Vitaliy A. Brunchukov

Введение. Адъювантная лучевая терапия (АДЛТ) является стандартным лечением рака молочной железы (РМЖ), однако ранние лучевые повреждения кожи (РЛПК) остаются частым осложнением, ухудшающим переносимость лечения и качество жизни. Поиск прогностических маркеров индивидуальной радиочувствительности нормальных тканей является актуальной задачей современной радиологии. Особый интерес вызывает иммуновоспалительный статус пациенток до АДЛТ, а также влияние предшествующей полихимиотерапии (ПХТ), способной модифицировать системное воспаление и Т-клеточный баланс. Цель. Оценить прогностическую значимость интегральных индексов воспаления (индекс системного воспаления (SIRI), индекс системного воспалительного ответа (SII) и совокупный системный индекс воспаления (AISI)) и параметров Т-клеточного иммунитета, включая иммунорегуляторный индекс (ИРИ) (CD4+/CD8+), в развитии РЛПК у пациенток с РМЖ при проведении АДЛТ, в том числе получавших ПХТ. Материалы и методы. 168 пациенток с РМЖ I–III стадии включены в исследование, получивших АДЛТ в режиме гипофракционирования. До начала ЛТ оценивали показатели периферической крови, рассчитывали SIRI, SII, AISI, определяли субпопуляционный состав Т-лимфоцитов (СD3+; CD4+; CD8+) методом проточной цитофлуориметрии. РЛПК оценивали клинически. Группы стратифицировали с учетом предшествующей ПХТ. Результаты. РЛПК зафиксированы у всех пациенток, II–III степень отмечена у 32,7 %. В общей группе базовые гематологические и иммунологические показатели находились в пределах референсных значений. Частота РЛПК не различалась между пациентками с ПХТ и без нее (33,3 против 35,4 % соответственно, р = 0,48). В группе ПХТ определялись более выраженный нейтрофильно-моноцитарный профиль и снижение показателей лимфоцитарного звена. У пациенток с ПХТ и РЛПК II–III степени отмечались более высокие значения SIRI, чем при I степени (р = 0,048; 1,069 [0,9199; 1,218] против 1,395 [1,024; 1,767]). Пороговые значения SIRI ≥ 1,25 и SII ≥ 540 ассоциировались с более чем трехкратным увеличением риска тяжелых РЛПК. Также выявлено снижение иммунорегуляторного индекса за счет относительного преобладания CD8+ клеток после ПХТ. Выводы. Исходный иммуновоспалительный профиль и субпопуляционный состав Т-клеток влияют на индивидуальную радиочувствительность кожи у пациенток с РМЖ. Учет данных показателей может быть полезен для персонализации АДЛТ.

Маркер неблагоприятного прогноза при раке молочной железы HLA-A*02 + HLA-B*44 + TNF-308A +

Background: Tracking the migration pathways of living cells after their introduction into a patient’s body is a topical issue in the field of cell therapy. Questions related to studying the possibility of long-term intravital biodistribution of mesenchymal stromal cells in the body currently remain open. Methods: Forty-nine laboratory animals were used in the study. Modeling of local radiation injuries was carried out, and the dynamics of the distribution of mesenchymal stromal cells labeled with [⁸⁹Zr]Zr-oxine in the rat body were studied. Results: the obtained results of the labelled cell distribution allow us to assume that this procedure could be useful for visualization of local radiation injury using positron emission tomography. However, further research is needed to confirm this assumption. Conclusions: intravenous injection leads to the initial accumulation of cells in the lungs and their subsequent redistribution to the liver, spleen, and kidneys. When locally injected into tissues, mesenchymal stromal cells are not distributed systemically in significant quantities.

Background: Radiation therapy (RT) is an important step in the treatment of primary breast cancer as it is one of the leading contributors to cancer incidence among women. Most patients with this disease acquire radiation-induced lymphopenia in the early post-radiation period; however, little is known about the effect of RT on the composition of lymphocyte populations in such patients. This study was aimed at investigating the effect of adjuvant remote RT—performed in the classical mode for patients with primary breast cancer—on the main components of cell-mediated immunity (major lymphocyte populations), including those in patients receiving chemotherapy. Methods: Between 2020 and 2022, 96 patients with stage I–III breast cancer were included in this study. All patients in the final stage of complex treatment received RT via a 3D conformal technique (3DCRT). The clinical target volume of this RT included the breast or chest wall and locoregional lymphatics. Flow cytometry was used to assess the levels and phenotypes of circulating lymphocytes before and after RT (no more than 7 days before and after RT). The evaluation of the impact of polychemotherapy (PCT) was conducted to determine whether it was a risk factor for the onset of radio-induced lymphopenia (RIL) in the context of RT. Results: When assessing the immune status in the general group of patients (n = 96), before the start of adjuvant external beam radiotherapy (EBRT), the average number of lymphocytes was 1.68 ± 0.064 × 10⁹/L; after the course of adjuvant EBRT, it decreased to 1.01 ± 0.044 × 10⁹/L (p < 0.001). When assessing the absolute indicators of cellular immunity in the general group of patients with BC after a course of adjuvant EBRT, significant dynamics were revealed by the changes in all cell populations of lymphocytes (paired t-test, p < 0.05). Conclusion: The adaptive immune system in breast cancer patients changed in the early post-radiation period. The absolute levels of B-, T- and natural killer cells significantly reduced after RT regardless of whether the patients previously underwent chemotherapy courses. RT for patients with primary breast cancer should be considered in clinical management because it significantly alters lymphocyte levels and should be considered when assessing antitumor immunity, as significant changes in T-cell immunity have been observed. In addition, the identified changes are critical if specific targeted therapy or immunotherapy is needed.

