Peripheral NF-κB dysregulation in people with schizophrenia drives inflammation: putative anti-inflammatory functions of NF-κB kinases

Elevations in plasma levels of pro-inflammatory cytokines and C-reactive protein (CRP) in patient blood have been associated with impairments in cognitive abilities and more severe psychiatric symptoms in people with schizophrenia. The transcription factor nuclear factor kappa B (NF-κB) regulates the gene expression of pro-inflammatory factors whose protein products trigger CRP release. NF-κB activation pathway mRNAs are increased in the brain in schizophrenia and are strongly related to neuroinflammation. Thus, it is likely that this central immune regulator is also dysregulated in the blood and associated with cytokine and CRP levels. We measured levels of six pro-inflammatory cytokine mRNAs and 18 mRNAs encoding NF-κB pathway members in peripheral blood leukocytes from 87 people with schizophrenia and 83 healthy control subjects. We then assessed the relationships between the alterations in NF-κB pathway genes, pro-inflammatory cytokine and CRP levels, psychiatric symptoms and cognition in people with schizophrenia. IL-1β and IFN-γ mRNAs were increased in patients compared to controls (both p < 0.001), while IL-6, IL-8, IL-18, and TNF-α mRNAs did not differ. Recursive two-step cluster analysis revealed that high levels of IL-1β mRNA and high levels of plasma CRP defined ‘high inflammation’ individuals in our cohort, and a higher proportion of people with schizophrenia were identified as displaying ‘high inflammation’ compared to controls using this method (p = 0.03). Overall, leukocyte expression of the NF-κB-activating receptors, TLR4 and TNFR2, and the NF-κB subunit, RelB, was increased in people with schizophrenia compared to healthy control subjects (all p < 0.01), while NF-κB-inducing kinase mRNAs IKKβ and NIK were downregulated in patients (all p < 0.05). We found that elevations in TLR4 and RelB appear more related to inflammatory status than to a diagnosis of schizophrenia, but changes in TNFR2 occur in both the high and low inflammation patients (but were exaggerated in high inflammation patients). Further, decreased leukocyte expression of IKKβ and NIK mRNAs was unique to high inflammation patients, which may represent schizophrenia-specific dysregulation of NF-κB that gives rise to peripheral inflammation in a subset of patients.

Phenotypic heterogeneity in individuals with MECOM variants in 2 families

MECOM encodes the transcriptional regulators, EVI1 and MDS1-EVI1, from two distinct transcription start sites. EVI1 plays important roles in hematopoiesis and stem cell self-renewal. Recently, our group and others revealed that individuals with MECOM variants present diverse hematological and skeletal defects, including radioulnar synostosis (RUS). In the present study, we analyzed two families suspected with MECOM-associated syndrome. In family 1, a MECOM splicing variant (c.2285+1G&gt;A) was identified in an individual with bone marrow failure (TRS4) without RUS and her mother, who had mild leukocytopenia, thrombocytopenia, and bilateral RUS. A copy neutral loss of heterozygosity decreasing the variant allele frequency was observed in the bone marrow of TRS4 and the peripheral blood leukocytes of her mother. However, TRS4 remained transfusion-dependent. In family 2, a MECOM variant (c.2208-4A&gt;G), which was predicted to cause a cryptic acceptor site that results in a 3-base insertion (an insertion of Ser) in the mRNA, was identified in the proband, with bone marrow failure; this variant was also observed in her brother and father, both of whom have skeletal malformations, but no cytopenia. RT-PCR using leukocytes revealed a transcript with a 3-bp insertion in the proband, her brother, and the father, suggesting that the transcript variant with a 3-bp insertion is independent of blood phenotype. Collectively, these results suggest the presence of intrafamilial clinical heterogeneity in both families with MECOM splicing variants. Somatic genetic event may complicate the understanding of clinical variability among family members.

