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Combination treatment with ethyl pyruvate and IGF-I exerts neuroprotective effects against brain injury in a rat model of neonatal hypoxic-ischemic encephalopathy.

Rong Z, Pan R, Chang L, Lee W - Int. J. Mol. Med. (2015)

Bottom Line: Under conditions of OGD, the LDH levels increased and neuronal viability decreased.Treatment with 0.5 mM EP or 25 ng/ml IGF‑I protected the neurons (P<0.05), exerting additive effects.This effect may be the result of reduced neuronal injury, and enhanced neurogenesis and maturation.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, P.R. China.

ABSTRACT
Neonatal hypoxic-ischemic (HI) brain injury causes severe brain damage in newborns. Following HI injury, rapidly accumulating oxidants injure neurons and interrupt ongoing developmental processes. The antioxidant, sodium pyruvate, has been shown to reduce neuronal injury in neonatal rats under conditions of oxygen glucose deprivation (OGD) and HI injury. In this study, we evaluated the effects of ethyl pyruvate (EP) and insulin‑like growth factor‑I (IGF‑I) alone or in combination in a similar setting. For this purpose, we used an in vitro model involving primary neonatal rat cortical neurons subjected to OGD for 2.5 h and an in vivo model involving unilateral carotid ligation in rats on post-natal day 7 with exposure to 8% hypoxia for 2.5 h. The cultured neurons were examined by lactate dehydrogenase (LDH) and cell viability assays. For the in vivo experiments, behavioral development was evaluated by the foot fault test at 4 weeks of recovery. 2,3,5‑Triphenyltetrazolium chloride monohydrate and cresyl violet staining were used to evaluate HI injury. The injured neurons were Fluoro‑Jade B-labeled, new neuroprecursors were double labeled with bromodeoxyuridine (BrdU) and doublecortin, new mature neurons were BrdU-labeled and neuronal nuclei were labeled by immunofluorescence. Under conditions of OGD, the LDH levels increased and neuronal viability decreased. Treatment with 0.5 mM EP or 25 ng/ml IGF‑I protected the neurons (P<0.05), exerting additive effects. Similarly, either the early administration of EP or delayed treatment with IGF‑I protected the neonatal rat brains against HI injury and improved neurological performance and these effects were also additive. This effect may be the result of reduced neuronal injury, and enhanced neurogenesis and maturation. On the whole, our findings demonstrate that the combination of the early administration of EP with delayed treatment with IGF‑I exerts neuroprotective effects against HI injury in neonatal rat brains.

No MeSH data available.


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Combined treatment with insulin-like growth factor-I (IGF-I) and ethyl pyruvate (EP) increases hypoxic-ischemic (HI) injury-induced neurogenesis. (A) High magnification view of the square in the hippocampus. Double immunostaining to identify the newborn neurons (panel a, merge) with doublecortin (DCX) immunostaining (panel b, red) and bromodeoxyuridine (BrdU) immunostaining (panel c, green) 72 h post-HI injury. Nuclei are stained with DAPI (panel d, blue). (B) Quantification of the number of newborn neurons (DCX+BrdU+). *P<0.05, **P<0.01 as indicated (n=6).
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f6-ijmm-36-01-0195: Combined treatment with insulin-like growth factor-I (IGF-I) and ethyl pyruvate (EP) increases hypoxic-ischemic (HI) injury-induced neurogenesis. (A) High magnification view of the square in the hippocampus. Double immunostaining to identify the newborn neurons (panel a, merge) with doublecortin (DCX) immunostaining (panel b, red) and bromodeoxyuridine (BrdU) immunostaining (panel c, green) 72 h post-HI injury. Nuclei are stained with DAPI (panel d, blue). (B) Quantification of the number of newborn neurons (DCX+BrdU+). *P<0.05, **P<0.01 as indicated (n=6).

Mentions: To examine the neuronal differentiation of BrdU+ cells, we double labeled BrdU+ cells with either DCX, a marker for immature neurons, or NeuN, a marker for mature neurons. At 72 h post-HI injury, the number of BrdU+DCX+ cells in the group subjected to HI injury was increased compared to that of the rats in the sham-operated group (Fig. 6). Although mainly distributed in the SGZ of the rats in the sham-operated group, the BrdU+DCX+ cells were also distributed in the granule cell layer (GCL) in the group subjected to HI (Fig. 6A, panel d), indicating that more BrdU+DCX+ cells may have migrated. Although the numbers of the BrdU+DCX+ cells in the group treated with EP were similar to those of the vehicle-treated group, the numbers were markedly increased in the IGF-I-treated group and the combined treatment group, indicating that IGF-I, but not EP, stimulated neurogenesis (Fig. 6B). At 4 weeks post-injury, the number of NeuN+BrdU+ neurons was also elevated (Fig. 7) in the group subjected to HI compared to the sham-operated group. Of note, the increase in the number of DCX+ cells (Fig. 6B) and NeuN+ cells (Fig. 7C), showing a similar pattern among the treatment groups, suggesting that IGF-I not only stimulated neurogenesis, but also neuronal survival and differentiation.


