Autoimmune hepatitis (AIH) is a progressive hepatocellular inflammation of unknown etiology, accompanied by the process of liver fibrosis. Damage in the liver is associated with the formation of highly reactive molecules that contain unpaired electrons (free radicals), which in turn induce lipid peroxidation, leading to the development of mitochondrial dysfunction. Mitochondrial DAMPs released from damaged hepatocyte mitochondria (with mtDNA as the main active component) directly activate hepatic stellate cells and cause liver scarring. Assessing cellular metabolism can be done by examining the activity of succinate dehydrogenase (SDH), a key enzyme involved in both the tricarboxylic acid cycle and OXPHOS. Aim. to examine indicators of cellular immunity and succinate dehydrogenase activity in lymphocyte populations at different stages of liver fibrosis in children with type I AIH.. Materials and methods. During hospitalization, 69 children with AIH type I, aged 12 to 18 years, Me=15.5 [12.8;16.9], were examined. 62 healthy children made up the comparison group. The stage of liver fibrosis was assessed using the FibroScan F502. The subpopulation composition of lymphocytes and the activity of succinate dehydrogenase in them were assessed using flow cytometry. The following lymphocyte populations were evaluated: T-lymphocytes, activated T-cells, B-lymphocytes, NK cells, CD16/56+CD3+ lymphocytes, cytotoxic T-lymphocytes, T-helpers, including small populations: activated T-helpers, regulatory T-cells, Th17 lymphocytes. Statistica 10.0 (USA) program was used for statistical data processing. Results. The study included 47 girls and 22 boys. Children with AIH were on standard therapy with glucocorticosteroids for 2.6 [1.1;5.4] years. In 53 children, AIH occurred in combination with gastroduodenitis (76.8% of observations), in 12 children – with gastroesophageal reflux disease (17% of observations). 7 children had comorbid autoimmune diseases or a combination of them: 5 patients had autoimmune thyroiditis (7% of cases), 2 children had vitiligo (3% of cases), as well as type 1 diabetes mellitus, juvenile rheumatoid arthritis and celiac disease in one case (1.5%), 3 children (5% of cases) had multiple autoimmune syndrome. In children with AIH, an increase in the relative content of T cells and T helper cells was detected. The relative number of NK cells and B cells was reduced in all stages of AF relative to the comparison group. A significant increase in the absolute number of Thact and Tregs was obtained at all stages of fibrosis relative to the comparison group, while the relative number of Thact (% CD4+) was significantly higher only at stages F0, F3 and F4, and the relative number of Tregs (% CD4+) was significantly higher only at stages F0 and F4. The proportion of Th17 lymphocytes (% CD4+) was significantly reduced relative to the comparison group at stages F0 and F2. From the F0 to F3 stage, the relative number of B cells decreased, and then significantly increased by the F4 stage. The proportion of T helper cells, Tregs and Thact in most patients was higher than the comparison group, however, no significant differences were found between different stages. The proportion of Th17 lymphocytes increased with increasing stage of liver fibrosis, the maximum values were detected at stage F4. A study of SDH revealed a significant decrease in the activity of this enzyme in all populations of lymphocytes. The greatest decrease in activity was detected in double negative CD3+CD4-CD8- cells by 18.5% and NK cells by 17.5% relative to the comparison group. In the populations of Th17 and Treg lymphocytes, SDH activity was significantly lower in the state of exacerbation of the disease, relative to in the state of remission. Analysis of SDH activity indicators depending on the stage of liver fibrosis revealed a nonlinear relationship: in the populations of T-helpers, T-cytotoxic and Thact – an increase to stage F2-3, and then a decrease to stage F4. SDH activity in the population of Th17 lymphocytes increased maximally at the F1 stage and subsequently decreased to the F4 stage. With increasing stage of liver fibrosis, SDH activity significantly increased in the Treg population, but did not reach normative values. Children with autoimmune hepatitis need clinical observation and strict monitoring of the development of the disease. Conclusion. The subpopulation composition of peripheral blood lymphocytes and the activity of succinate dehydrogenase in lymphocyte populations objectively reflect the severity of the patient’s condition at the time of examination and can be used for additional laboratory assessment of the progression of liver fibrosis. The identified signs of mitochondrial dysfunction justify the use of correction of metabolic disorders as a therapy that improves the basic treatment of patients with type 1 AIH.