Immune Aging and How It Works for Inflammation and Fibrosis

Almost all mature cells that undergo apoptosis in an age-dependent or an accidental manner are completely recovered in tissue-specific microenvironments without any physiological changes. After peripheral blood leukocytes are released into the local region, fibroblast cells and new blood vessels commonly proliferate during wound healing. Inducible repair tools mainly supplied from blood vessels are cleared by peripheral blood phagocytic macrophages. Finally, hematopoietic stem cell (HSC)-derived precursor cells migrate from bone marrow (BM) to the microenvironment to rebuild damaged tissues (the mature immune system). In contrast to the mature immune system, the effects of aging on HSCs (long-term HSCs) and peripheral blood lymphocytes (long-term PBLs) are not clearly understood in the BM and thymus niches with tissue-specific microenvironments with some physiological changes (the aged BM niche) for incomplete rebuilding of damaged tissues (the aged immune system). In this review, the roles of the aged immune system in both a delay of acute inflammation and the development of chronic inflammation or fibrosis are discussed.

Tumor-Associated Regulatory T Cell Expression of LAIR2 Is Prognostic in Lung Adenocarcinoma

Cancer development requires a permissive microenvironment that is shaped by interactions between tumor cells, stroma, and the surrounding matrix. As collagen receptors, the leukocyte-associated immunoglobulin-like receptor (LAIR) family allows the immune system to interact with the extracellular matrix. However, little is known about their role in regulating tumor immunity and cancer progression. Methods: Genetic analysis of resected human lung adenocarcinoma was correlated to clinical-pathological characteristics, gene ontologies, and single cell RNA sequencing (scRNASeq). LAIR2 production was determined in subsets of immune cells isolated from blood leukocytes and lung adenocarcinoma tumor. Functional assays were used to determine the role of LAIR2 in tumorigenesis. Results: LAIR2 expression was adversely prognostic in lung adenocarcinoma. LAIR2 was preferentially produced by activated CD4+ T cells and enhanced in vitro tumor invasion into collagen. scRNASeq analysis of tumor infiltrating T cells revealed that LAIR2 expression co-localized with FOXP3 expressing cells and shared a transcriptional signature with tumor-associated regulatory T (Treg) cells. A CD4+LAIR2+ Treg gene signature was prognostically significant in the TCGA dataset (n = 439; hazard ratio (HR) = 1.37; 95% confidence interval (CI), 1.05–1.77, p = 0.018) and validated in NCI Director’s Challenge lung adenocarcinoma dataset (n = 488; HR = 1.54; 95% CI, 1.14–2.09, p = 0.0045). Conclusions: Our data support a role for LAIR2 in lung adenocarcinoma tumorigenesis and identify a CD4+ LAIR2+ Treg gene signature in lung adenocarcinoma prognosis. LAIR2 provides a novel target for development of immunotherapies.

Severity of the acute phase response in experimental ulcerative colitis under conditions of the use of vitamin D3 in a novel rectal suppository

