Presence of Increased Mast Cells in Infants and Children with Volume and Variety Limited Intake

Background: Reports indicate patients with feeding difficulties demonstrate signs of inflammation on biopsies, notably eosinophilia, but it is unknown whether mast cell density contributes to variety or volume limitation symptoms. The aim of our study was to evaluate eosinophil and mast cell density of EGD biopsies in pediatric patients with symptoms of decreased volume or variety of ingested foods. Methods: We conducted a single-center, retrospective chart review of EMRs for all new feeding clinic patients between 0 and 17 years of age. Patients were categorized by symptoms at the initial visit as well as eosinophil and mast cell densities in those with EGD biopsies. Ten patients were identified as controls. Results: We identified 30 patients each with volume and variety limitation. Antral mast cell density was increased in 32.1% of variety-limited patients, 37.5% of volume limited patients, and in no controls; Duodenal mast cell density was increased in 32.1% of variety-limited patients, 40.6% of volume-limited patients, and in no controls. Conclusions: In both variety- and volume-limited patients, antral and duodenal mast cell densities were increased. These associations warrant further investigation of the mechanism between mast cells and development of feeding difficulties, allowing more targeted pediatric therapies.

Spatial and Temporal Heterogeneity of Tumor-Infiltrating Lymphocytes in Advanced Urothelial Cancer

Checkpoint inhibitors targeting PD-(L)1 induce objective responses in 20% of patients with metastatic urothelial cancer (UC). CD8+ T cell infiltration has been proposed as a putative biomarker for response to checkpoint inhibitors. Nevertheless, data on spatial and temporal heterogeneity of tumor-infiltrating lymphocytes in advanced UC are lacking. The major aims of this study were to explore spatial heterogeneity for lymphocyte infiltration and to investigate how the immune landscape changes during the disease course. We performed multiplex immunohistochemistry to assess the density of intratumoral and stromal CD3+, CD8+, FoxP3+ and CD20+ immune cells in longitudinally collected samples of 49 UC patients. Within these samples, spatial heterogeneity for lymphocyte infiltration was observed. Regions the size of a 0.6 tissue microarray core (0.28 mm2) provided a representative sample in 60.6 to 71.6% of cases, depending on the cell type of interest. Regions of 3.30 mm2, the median tumor surface area in our biopsies, were representative in 58.8 to 73.8% of cases. Immune cell densities did not significantly differ between untreated primary tumors and metachronous distant metastases. Interestingly, CD3+, CD8+ and FoxP3+ T cell densities decreased during chemotherapy in two small cohorts of patients treated with neoadjuvant or palliative platinum-based chemotherapy. In conclusion, spatial heterogeneity in advanced UC challenges the use of immune cell infiltration in biopsies as biomarker for response prediction. Our data also suggests a decrease in tumor-infiltrating T cells during platinum-based chemotherapy.

Optimized Cell Mixing Facilitates the Reproducible Bioprinting of Constructs with High Cell Viability

Bioprinting with cell-laden hydrogels (bioink) requires the careful mixing of cells with the hydrogel carrier to ensure that the bioink is homogeneous and functional, and the printing results are reproducible. Bioink preparation is therefore a critical process step that must accommodate the specific rheological properties of different bioinks. Here, we developed a reproducible method for the optimized mixing of cells and hydrogel carriers that can be integrated into current bioprinting processes. First, we tested and optimized different mixing devices for their effect on bioink homogeneity and rheological properties, resulting in a low-shear process for the preparation of homogenous bioinks. Based on these findings, we evaluated the impact of different cell densities on the rheological profile of bioinks according to shear and temperature, and estimated the impact of shear stress intensity and duration on 1.1B4 cells. Finally, we integrated the optimized mixing method into a current printing process and monitored the printed construct for 14 days to confirm cell viability. We found that the cell viability in the printed cell-laden constructs remained in excess of 91% after 14 days.

Self-generated oxygen gradients control collective aggregation of photosynthetic microbes

For billions of years, photosynthetic microbes have evolved under the variable exposure to sunlight in diverse ecosystems and microhabitats all over our planet. Their abilities to dynamically respond to alterations of the luminous intensity, including phototaxis, surface association and diurnal cell cycles, are pivotal for their survival. If these strategies fail in the absence of light, the microbes can still sustain essential metabolic functionalities and motility by switching their energy production from photosynthesis to oxygen respiration. For suspensions of motile C. reinhardtii cells above a critical density, we demonstrate that this switch reversibly controls collective microbial aggregation. Aerobic respiration dominates over photosynthesis in conditions of low light, which causes the microbial motility to sensitively depend on the local availability of oxygen. For dense microbial populations in self-generated oxygen gradients, microfluidic experiments and continuum theory based on a reaction–diffusion mechanism show that oxygen-regulated motility enables the collective emergence of highly localized regions of high and low cell densities.

