New Application of the Commercially Available Dye Celestine Blue B as a Sensitive and Selective Fluorescent "Turn-On" Probe for Endogenous Detection of HOCl and Reactive Halogenated Species.

Hypochlorous acid (HOCl) derived from hydrogen peroxide and chloride anion by myeloperoxidase (MPO) plays a significant role in physiological and pathological processes. Herein we report a phenoxazine-based fluorescent probe Celestine Blue B (CB) that is applicable for HOCl detection in living cells and for assaying the chlorinating activity of MPO. A remarkable selectivity and sensitivity (limit of detection is 32 nM), along with a rapid "turn-on" response of CB to HOCl was demonstrated. Furthermore, the probe was able to detect endogenous HOCl and reactive halogenated species by fluorescence spectroscopy, confocal microscopy, and flow cytometry techniques. Hence, CB is a promising tool for investigating the role of HOCl in health and disease and for screening the drugs capable of regulating MPO activity.

Functional link between ferroxidase activity of ceruloplasmin and protective effect of apo-lactoferrin: studying rats kept on a silver chloride diet.

Strongly pronounced argyrosis caused by adding AgCl to the feed of laboratory rats efficiently mimics the deficiency of ceruloplasmin (CP) ferroxidase activity. Bringing the concentration of AgCl in the feedstuff of lactating rats to 250 mg % and keeping their progeny (Ag-rats) for 3 months on the same silver-containing feed provided the serum iron content 1.4 times lower than that in the control group. Besides, the ferroxidase activity of CP dropped to zero. In CP purified from sera of Ag-rats two copper ions were substituted with two silver ions. Using rat models of both post-hemorrhagic and hemolytic anemia we showed that the deficiency of CP ferroxidase activity in Ag-rats affects the iron content in serum, though does not prevent the recovery of hemoglobin level accompanied by exhaustion of iron caches in liver and spleen. When apo-lactoferrin (apo-LF) was administered to Ag-rats suffering from either post-hemorrhagic or hemolytic anemia, both hemoglobin and serum iron were restored more rapidly than in the control animals. In independent experiments Ag-rats were compared with those fed on regular diet and the former displayed a prolonged 3-day stabilization of hypoxia-inducible factors 1 and 2 alpha (HIF-1a and HIF-2a) along with an increased serum concentration of erythropoietin. Introduction to Ag-rats of active CP separately or together with apo-LF reduced that effect to 1 day only. It is concluded that saturation of apo-LF with iron, provided by active CP, can strongly affect its protective capacity.