Цель - изучение влияния витамина D3 в составе оригинальных ректальных суппозиториев на выраженность острофазового ответа при экспериментальном язвенном колите Методика. Эксперимент выполнен на 49 половозрелых крысах-самцах Wistar. Язвенный колит (ЯК) моделировали двухэтапным (накожным, а затем ректальным) применением 3% спиртового раствора оксазолона. Оригинальные суппозитории на основе смеси полиэтиленгликолей массой 300 мг, содержащие 1500 ME витамина D3, вводили per rectum каждые 12 ч в течение 6 сут. Исследования проводили на 2-е, 4-е и 6-е сут заболевания. Клиническую картину оценивали по адаптированной для крыс шкале Disease activity index (DAI) с учетом массы тела, консистенции и наличия крови в каловых массах. Количество лейкоцитов и нейтрофилов определяли на гематологическом анализаторе и в мазках крови. Функциональную активность нейтрофилов исследовали по поглощению частиц монодисперсного латекса в спонтанном и индуцированном НСТ-тесте. Концентрацию в сыворотке крови C-реактивного белка (С-РБ), IL-6 и IL-8 определяли с помощью иммуноферментного анализа, экспрессию TNF-α в стенке толстой кишки - иммуногистохимическим методом. Результаты. При экспериментальном ЯК на 2-е, 4-е и 6-е сут наряду с прогрессивным повышением DAI зафиксированы изменения показателей острофазового ответа (ООФ): увеличивается экспрессия в толстой кишке TNF-α с максимальным уровнем на 4-е и 6-е сут, повышается концентрация в сыворотке крови IL-6 с максимальным уровнем на 4-е, IL-8 - на 6-е сут; в сыворотке возрастает концентрация С-РБ максимально на 6-е сут, в крови увеличивается общее количество лейкоцитов, число палочкоядерных и сегментоядерных нейтрофилов с максимальным уровнем на 2-е и 4-е сут. Возрастает поглотительная и НСТ-редуцирующая способность нейтрофилов крови с максимальными значениями на 4-е и 6-е сут эксперимента. Применение при экспериментальном ЯК ректальных суппозиториев с витамином D3 приводит на 4-е и 6-е сут наблюдения к снижению DAI, экспрессии TNF-α в толстой кишке и количества нейтрофилов, на 2-е, 4-е и 6-е сут - к уменьшению концентрации IL-6 и С-РБ в сыворотке, поглотительной и НСТ-редуцирующей способности нейтрофилов крови, на 6-е сут - к снижению концентрации IL-8 в сыворотке и общего количества лейкоцитов в крови. Заключение. Применение при экспериментальном ЯК оригинальных ректальных суппозиториев с витамином D3 приводит к снижению индекса активности болезни. Выраженность клинических проявлений ослабевает по мере снижения экспрессии в кишке TNF-α, снижения концентрации в сыворотке IL-6, количества в крови лейкоцитов и нейтрофилов, уменьшения НСТ-редуцирующей способности нейтрофилов. Aim. To study the effect of vitamin D3 in rectal suppositories on severity of the acute phase response (APR) in experimental ulcerative colitis (UC). Methods. Experiments were performed on 49 mature male Wistar rats. UC was induced by cutaneous followed by rectal applications of a 3% oxazolone alcohol solution. Novel polyethylene glycol suppositories (300 mg) containing 1500 IU of vitamin D3 were injected per rectum every 12 hrs for 6 days. Data were collected on days 2, 4, and 6 of UC. The clinical picture was assessed according to the Disease Activity Index (DAI) scale adapted for rats, taking into account body weight, consistency of, and the presence of blood in the feces. The number of blood leukocytes and neutrophils was determined on a hematological analyzer and in blood smears. The functional activity of neutrophils isolated from the blood was evaluated by the absorption of monodisperse latex particles and with the spontaneous and induced NBT test. Serum concentrations of C-reactive protein (CRP), IL-6 and IL-8 were measured by enzyme immunoassay. TNF-α expression in the colon wall was measured by an immunohistochemical method. Results. The following changes in APR variables along with a progressive increase in DAI were recorded on days 2, 4, and 6. Expression of colon TNF-α increased, with maximal values on days 4 and 6. Serum IL-6 increased and reached a maximal value on day 4; and serum IL-8 increased, with a maximum on day 6. Serum CRP increased, with a maximum on day 6. The total number of blood leukocytes and the number of stab and segmented neutrophils increased, with maximal values on days 2 and 4. The absorption and NBT-reducing ability of blood neutrophils increased, with maximal values on days 4 and 6. The use of rectal suppositories with vitamin D3 in experimental UC resulted in decreases in DAI, colon TNF-α expression, and the number of blood neutrophils on days 4 and 6; decreases in serum concentrations of IL-6 and CRP, and the absorption and NBT-reducing ability of blood neutrophils on days 2, 4, and 6; and decreases in the serum concentration of IL-8 and in the total number of blood leukocytes on day 6. Severity of clinical manifestations was alleviated with decreases in the expression of colon TNF-α, the serum concentration of IL-6, the number of blood leukocytes and neutrophils, and the NBT-reducing ability of neutrophils decreases. Conclusion. Application of the original rectal suppositories with vitamin D3 every 12 hours in experimental UC leads to a decrease in the disease activity index. The severity of clinical manifestations weakens as TNF-α expression, serum IL-6 concentration, number of leukocytes and neutrophils in blood, and decrease of neutrophil NST-reducing capacity decrease.

