[Analysis of the work of the Immunohistochemical Quality Control Center of the Russian Medical Academy of Continuous Professional Education of the Ministry of Health of Russia in 2023]. / Analiz raboty Tsentra kontrolya kachestva immunogistokhimicheskikh issledovanii FGBOU DPO «RMANPO¼ Minzdrava Rossii v 2023 godu.

The article demonstrates a detailed analysis of the results of the rounds of quality control of immunohistochemical studies conducted by the Central Committee of the Immunohistochemical Quality Control Center of the Russian Medical Academy of Continuous Professional Education of the Ministry of Health of Russia in 2023. Typical shortcomings and errors in the immunohistochemical examination of various tumors have been identified and ways to eliminate them are given. Particular attention is paid to defining a panel of standard breast cancer markers and eliminating the shortcomings of immunohistochemical examination of markers of accompanying diagnosis.

A role of methionines in the functioning of oxidatively modified fibrinogen.

Posttranslational modifications in fibrinogen resulting from induced oxidation or oxidative stress in the organism can have deleterious influence on optimal functioning of fibrinogen, causing a disturbance in assembly and properties of fibrin. The protective mechanism supporting the ability of fibrinogen to function in ROS-generating environment remains completely unexplored. The effects of very low and moderately low HOCl/-OCl concentrations on fibrinogen oxidative modifications, the fibrin network structure as well as the kinetics of both fibrinogen-to-fibrin conversion and fibrin hydrolysis have been explored in the current study. As opposed to 25 Μm, HOCl/-OCl, 10 µM HOCl/-OCl did not affect the functional activity of fibrinogen. It is shown for the first time that a number of Met residues, AαMet476, AαMet517, AαMet584, BßMet367, γMet264, and γMet94, identified in 10 µM HOCl/-OCl fibrinogen by the HPLC-MS/MS method, operate as ROS scavengers, performing an important antioxidant function. In turn, this indicates that the fibrinogen structure is adapted to the detrimental action of ROS. The results obtained in our study provide evidence for a protective mechanism responsible for maintaining the structure and functioning of fibrinogen molecules in the bloodstream under conditions of mild and moderate oxidative stress.

[External quality control of immunohistochemical studies in pathomorphology: tasks, problems, solutions, development prospects]. / Vneshnii kontrol' kachestva immunogistokhimicheskikh issledovanii v patomorfologii: zadachi, problemy, puti resheniya, perspektivy razvitiya.

In 2022, the Quality Control Center for Immunohistochemical Studies of the Russian Medical Academy of Continuing Professional Education conducted 12 rounds of markers for breast, lung, prostate, bladder cancer with the participation of 83 laboratories. For the first time, a round was held to control the method of in situ hybridization in the diagnosis of breast cancer, and a digital round. Typical problems in carrying out immunohistochemical studies in oncomorphology have been identified and the importance of participation of laboratories in external control has been shown.

[Results of the work of the Russian Medical Academy of Continuous Professional Education Center for quality control of immunohistochemical studies for 2019-2021]. / Itogi raboty Tsentra kontrolya kachestva immunogistokhimicheskikh issledovanii FGBOU DPO «RMANPO¼ za 2019­2021 gg.

In 2019-2021 the Russian Medical Academy of Continuous Professional Education Center for quality control of immunohistochemical studies conducted rounds on the most used tumour markers of various localizations. The deficiencies in the conduct of immunohistochemical studies were identified and the importance of the participation of medical organizations in measures to improve the quality control of immunohistochemical studies in oncomorphology was shown.

Radioprotective Effect of Nanocerium by Irradiation of Mice with Carbon Ions in Medium and Lethal Doses.

The data of the study of the radioprotective properties of nanocerium (nCeO2) after total irradiation of mice with carbon ions in medium and lethal doses according to the micronucleus test and the criterion of 30-day survival are presented. A significant protective effect of nCeO2 upon irradiation at medium doses was observed at per os administration for 5 days before irradiation (that is, at long-term prophylactic use). Mouse survival data showed no protective effect of per os administration of nCeO2 in contrast to the micronucleus test results. After injections of both nCeO2 and saline solution 24 h before or immediately after irradiation, the radioprotective effect was detected using both methods. The data obtained revealed the dependence of the observed effects on the mode and time of nCeO2 administration, the influence of the solvent, the level of doses and the quality of radiation, as well as demonstrated the possibility of using nanocerium preparations to protect organisms from radiation with high LET values and the importance of further studies of the radioprotective properties of new nanomaterials.

Peroxide-Induced Damage to Plasminogen Molecules.

