Human MAP4 peptide

1. Neuropharmacological actions of all the possible stereoisomers of 3', 3'-difluoro-2-(carboxycyclopropyl)glycine (3',3'-difluoro-CCG) were compared with those of the corresponding 2-(carboxycyclopropyl)glycine (CCG) isomers in the isolated spinal cord of newborn rats. (2S,1'S,2'S)- and (2S,1'R,2'S)-2(2-carboxy-3,3-difluorocyclopropyl)glycin (L-F(2)CCG-I and L-F(2)CCG-IV) were the most potent in causing depolarization, their threshold concentrations being approximately 1 mu M. 2. The depolarization evoked by L-F(2)CCG-I (30 mu M) was depressed by (+)-alpha-methyl-4-carboxyphenylglycine (MCPG, 1 mM (n=4)) to 17 +/- 3% of the control: this depolarizing action was not decreased by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 100 mu M), and only slightly decreased by high concentrations of D-2-amino-5-phosphonopentanoic acid (D-AP5, 100 mu M), suggesting that L-F(2)CCG-I activates mainly metabotropic glutamate receptors. 3. L-F(2)CCG-I preferentially depressed the monosynaptic component of the spinal reflex approximately 3 times more effectively than (2S,1'S,2'S)-2-(carboxycyclopropyl)glycine (L-CCG-I). The depressant action of L-F(2)CCG-I (0.2 mu M-0.7 mu M) on monosynaptic excitation was antagonized by (2S,1'S,2'S)-2-methyl-2-(carboxycyclopropyl)glycine (MCCG, 0.3 mM-1 mM) and (S)-2-amino-2-methyl-4-phosphonobutanoic acid (MAP4, 0.3 mM). 4. DL-alpha-Aminopimelate (10 and 100 mu M) selectively potentiated the depression of monosynaptic excitation caused by L-CCG-I (0.2 mu M) and L-F(2)CCG-I (0.1 mu M). The actions of (2S,1'R,2'R,3'R)-2-(2,3-dicarboxycyclopropyl)glycine (DCG-IV) (50 nM-0.2 mu M), L-2-amino-4-phosphonobutanoic acid (L-AP4) 0.3-1 mu M), (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid ((1S,3R)-ACPD) (1-7 mu M) and baclofen (0.1-0.7 mu M) were unaffected by DL-alpha-aminopimelate. The threshold concentration for the potentiating actions of DL-alpha-aminopimelate was 3 mu M. 5. The depolarization induced by quisqualate (3 mu M, 10 s application) was increased to 115 +/- 2% and 137 +/- 5% of the control values during combined application of quisqualate with either 30 mu M or 100 mu DL-alpha-aminopimelate, respectively. 6. Following the application and subsequent washout of L-F(2)CCG-I, DL-alpha-aminopimelate (3-100 mu M) decreased the amplitude of the monosynaptic component of spinal reflexes in a concentration-dependent manner, indicating a 'priming' effect of L-F(2)CCG-I.

Background The presence of eosinophils in the airway is associated with asthma severity and risk of exacerbations. Cell-free eosinophil granules are found in tissues in eosinophilic diseases, including asthma. This suggests that eosinophils have lysed and released cellular content, likely harming tissues. Objective The present study explores the mechanism of CD32- and alpha M ss 2 integrin-dependent eosinophil cytolysis of IL3-primed blood eosinophils seeded on heat-aggregated immunoglobulin G (HA-IgG). Methods Cytoskeletal events and signalling pathways potentially involved in cytolysis were assessed using inhibitors. The level of activation of the identified events and pathways involved in cytolysis was measured. In addition, the links between these identified pathways and changes in degranulation (exocytosis) and adhesion were analysed. Results Cytolysis of IL3-primed eosinophils was dependent on the production of reactive oxygen species (ROS) and downstream phosphorylation of p-38 MAPK. In addition, formation of microtubule (MT) arrays was necessary for cytolysis and was accompanied by changes in MT dynamics as measured by phosphorylation status of stathmin and microtubule-associated protein 4 (MAP4), the latter of which was regulated by ROS production. Reduced ROCK signalling preceded cytolysis, which was associated with eosinophil adhesion and reduced migration. Conclusion and Clinical Relevance In this CD32- and alpha M ss 2 integrin-dependent adhesion model, lysing eosinophils exhibit reduced migration and ROCK signalling, as well as both MT dynamic changes and p-38 phosphorylation downstream of ROS production. We propose that interfering with these pathways would modulate eosinophil cytolysis and subsequent eosinophil-driven tissue damage.