Combination treatment with ethyl pyruvate and IGF-I exerts neuroprotective effects against brain injury in a rat model of neonatal hypoxic-ischemic encephalopathy.

Rong Z, Pan R, Chang L, Lee W - Int. J. Mol. Med. (2015)

Combined treatment with insulin-like growth factor-I (IGF-I) and ethyl pyruvate (EP) increases hypoxic-ischemic (HI) injury-induced neurogenesis. (A) High magnification view of the square in the hippocampus. Double immunostaining to identify the newborn neurons (panel a, merge) with doublecortin (DCX) immunostaining (panel b, red) and bromodeoxyuridine (BrdU) immunostaining (panel c, green) 72 h post-HI injury. Nuclei are stained with DAPI (panel d, blue). (B) Quantification of the number of newborn neurons (DCX+BrdU+). *P<0.05, **P<0.01 as indicated (n=6).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494588&req=5

f6-ijmm-36-01-0195: Combined treatment with insulin-like growth factor-I (IGF-I) and ethyl pyruvate (EP) increases hypoxic-ischemic (HI) injury-induced neurogenesis. (A) High magnification view of the square in the hippocampus. Double immunostaining to identify the newborn neurons (panel a, merge) with doublecortin (DCX) immunostaining (panel b, red) and bromodeoxyuridine (BrdU) immunostaining (panel c, green) 72 h post-HI injury. Nuclei are stained with DAPI (panel d, blue). (B) Quantification of the number of newborn neurons (DCX+BrdU+). *P<0.05, **P<0.01 as indicated (n=6).
Mentions: To examine the neuronal differentiation of BrdU+ cells, we double labeled BrdU+ cells with either DCX, a marker for immature neurons, or NeuN, a marker for mature neurons. At 72 h post-HI injury, the number of BrdU+DCX+ cells in the group subjected to HI injury was increased compared to that of the rats in the sham-operated group (Fig. 6). Although mainly distributed in the SGZ of the rats in the sham-operated group, the BrdU+DCX+ cells were also distributed in the granule cell layer (GCL) in the group subjected to HI (Fig. 6A, panel d), indicating that more BrdU+DCX+ cells may have migrated. Although the numbers of the BrdU+DCX+ cells in the group treated with EP were similar to those of the vehicle-treated group, the numbers were markedly increased in the IGF-I-treated group and the combined treatment group, indicating that IGF-I, but not EP, stimulated neurogenesis (Fig. 6B). At 4 weeks post-injury, the number of NeuN+BrdU+ neurons was also elevated (Fig. 7) in the group subjected to HI compared to the sham-operated group. Of note, the increase in the number of DCX+ cells (Fig. 6B) and NeuN+ cells (Fig. 7C), showing a similar pattern among the treatment groups, suggesting that IGF-I not only stimulated neurogenesis, but also neuronal survival and differentiation.

Bottom Line: Under conditions of OGD, the LDH levels increased and neuronal viability decreased.Treatment with 0.5 mM EP or 25 ng/ml IGF‑I protected the neurons (P<0.05), exerting additive effects.This effect may be the result of reduced neuronal injury, and enhanced neurogenesis and maturation.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, P.R. China.

ABSTRACT
Neonatal hypoxic-ischemic (HI) brain injury causes severe brain damage in newborns. Following HI injury, rapidly accumulating oxidants injure neurons and interrupt ongoing developmental processes. The antioxidant, sodium pyruvate, has been shown to reduce neuronal injury in neonatal rats under conditions of oxygen glucose deprivation (OGD) and HI injury. In this study, we evaluated the effects of ethyl pyruvate (EP) and insulin‑like growth factor‑I (IGF‑I) alone or in combination in a similar setting. For this purpose, we used an in vitro model involving primary neonatal rat cortical neurons subjected to OGD for 2.5 h and an in vivo model involving unilateral carotid ligation in rats on post-natal day 7 with exposure to 8% hypoxia for 2.5 h. The cultured neurons were examined by lactate dehydrogenase (LDH) and cell viability assays. For the in vivo experiments, behavioral development was evaluated by the foot fault test at 4 weeks of recovery. 2,3,5‑Triphenyltetrazolium chloride monohydrate and cresyl violet staining were used to evaluate HI injury. The injured neurons were Fluoro‑Jade B-labeled, new neuroprecursors were double labeled with bromodeoxyuridine (BrdU) and doublecortin, new mature neurons were BrdU-labeled and neuronal nuclei were labeled by immunofluorescence. Under conditions of OGD, the LDH levels increased and neuronal viability decreased. Treatment with 0.5 mM EP or 25 ng/ml IGF‑I protected the neurons (P<0.05), exerting additive effects. Similarly, either the early administration of EP or delayed treatment with IGF‑I protected the neonatal rat brains against HI injury and improved neurological performance and these effects were also additive. This effect may be the result of reduced neuronal injury, and enhanced neurogenesis and maturation. On the whole, our findings demonstrate that the combination of the early administration of EP with delayed treatment with IGF‑I exerts neuroprotective effects against HI injury in neonatal rat brains.

No MeSH data available.


Related in: MedlinePlus