Effects of recombinant human lactoferrin on calcium signaling and functional responses of human neutrophils.

Lactoferrin is a non-heme iron-binding glycoprotein with multiple health-beneficial functions including antimicrobial, antioxidant, anticarcinogenic, and immunomodulatory effects. There is emerging evidence that neutrophils may serve as targets of lactoferrin in vivo, and here we show how recombinant human lactoferrin (rhLf) can contribute to this regulation. Indeed, our results demonstrate that rhLf binds efficiently to human neutrophils and induces a variety of early cellular responses such as mobilization of intracellular Ca2+, remodeling of actin cytoskeleton, and degranulation (release of lysozyme and myeloperoxidase). In addition, rhLf facilitates lectin-induced H2O2 production and stabilization of lectin-induced cellular aggregates. The role of calcium signaling seems to be essential for rhLf-induced activation of neutrophils, as Ca2+-chelators inhibit degranulation response while lectin-induced H2O2 production correlates significantly with cytoplasmic Ca2+ elevation. Taken together, our findings justify that rhLf can activate neutrophil functions in a calcium-dependent manner and hence, can potentiate innate immune responses.

Protective Effect of Dinitrosyl Iron Complexes with Glutathione in Red Blood Cell Lysis Induced by Hypochlorous Acid.

Hypochlorous acid (HOCl), one of the major precursors of free radicals in body cells and tissues, is endowed with strong prooxidant activity. In living systems, dinitrosyl iron complexes (DNIC) with glutathione ligands play the role of nitric oxide donors and possess a broad range of biological activities. At micromolar concentrations, DNIC effectively inhibit HOCl-induced lysis of red blood cells (RBCs) and manifest an ability to scavenge alkoxyl and alkylperoxyl radicals generated in the reaction of HOCl with tert-butyl hydroperoxide. DNIC proved to be more effective cytoprotective agents and organic free radical scavengers in comparison with reduced glutathione (GSH). At the same time, the kinetics of HOCl-induced oxidation of glutathione ligands in DNIC is slower than in the case of GSH. HOCl-induced oxidative conversions of thiolate ligands cause modification of DNIC, which manifests itself in inclusion of other ligands. It is suggested that the strong inhibiting effect of DNIC with glutathione on HOCl-induced lysis of RBCs is determined by their antioxidant and regulatory properties.

Immunoreactivity changes of human serum albumin and alpha-1-microglobulin induced by their interaction with dendrimers.

Dendrimers are hyperbranched polymers for delivery of therapeutic genetic material to cancer cells. The fine tuning chemical modifications of dendrimers allow for the modification of the composition. The architecture and the properties of dendrimers are key factors to improve their in vitro and in vivo properties such as biocompatibility with cells and tissues and their pharmacokinetic/pharmacodynamic behavior. The side effects of dendrimers on structure and function of proteins is an important question that must be addressed. We herein describe the effect of newly synthesized piperidine-based cationic phosphorous dendrimers of 2 generations and commercial cationic, neutral and anionic poly(amidoamine) (PAMAM) dendrimers of 4th generation on immunochemical properties of 2 serum proteins: human serum albumin (HSA) and alpha-1-microglobulin (A1M). Both can bind and transfer ligands in blood, including hormones, fatty acids, toxins and drugs, and have immunoreactivity properties. Comparing the effects of piperidinium-terminated phosphorus and cationic, neutral and anionic PAMAM dendrimers on HSA and A1M, we conclude that, in the case of equimolar complexes, these dendrimers had no significant effect on immunoreactivity of proteins. In contrast, the formation of complexes in which a protein is fully bound to dendrimers leads to partial (1.2-2.3 times) reduction in protein immunoreactivity. The most important fact is that dendrimer-induced change in immunoreactivity of proteins is not complete, even if the protein is entirely bound by dendrimers. This means that the application of dendrimers in vivo will not totally hamper the immunoreactivity of these proteins and antibodies.

Dendronization of gold nanoparticles decreases their effect on human alpha-1-microglobulin.

Gold nanoparticles are new kinds of nanomaterials. Their large surface-to-volume ratio, stability, excellent biocompatibility, low toxicity and functionality make them very attractive for biomedical applications. Therefore we have analyzed how dendronized gold nanoparticles interact with human alpha-1-microglobulin. This is a glycoprotein of ∼30kDa present in blood plasma and some tissues of the human body. Comparing 3 nanoparticles with different dendronization, we conclude that the effect of a nanoparticle on the structure of alpha-1-microglobulin significantly decreased with second and third generations dendrons as a result of less exposure of the metal cores in the nanoparticles. These interactions indicate weak changes in the immunochemical properties of the protein, whereas the dendron coating had no effect. Thus, dendronization of gold nanoparticles helps to modify their binding properties by shielding them from interactions with plasma proteins.