[Experimental study on the damage of immature brain induced by chronic treatment with exogenous glucocorticosteroid].
OBJECTIVE
To explore the possibility of brain damage induced by exogenous glucocorticosteroid (GC) at therapeutic level during early life.


METHODS
Totally 192 healthy Sprague-Dawley (SD) rats were selected and divided into three groups including PD7, PD15 and PD60 corresponding to three age-groups in human, i.e., the full-term newborn, one-year-old infant and adult, respectively. According to the therapeutic regime for infantile spasms, the dose of prednisone and ACTH was designed as 6 mg/(kg.d) and 150 U/(m(2).d) respectively. SD rats of different age-groups were treated with prednisone or ACTH and normal saline as the control for 4 days. The specimens were collected on Day 4 or 3 weeks after treatment. Body and brain weights were measured when the rats were sacrificed. Histological studies on the tissues of frontal lobe and hippocampus were performed by Nissl staining. Ultrastructural changes of brain were observed by the transmission electron microscopy. Expression of apoptosis-related proteins Bcl-2 and Bax in neurons was detected by immunohistochemistry. Neuronal apoptosis was detected by TUNEL. Mitochondria membrane potential of neurons in frontal lobe and hippocampus were detected by flow cytometry, and the Caspase-3 activity was detected by spectrophotometric assay.


RESULTS
(1) Significant reduction of brain weight [prednisone group of PD7: (0.78 +/- 0.08) g, control group: (0.89 +/- 0.08) g] and decrease of neurons numbers [(77.7 +/- 4.7) neurons per VF, control group: (102.3 +/- 5.9) neurons per VF] with cellular ultrastructural abnormalities in the frontal cortex were observed in infantile rats, including PD7 and PD15 groups. However none of the differences in brain weight and frontal cerebral cortex neurons were detected 3 weeks after drug discontinuation in PD7 group. (2) Compared with NS control, the numbers of TUNEL-positive cell (prednisone group of PD7: (114.8 +/- 8.1) cells per VF, control group: (56.3 +/- 5.6) cells per VF], Caspase-3 activity (345.3 +/- 13.0, control group: 166.3 +/- 6.4) and Bax protein expression (0.27 +/- 0.02, control group: 0.15 +/- 0.06) in neurons of the infant rats increased significantly but their potentials of mitochondrial membrane (112.9 +/- 8.6, control group: 140.5 +/- 5.6) were reduced remarkably, especially in PD7 group.


CONCLUSIONS
(1) Long-term use of ACTH or prednisone could induce brain damage. The younger the rats were, the more liable they were to the damage and more severe the damages were, and the less possibly to recover completely from the damage. (2) The pathogenesis of the brain injury induced by GC is primarily due to up-regulated Bax protein expression resulted from the enhanced ratio of Bax to Bcl-2 and increased mitochondria membrane potential, followed by Caspase enzymes activation and irreversible excessive apoptosis.
Cao
Jie
J
Clinical Electroneurophysiology Centre, Children's Hospital, Chongqing Medical University, Chongqing 400014, China.
Cai
Fang-cheng
FC
Chen
Jin
J
Zhang
Xiao-ping
XP
D000818:Animals; D000831:Animals, Newborn; D001921:Brain / Q000254:growth & development / Q000378:metabolism* / Q000473:pathology; D053148:Caspase 3 / Q000378:metabolism; D005938:Glucocorticoids / Q000009:adverse effects*; D008297:Male; D009474:Neurons / Q000378:metabolism*; D011241:Prednisone / Q000009:adverse effects; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D051028:bcl-2-Associated X Protein
D005938:Glucocorticoids; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; C505410:Casp3 protein, rat; D053148:Caspase 3; D011241:Prednisone