Corneal dendritic cells and the subbasal nerve plexus following neurotoxic treatment with oxaliplatin or paclitaxel

Immune cell infiltration has been implicated in neurotoxic chemotherapy for cancer treatment. However, our understanding of immune processes is still incomplete and current methods of observing immune cells are time consuming or invasive. Corneal dendritic cells are potent antigen-presenting cells and can be imaged with in-vivo corneal confocal microscopy. Corneal dendritic cell densities and nerve parameters in patients treated with neurotoxic chemotherapy were investigated. Patients treated for cancer with oxaliplatin (n = 39) or paclitaxel (n = 48), 3 to 24 months prior to assessment were recruited along with 40 healthy controls. Immature (ImDC), mature (MDC) and total dendritic cell densities (TotalDC), and corneal nerve parameters were analyzed from in-vivo corneal confocal microscopy images. ImDC was increased in the oxaliplatin group (Median, Md = 22.7 cells/mm2) compared to healthy controls (Md = 10.1 cells/mm2, p = 0.001), but not in the paclitaxel group (Md = 10.6 cells/mm2). ImDC was also associated with higher oxaliplatin cumulative dose (r = 0.33, p = 0.04) and treatment cycles (r = 0.40, p = 0.01). There was no significant difference in MDC between the three groups (p > 0.05). Corneal nerve parameters were reduced in both oxaliplatin and paclitaxel groups compared to healthy controls (p < 0.05). There is evidence of elevation of corneal ImDC in oxaliplatin-treated patients. Further investigation is required to explore this potential link through longitudinal studies and animal or laboratory-based immunohistochemical research.

Metabolome and microbiome of chronic periapical periodontitis in permanent anterior teeth: a pilot study

Background Periapical periodontitis is a common oral inflammatory disease that affects periapical tissues and is caused by bacteria in the root canal system. The relationship among the local metabolome, the inflammatory grade, and the type and abundance of microorganisms associated with periapical periodontitis is discussed in this study. Methods The inflammatory grades of periapical samples from 47 patients with chronic periapical periodontitis in permanent anterior teeth were determined based on the immune cell densities in tissues subjected to haematoxylin and eosin staining. The metabolome was evaluated using ultrahigh-performance liquid chromatography-quadrupole time-of-flight mass spectrometry, followed by principal component analysis and orthogonal partial least squares discriminant analysis. The microbiome was accessed using 16 S rRNA high-throughput sequencing. The differences in the metabolomes and microbiomes of the periapical periodontitis samples were assessed using Spearman’s correlation analysis. Result N-acetyl--glucosamine, -tryptophan, -phenylalanine, and 15 other metabolites were identified by the comparison between samples with severe inflammation and mild or moderate inflammation. Four amino acid metabolism pathways and one sugar metabolism pathway were associated with the inflammatory grade of periapical periodontitis. The abundance of Actinomycetes was negatively correlated with the abundance of glucosamine (GlcN), while the abundance of Tannerella was positively correlated with the abundance of -methionine. Conclusions The local metabolome of periapical periodontitis is correlated with the inflammatory grade. The abundance of the local metabolites GlcN and -methionine is correlated with the abundance of the major microorganisms Actinomycetes and Tannerella, respectively.

Novel Platform for Regulation of Extracellular Vesicles and Metabolites Secretion from Cells Using a Multi-Linkable Horizontal Co-Culture Plate

Microfluidics is applied in biotechnology research via the creation of microfluidic channels and reaction vessels. Filters are considered to be able to simulate microfluidics. A typical example is the cell culture insert, which comprises two vessels connected by a filter. Cell culture inserts have been used for years to study cell-to-cell communication. These systems generally have a bucket-in-bucket structure and are hereafter referred to as a vertical-type co-culture plate (VTCP). However, VTCPs have several disadvantages, such as the inability to simultaneously observe samples in both containers and the inability of cell-to-cell communication through the filters at high cell densities. In this study, we developed a novel horizontal-type co-culture plate (HTCP) to overcome these disadvantages and confirm its performance. In addition, we clarified the migration characteristics of substances secreted from cells in horizontal co-culture vessels. It is generally assumed that less material is exchanged between the horizontal vessels. However, the extracellular vesicle (EV) transfer was found to be twice as high when using HTCP. Other merits include control of the degree of co-culture via the placement of cells. We believe that this novel HTCP container will facilitate research on cell-to-cell communication in various fields.

High Densities of a Prochlorophyte (Unresolved Species) Inhibit Grazing by the Herbivorous Copepod Parvocalanus crassirostris

Harmful algal blooms (HABs) are increasing in frequency and severity, underscoring the importance of understanding potential top-down controls of blooms. In the Indian River Lagoon (IRL), a shallow subtropical estuary, one harmful bloom was co-dominated by an unresolved prochlorophyte in the Family Prochlorothricaceae, which reached densities of &gt; 106 cells ml–1 engendering the term “Superbloom.” Experiments were conducted to evaluate grazing rates and the potential for top-down control by an abundant herbivorous copepod, Parvocalanus crassirostris, on the prochlorophyte. Those grazing rates were lower than the rates on a palatable alternative algal food, Isochrysis galbana, when both algal species were presented in monocultures with identical densities. Grazing on the prochlorophyte decreased or ceased at densities over 4.8 × 105 cells ml–1. When the prochlorophyte and the palatable alternative each comprised half of the total density, both species were consumed, but grazing on I. galbana was reduced compared to the grazing rates in a monoculture of this species, especially at higher cell densities. Copepod mortality was observed in treatments with high concentrations of the prochlorophyte, and these treatments contained mucilage. Experiments simulating viscosities produced by prochlorophyte mucilage yielded results consistent with the original grazing experiments (i.e., copepods showed lower grazing rates and higher mortality rates in higher viscosity treatments). Results reveal potential limitations of top-down controls by this grazer on prochlorophyte blooms and HABs that produce mucilage.