Peculiarities of interaction of high and low virulent strains of tick-borne Encephalitis virus of the Far Eastern subtype with human blood leukocytes

In this work, it was necessary to show the first stage of the interaction of the tick-borne encephalitis virus with blood cells, during which the selection of virus strains capable of successful assembly of virions occurs.The aim of the work: to show ex vivo the features of interaction of strains TBEV with different molecular genetic structure and virulence with human blood leukocytes.Materials and methods. The donor’s venous blood was infected ex vivo with the TBEV using the highly virulent strain Dal’negorsk and the low-virulent strain Primorye-437. Virus accumulation in experimental blood samples was observed after 15 min, 30 min, 1 h, and 24 h of exposition. The indication of the virus, shown by different methods of laboratory diagnostics (ELISA, PCR, IFA, titer of the infectious virus), made it possible to identify the characteristic features of the initial stage of the infectious process caused by these strains TBEV.Conclusion. The highly pathogenic strain Dal‘ has demonstrated the ability to quickly penetrate into leukocytes in 15 minutes, and, therefore, at the initial stage of the infectious process, to implement the mechanisms of its “escape” from immune supervision. The low-virulent strain P-437, on the contrary, showed the ability to stay on the surface of leukocytes for a long time, penetrating into them only after 24 hours of exposure, immediately influencing the cells of the immune system, which can lead to rapid elimination of the virus from the body.

Diagnosing Alzheimer’s Disease from Circulating Blood Leukocytes Using a Fluorescent Amyloid Probe

Background: Toxic amyloid-β (Aβ) peptides aggregate into higher molecular weight assemblies and accumulate not only in the extracellular space, but also in the walls of blood vessels in the brain, increasing their permeability, and promoting immune cell migration and activation. Given the prominent role of the immune system, phagocytic blood cells may contact pathological brain materials. Objective: To develop a novel method for early Alzheimer’s disease (AD) detection, we used blood leukocytes, that could act as “sentinels” after trafficking through the brain microvasculature, to detect pathological amyloid by labelling with a conformationally-sensitive fluorescent amyloid probe and imaging with confocal spectral microscopy. Methods: Formalin-fixed peripheral blood mononuclear cells (PBMCs) from cognitively healthy control (HC) subjects, mild cognitive impairment (MCI) and AD patients were stained with the fluorescent amyloid probe K114, and imaged. Results were validated against cerebrospinal fluid (CSF) biomarkers and clinical diagnosis. Results: K114-labeled leukocytes exhibited distinctive fluorescent spectral signatures in MCI/AD subjects. Comparing subjects with single CSF biomarker-positive AD/MCI to negative controls, our technique yielded modest AUCs, which improved to the 0.90 range when only MCI subjects were included in order to measure performance in an early disease state. Combining CSF Aβ 42 and t-Tau metrics further improved the AUC to 0.93. Conclusion: Our method holds promise for sensitive detection of AD-related protein misfolding in circulating leukocytes, particularly in the early stages of disease.

