ABSTRACT
<p><b>OBJECTIVE</b>To evaluate the role of water channel AQP4 in NMDA-induced brain injury in mice.</p><p><b>METHODS</b>In AQP4 gene knockout (AQP4(-/-)) mice, brain injury was induced by microinjection of NMDA into the cortex. The injured area was determined by toluidine blue staining, degenerated neurons were detected by Fluro-Jade B staining, and increased blood-brain barrier (BBB) permeability was evaluated by IgG immunostaining.</p><p><b>RESULT</b>Compared with wild-type mice, AQP4(-/-) mice exhibited increased cortical lesion area, aggravated neuron degeneration, and increased BBB disruption after NMDA microinjection.</p><p><b>CONCLUSION</b>AQP4 may play a protective role in NMDA-induced brain injury in mice.</p>
Subject(s)
Animals , Mice , Aquaporin 4 , Genetics , Physiology , Blood-Brain Barrier , Pathology , Brain , Pathology , Mice, Knockout , N-Methylaspartate , ToxicityABSTRACT
<p><b>OBJECTIVE</b>To prepare and identify a polyclonal antibody (pAb) against GPR17, a novel cysteinyl leukotriene receptor.</p><p><b>METHODS</b>Rabbits were immunized with KLH-coupled GPR17 peptide to prepare the pAb. The titer of the pAb in rabbit plasma was detected by indirect ELISA, and the specificity of the pAb was tested by antigen blockade. GPR17 tissue distribution was detected by Western blot with the pAb.</p><p><b>RESULTS</b>The pAb showed a titer as high as 1:16 364,and was not cross-reacted with the antigens of CysLT(1) and CysLT(2) receptors. A higher expression of GPR17 in the rat brain and heart was detected using the newly prepared pAb. The molecular weigh of GPR17 protein was about 43 kD.</p><p><b>CONCLUSION</b>The prepared GPR17 pAb has high sensitivity and specificity,and can be used in Western blot for detecting GPR17.</p>
Subject(s)
Animals , Humans , Rabbits , Rats , Antibodies, Monoclonal , Allergy and Immunology , Enzyme-Linked Immunosorbent Assay , Receptors, G-Protein-Coupled , Allergy and Immunology , Receptors, Leukotriene , Allergy and ImmunologyABSTRACT
<p><b>OBJECTIVE</b>To prepare and identify a polyclonal antibody against cysteinyl leukotriene receptor (CysLT(2)receptor).</p><p><b>METHODS</b>Rabbits were immunized with KLH-coupled CysLT(2) receptor peptide to prepare the polyclonal antibody (pAb). The titer of the pAb in rabbit plasma was detected by indirect ELISA, and the specificity of the pAb was tested by antigen blockade. The tissue distribution of CysLT(2) receptor was detected by Western blot and immunohistochemistry with the prepared pAb.</p><p><b>RESULT</b>The pAb showed a titer higher than 1/1047296, and was specific to CysLT(2) receptor, without cross-reaction with the antigens of CysLT(1) receptor and GPR17. A higher expression of CysLT(2) receptor in kidney, brain and lung of rats and mice was detected by Western blot analysis using the prepared pAb. The molecular weight of CysLT(2) receptor protein was about 40 kD. Immunohistochemical examination showed that CysLT(2) receptor was expressed mainly in the neuron, and partly in astrocytes in rat brain.</p><p><b>CONCLUSION</b>The prepared CysLT(2) receptor pAb has high sensitivity and specificity, and can be used in Western blot and immunohistochemistry.</p>
Subject(s)
Animals , Mice , Rabbits , Rats , Antibodies, Monoclonal , Allergy and Immunology , Brain , Metabolism , Kidney , Metabolism , Lung , Metabolism , Rats, Sprague-Dawley , Receptors, Leukotriene , Allergy and Immunology , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To determine whether the skilled reaching test is an objective method for evaluating long-term neurological deficits after focal cerebral ischemia in mice.</p><p><b>METHODS</b>In a reaching box, mice were trained to reach food pellets with their left forelimb through a 0.5 cm slit for 3 weeks. Then focal cerebral ischemia was induced by occluding the right middle cerebral artery, and the percentage of success in obtaining food was observed for 4 weeks. In comparison, the neurological deficit score, the holding angle in an inclined board test, and right turns in a corner test were simultaneously performed. At the end of the experiments, brain infarcts and neuron densities were determined.</p><p><b>RESULT</b>After focal cerebral ischemia, the percentage of success in the reaching test was reduced, the right turns in the corner test were increased, the neurological deficit score was increased, and the holding angle in the inclined board test was reduced as well. The holding angle recovered 5 d after ischemia, whereas other 3 indicators remained abnormal until 4 weeks. At the end of the experiments, the brain infarct volumes were increased, and the neuron densities in the cortex, hippocampal CA1 region and striatum were reduced in ischemic mice.</p><p><b>CONCLUSION</b>The skill reaching test is an objective and stable method for evaluating long-term neurological deficits after focal cerebral ischemia in mice.</p>