Korean J Physiol Pharmacol.  2018 Jan;22(1):101-110. 10.4196/kjpp.2018.22.1.101.

Caffeic acid phenethyl ester protects against photothrombotic cortical ischemic injury in mice

• Hwang SA
• Kim CD
• Lee WS

• KMID: 2398560
• DOI: http://doi.org/10.4196/kjpp.2018.22.1.101

Abstract

In this study, we aimed to investigate the neuroprotective effects of caffeic acid phenethyl ester (CAPE), an active component of propolis purified from honeybee hives, on photothrombotic cortical ischemic injury in mice. Permanent focal ischemia was achieved in the medial frontal and somatosensory cortices of anesthetized male C57BL/6 mice by irradiation of the skull with cold light laser in combination with systemic administration of rose bengal. The animals were treated with CAPE (0.5-5 mg/kg, i.p.) twice 1 and 6 h after ischemic insult. CAPE significantly reduced the infarct size as well as the expression of tumor necrosis factor-α, hypoxiainducible factor-1α, monocyte chemoattractant protein-1, interleukin-1α, and indoleamine 2,3-dioxygenase in the cerebral cortex ipsilateral to the photothrombosis. Moreover, it induced an increase in heme oxygenase-1 immunoreactivity and interleukin-10 expression. These results suggest that CAPE exerts a remarkable neuroprotective effect on ischemic brain injury via its anti-inflammatory properties, thereby providing a benefit to the therapy of cerebral infarction.

Keyword

Caffeic acid phenethyl ester; Cerebral ischemia; Chemokine; Cytokine; Neuroinflammation

MeSH Terms

Animals
Brain Injuries
Brain Ischemia
Cerebral Cortex
Cerebral Infarction
Chemokine CCL2
Heme Oxygenase-1
Humans
Indoleamine-Pyrrole 2,3,-Dioxygenase
Interleukin-10
Ischemia
Male
Mice*
Necrosis
Neuroprotective Agents
Propolis
Rose Bengal
Skull
Urticaria
Chemokine CCL2
Heme Oxygenase-1
Indoleamine-Pyrrole 2,3,-Dioxygenase
Interleukin-10
Neuroprotective Agents
Propolis
Rose Bengal

Figure

• 

  Fig. 1 Effect of caffeic acid phenethyl ester (CAPE) on infarct size.(A) Representative photographs showing the ischemic lesion. Male C57BL/6 mice were treated with CAPE (0.5–5 mg/kg, i.p.) twice 1 and 6 h after ischemic insult, and were sacrificed 24 h after photothrombotic cortical ischemia. The pale non-stained area indicates the lesion of ischemic injury, while the deep red-colored area indicates the viable tissue. CAPE elicited the reduction in the infarct area (B) and infarct volume (C). The numbers in parentheses and columns indicate the numbers of animals. Data are presented as mean±SEM. *p<0.05; **p<0.01; ***p<0.001 vs. vehicle group.

• 

  Fig. 2 Immunohistochemical and Western blot analyses of expression of tumor necrosis factor-α (TNF-α).(A) Representative photomicrographs showing TNF-α positive cells (left panel) and number of TNF-α positive cells (right panel) in the cortical ischemic core region 24 h after photothrombosis. Treatment with CAPE 5 mg/kg significantly reduced the immunohistochemical staining of TNF-α positive cells compared to the vehicle group. Five animals were used in each group. Scale bar=100 µm. (B) Western blot analysis revealed that TNF-α expression was reduced by CAPE in a dose-dependent manner. Data are presented as mean±SEM. *p<0.05; ***p<0.001 vs. vehicle group.

• 

  Fig. 3 Immunohistochemical analysis of expression of hypoxia-inducible factor-1α (HIF-1α) and monocyte chemoattractant protein-1 (MCP-1).Representative photomicrographs showing HIF-1α and MCP-1 positive cells (left panels of A and B, respectively) and number of HIF-1α and MCP-1 positive cells (right panel panels of A and B, respectively) in the cortical ischemic core region 24 h after photothrombosis. Treatment with CAPE dose-dependently reduced the immunohistochemical staining of HIF-1α and MCP-1 positive cells compared to the vehicle group. Scale bar=100 µm. The numbers in columns indicate the numbers of animals. Data are presented as mean±SEM. *p<0.05; ***p<0.001 vs. vehicle group.

• 

  Fig. 4 Immunohistochemical analysis of expression of heme oxygenase-1 (HO-1).Representative photomicrographs showing HO-1 positive cells (left panel) and number of HO-1 positive cells (right panel) in the cortical ischemic core region 24 h after photothrombosis. CAPE dose-dependently increased the number of HO-1 positive cells compared to the vehicle group. Scale bar=100 µm. The numbers in columns indicate the numbers of animals. Data are presented as mean±SEM. *p<0.05; ***p<0.001 vs. vehicle group.

• 

  Fig. 5 Immunohistochemical and Western blot analyses of expression of indoleamine 2,3-dioxygenase (IDO).(A) Representative photomicrographs showing IDO positive cells (left panel) and number of IDO positive cells (right panel) in the cortical ischemic core region 24 h after photothrombosis. Treatment with CAPE 5 mg/kg significantly reduced the immunohistochemical staining of IDO positive cells compared to the vehicle group. Scale bar=100 µm. (B) Western blot analysis revealed that IDO expression was significantly reduced by CAPE 5 mg/kg. The numbers in columns indicate the numbers of animals. Data are presented as mean±SEM. **p<0.01; ***p<0.001 vs. vehicle group.

• 

  Fig. 6 Effect of CPAE on production of IL-1α and IL-10.The cerebral cortex ipsilateral to the photothrombosis was obtained 24 h after photothrombotic cortical ischemia, and was homogenized in lysis buffer. The supernatant of centrifuged sample was used for measuring the levels of IL-1α and IL-10 using ELISA (left and right panels, respectively). CAPE reduced the production of IL-1α (left panel) and enhanced production of IL-10 (right panel) in the ischemic region in a dose-dependent manner. Four animals were used in each group. ***p<0.001 vs. vehicle group.

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