Plasminogen is a zymogenic form of plasmin, an enzyme that plays a fundamental role in the dissolution of fibrin clots as well as in many other physiological processes. For the first time, by the method of gas chromatography-mass spectrometry, post-translational modifications in the primary structure of plasminogen treated with physiologically relevant amounts of hydrogen peroxide were identified. It was found that methionine and tryptophan residues located in different structural regions of plasminogen served as targets of the oxidant. Plasminogen oxidation caused a dose-dependent effect in decreasing the fibrinogenolytic activity of plasmin evidenced by the formation of fibrinogen degradation products. The possible antioxidant role of methionines in the oxidative modification of plasminogen is discussed.

Effect of Hypochlorite- and Peroxide-Induced Oxidation of Fibrinogen on the Fibrin Structure.

Using the methods of dynamic and elastic light scattering and confocal laser scanning microscopy, the damage in the spatial fibrin structure during peroxide- and hypochlorite-induced oxidation of fibrinogen was studied. Peroxide had a weak effect on the structural organization of fibrin, whereas hypochlorite caused the formation of abnormal fibrin with reduced individual fiber diameter and decreased porosity. Measurements of the size distributions of the native and oxidized fibrinogen revealed a decrease in the hydrodynamic size of the oxidized fibrinogen molecule with an increase in the concentration of oxidizers. These results indicate that the hydrophobicity of fibrinogen surface increased and its colloidal stability decreased. The possible role of oxidative sites in the assembly of structurally abnormal fibrin is analyzed.

The Nature of Resistance of the Coagulation Factor XIII Structure to Hypochlorite-Induced Oxidation.

The damage to blood coagulation factor XIII (FXIII) at different stages of its enzymatic activation under the action of various physiological amounts of hypochlorite ion was studied. The results obtained by HPLC-MS/MS, SDS-PAGE, and colorimetry showed that, during the conversion of FXIII to FXIIIa, the vulnerability of FXIII to hypochlorite-induced oxidation increased. FXIII oxidized with 150 µM hypochlorite completely retained its enzymatic activity inherent to the intact protein, whereas FXIIIa treated with 50 µM hypochlorite showed sharply reduced enzymatic activity. It was shown that a number of methionine and cysteine residues on the catalytic subunit can perform antioxidant function; additionally, the regulatory subunits of FXIII-B contribute to the antioxidant protection of the catalytic center of the FXIII-A subunit, which, together with the tight packing of the tetrameric structure of the FXIII proenzyme, are the three factors that provide high protein resistance to the oxidizing agent.

The Structural-Functional Damage of Fibrinogen Oxidized by Hydrogen Peroxide.

The effect of peroxide-induced oxidation of fibrinogen on modification of its primary structure and functional properties was investigated. The oxidation sites were shown to be Met, Trp, and His residues. Using the DLS method, it was found that the oxidative modification of fibrinogen results in the change of microrheological characteristics of fibrin network. The fibrinogen oxidation diminishes its tolerance to plasmin hydrolysis and deteriorates the factor XIIIa ability to stabilize the fibrin gel.

Hypochlorite-Induced Damage of Plasminogen Molecules: Structural-Functional Disturbance.

Plasminogen, the precursor of plasmin, is a serine protease that plays a fundamental role in the intravascular thrombolysis. For the first time, by using high-resolution mass spectrometry, data on the oxidative modifications of the plasminogen molecule under induced oxidation were obtained. The FTIR data show that, under oxidation on the protein, its secondary structure also undergoes the rearrangements. The high tolerance of plasminogen to oxidation can be due to both the closed conformation and the ability of some Met residues to serve as ROS trap.

Peroxide-Induced Oxidative Modification of Hemoglobin.

The oxidative modification of human hemoglobin (Hb) treated with hydrogen peroxide was investigated. Using the mass spectrometry method, the oxidized amino acid residues of the hemoglobin molecule were detected: αTrp14, αTyr24, αArg31, αMet32, αTyr42, αHis45, αHis72, αMet76, αPro77, αLys90, αCys104, αTyr140, ßHis2, ßTrp15, ßTrp37, ßMet55, ßCys93, ßCys112, ßTyr130, ßLys144, and ßHis146. The antioxidant potential of the Hb molecule in the intracellular space and in the blood plasma is discussed.

Hypochlorite-Induced Oxidative Modification of Fibrinogen.

Oxidation of fibrinogen with hypochlorite inhibited the fibrin network self-assembly even at the lowest concentration of the oxidant. The analysis of the results of protein electrophoresis at this hypochlorite concentration showed the absence of fragmentation of the protein and covalent cross-linking of its chains. The study of the areas responsible for the conversion of fibrinogen into fibrin by mass spectrometry showed that they are not subject to oxidative damage. However, we identified oxidized amino acid residues, which could affect the protofibril aggregation.

