J Vet Sci.  2015 Sep;16(3):245-251. 10.4142/jvs.2015.16.3.245.

Comparison of pharmacological and genetic inhibition of cyclooxygenase-2: effects on adult neurogenesis in the hippocampal dentate gyrus

• Nam SM
• Kim JW
• Yoo DY
• Choi JH
• Kim W
• Jung HY
• Won MH
• Hwang IK
• Seong JK
• Yoon YS

• KMID: 2344296
• DOI: http://doi.org/10.4142/jvs.2015.16.3.245

Abstract

Inducible cyclooxygenase-2 (COX-2) has received much attention because of its role in neuro-inflammation and synaptic plasticity. Even though COX-2 levels are high in healthy animals, the function of this factor in adult neurogenesis has not been clearly demonstrated. Therefore, we performed the present study to compare the effects of pharmacological and genetic inhibition of COX-2 on adult hippocampal neurogenesis. Physiological saline or the same volume containing celecoxib was administered perorally every day for 5 weeks using a feeding needle. Compared to the control, pharmacological and genetic inhibition of COX-2 reduced the appearance of nestin-immunoreactive neural stem cells, Ki67-positive nuclei, and doublecortin-immunoreactive neuroblasts in the dentate gyrus. In addition, a decrease in phosphorylated cAMP response element binding protein (pCREB) at Ser133 was observed. Compared to pharmacological inhibition, genetic inhibition of COX-2 resulted in significant reduction of neural stem cells, cell proliferation, and neuroblast differentiation as well as pCREB levels. These results suggest that COX-2 is part of the molecular machinery that regulates neural stem cells, cell proliferation, and neuroblast differentiation during adult hippocampal neurogenesis via pCREB. Additionally, genetic inhibition of COX-2 strongly reduced neural stem cell populations, cell proliferation, and neuroblast differentiation in the dentate gyrus compared to pharmacological inhibition.

Keyword

cell proliferation; cyclooxygenase-2; hippocampus; neural stem cells; neuroblast differentiation

MeSH Terms

Animals
Celecoxib/*pharmacology
Cell Differentiation/drug effects/physiology
Cell Proliferation/drug effects/physiology
Cyclooxygenase 2/*genetics/metabolism
Cyclooxygenase 2 Inhibitors/*pharmacology
Dentate Gyrus/drug effects/*physiology
Male
Mice
Mice, Knockout
Neural Stem Cells/drug effects/physiology
Neurogenesis/drug effects
Celecoxib
Cyclooxygenase 2
Cyclooxygenase 2 Inhibitors

Figure

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  Fig. 1 Changes in body weight (A) and food intake (B) of vehicle (V), celecoxib-treated (COX-I), and cyclooxygenase-2 (COX-2) knockout (COX-2-KO) mice (n = 5 per group).

• 

  Fig. 2 COX-2 immunoreactivity in the dentate gyrus of the V (A), COX-I (B), and COX-2-KO (C) groups. In the V animals, COX-2 immunoreactivity (arrows) was observed in the granule cell layer (GCL) and polymorphic layer (PL) of the dentate gyrus. In the COX-I and COX-2-KO groups, COX-2 immunoreactivity was weak compared to that in the V group. (D) The relative optical density (ROD) of the V, COX-I, and COX-2-KO groups (n = 5 per group) expressed as a percentage of the value for COX-2 immunoreactivity in the dentate gyrus of the V group per section; *p < 0.05, indicating a significant difference relative to the V group. All data are presented as the mean values ± standard error of the mean (SEM). ML: molecular layer, GCL: granule cell layer, PoL: polymorphic layer. Scale bar = 25 µm.

• 

  Fig. 3 Immunohistochemical staining for nestin expression in the dentate gyrus of the V (A), COX-I (B), and COX-2-KO (C) groups. Nestin-positive cells and fibers were observed in the granular cell layer (GCL) and subgranular zone of the dentate gyrus. Note that nestin-expressing cells and fibers were weakly detected in the COX-I and COX-2-KO groups compared to the V group. The degree of nestin reduction was prominent in the COX-2-KO mice compared to the COX-I group. (D) The ROD expressed as a percentage of the value for nestin immunoreactivity in the dentate gyrus per section of the V, COX-I, and COX-2-KO groups (n = 5 per group; *p < 0.05, indicating a significant difference relative to V group). All data are presented as the mean ± SEM. Scale bar = 50 µm.

• 

  Fig. 4 Immunohistochemical staining for Ki67 in the dentate gyrus of the V (A), COX-I (B), and COX-2-KO (C) groups. Ki67-positive nuclei were observed in the subgranular zone (arrows) of the dentate gyrus. Few Ki67-positive nuclei were found in the COX-2-KO group compared to the V and COX-I groups. In particular, the number of Ki67-positive nuclei was noticeably decreased in the COX-2-KO group compared to the other groups. (D) The mean number of Ki67-positive nuclei per section in all groups (n = 5 per group; *p < 0.05, indicating a significant difference relative to the V group). All data are shown as the mean ± SEM. Scale bar = 50 µm.

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  Fig. 5 Immunohistochemical staining for DCX in the dentate gyrus of the V (A and B), COX-I (C and D), and COX-2-KO (E and F) groups. DCX-immunoreactive neuroblasts were detected in the subgranular zone (arrows) of the dentate gyrus. Note that the number of cell bodies and dendrites of DCX-positive neuroblasts were prominently decreased in the COX-2-KO group compared to the V and COX-I groups. (G and H) The mean number of DCX-immunoreactive neuroblasts per section (G) and ROD expressed as a percentage of DCX immunoreactivity per section (H) in the dentate gyrus of the V, COX-I, and COX-2-KO groups relative to the V group (n = 5 per group; ap < 0.05, indicating a significant difference compared to the V group; bp < 0.05, a significant difference relative to the COX-I group). All data are presented as the mean ± SEM. Scale bars = 50 µm (A, C, and E) or 25 µm (B, D, and F).

• 

  Fig. 6 Immunohistochemical staining for cAMP response element binding protein phosphorylated at Ser133 (pCREB) in the dentate gyrus of the V (A), COX-I (B), and COX-2-KO (C) groups. Note that the number of pCREB-positive nuclei (arrows) decreased in both the COX-I and COX-2-KO groups compared to the V group. The number of pCREB-immunoreactive nuclei in the COX-2-KO group was more noticeably decreased compared to the COX-I groups. (D) The mean number of pCREB-positive nuclei per section in all groups (n = 5 per group; ap < 0.05, indicating a significant difference compared to the V group; bp < 0.05, a significant difference relative to the COX-I group). All data are presented as the mean ± SEM. Scale bar = 50 µm.

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