Reflection confocal imaging of type I and type III isozymes of hexokinase in PC12 cells

Reflection confocal microscopy was used to determine the intracellular distribution of Type-I and III isozymes of hexokinase (ATP:D-hexose 6-phosphotransferase, EC 2.7.1.1) in PC12 cells; detection was by staining with diaminobenzidine as a substrate for horseradish peroxidase-conjugated antimouse immunoglobulins bound to isozyme-specific monoclonal antibodies. With both isozymes, detection of the staining pattern was significantly enhanced by reflection confocal imaging compared with viewing with transmitted brightfield optics. For Type I, prominent staining of cytoplasmic organelles having a distribution consistent with that of mitochondria was noted. For Type III, intense staining at the nuclear periphery was observed. A distinct punctate pattern along the nuclear surface implied a nonuniform distribution of the Type III hexokinase and may represent preferential association with nuclear pore structures. A study of technical factors involved in optimizing the reflection image was conducted. We demonstrate that both the choice of objective and the thickness of the mounting medium are critical to successful imaging, and we describe a simple test for assessing the suitability of objectives in any system.     

Modifications induced by zinc,cadmium and mercury acetates on hexokinase in aqueous solutions

Preferential interactions of hexokinase with solvent were studied in aqueous solutions of Zn²⁺, Cd²⁺ and Hg²⁺ acetates. In addition, these preferential interactions were correlated with protein solubility in the solvent media and the influence of salts on the tertiary structure of hexokinase using viscometric studies. The viscometric results show that hexokinase can undergo a conformational change at 2·5mM of Zn²⁺ and Cd²⁺ salts, whereas the effect of Hg²⁺ acetate on hexokinase is evident at ten times smaller concentrations. The interaction parameter is always positive for all the salts studied, indicating that interaction of the salt and protein is thermodynamically unfavourable, although this effect is lower for the Zn²⁺ salt. The transfer free energy parameter and solubility data show that for hexokinase zinc acetate is a salting-in agent, mercury acetate is a salting-out agent and cadmium acetate cannot be clearly classified. © 1998 Society of Chemical Industry.     

鲨肝刺激物质降血糖作用机制的研究

目的:研究鲨肝刺激物质的降血糖作用机制。方法:采用四氧嘧啶糖尿病小鼠模型, 观察鲨肝刺激物质对糖尿病小鼠空腹血糖、果糖胺、胰岛素、肝糖原、己糖激酶、过氧化脂质等生化指标的影响。采用离体培养原代小鼠肝细胞, 研究鲨肝刺激物质对四氯化碳、对乙酰氨基酚所致肝细胞损伤的作用。结果:鲨肝刺激物质显著降低糖尿病小鼠空腹血糖和果糖胺水平, 增加血清胰岛素、肝糖原含量, 提高己糖激酶活性, 减轻四氯化碳、对乙酰氨基酚对原代小鼠肝细胞的损伤。结论:鲨肝刺激物质降血糖作用机制与保护胰岛和肝细胞功能、提高己糖激酶活性、促进肝糖原合成和抗氧化损伤作用密切相关。