[Changes in biomechanical properties of the ocular fibrous tunic in patients with primary open-angle glaucoma in response to drug-induced stress-relief tests]. / Biomekhanicheskie parametry fibroznoi obolochki glaza na fone provedeniia razgruzochnykh prob pri pervichnoi otkrytougol'noi glaukome.

AIM: To determine the effect of drug-induced stress-relief tests on biomechanical properties of the ocular fibrous tunic in eyes with early, moderate, or advanced primary open-angle glaucoma (POAG). MATERIAL AND METHODS: A total of 202 eyes of 150 patients with POAG of different severity (early, moderate, or advanced) and 36 eyes of 30 healthy controls were examined. The mean patient age was 62±8.2 years. All groups were standardized by age, sex, and the range of corneal-compensated intraocular pressure (IOP) at baseline (21-30 mmHg). All patients underwent a standard ophthalmic examination, including visometry, biomicroscopy, gonioscopy, ophthalmoscopy, and Humphrey visual field assessment. Corneal hysteresis (CH), corneal resistance factor (CRF), CH-CRF difference, corneal-compensated IOP (IOPcc), and Goldmann-related IOP (IOPg) were measured with Ocular Response Analyzer (ORA, Reichert, USA). The axial eye length was measured on an ultrasonic A-scan (Ocuscan RxP, Alcon, USA) in the 10 MHz mode. RESULTS: CH and CRF variability analysis was conducted to assess changes in biomechanical properties of the fibrous tunic due to an IOP decrease and a tendency toward compensation. For the first time, the ratio between CH and CRF changes (ΔCH/ΔCRF) was used to evaluate biomechanical properties of the fibrous tunic. This ratio provides understanding of the significance of CH and CRF changes after reduction of IOP. In early glaucoma patients, it differed inconsiderably from the control group (p>0.05) and averaged 1.62±0.9. In moderate glaucoma, CH changes were more pronounced than those of CRF: ΔCH/ΔCRF - 2.03±2.41 (under conservative treatment) and 2.12±1.07 (without treatment). In advanced glaucoma an opposite pattern was observed: the CH/CRF ratio got closer to 1.0 largely due to CRF changes, while CH changes became much less pronounced (or even negative, in some cases): ΔCH/ΔCRF - 0.27±0.32 (under conservative treatment), 0.16±1.29 (without treatment). CONCLUSION: While the IOP decreased as a result of drug-induced stress-relief tests, CH and CRF changes differed between the groups depending on the stage of POAG. In early and moderate glaucoma, CH undergoes greater changes than CRF. In advanced glaucoma, CRF changes are more pronounced, and, hence, the ΔCH/ΔCRF ratio shifts reliably as glaucoma progresses. The ratio has also been shown to depend on the duration of the disease and on whether or not eye drops were prescribed.

Identification of OAT1/OAT3 as Contributors to Cisplatin Toxicity.

Cisplatin is among the most widely used anticancer drugs and known to cause a dose-limiting nephrotoxicity, which is partially dependent on the renal uptake carrier OCT2. We here report a previously unrecognized, OCT2-independent pathway of cisplatin-induced renal injury that is mediated by the organic anion transporters OAT1 and OAT3. Using transporter-deficient mouse models, we found that this mechanism regulates renal uptake of a mercapturic acid metabolite of cisplatin that acts as a precursor of a potent nephrotoxin. The function of these two transport systems can be simultaneously inhibited by the tyrosine kinase inhibitor nilotinib through noncompetitive mechanisms, without compromising the anticancer properties of cisplatin. Collectively, our findings reveal a novel pathway that explains the fundamental basis of cisplatin-induced nephrotoxicity, with potential implications for its therapeutic management.

Oxidation-induced modification of the fibrinogen polypeptide chains.

By using the mass-spectrometry method, the oxidative modifications of the fibrinogen Aα, Bß, and γ polypeptide chains induced by its oxidation have been studied. The αC-region has been proven to be the most vulnerable target for the oxidizer (ozone) as compared with the other structural elements of the Aα chain. The Bß chain mapping shows that the oxidative sites are localized within all the structural elements of the chain in which the ß-nodule exhibits high susceptibility to oxidation. The γ chains are the least vulnerable to the oxidizer action. The results obtained demonstrate convincingly that the self-assembly centers dealing with interactions of knob "A": hole "a" are not involved in oxidative modification. It is concluded that the numerous oxidative sites revealed are mainly responsible for inhibiting lateral aggregation of protofibrils. The part of amino acid residues subjected to oxidation is supposed to carry out the antioxidant function.