Abstract Background Upadacitinib (UPA) is a novel selective JAK-1 inhibitor that has demonstrated efficacy in the treatment of ulcerative colitis (UC) and Crohn's disease (CD) and has received regulatory approval in adults only. Current data on the efficacy of this drug in children are limited. We report our experience in the treatment of pediatric patients with UC and CD with failure of immunosuppressive and biologic therapies. The aim of this study was to evaluate short-term effectiveness and safety UPA therapy in children with IBD. Methods Disease activity was monitored using clinical PUCAI/PCDAI scores, faecal calprotectin (FC), C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR), platelet. Hemoglobin (Hb), leukocyte, total cholesterol, prothrombin time, INR - international normalized ratio, fibrinogen, activated partial thromboplastin time, thrombin time were evaluated to assess side effects. At baseline biochemical and previous therapies were recorded. Results We performed a prospective analysis of clinical outcomes from January to October 2023 during UPA therapy 11 children aged 12 to 16 years (4 boys, 7 girls) 3 with UС and 8 with CD. All 11 children had received immunosuppressive and biological therapies prior to UPA prescription without success. UPA induction was administered in UC for 8 weeks and CD for 12 weeks at a dose of 45 mg once a day, then 15 mg once a day as maintenance. At the beginning 2 (18%) of 11 children had high PUCAI/PCDAI disease activity and remain 9 (82%) had mild disease activity. Elevated white blood cells was determined in 3 (27%) out of 11 children, platelet increase in 9 (82%) children, and Hb less than 110 g/L in 6 (54%) children, elevation of ESR and CRP was observed in 7 (63%) subjects. FC was elevated in 5 (62.5%) out of 8 children. Total cholesterol was normal in all patients, coagulation tests showed increased fibrinogen in 3 out of 11 children. After induction 3 (27%) of 11 children achieved remission and 8 (73%) had clinical response according to the PUCAI/PCDAI score. Elevated white blood cells was detected in 2 (18%) and decrease Hb persisted in 3 (27%) out of 11 children, elevation of ESR persisted in 5 (45%) and CRP in 6 (54%) of 11 children. FC was elevated in 6 (75%) out 8 patients. Total cholesterol and the coagulation tests remained within normal range. Side effects were not observed and UPA maintenance treatment was continued in all children. Conclusion In this real-world experience in drug-resistant pediatric patients with UC and CD we verified the efficacy and safety of UPA, but longer follow-up is needed.

NF-kB is a universal transcription factor located in the cell cytoplasm that translocates into the nucleus when it is activated. This leads to the synthesis of proinflammatory cytokines, chemokines, growth factors and activation of cell effector functions. The aim of this study was to assess the level of NF-kB translocation in lymphocyte populations during treatment with infliximab (IFX) in children with IBD. In total, 31 children aged 12–18 years were examined: 21 patients with IBD (12 UC, 9 CD) (Group 1) and 10 clinically healthy children (Group 2). Blood samples were taken from Group 1 patient before the infusion of IFX and next day after infusion. Intracellular NF-kB localisation was quantitatively analysed in lymphocyte populations by Amnis NF-kB kit by ImageStreamX MKII (Amnis). For populations of CD3 + CD4 + (Th), CD3 + CD8 + (Tc), CD3-CD19 + (B cell), CD3-CD16 / 56 + (NK), CD3 + CD4 + CD161 + (Th17), CD3 + CD4 + CD25highCD127low (Treg) was estimated the total number of cells and the percentage of cells with NF-kB translocation (the level of activation). The IDEAS image analysis software was applied. Similarity score algorithm was used for assess events of translocation. In Group 1 on IFX treatment with clinical and endoscopic relapse there is an increase in cells with NF-kB translocation in the main populations of lymphocytes compared with a Group 2, mostly in Th (37.7 ± 2.7% vs. 14.9 ± 1.0, p < 0.01) and Tc (39.4 ± 3.0% vs. 15.3 ± 1.5, p < 0.01). The total number of Th17 (% of CD4 +) in Group 1 before the IFX infusion averaged 34.6%, and Treg (% of CD4 +) – 11.6%, which exceeded the level of these populations in Group 2 (Th17 18.6%; Treg 7.8%). At the same time, NF-kB translocation level of in the Treg population did not differ from Group 2, and in the population of Th17 lymphocytes the level of NF-kB translocation was 1.7 times higher. It was observed a 1.4 times decrease in the activation level of Th17 and 2 times increase in the activation of Treg while the quantity of those lymphocyte populations were stable. The level of translocation NF-kB, measured by the ImageStream platform, is a rapidly changing index that allows to evaluate the functional activity of the lymphocyte populations at the time of the laboratory investigation. The level of NF-kB translocation in the lymphocyte populations, which is an important marker of the autoimmune activity in IBD (Treg, Th17) reflects the body's response to the administration of IFX. An increase in Treg activity and a decrease in Th17 activity under the action of a TNFα blocker explains the molecular mechanism leading to a weakening of the inflammatory process.

