Cyclopenta[ b]indole Derivative Inhibits Aurora B in Primary Cells

The Aurora family of kinases is closely involved in regulating cell division. Inhibition of Aurora A and B with small molecules is currently being investigated in clinical trials for the treatment of different cancers. It has also been evaluated as a treatment option against different autoimmune diseases in preclinical studies. Here, we present a cyclopenta[b]indole derivative capable of inhibiting Aurora B selectively in kinase assays. To evaluate the Aurora B inhibition capacity of the compound, we used a kinase IC50 assay as well as a suppression assay of proliferating primary cells. In addition, we examined if the cells had gained a phenotype characteristic for Aurora B inhibition after treatment with the compound. We found that the compound selectively inhibited Aurora B (IC50 = 1.4 μM) over Aurora A (IC50 > 30 μM). Moreover, the compound inhibited proliferating PBMCs with an IC50 = 4.2 μM, and the cells displayed reduced phosphorylation of histone H3 as well as tetraploidy, consistent with Aurora B inhibition

Cultivation of the causative agent of human neoehrlichiosis from clinical isolates identifies vascular endothelium as a target of infection

ABSTRACTCandidatus (Ca.) Neoehrlichia mikurensis is the cause of neoehrlichiosis, an emerging tick-borne infectious disease characterized by fever and vascular events. The bacterium belongs to the Anaplasmataceae, a family of obligate intracellular pathogens, but has not previously been cultivated, and it is uncertain which cell types it infects. The goals of this study were to cultivate Ca. N. mikurensis in cell lines and to identify possible target cells for human infection. Blood components derived from infected patients were inoculated into cell lines of both tick and human origin. Bacterial growth in the cell cultures was monitored by real-time PCR and imaging flow cytometry. Ca. N. mikurensis was successfully propagated from the blood of immunocompromised neoehrlichiosis patients in two Ixodes spp. tick cell lines following incubation periods of 7–20 weeks. Human primary endothelial cells derived from skin microvasculature as well as pulmonary artery were also susceptible to infection with tick cell-derived bacteria. Finally, Ca. N. mikurensis was visualized within circulating endothelial cells of two neoehrlichiosis patients. To conclude, we report the first successful isolation and propagation of Ca. N. mikurensis from clinical isolates and identify human vascular endothelial cells as a target of infection

Functional and Phenotypic Studies of Eosinophilic Granulocytes in Patients with Eosinophilic Esophagitis

Eosinophilic Esophagitis (EoE) is a chronic inflammatory disorder of unknown etiology, in which the esophagus is infiltrated by eosinophils and T cells. Topical corticosteroids are one of the treatment options for patients with EoE. The function of eosinophils in EoE is unknown, here we hypothesize that eosinophils serve as immunoregulatory cells. The eosinophils in the blood of untreated adult patients with EoE have a distinct phenotype. The aims of this thesis were to explore whether the eosinophilic phenotype of untreated patients with EoE can be reverted to the healthy phenotype by topical corticosteroid treatment, and to examine whether blood eosinophils from children with EoE have a distinct phenotype, different from that of healthy children. Moreover, we tested the hypothesis that eosinophils, similar to regulatory T cells, can diminish T cell proliferation and express FOXP3. The role of the eosinophilic protein galectin-10 in mediating immunosuppression was also investigated. This thesis demonstrates that the EoE phenotype of blood eosinophils is not restored by topical corticosteroid treatment, except with respect to CD18. We also show that eosinophils from patients with EoE have an immunoregulatory phenotype, i.e., increased levels of FOXP3 and galectin-10. Moreover, eosinophils from healthy subjects and patients with EoE are able to suppress T cell proliferation in vitro, in part via galectin-10. We show that eosinophils exposed to activated T cells release galectin-10 via DNA nets and appear to transfer this protein to T cells through synapses. Two subsets of eosinophils emerge after co-culturing. Finally, we demonstrate that the blood eosinophils of children with EoE have a distinct phenotype, different from that of healthy children and that of adults with EoE. Importantly, we reveal marked age-related differences regarding the molecular patterns displayed by the blood eosinophils of healthy donors. Our finding that eosinophils from patients with EoE have upregulated immunoregulatory molecules could indicate that the function of eosinophils in EoE is to reduce a T cell-mediated inflammation in the esophagus

Functional and Phenotypic Studies of Eosinophilic Granulocytes in Patients with Eosinophilic Esophagitis

Eosinophilic Esophagitis (EoE) is a chronic inflammatory disorder of unknown etiology, in which the esophagus is infiltrated by eosinophils and T cells. Topical corticosteroids are one of the treatment options for patients with EoE. The function of eosinophils in EoE is unknown, here we hypothesize that eosinophils serve as immunoregulatory cells. The eosinophils in the blood of untreated adult patients with EoE have a distinct phenotype. The aims of this thesis were to explore whether the eosinophilic phenotype of untreated patients with EoE can be reverted to the healthy phenotype by topical corticosteroid treatment, and to examine whether blood eosinophils from children with EoE have a distinct phenotype, different from that of healthy children. Moreover, we tested the hypothesis that eosinophils, similar to regulatory T cells, can diminish T cell proliferation and express FOXP3. The role of the eosinophilic protein galectin-10 in mediating immunosuppression was also investigated. This thesis demonstrates that the EoE phenotype of blood eosinophils is not restored by topical corticosteroid treatment, except with respect to CD18. We also show that eosinophils from patients with EoE have an immunoregulatory phenotype, i.e., increased levels of FOXP3 and galectin-10. Moreover, eosinophils from healthy subjects and patients with EoE are able to suppress T cell proliferation in vitro, in part via galectin-10. We show that eosinophils exposed to activated T cells release galectin-10 via DNA nets and appear to transfer this protein to T cells through synapses. Two subsets of eosinophils emerge after co-culturing. Finally, we demonstrate that the blood eosinophils of children with EoE have a distinct phenotype, different from that of healthy children and that of adults with EoE. Importantly, we reveal marked age-related differences regarding the molecular patterns displayed by the blood eosinophils of healthy donors. Our finding that eosinophils from patients with EoE have upregulated immunoregulatory molecules could indicate that the function of eosinophils in EoE is to reduce a T cell-mediated inflammation in the esophagus

