Abstract

 
 
 
 Purpose: To investigate the neuroprotective efficacy of plant polyphenol luteolin against hypoxic ischemia (HI)-induced neuronal injury in neonatal rat pups.
 Methods: Postnatal rat pups (aged 7 days) were subjected to HI insult, and then treated with luteolin at a dose of 1.2 mg/kg body weight (bwt) for up to 4 days after HI injury. Following the sacrifice of the pups, the brain tissues were subjected to histological examination (H & E staining), as well as biochemical and antioxidant assays. Moreover, levels of cell death regulator proteins were determined by enzyme-linked immunosorbent assay (ELISA).
 Results: There was significant increase in the tissue levels of reactive oxygen species (ROS) and malondialdehyde (MDA), but marked decrease in mitochondrial membrane potential (ΔΨm) in the HI rat pups (p < 0.05). Furthermore, HI decreased the brain tissue levels of the antioxidants, SOD, CAT, GPX and GSH, in the postnatal pups. However, luteolin treatment significantly reversed the abnormal increase in the levels of ROS and MDA, but reduced the loss in mitochondrial membrane potential (ΔΨm), while increasing the levels of SOD, CAT GPX and GSH (p < 0.05). Furthermore, HI significantly increased the brain expression levels of the pro-apoptotic markers, i.e., cytosolic cytochrome C (cyt c), caspase-3 and caspase-9 in the pups exposed to HI, indicating neuronal cell death. However, the HI- induced increase in the expression levels of these pro-apoptotic factors was reduced by luteolin treatment.
 Conclusion: These results suggest that luteolin protects postnatal rat pups from hypoxic ischemic- induced brain damage (neuronal cell death) due to its antioxidant and free radical-scavenging activities. Therefore, luteolin may be a potential neuroprotective agent in the management of HI-associated complications.
 
 
 
Highlights
Perinatal hypoxic ischemic (HI) encephalopathy which is characterized by cognitive, retarded growth, motor and sensory problems, is a principal cause of disability in infants [1]
Total brain tissue homogenates from HI induced hemisphere were used for reactive oxygen species (ROS) estimation
Brain tissue samples from HI-induced pups showed significant increases in the levels of ROS and MDA (p < 0.05), while treatment with luteolin significantly reduced the abnormal increases in ROS in HI group brain tissue samples (p < 0.05)
Summary
Perinatal hypoxic ischemic (HI) encephalopathy which is characterized by cognitive, retarded growth, motor and sensory problems, is a principal cause of disability in infants [1]. Inhalation of 8 % oxygen-adjusted nitrogen is used widely to induce HI in rat pups This protocol was followed from previous studies, but with slight modifications [12]. The lipid peroxidation product malondialdehyde (MDA) was estimated using the method described in a previous report [14] In this method, 200 μL supernatant from brain tissue homogenate was added to 50 μL of 8.1% sodium dodecyl sulfate in a vial, and the vial was vortexed for few seconds. The concentration of nitrate by-products in brain tissue homogenates was measured using total nitric oxide assay kit from Beyotime, Jiangsu, China). A portion of brain tissue from each experimental group of pups was incubated with paraformaldehyde at 4 ̊C and dehydrated with graded concentrations of ethanol.
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