I.I. Nikolskaya

Carbohydrates regulate the dimerization of angiotensin‐converting enzyme

Summary. Regulation of the catalytic activity and supramolecular structure of angiotensin‐converting enzyme was studied in reverse micelles of Aerosol OT in octane as biomembrane model. The kinetic experiments and the sedimentation analysis demonstrated that the enzyme can function both in monomeric and dimeric form. The degree of dimerization was strongly dependent on the concentration and structure of mono‐ and disaccharides added to the media, indicating the specific role of carbohydrates in forming the supramolecular structure of angiotensin‐converting enzyme. The existence of carbohydrate‐binding center on the enzyme molecule is proposed.

Thermo-sensitive gel for prolongation of ophthalmic drug action

A new thermo-sensitive ophthalmic formulation has been developed on the basis of poly-N-isopropylacrylamide (poly-NIPAA) gel suspension. Gel particles effectively entrap the substances from aqueous solution at 20°C (hydrophilic state of a polymer) and form a thin film after contact with the surface of rabbit eye cornea when the gel becomes hydrophobic in character. In vitro time-consuming release of entrapped compounds from poly-NIPAA gel after a rise of temperature from 20°C to physiological temperature 37°C depends on the nature of the substance. The studies in vivo demonstrate that proxodolol (an β 1,2 α 1 -adrenoblocker, lowering intraocular pressure) in the form of poly-NIPAA gel suspension exhibits larger physiological effect on the intraocular pressure in comparison with the effect of the same drug in the form of traditional eye drops. Prolongation of action of physiologically active compounds in the form of thermo-sensitive gel suspension is expected to be sufficiently longer for more hydrophobic substances or formulation with higher molecular weight.

Superoxide Dismutase 1 Nanozyme for Treatment of Eye Inflammation

Use of antioxidants to mitigate oxidative stress during ocular inflammatory diseases has shown therapeutic potential. This work examines a nanoscale therapeutic modality for the eye on the base of antioxidant enzyme, superoxide dismutase 1 (SOD1), termed “nanozyme.” The nanozyme is produced by electrostatic coupling of the SOD1 with a cationic block copolymer, poly(L-lysine)-poly(ethyleneglycol), followed by covalent cross-linking of the complexes with 3,3′-dithiobis(sulfosuccinimidylpropionate) sodium salt. The ability of SOD1 nanozyme as well as the native SOD1 to reduce inflammatory processes in the eye was examined in vivo in rabbits with immunogenic uveitis. Results suggested that topical instillations of both enzyme forms demonstrated anti-inflammatory activity; however, the nanozyme was much more effective compared to the free enzyme in decreasing uveitis manifestations. In particular, we noted statistically significant differences in such inflammatory signs in the eye as the intensities of corneal and iris edema, hyperemia of conjunctiva, lens opacity, fibrin clots, and the protein content in aqueous humor. Clinical findings were confirmed by histological data. Thus, SOD1-containing nanozyme is potentially useful therapeutic agent for the treatment of ocular inflammatory disorders.

The in vitro cross-effects of inhibitors of renin-angiotensin and fibrinolytic systems on the key enzymes of these systems

The effects of hypotensive agents (captopril, enalaprilat, and lisinopril) on the activities of components of the fibrinolytic system (FS) and the effects of antifibrinolytic agents (6-aminohexanoic acid (6-AHA) and tranexamic acid (t-AMCHA)) on the activities of angiotensin converting enzyme (ACE) were studied in vitro. Enalaprilat did not affect the FS activity. Captopril considerably inhibited the amidase activities of urokinase (u-PA), tissue plasminogen activator (t-PA), and plasmin ([I]50 (2.0−2.6) ± 0.1 mM), and the activation of Glu-plasminogen by t-PA and u-PA ([I]50 (1.50−1.80) ± 0.06 mM), which may be due to the presence of a mercapto group in the inhibitor molecule. Lisinopril did not affect the amidase activities of FS enzymes, but stimulated Glu-plasminogen activation by u-PA and inhibited activation fibrin-bound Glu-plasminogen by t-PA ([I]50 (12.0 ± 0.5) mM). Presumably, these effects can be explained by the presence in lisinopril of a Lys side residue, whose binding to lysine-binding Glu-plasminogen centers resulted, on the one hand, in the transformation from its closed conformation to a semi-open one and, on the other hand, in its desorption from fibrin. Unspecific inhibition of the activity of ACE, a key enzyme of the renin-angiotensin system, in the presence of 6-AHA and t-AMCHA ([I]50 10.0 ± 0.5 and 7.5 ± 0.4 mM, respectively) was found. A decrease in the ACE activity along with the growth of the fibrin monomer concentration was revealed. The data demonstrate that, along with endogenous mediated interaction between FS and RAS, relations based on the direct interactions of exogenous inhibitors of one system affecting the activities of components of another system can take place.

The Inclusion of Timolol and Lisinopril in Calcium Phosphate Particles Covered by Chitosan: Application in Ophthalmology

Chitosan-covered calcium phosphate nanoparticles with the mean dynamic radii of 30 to 130 nm and ζ-potential of +22 ± 4 mV containing timolol and lisinopril are prepared and characterized. These particles are formed by the amorphous phase represented by amorphous calcium phosphate Cax(PO4)y · zH2O and the crystalline phase represented by hydrated calcium hydrophoshate (brushite) CaHPO4 · 2H2O. The experiments in vivo demonstrated that the inclusion of timolol into calcium phosphate particles covered with chitosan substantially prolonged its effect on the intraocular pressure.

Calcium phosphate particles containing superoxide dismutase are a promising agent for the treatment of eye diseases accompanied by oxidative stress

A method for the preparation of calcium phosphate particles is optimized. Particles with entrapped superoxide dismutase are obtained. The size, surface charge, and stability of the calcium phosphate particles are determined under different conditions. The kinetics of the enzyme release from the particles and the influence of the drug release on the size and surface charge of the particles are investigated. On the rabbit model of immunogenic uveitis (the model of an inflammatory process in the eye accompanied by oxidative stress), it is shown that superoxide dismutase enclosed in calcium phosphate particles has a higher antiinflammatory effect than superoxide dismutase in an aqueous solution.