Differential subcellular distribution and colocalization of the microsomal and soluble epoxide hydrolases in cultured neonatal rat brain cortical astrocytes
| Autor: | Bruce D. Hammock, Seema Rawal, Christophe Morisseau, Amruthesh C. Shivachar |
|---|---|
| Rok vydání: | 2009 |
| Předmět: |
Epoxide hydrolase 2
Article Rats Sprague-Dawley Cellular and Molecular Neuroscience Western blot Glial Fibrillary Acidic Protein medicine Animals Cells Cultured Cerebral Cortex Epoxide Hydrolases Glial fibrillary acidic protein biology medicine.diagnostic_test Chemistry Subcellular localization Molecular biology Rats Blot Animals Newborn Biochemistry Astrocytes Microsomal epoxide hydrolase cardiovascular system biology.protein Cell fractionation Subcellular Fractions |
| Zdroj: | Journal of Neuroscience Research. 87:218-227 |
| ISSN: | 1097-4547 0360-4012 |
| DOI: | 10.1002/jnr.21827 |
| Popis: | The microsomal epoxide hydrolase (mEH) and soluble epoxide hydrolase (sEH) enzymes exist in a variety of cells and tissues, including liver, kidney and testis. However, very little is known about brain epoxide hydrolases. Here we report the expression, localization and subcellular distribution of mEH and sEH in cultured neonatal rat cortical astrocytes by immunocytochemistry, subcellular fractionation, western blotting and radiometric enzyme assays. Our results showed a diffused immunofluorescence pattern for mEH, which co-localized with the astroglial cytoskeletal marker, glial fibrillary acidic protein (GFAP). The GFAP-positive cells also expressed sEH which was mainly localized in the cytoplasm especially in and around the nucleus. Western blot analyses, revealed a distinct protein band with a molecular mass of ~50 kDa, the signal intensity of which increased about 1.5-fold in the microsomal fraction over the whole cell lysate and other subcellular fractions. The polyclonal anti-human sEH rabbit serum recognized a protein band with a molecular mass similar to that of purified sEH protein (~62 kDa), and the signal intensity increased 1.7-fold in the 105,000×g supernatant fraction over the cell lysate. Although the corresponding mEH enzyme activities generally corroborated with the immunocytochemical and western blotting data a low sEH enzyme activity was detected especially in the total cell lysate and in the soluble fractions. These results suggest that rat brain cortical astrocytes differentially co-express mEH and sEH enzymes. The differential subcellular localization of mEH and sEH may play a role in the cerebrovascular functions that are known to be affected by brain-derived vasoactive epoxides. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Plný text