Our aim was to identify the value of the laboratory criteria such as residual level of infliximab (IFX) in blood, antibodies to IFX and circulating cytokine levels in the prognosis of the effectiveness of the therapy in children with IBD. Were included in the study 75 children with IBD (31 patients with UC and 44 patients with CD) aged 4–18 years who were treated with IFX. Clinical response was evaluated according PUCAI (UC) and PCDIA (CD) scores. Blood samples were taken 8 weeks after the last infusion of IFX. Residual levels of IFX (Q-IFX) in serum and IFX antibodies (ATI) were assessed by enzyme immunoassay using Shikari Q-INFLIXI, Q-ATI (Turkey) kits. The cytokine levels were measured by multiplex analysis using HumanThl7 MagneticBead Panel (MilliplexMapKit, Germany). Evaluation of the statistical significance was performed using nonparametric Mann–Whitney test and ROC-analysis. There were observed increase in the inflammatory activity according to PUCAI and PCDIA scores (p = 0.000) in children with the loss of response to IFX. In patients with the loss of the effect to IFX (Group 1) there was a significant decrease Q-IFX compared with a group of children with persistent positive effect (Group 2) in both diseases CD (p = 0.002) and UC (p = 0.019). ROC analysis showed that the cut-off level for patients with UC is 2.55 μg/ml (AUC = 0.813; sensitivity (Se) 64%, specificity (Sp) 92%), and for children with CD 2.21 μg/ml (AUC=0.813; Se 79%, Sp 78%). In the examined patients, IFX antibodies were detected in 17% cases, and the fast formation of IFX antibodies were associated to the younger age of children (R = 0.58). In one patient with a persistent positive effect for 5 years of therapy, the values of Q-IFX were in the range from 4.9 to 9.4 μg/ml in the absence of IFX antibodies. Cytokine analysis revealed significant differences between examined groups in the level of proinflammatory cytokines: IL-23, IL-27, IL-22, INF-γ, TNFα. ROC analysis revealed good quality TNFα as the separation model, the cut-off level was 13.4 pg/ml (AUC = 0.843; Se = 77%, Sp = 79%). The reduction of the Q-IFX in children with UC below 2.55 μg/ml and in children with CD below 2.21 μg/ml, leads to the decrease of the therapy effect and can adduct to the exacerbation of the disease. These findings correlate with the results obtained in adults (>2 µg/ml, C. Moore et al., 2016). TNFα level (>13.4 pg/ml) can serve as the laboratory criterion of loss of effect from IFX. Elevated levels of proinflammatory cytokines correlates with the lower Q-IFX and loss of the therapy effect.

Epidemiology S319 immune modulatory medicines.We observed no association with AZA or other medication.Conclusions: In conclusion; The prevalence of anemia and iron deficiency in an out-patient IBD cohort was found to be relatively high.Anemia and iron deficiency were seemed to be associated with ileal involvment in CD, there was a trend towards significance in extensive colitis in UC.