Modification of human serum albumin under induced oxidation.

For the first time, by using the complex of physicochemical methods (mass-spectrometry, differential scanning calorimetry, spectrofluorimetry, method of spectral and fluorescent probes, dynamic light scattering, and UV spectrophotometry), the oxidation-mediated modification of chemical and spatial structure of albumin has been studied. All albumin structural regions are subjected to oxidation, methionine and aromatic amino acids primarily involved in oxidation. The albumin melting shows a decrease in thermal stabilization of the structure and changing of aggregation upon oxidation. The change in physical and chemical properties of albumin under different quantity of the oxidizer has been analyzed.

[New horizons in the use of biological agents during pregnancy in patients with rheumatic disease]. / Novye gorizonty primeneniia genno-inzhenernykh biologicheskikh preparatov pri beremennosti u bol'nykh revmaticheskimi zabolevaniiami.

Pregnancy in the presence of rheumatic diseases (RD) and adequate therapy before planned conception, during gestation, and after delivery during lactation is challenging. Advances in the treatment of RD are largely due to the clinical introduction of a new class of biological agents (BAs). There are less than two decades of experience in using BAs in rheumatology and to date there are no unified standards and accepted rules governing their use during pregnancy. According to the current requirements, information on a medicine should be given in three sections: 1) pregnancy; 2) lactation, and 3) use in men and women who are planning concept (the latter section has appeared for the first time). The present article summarizes data on the possible use of BAs in patients with RD during pregnancy planning, pregnancy, and breastfeeding.

Influence of OATP1B1 Function on the Disposition of Sorafenib-ß-D-Glucuronide.

The oral multikinase inhibitor sorafenib undergoes extensive UGT1A9-mediated formation of sorafenib-ß-D-glucuronide (SG). Using transporter-deficient mouse models, it was previously established that SG can be extruded into bile by ABCC2 or follow a liver-to-blood shuttling loop via ABCC3-mediated efflux into the systemic circulation, and subsequent uptake in neighboring hepatocytes by OATP1B-type transporters. Here we evaluated the possibility that this unusual process, called hepatocyte hopping, is also operational in humans and can be modulated through pharmacological inhibition. We found that SG transport by OATP1B1 or murine Oatp1b2 was effectively inhibited by rifampin, and that this agent can significantly increase plasma levels of SG in wildtype mice, but not in Oatp1b2-deficient animals. In human subjects receiving sorafenib, rifampin acutely increased the systemic exposure to SG. Our study emphasizes the need to consider hepatic handling of xenobiotic glucuronides in the design of drug-drug interaction studies of agents that undergo extensive phase II conjugation.

Modification of the catalytic subunit of plasma fibrin-stabilizing factor under induced oxidation.

For the first time, by using mass-spectrometry method, the oxidation-mediated modification of the catalytic FXIII-A subunit of plasma fibrin-stabilizing factor, pFXIII, has been studied. The oxidative sites were identified to belong to all structural elements of the catalytic subunit: the ß-sandwich (Tyr104, Tyr117, and Cys153), the catalytic core domain (Met160, Trp165, Met266, Cys328, Asp352, Pro387, Arg409, Cys410, Tyr442, Met475, Met476, Tyr482, and Met500), the ß-barrel 1 (Met596), and the ß-barrel 2 (Met647, Pro676, Trp692, Cys696, and Met710), which correspond to 3.9%, 1.11%, 0.7%, and 3.2%, respectively, of oxidative modifications as compared to the detectable amounts of amino acid residues in each of the structural domains. Lack of information on some parts of the molecule may be associated with the spatial unavailability of residues, complicating analysis of the molecule. The absence of oxidative sites localized within crucial areas of the structural domains may be brought about by both the spatial inaccessibility of the oxidant to amino acid residues in the zymogen and the screening effect of the regulatory FXIII-B subunit.

The oxidative modification of cellular fibrin-stabilizing factor.

For the first time, the induced oxidative modification of cellular fibrin-stabilizing factor (cFXIII) has been studied. According to the electrophoresis analysis, the conversion of oxidized cFXIII into FXIIIa resulted in producing the enzyme that significantly lost the initial enzymatic activity. At the same time, FXIIIa subjected to induced oxidation was completely devoid of enzymatic activity. The results of FTIR spectroscopy showed that the oxidation of cFXIII or FXIIIa was accompanied by profound changes both in chemical and spatial structures of the protein. The results of this study are in good agreement with our earlier assumption regarding the antioxidant role of the regulatory subunits B of plasma fibrin-stabilizing factor.