of DHCA and G-CSF on Bcl-2, pAkt and pJAK2 immunoreactivities in the hippocampus of newborn brain. Bars represent the means ± SEM for the density of the bands for six independent experiments.

Figure 3

Effect of DHCA and G-CSF on Bcl-2, pAkt and pJAK2 immunoreactivities in the striatum of newborn brain. Bars represent the means ± SEM for the density of the bands for six independent experiments.

Figure 4

Effect of DHCA and G-CSF on Caspase3, Fas, Fas-L and DR6 immunoreactivities in striatum of newborn brain. Bars represent the means ± SEM for the density of the bands for six independent experiments.