Exploring a cascade Heck-Suzuki reaction based route to kinase inhibitors using design of experiments

Design of Experiments (DoE) has been used to optimize a diversity oriented palladium catalyzed cascade Heck-Suzuki reaction for the construction of 3-alkenyl substituted cyclopenta[b]indole compounds. The obtained DoE model revealed a reaction highly dependent on the ligand. Guided by the model, an optimal ligand was chosen that selectively delivered the desired products in high yields. The conditions were applicable with a variety of boronic acids and were used to synthesize a library of 3-alkenyl derivatized compounds. Focusing on inhibition of kinases relevant for combating melanoma, the library was used in an initial structure-activity survey. In line with the observed kinase inhibition, cellular studies revealed one of the more promising derivatives to inhibit cell proliferation via an apoptotic mechanism

Cytokine responses of immunosuppressed and immunocompetent patients with Neoehrlichia mikurensis infection

Purpose The tick-borne bacterium Neoehrlichia mikurensis causes the infectious disease neoehrlichiosis in humans. Vascular endothelium is one of the target cells of the infection. Neoehrlichiosis patients with compromised B cell immunity present with more severe inflammation than immunocompetent patients. The aim of this study was to compare the cytokine profiles of immunocompetent and immunosuppressed patients with neoehrlichiosis. Methods Blood samples from Swedish and Norwegian immunosuppressed (N = 30) and immunocompetent (N = 16) patients with neoehrlichiosis were analyzed for the levels of 30 cytokines, using a multiplex cytokine assay and ELISA. A gender-matched healthy control group (N = 14) was analyzed in parallel. Data were analyzed using the multivariate method OPLS-DA. Results The multiplex cytokine analyses generated more cytokine results than did the uniplex ELISA analyses. Multivariate analysis of the multiplex cytokine results established that increased levels of FGF2, GM-CSF, CXCL10, and IFNγ were associated with immunosuppressed patients, whereas increased levels of IL-15 and VEGF were associated with immunocompetent neoehrlichiosis patients. When multivariate analysis findings were confirmed with uniplex ELISA, it was found that both groups of patients had similarly elevated levels of VEGF, FGF2 and IFNγ. In contrast, the immunosuppressed patients had clearly elevated levels of CXCL10, CXCL13 and BAFF, whereas the immunocompetent patients had the same levels as healthy controls. Conclusion Pro-angiogenic and type 1 cytokines were produced as part of the host response of neoehrlichiosis independent of immune status, whereas immunosuppressed neoehrlichiosis patients produced cytokines required for B cell-mediated defense.Funding Agencies: University of Gothenburg; European Union through the European Regional Development Fund, European Commission; Interreg North Sea Region Programme [J-No.: 38-2-7-19]; Swedish government; county councils; ALF-agreement ALF Research Fund [ALFGBG-827291]; Cancer and Allergy Foundation [2020-10154]; Swedish Research Council, European Commission [K200858X-14631-06-3, 2020-01287]</p

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Additional file 3. Percentage of RSV specific CD3+CD4+ and CD3+CD8+ responses

Collagenous Gastritis in Children : Incidence, Disease Course, and Associations With Autoimmunity and Inflammatory Markers

INTRODUCTION: Collagenous gastritis (CG), a rare disorder of unknown etiology, has been postulated to have immune-mediated mechanisms. We investigated (i) the incidence and prevalence of CG in a pediatric population; (ii) the clinical, endoscopic, and histologic characteristics of childhood-onset CG; and (iii) the evidence for autoimmunity and/or inflammatory activity in these patients. METHODS: Clinical, endoscopic, and histologic data were reviewed longitudinally in a population-based Swedish cohort of 15 patients with childhood-onset CG diagnosed in the period 2008-2019. A set of 11 autoantibodies, 4 blood inflammatory biomarkers, and the human leukocyte antigen DQ2/DQ8 genotype was analyzed cross-sectionally. RESULTS: The incidence rate of childhood-onset CG was 0.25/100,000 person-years, with an incidence rate ratio of girls to boys of 4.2 (95% confidence interval, 1.2-15). The prevalence of CG was 2.1/100,000 in children aged younger than 18 years. The endoscopic and histologic findings remained pathologic in all the examined patients during a median follow-up of 4.4 years. Many patients had heredity for autoimmune disorders (47%) and/or tested positive for autoantibodies (40%) or human leukocyte antigen DQ2/DQ8 (53%). No associated autoimmune comorbidities were observed. The serum levels of calprotectin and amyloid A were increased in 10/15 (67%) and 5/15 (33%) of the patients, respectively, whereas plasma C-reactive protein levels were normal in all, but 1 patient. DISCUSSION: The results indicate that childhood-onset CG is rare and has a chronic disease course. Although signs of autoimmune predisposition are frequent, early development of autoimmune comorbidities seems seldom. Serum calprotectin and amyloid A represent novel candidate biomarkers of inflammatory activity in CG (see Visual Abstract, Supplementary Digital Content 4, http://links.lww.com/CTG/A349)

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Additional file 4. Workflow to perform viSNE analysis on antigen-specific cells