Breast milk fatty acids may link innate and adaptive immune regulation: analysis of soluble CD14, prostaglandin E2, and fatty acids

In addition to its role in sensing intraluminal microbial antigens, soluble (s)CD14 may regulate immune responses by its lesser known function as a lipid carrier with possible influences in the production of fatty acid-derived eicosanoids. We investigated the interrelations of fatty acids, prostaglandin E2 (PGE2), and sCD14 and their role in infant atopic eczema during the first year of life. Serum and breast milk samples from mothers and serum samples from their infants were collected at infant's age 3 mo and analyzed for sCD14 and PGE2 concentrations and for fatty acid compositions. The main correlation of sCD14 was with arachidonic acid (20:4n-6) (AA). Dihomo-gamma-linolenic acid (20:3n-6) (DHGLA) and the ratio of n-6 to n-3 fatty acids correlated positively and docosahexaenoic acid (22:6n-3) (DHA) and sum of n-3 fatty acid negatively with PGE2 in mother's serum and linoleic acid (LA) negatively with PGE2 in breast milk. Soluble CD14 tended to be higher and LA, total polyunsaturated fatty acid (PUFA), and sum of n-6 fatty acids were lower in breast milk received by infants with atopic eczema compared with those without. These results suggest that fatty acids contribute to the regulation of innate and adaptive immune responses and link intraluminal exposures, mother's diet, and microbes.     

ALDH1A1-related stemness in high-grade serous ovarian cancer is a negative prognostic indicator but potentially targetable by EGFR/mTOR-PI3K/aurora kinase inhibitors

Poor chemotherapy response remains a major treatment challenge for high-grade serous ovarian cancer (HGSC). Cancer stem cells are the major contributors to relapse and treatment failure as they can survive conventional therapy. Our objectives were to characterise stemness features in primary patient-derived cell lines, correlate stemness markers with clinical outcome and test the response of our cells to both conventional and exploratory drugs. Tissue and ascites samples, treatment-naive and/or after neoadjuvant chemotherapy, were prospectively collected. Primary cancer cells, cultured under conditions favouring either adherent or spheroid growth, were tested for stemness markers; the same markers were analysed in tissue and correlated with chemotherapy response and survival. Drug sensitivity and resistance testing was performed with 306 oncology compounds. Spheroid growth condition HGSC cells showed increased stemness marker expression (including aldehyde dehydrogenase isoform I; ALDH1A1) as compared with adherent growth condition cells, and increased resistance to platinum and taxane. A set of eight stemness markers separated treatment-naive tumours into two clusters and identified a distinct subgroup of HGSC with enriched stemness features. Expression of ALDH1A1, but not most other stemness markers, was increased after neoadjuvant chemotherapy and its expression in treatment-naive tumours correlated with chemoresistance and reduced survival. In drug sensitivity and resistance testing, five compounds, including two PI3K-mTOR inhibitors, demonstrated significant activity in both cell culture conditions. Thirteen compounds, including EGFR, PI3K-mTOR and aurora kinase inhibitors, were more toxic to spheroid cells than adherent cells. Our results identify stemness markers in HGSC that are associated with a decreased response to conventional chemotherapy and reduced survival if expressed by treatment-naive tumours. EGFR, mTOR-PI3K and aurora kinase inhibitors are candidates for targeting this cell population. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Serglycin inhibits the classical and lectin pathways of complement via its glycosaminoglycan chains: implications for multiple myeloma

Serglycin (SG) is a proteoglycan expressed by hematopoietic cells and is constitutively secreted by multiple myeloma (MM) cells. SG participates in the regulation of various inflammatory events. We found that SG secreted by human MM cell lines inhibits both the classical and lectin pathways of complement, without influencing alternative pathway activity. The inhibitory effect of SG is due to direct interactions with C1q and mannose-binding lectin (MBL). C1q-binding is mediated through the glycosaminoglycan moieties of SG, whereas MBL binds additionally to SG protein core. Interactions between SG and C1q as well as MBL are diminished in the presence of chondroitin sulfate type E. In addition, we localized the SG-binding site to the collagen-like stalk of C1q. Interactions between SG and C1q as well as MBL are ionic in character and only the interaction with MBL was found to be partially dependent on the presence of calcium. We found the serum levels of SG to be elevated in patients with MM compared to healthy controls. Moreover, we found that SG expressed from myeloma plasma cells protects these cells from complement activation induced by treatment with anti-thymocyte immunoglobulins. This might protect myeloma cells during immunotherapy and promote survival of malignant cells.     

Integrative analysis of microRNA, mRNA and aCGH data reveals asbestos- and histology-related changes in lung cancer

Lung cancer has the highest mortality rate of all of the cancers in the world and asbestos-related lung cancer is one of the leading occupational cancers. The identification of asbestos-related molecular changes has long been a topic of increasing research interest. The aim of this study was to identify novel asbestos-related molecular correlates by integrating miRNA expression profiling with previously obtained profiling data (aCGH and mRNA expression) from the same patient material. miRNA profiling was performed on 26 tumor and corresponding normal lung tissue samples from highly asbestos-exposed and non-exposed patients, and on eight control lung tissue samples. Data analyses on miRNA expression, and integration of miRNA and previously obtained mRNA data were performed using Chipster. A separate analysis was used to integrate miRNA and previously obtained aCGH data. Both known and new lung cancer-associated miRNAs and target genes with inverse correlation were discovered. Furthermore, DNA copy number alterations (e.g., gain at 12p13.31) were correlated with the deregulated miRNAs. Specifically, thirteen novel asbestos-related miRNAs (over-expressed: miR-148b, miR-374a, miR-24-1*, Let-7d, Let-7e, miR-199b-5p, miR-331-3p, and miR-96 and under-expressed: miR-939, miR-671-5p, miR-605, miR-1224-5p and miR-202) and inversely correlated target genes (e.g., GADD45A, LTBP1, FOSB, NCALD, CACNA2D2, MTSS1, EPB41L3) were identified. In addition, over-expression of the well known squamous cell carcinoma-associated miR-205 was linked to down-regulation of the DOK4 gene. The miRNAs/genes presented here may represent interesting targets for further investigation and could eventually have potential diagnostic implications.     

