Unraveling the Host-Selective Toxic Interaction of Cassiicolin with Lipid Membranes and Its Cytotoxicity.

Cassiicolin (Cas), a toxin produced by Corynespora cassiicola, is responsible for Corynespora leaf fall disease in susceptible rubber trees. Currently, the molecular mechanisms of the cytotoxicity of Cas and its host selectivity have not been fully elucidated. Here, we analyzed the binding of Cas1 and Cas2 to membranes consisting of different plant lipids and their membrane disruption activities. Using high-speed atomic force microscopy and confocal microscopy, we reveal that the binding and disruption activities of Cas1 and Cas2 on lipid membranes are strongly dependent on the specific plant lipids. The negative phospholipids, glycerolipids, and sterols are more sensitive to membrane damage caused by Cas1 and Cas2 than neutral phospholipids and betaine lipids. Mature Cas1 and Cas2 play an essential role in causing membrane disruption. Cytotoxicity tests on rubber leaves of Rubber Research Institute of Vietnam (RRIV) 1, RRIV 4, and Prang Besar (PB) 255 clones suggest that the toxins cause necrosis of rubber leaves, except for the strong resistance of PB 255 against Cas2. Cryogenic scanning electron microscopy analyses of necrotic leaf tissues treated with Cas1 confirm that cytoplasmic membranes are vulnerable to the toxin. Thus, the host selectivity of Cas toxin is attained by the lipid-dependent binding activity of Cas to the membrane, and the cytotoxicity of Cas arises from its ability to form biofilm-like structures and to disrupt specific membranes.

Metabolomic analysis and mass spectrometry imaging after neonatal stroke and cell therapies in mouse brains.

Ischemic brain injury provokes complex, time-dependent downstream pathways that ultimately lead to cell death. We aimed to demonstrate the levels of a wide range of metabolites in brain lysates and their on-tissue distribution following neonatal stroke and cell therapies. Postnatal day 12 mice underwent middle cerebral artery occlusion (MCAO) and were administered 1 × 105 cells after 48 h. Metabolomic analysis of the injured hemisphere demonstrated that a variety of amino acids were significantly increased and that tricarboxylic acid cycle intermediates and some related amino acids, such as glutamate, were decreased. With the exception of the changes in citric acid, neither mesenchymal stem/stromal cells nor CD34+ cells ameliorated these changes. On-tissue visualization with matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) imaging revealed that the signal intensity of glutamate was significantly decreased in the infarct area, consistent with the metabolomic analysis, while its intensity was significantly increased in the peri-infarct area after MCAO. Although cell therapies did not ameliorate the changes in metabolites in the infarct area, mesenchymal stem cells ameliorated the increased levels of glutamate and carnitine in the peri-infarct area. MALDI-MS imaging showed the location-specific effect of cell therapies even in this subacute setting after MCAO. These methodologies may be useful for further investigation of possible treatments for ischemic brain injury.