Each person is inevitably exposed to low doses of ionizing radiation (LDIR) throughout their life. The research results of LDIR effects are ambiguous and an accurate assessment of the risks associated with the influence of LDIR is an important task. Mesenchymal stromal cells (MSCs) are the regenerative reserve of an adult organism; because of this, they are a promising model for studying the effects of LDIR. The qualitative and quantitative changes in their characteristics can also be considered promising criteria for assessing the risks of LDIR exposure. The MSCs from human connective gingiva tissue (hG-MSCs) were irradiated at doses of 50, 100, 250, and 1000 mGy by the X-ray unit RUST-M1 (Russia). The cells were cultured continuously for 64 days after irradiation. During the study, we evaluated the secretory profile of hG-MSCs (IL-10, IDO, IL-6, IL-8, VEGF-A) using an ELISA test, the immunophenotype (CD45, CD34, CD90, CD105, CD73, HLA-DR, CD44) using flow cytometry, and the proliferative activity using the xCelligence RTCA cell analyzer at the chosen time points. The results of study have indicated the development of stimulating effects in the early stages of cultivation after irradiation using low doses of X-ray radiation. On the contrary, the effects of the low doses were comparable with the effects of medium doses of X-ray radiation in the long-term periods of cultivation after irradiation and have indicated the inhibition of the functional activity of MSCs.

Introduction. Tumor necrosis factor (TNF) plays an important role in the pathogenesis of breast cancer (BC). The -308(G/A)TNF polymorphism allele -308A is part of the autoimmune ancestral haplotype AH8.1 associated with elevated serum sTNF levels. Previously, we showed a decrease in overall survival for carriers -308A/AH8.1–. The aim of the study was to assess the effect of haplotype AH8.1 on the level of sTNF and clinical and morphological characteristics of BC patients. Materials and methods. The study included 100 with adjuvant radiotherapy treatment. The concentration of sTNF was determined by the ELISA. Additionally, 30 archival DNA samples of primary BC patients carrying the allele -308A with a known sTNF level were genotyped. Alleles -308(G/A)TNF and marker alleles of haplotype AH8.1 (HLA-A*01, HLA-B*08 and HLA-DRB1*03) were determined by allele-specific PCR. Results and discussion. Based on the polymorphism of -308(G/A)TNF and marker alleles of haplotype AH8.1, three comparison groups were identified: carriers -308GG of the TNF gene independently of haplotype AH8.1 (76%) — were used as a control, carriers -308A/AH8.1– (9%) and -308A/M(AH8.1)+ carriers of at least one haplotype AH8.1 allele (15%). The comparison groups had no statistically significant differences regarding the stage of the disease, the degree of malignancy, histological type and receptor status of the tumor. The level of sTNF was significantly higher in the genetic group -308A/AH8.1– both before surgery and before and after adjuvant radiotherapy. Conclusion. In BC patients’ carriers of the TNF gene allele -308A, the autoimmune haplotype AH8.1 has a normalizing effect on the production of sTNF to a level comparable to the level of cytokine in the group of carriers of the common -308GG genotype. A genetic group -308A/AH8.1– with an increased level of sTNF has been identified, for which, bearing in mind its prognostic unfavorability, anti-TNF therapy can presumably be offered. The results obtained can be used in the framework of personalized medicine.

Tumor necrosis factor (TNF) plays an important role in the pathogenesis of breast cancer (BC). The -308(G/A)TNF polymorphism allele -308A is part of the au