The microenvironment of proliferative diabetic retinopathy supports lymphatic neovascularization

Proliferative diabetic retinopathy (PDR) is a major diabetic microvascular complication characterized by pathological angiogenesis. Several retinopathy animal models have been developed to study the disease mechanisms and putative targets. However, knowledge on the human proliferative disease remains incomplete, relying on steady‐state results from thin histological neovascular tissue sections and vitreous samples. New translational models are thus required to comprehensively understand the disease pathophysiology and develop improved therapeutic interventions. We describe here a clinically relevant model, whereby the native multicellular PDR landscape and neo(fibro)vascular processes can be analysed ex vivo and related to clinical data. As characterized by three‐dimensional whole‐mount immunofluorescence and electron microscopy, heterogeneity in patient‐derived PDR neovascular tissues included discontinuous capillaries coupled with aberrantly differentiated, lymphatic‐like and tortuous endothelia. Spatially confined apoptosis and proliferation coexisted with inflammatory cell infiltration and unique vascular islet formation. Ex vivo‐cultured explants retained multicellularity, islet patterning and capillary or fibrotic outgrowth in response to vitreoretinal factors. Strikingly, PDR neovascular tissues, whose matched vitreous samples enhanced lymphatic endothelial cell sprouting, contained lymphatic‐like capillaries in vivo and developed Prox1⁺ capillaries and sprouts with lymphatic endothelial ultrastructures ex vivo. Among multiple vitreal components, vascular endothelial growth factor C was one factor found at lymphatic endothelium‐activating concentrations. These results indicate that the ischaemia‐induced and inflammation‐induced human PDR microenvironment supports pathological neolymphovascularization, providing a new concept regarding PDR mechanisms and targeting options. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Ecto‐5′‐nucleotidase/CD73 enhances endothelial barrier function and sprouting in blood but not lymphatic vasculature

CD73/ecto‐5′‐nucleotidase is a key enzyme in the regulation of purinergic signaling and inflammatory reactions. It hydrolyzes extracellular AMP into adenosine, which dampens immune cell activation, and reduces leukocyte trafficking. By comparing CD73 expression and function in mononuclear and endothelial cells (ECs) of blood and lymph, we show that extracellular purines and CD73 activity have differential effects in these two vascular systems. We found that CD8‐positive T lymphocytes and CD19‐positive B lymphocytes in human lymph expressed high levels of CD73 and other purinergic enzymes and adenosine receptors. Soluble CD73 was less abundant in human lymph than in serum, whereas CD73 activity was higher in afferent lymphatic ECs than in blood ECs. Adenosine signaling improved barrier function and induced sprouting of human blood, but not lymphatic, ECs in vitro. Similarly, using CD73‐deficient mice we found that CD73 controls only blood vascular permeability at selected lymphoid organs under physiological conditions. Thus, both vascular and lymphatic arms of the immune system synthesize the components of purinergic signaling system, but surprisingly they use CD73 differentially to control endothelial permeability and sprouting.     

Enhanced intracellular replication of Salmonella enteritidis in HLA–B27–expressing human monocytic cells: Dependency on glutamic acid at position 45 in the B pocket of HLA–B27

Objective

To reveal the cause of the impaired elimination of Salmonella enteritidis in HLA–B27–transfected human monocytic cells and to study whether the B pocket of HLA–B27 contributes to these modulatory effects.

Methods

Stable U937 cell transfectants expressing HLA–A2, B27, or different forms of B27 with amino acid substitutions in the B pocket were prepared. Mock‐transfected cells were prepared using the antibiotic resistance vector (pSV2neo) alone. Cells were differentiated, infected with S enteritidis, and the number of live intracellular S enteritidis organisms was determined using the colony‐forming unit method. To visualize intracellular S enteritidis, the bacteria were transformed with green fluorescent protein (GFP), and studied by confocal microscopy.

Results

Cells expressing wild‐type HLA–B27 were more permissive of intracellular replication of S enteritidis compared with mock‐transfected or A2‐transfected controls. Cells expressing B27 with an altered B pocket composition having either 6 amino acid substitutions (B27.A2B; substitutions H9F, T24A, E45M, I66K, C67V, and K70H) or a single substitution (B27.E45M) were no longer permissive of S enteritidis replication. In contrast, cells expressing B27 with the single substitution of F for H at position 9 (B27.H9F) retained their permissiveness. Studies using GFP‐transformed S enteritidis confirmed that the increase in the amount of intracellular bacteria in B27‐expressing cells was due to replication of the bacteria.

Conclusion

Our data indicate that HLA–B27 expression modulates the host–microbe interaction that results in an impaired capacity of monocytes to resist intracellular replication of S enteritidis. The phenotype is dependent on glutamic acid at position 45 in the B pocket and, thus, may be due to properties of the B27 heavy chain that are related to this residue. The ability of HLA–B27 to confer susceptibility to Salmonella‐triggered reactive arthritis may occur, at least in part, through these modulatory effects.