ABSTRACT: 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.
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.
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).
In the piglet model of hypoxic brain injury, granulocyte-colony stimulating factor decreases proapoptotic signaling, particularly in the striatum.
The Journal of thoracic and cardiovascular surgery 02/2012; 143(6):1436-42. · 3.41 Impact Factor
ABSTRACT: To compare the effects of pH-stat and α-stat management before deep hypothermic circulatory arrest followed by a period of low-flow (two rates) cardiopulmonary bypass on cortical oxygenation and selected regulatory proteins: Bax, Bcl-2, Caspase-3, and phospho-Akt.
Piglets were placed on cardiopulmonary bypass, cooled with pH-stat or α-stat management to 18 °C over 30 mins, subjected to 30-min deep hypothermic circulatory arrest and 1-hr low flow at 20 mL/kg/min (LF-20) or 50 mL/kg/min (LF-50), rewarmed to 37 °C, separated from cardiopulmonary bypass, and recovered for 6 hrs.
Newborn piglets, 2-5 days old, assigned randomly to experimental groups.
Cortical oxygen was measured by oxygen-dependent quenching of phosphorescence; proteins were measured by Western blots. The means from six experiments ± sem are presented as % of α-stat. Significance was determined by Student's t test. For LF-20, cortical oxygenation was similar for α-stat and pH-stat, whereas for LF-50, it was significantly better using pH-stat. For LF-20, the measured proteins were not different except for Bax in the cortex (214 ± 24%, p = .006) and hippocampus (118 ± 6%, p = .024) and Caspase 3 in striatum (126% ± 7%, p = .019). For LF-50, in pH-stat group: In cortex, Bax and Caspase-3 were lower (72 ± 8%, p = .001 and 72 ± 10%, p = .004, respectively) and pAkt was higher (138 ± 12%, p = .049). In hippocampus, Bcl-2 and Bax were not different but pAkt was higher (212 ± 37%, p = .005) and Caspase 3 was lower (84 ± 4%, p = .018). In striatum, Bax and pAkt did not differ, but Bcl-2 increased (146 ± 11%, p = .001) and Caspase-3 decreased (81 ± 11%, p = .042).
In this deep hypothermic circulatory arrest-LF model, when flow was 20 mL/kg/min, there was little difference between α-stat and pH-stat management. However, for LF-50, pH-stat management resulted in better cortical oxygenation during recovery and Bax, Bcl-2, pAk, and Caspase-3 changes were consistent with lesser activation of proapoptotic signaling with pH-stat than with α-stat.
Pediatric Critical Care Medicine 03/2011; 12(2):e79-86. · 3.13 Impact Factor
ABSTRACT: To determine the effect of recovery with mild hypothermia after cardiopulmonary bypass (CPB) and deep hypothermic circulatory arrest (DHCA) on the activity of selected key proteins involved in initiation (Bax, Caspase-3) or inhibition of apoptotic injury (Bcl-2, increased ratio Bcl-2/Bax) in the brain of newborn piglets.
The piglets were placed on CPB, cooled with pH-stat management to 18 degrees C, subjected to 30 min of DHCA followed by 1h of low flow at 20 ml/kg/min, rewarmed to 37 degrees C (normothermia) or to 33 degrees C (hypothermia), separated from CPB, and monitored for 6h. Expression of above proteins was measured in striatum, hippocampus and frontal cortex by Western blots. The results are mean for six experiments+/-SEM.
There were no significant differences in Bcl-2 level between normothermic and hypothermic groups. The Bax levels in normothermic group in cortex, hippocampus and striatum were 94+/-9, 136+/-22 and 125+/-34 and decreased in the hypothermic group to 59+/-17 (p=0.028), 70+/-6 (p=0.002) and 48+/-8 (p=0.01). In cortex, hippocampus and striatum Bcl-2/Bax ratio increased from 1.23, 0.79 and 0.88 in normothermia to 1.96, 1.28 and 2.92 in hypothermia. Expression of Caspase-3 was 245+/-39, 202+/-74 and 244+/-31 in cortex, hippocampus and striatum in the normothermic group and this decreased to 146+/-24 (p=0.018), 44+/-16 (p=7 x 10(-7)) and 81+/-16 (p=0.01) in the hypothermic group.
In neonatal piglet model of cardiopulmonary bypass with circulatory arrest, mild hypothermia during post bypass recovery provides significant protection from cellular apoptosis, as indicated by lower expression of Bax and Caspase-3 and an increased Bcl-2/Bax ratio. The biggest protection was observed in striatum probably by decreasing of neurotoxicity of striatal dopamine.
European journal of cardio-thoracic surgery: official journal of the European Association for Cardio-thoracic Surgery 03/2009; 35(4):577-81; discussion 581. · 2.40 Impact Factor