Purpose of the study. The aim of this work was to study the effect of conservation in various cryoprotectants on the biomechanical and morphological properties of the umbilical cord (UCA) arteries.Materials and methods. The material for this work was 12 umbilical arteries, which were divided into 4 study groups: (1) native UCA – control without freezing; (2) APC frozen in 5 % dextran with the addition of dimethyl sulfoxide (DMSO) to 10 % and 7.5 % human serum albumin (HSA); (3) APC frozen in glycerin and (4) UCA frozen in propanediol. For all groups, morphological and biomechanical studies were carried out. The statistical data were processed using the GraphPad Prism 9.2.0 program. Analysis of variance was used to find significant differences.Results. Freezing with 5 % dextran, 10 % DMSO and 7.5 % human albumin did not lead to significant changes in the morphology of vascular tissue compared to control. The conserved vascular tissue retains a moderate level of type I collagen, regardless of the method of conservation. The presence of statistically significant differences between the control and frozen UCAs was shown for Young's modulus (p < 0.05; 6.9 [6.8; 7.0] for the native artery; 2.3 [2.0; 2.6] for the artery preserved with the use of dextran, DMSO and human albumin; 2.3 [1.4; 3.2] for the artery preserved in glycerin and 0.8 [0.6; 1.0] – in propanediol, respectively).Conclusion. 1) Freezing and storage of UCA at 5 % dextran, 10 % DMSO and 7.5 % HSA does not lead to a significant change in the morphology of vascular membranes and cell nuclei; 2) The preservation of type I collagen according to the results of IHC is qualitatively reduced from a conditional indicator of +2 to +1 for frozen agroindustrial complexes compared with control ones; 3) Conservation leads to a significant decrease in the modulus of elasticity of the walls of the agroindustrial complex compared with control vessels (p < 0.05).

Background: The search for an effective therapy for local radiation injuries (LRI) is urgent; one option is mesenchymal stem cells (MSC) derived from the placenta and their conditioned medium for the regenerative processes of the skin. Methods: We used 80 animals, randomly assigned to four groups: control (C) animals that did not receive therapy; control with the introduction of culture medium concentrate (CM); introduction of MSCs (PL); introduction of CMPL. LRI modeling was performed on an X-ray machine at a dose of 110 Gy. Histological and immunohistochemical tests were performed. Results: On the 112th day, the area of the open wound surface in the CMPL group was 6.7 times less than in the control group. Complete healing of the open wound surface of the skin in the CM group was observed in 40%, in CMPL 60%, in the PL group 20%, and in the C group there were no animals with a prolonged wound defect. A decrease in inflammatory processes was observed in the CMPL group. Conclusions: the use of a concentrate of conditioned MSCs (CMPL group) in severe LRI in laboratory animals accelerates the transition of the wound process to the stage of regeneration and epithelization.

Purpose: To study the regeneration processes in the treatment of radiation skin lesions with the mesenchymal stem cells (MSC) derived from human gingiva and their conditional medium concentrate (CCM) during animal studies. Material and methods: The study includes 80 white male Wistar rats weighing 210 ± 30 g at the age of 8–12 weeks, randomized into 4 groups (20 animals in each): control group (C), animal did not receive treatment; control with the introduction of the conditional medium concentrate (CCM) three times on days 1, 14 and 21; the introduction of MSC in a dose of 2 million cells per 1 kg three times on days 1, 14 and 21; the introduction of CCM in the estimated dose of 2 million cells per 1 kg three times on days 1, 14 and 21. Radiation burn simulation was performed (using on an X-ray unit at a dose of 110 Gy) and each animal was observed 17 times: at days 1, 7, 14, 21, 28, 35, 42, 49, 56, 63, 70, 77, 84, 91, 98, 105 and 112. Histological (stained with hematoxylin-eosin) and immunohistochemical (CD31, CD68, and VEGF) studies were performed. MSC was cultivated according to the standard procedure up to passages 3–5, the conditioned medium was collected and concentrated 10 times. The MSC immunophenotype (CD34, CD45, CD90, CD105, CD73, HLA-DR) and viability (7-ADD) were determined using flow cytometry. Results: Under the assessment of the animal skin on the day 7 in the CCM group, the area was significantly larger compared to the C, MSC, CM groups (р ≤ 0.05). In the CM group on the day 14 the area of the open wound surface and ulcers from day 28 to day 42 was significantly less, compared with the C, MSC and CCM groups (р ≤ 0.05). In group C, from 42 to 77 days of observation, an increase in the area of skin ulcers was observed compared with the CM and CCM groups (р ≤ 0.05). On the day 112, healing of skin ulcers in the CM group was observed in 40 %, in the MSC group in 60 %, and only in 20 % of animals in the CCM group, and in the C group it was not registered. Expression of VEGF marker on endothelial cells and stromal cells was observed in groups C and CM on day 28 and in groups MSCs and CCM on day 112. On the 28th day in the MSC group, the average number of vessels (CD31) in the field of view was 6.0, and on day 112 it was 12.75, р ≤ 0.05, in the CCM group – 19.10 and 28.6, respectively, р ≤ 0.05. An increase in the number of macrophages (CD68) was found in group C from 28 to 112 days (11.6 and 24.73, р ≤ 0.05), and in the CM group the decrease was 22.1 and 13.07, respectively, р ≤ 0.05. Conclusion: Thus, all used treatment modes of radiation skin lesions, including 3-fold administration of CM, MSC and CCM at a dose of 2 million cells per 1 kg, were effective and resulted in a reduction in the damage area, accelerated ulcer healing, and improvement of the regenerative processes. In addition, the use of MSCs led to the improvement of inflammatory processes’ vascularization and reduction in the radiation skin lesions.