Glucocorticoid regulation of clinical and functional manifestations of asthma in patients with different airway response to hyposmolar stimulus during anti-inflammatory therapy

Introduction. The role of an alternative adenylate cyclase pathway of hormonal signal transmission under the action of synthetic glucocorticosteroids with the participation of endogenous stress-limiting activity of the adrenal cortex in conjunction with the adaptive capabilities of airway homeostasis in patients with asthma under conditions of osmotic stress has not been studied at present. Aim. To assess the dynamics of cortisol and cyclic adenosin monophosphate (cAMP) in asthma patients with different airway responses to hypoosmolar stimuli when using anti-inflammatory combination therapy with inhaled corticosteroids/long-acting β2 -agonists (ICS/LABA). Materials and methods. 96 patients diagnosed with asthma received combined anti-inflammatory therapy with ICS/LABA for 24 weeks. Group 1 included patients (n=18) with airway hyperresponsiveness to hypoosmolar stimulus, group 2 (n=78) – with no reaction of the bronchi to a 3-minute ultrasonic inhalation of distilled water. At baseline and at the end of treatment, the lung function was studied; to assess the regulatory function of glucocorticoids using non-genomic signaling pathways, the levels of cortisol in blood serum and cAMP in blood leukocytes were determined. Results. Patients of group 1 in comparison with the second one initially had a lower FEV1 ‒ 88.2±5.3 and 98.5±1.7%, respectively (p<0.05), after treatment in both groups there was a slight tendency to an increase in FEV1 (98.5±5.7 and 101.4±2.5%, respectively, p>0.05). The concentration of cortisol and cAMP at baseline and after 24 weeks of therapy in patients of group 1 was 588.7±32.0 and 495.0±48.7 nmol/L, 61.7±5.1 and 76.5±5, 2 pmol/106 cells (p<0.01); in group 2 − 610.5±20.1 and 522.2±15.60 nmol/L (p<0.001), 76.2±2.2 and 90.6±2.5 pmol/106 cells (p<0.001). Conclusion. In asthma patients with airway osmotic hyperresponsiveness, persistent adaptation to osmotic stress is traced, which is combined with a more significant impairment of the lung function and indicates insufficient therapeutic control over glucocorticoid regulation of osmotic stress by the selected volume of ICS/LABA therapy.

Features of cytokine spectrum in chronic urticarial

Urticaria is a serious medical and social problem due to its high prevalence, lack of unified approaches to diagnosis and treatment, with high financial costs for therapy and rehabilitation. Long-term recurrent course of the disease, resistance to traditional methods of therapy lead to a significant decrease in the quality of life of patients with chronic urticaria. Itching accompanying this disease leads to deterioration in the patient’s general well-being, frequent sleep disturbances and, as a result, significant decrease in working capacity. Up to the present moment, etiopathogenesis of urticaria is a complex challenge due to the multivector nature of cytokine response, interference of protides of the complement system, patterns of kininbradykinin interference, peculiar expression of the immune response. The problem of current population is lipotrophy – chronic, heterogeneous, cytokine mediating, progressive inflammatory disease attributed by abnormal accumulation of excessive adipose tissue. Adipose tissue, being a sporadic organ of endocrine system secretes multiple hormone-like substances, mediators, cytokines and chemokines which have been given a common name, i.e., adipokines or adipocytokines. True signs of destructive parenchymal changes of liver in the form of increasing bilirubin and AST, decreasing level of vitamin D in patients with chronic recurrent urticarial in presence of obesity have been revealed during the study performed. The action of cytokines, as mediators of intercellular interaction is closely related to the physiological and pathophysiological responses of the body with modulation of both local and systemic defense mechanisms. It is assumed that the cytokine status of patients with chronic urticaria is dominated by cytokines that increase allergic inflammation of the skin. Analysis of 12 T regulatory biomarker concentrations revealed increased concentrations of IL-10, IL-19, IL-20, IL-27, IL-35, IFNλ2 and IFNλ1 in blood serum of patients with chronic urticaria. It was found that in the group of patients with chronic urticaria and increased body mass index (BMI), the level of all investigated T regulatory cytokines is lower than in the patients with normal BMI, except for IL-10. Decreased levels of biologically active IFN I (α/β) and, especially, IFN II (γ) types of blood leukocytes in patients with chronic urticaria were revealed. The levels of 12 Treg cytokines were determined in blood serum of patients with chronic urticaria, showing trend for imbalance of Treg cytokines: IL-10, IL-19, IL-20, IL-27, IL-35, IFNλ2 and IFNλ1.