Katarzyna Wiglusz

Interactions of human serum albumin with meloxicam: Characterization of binding site

Human serum albumin (HSA) is the most prominent protein in plasma. The three-domain design of HSA provides a variety of binding sites for many ligands, including heme, bilirubin and drugs. Here, we report the effect of new generation, non-steroidal anti-inflammatory drug (NSAID) meloxicam on the albumin conformation and ligand binding. In the present work the interaction of meloxicam with HSA in aqueous solution at physiological pH has been investigated through circular dichroism and fluorescence spectroscopy. The strong quenching of the fluorescence clearly indicated that the binding of the drug to HSA changed the microenvironment of tryptophan residue and the tertiary structure of HSA. This was confirmed by the destabilization of the warfarin binding site. CD and fluorescence spectroscopic results showed marked reductions (about 40% decrease in the CD Cotton effect intensity, and approximately 15% decrease of the fluorescence intensity) in the affinity of albumin for bilirubin upon meloxicam binding. The strong inhibition of warfarin and ANS bound to protein after meloxicam modification compared with aspirin confirms that the binding site of both drugs is not the same.

Li+ activated nanohydroxyapatite doped with Eu3+ ions enhances proliferative activity and viability of human stem progenitor cells of adipose tissue and olfactory ensheathing cells. Further perspective of nHAP:Li+, Eu3+ application in theranostics

Spinal cord injuries (SCI) often require simultaneous regeneration of nerve tissue and bone. Hydroxyapatites are described as bioresorbable materials with proper biocompatibility and osteoconductivity, therefore its application for spinal surgery is considered. In this paper, we present repeatable method for developing nanocrystalline calcium hydroxyapatites structurally modified with Li+ ions (nHAP:Li+). Obtained biomaterials were profoundly characterized in terms of their physicochemical properties. Moreover, we have shown that nHAP:Li+ doped with europium (Eu3+) may serve as a theranostic agent, what additionally extend its potential usage for SCI treatment. The biocompatibility of nHAP:Li+ was determined using human olfactory ensheathing cells (hOECs) and adipose tissue-derived multipotent stromal cells (hASCs). Both population of cells are eagerly applied for cell-based therapies in SCI, mainly due to their paracrine activity. The extensive in vitro studies showed that nHAP:Li+ promotes the cells proliferation, viability and cell-cell interactions. Obtained results provides encouraging approach that may have potential application in regenerative medicine and that could fulfil the promise of personalized medicine - important in SCI treatment.

Multifunctional nanocrystalline calcium phosphates loaded with Tetracycline antibiotic combined with human adipose derived mesenchymal stromal stem cells (hASCs).

Osteoconductive drug delivery system composed of nanocrystalline calcium phosphates (Ca10(PO4)6(OH)2/β-Ca3(PO4)2) co-doped with Yb(3+)/Er(3+) ions loaded with Tetracycline antibiotic (TC) was developed. Their effect on human adipose derived mesenchymal stromal stem cells (hASCs) as a potential reconstructive biomaterial for bone tissue regeneration was studied. The XRD and TEM measurements were used in order to determine the crystal structure and morphology of the final products. The characteristics of nanocomposites with the TC and hASCs as potential regenerative materials as well as the antimicrobial activity of the nanoparticles against: Staphylococcus aureus ATCC 25923 as a model of the Gram-positive bacteria, Escherichia coli ATCC 8739 of the Gram-negative bacteria, were shown. These combinations can be a promising material for theranostic due to its regenerative, antimicrobial and fluorescent properties.

The effect of glycation on bovine serum albumin conformation and ligand binding properties with regard to gliclazide

Albumin, the major serum protein, plays a variety of functions, including binding and transporting endogenous and exogenous ligands. Its molecular structure is sensitive to different environmental modifiers, among which glucose is one of the most significant. In vivo albumin glycation occurs under physiological conditions, but it is increased in diabetes. Since bovine serum albumin (BSA) may serve as a model protein in in vitro experiments, we aimed to investigate the impact of glucose-mediated BSA glycation on the binding capacity towards gliclazide, as well as the ability of this drug to prevent glycation of the BSA molecule. To reflect normo- and hyperglycemia, the conditions of the glycation process were established. Structural changes of albumin after interaction with gliclazide (0-14μM) were determined using fluorescence quenching and circular dichroism spectroscopy. Moreover, thermodynamic parameters as well as energy transfer parameters were determined. Calculated Stern-Volmer quenching constants, as well as binding constants for the BSA-gliclazide complex, were lower for the glycated form of albumin than for the unmodified protein. The largest, over 2-fold, decrease in values of binding parameters was observed for the sample with 30mM of glucose, reflecting the poorly controlled diabetic state, which indicates that the degree of glycation had a critical influence on binding with gliclazide. In contrast to significant changes in the tertiary structure of BSA upon binding with gliclazide, only slight changes in the secondary structure were observed, which was reflected by about a 3% decrease of the α-helix content of glycated BSA (regardless of glucose concentration) in comparison to unmodified BSA. The presence of gliclazide during glycation did not affect its progress. The results of this study indicate that glycation significantly changed the binding ability of BSA towards gliclazide and the scale of these changes depended on glucose concentration. It may have a direct impact on the free drug fraction and its pharmacokinetic behavior, including the risk of hypoglycemic episodes or unexpected interactions with other ligands. The use of BSA in examining binding effects upon glycation seems to be good model for preliminary research and may be used to identify a potential drug response in a diabetic state.

Multispectroscopic studies of the interaction of folic acid with glycated human serum albumin

Abstract The interaction between glycated human serum albumin (gHSA) and folic acid (FA) was investigated by various spectroscopic techniques, such as fluorescence, circular dichroism, UV–vis absorption spectroscopy and electrophoretic light scattering technique. These methods characterize the binding properties of an albumin–folic acid system. The binding constants values (Ka) at 300 and 310 K are about 104 M−1. The standard enthalpy change (ΔH) and the standard entropy change (ΔS) were calculated to be ∼−20 kJ mol−1 and ∼16 J mol−1 K−1, respectively, which indicate characteristic electrostatic interactions between gHSA and folic acid. The CD studies showed that there are no significant conformational changes in the secondary structure of the protein. Moreover, the zeta potential measurements proved that under physiological conditions the gHSA–folic acid complex shows instability. No significant changes in the secondary structure of the protein and reversible drug binding are the desirable effect from pharmacological point of view. Communicated by Ramaswamy H. Sarma