| Localization of Cyclooxygenase Isozymes in Dermal Wound Healing in Mouse. |
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Jun Hyeok Koh, Kwang Seog Kim, Dae Young Kim, Sam Yong Lee, Bek Hyun Cho, Chun Sang Bae |
1Department of Plastic and Reconstructive Surgery, Chonnam National University Medical School, Kwangju, Korea. ir01212@hanmail. net
2Department of Anatomy, Chonnam National University Medical School, Kwangju, Korea. |
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| Abstract |
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Cyclooxygenase(COX)-1 and COX-2 expression in dermal wound healing of mouse was detected by immunohistochemistry and Western blot analysis. In order to gain more information on the functional importance of COX-1 and COX-2 in dermal wound healing, we analysed COX-1 and COX-2 protein levels using the Western blotting technique. In addition, we used immunohistochemistry to determine the cellular localization of the protein products.
The collected skins were rapidly frozen and kept at -70degrees Cuntil assayed. Each frozen skin was lysed with 0.5 ml of ice-cold solution. Large tissue debris and nuclear fragments were removed by two low-speed centrifugations and the resulting supernatant fraction was used for blots. The skin extracts were stored below -20degrees Cfor further experiments.
By Western blotting, compared to the activity of COX-2 in normal skin, its activity was increased at days 1, 4, 8, and 12 and was maximal at 1 day after incisional wound of mouse skin whereas COX-1 was barely detectable.
In normal skin, COX-1 immunostaining was observed among the basal cells of epidermis whereas COX-2 immunostaining was detected in the more differentiated, suprabasal keratinocytes. At post-incision 1-4 days, COX-2 staining was particularly prominent in the inflammatory cells, and at day 8, many macrophage-like cells were stained positively. COX-2 immunoreactive fibroblast, macrophage-like cells, and newly formed vascular endothelial cells were increased in number at 12 days after incision.
These data suggest that COX-2 is constitutively expressed, just as is COX-1, in epidermis and is associated with keratinocyte differentiation. In addition, these findings support the well-established role for COX-2, the prostaglandins that they generate, as mediators of inflammatory response. |
| Keywords:
Cyclooxygenase (COX)-1; Cyclooxygenase (COX)-2; Immunohistochemistry; Western blot analysis; Localization |
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