POSaM: a fast, flexible, open-source, inkjet oligonucleotide synthesizer and microarrayer

DNA arrays are valuable tools in molecular biology laboratories. Their rapid acceptance was aided by the release of plans for a pin-spotting microarrayer by researchers at Stanford. Inkjet microarraying is a flexible, complementary technique that allows the synthesis of arrays of any oligonucleotide sequences de novo. We describe here an open-source inkjet arrayer capable of rapidly producing sets of unique 9,800-feature arrays

Modulation of Immune Checkpoints by Chemotherapy in Human Colorectal Liver Metastases.

Metastatic colorectal cancer (CRC) is a major cause of cancer-related death, and incidence is rising in younger populations (younger than 50 years). Current chemotherapies can achieve response rates above 50%, but immunotherapies have limited value for patients with microsatellite-stable (MSS) cancers. The present study investigates the impact of chemotherapy on the tumor immune microenvironment. We treat human liver metastases slices with 5-fluorouracil (5-FU) plus either irinotecan or oxaliplatin, then perform single-cell transcriptome analyses. Results from eight cases reveal two cellular subtypes with divergent responses to chemotherapy. Susceptible tumors are characterized by a stemness signature, an activated interferon pathway, and suppression of PD-1 ligands in response to 5-FU+irinotecan. Conversely, immune checkpoint TIM-3 ligands are maintained or upregulated by chemotherapy in CRC with an enterocyte-like signature, and combining chemotherapy with TIM-3 blockade leads to synergistic tumor killing. Our analyses highlight chemomodulation of the immune microenvironment and provide a framework for combined chemo-immunotherapies

A wellness study of 108 individuals using personal, dense, dynamic data clouds.

Personal data for 108 individuals were collected during a 9-month period, including whole genome sequences; clinical tests, metabolomes, proteomes, and microbiomes at three time points; and daily activity tracking. Using all of these data, we generated a correlation network that revealed communities of related analytes associated with physiology and disease. Connectivity within analyte communities enabled the identification of known and candidate biomarkers (e.g., gamma-glutamyltyrosine was densely interconnected with clinical analytes for cardiometabolic disease). We calculated polygenic scores from genome-wide association studies (GWAS) for 127 traits and diseases, and used these to discover molecular correlates of polygenic risk (e.g., genetic risk for inflammatory bowel disease was negatively correlated with plasma cystine). Finally, behavioral coaching informed by personal data helped participants to improve clinical biomarkers. Our results show that measurement of personal data clouds over time can improve our understanding of health and disease, including early transitions to disease states

Maximum static inspiratory and expiratory pressures with different lung volumes

BACKGROUND: Maximum pressures developed by the respiratory muscles can indicate the health of the respiratory system, help to determine maximum respiratory flow rates, and contribute to respiratory power development. Past measurements of maximum pressures have been found to be inadequate for inclusion in some exercise models involving respiration. METHODS: Maximum inspiratory and expiratory airway pressures were measured over a range of lung volumes in 29 female and 19 male adults. A commercial bell spirometry system was programmed to occlude airflow at nine target lung volumes ranging from 10% to 90% of vital capacity. RESULTS: In women, maximum expiratory pressure increased with volume from 39 to 61 cmH(2)O and maximum inspiratory pressure decreased with volume from 66 to 28 cmH(2)O. In men, maximum expiratory pressure increased with volume from 63 to 97 cmH(2)O and maximum inspiratory pressure decreased with volume from 97 to 39 cmH(2)O. Equations describing pressures for both sexes are: P(e)/P(max )= 0.1426 Ln( %VC) + 0.3402 R(2 )= 0.95 P(i)/P(max )= 0.234 Ln(100 - %VC) - 0.0828 R(2 = )0.96 CONCLUSION: These results were found to be consistent with values and trends obtained by other authors. Regression equations may be suitable for respiratory mechanics models

Tumor slice culture as a biologic surrogate of human cancer.

Background: The tumor microenvironment (TME) is critical to every aspect of cancer biology. Organotypic tumor slice cultures (TSCs) preserve the original TME and have demonstrated utility in predicting drug sensitivity, but the association between clinicopathologic parameters and Methods: One hundred and eight fresh tumor specimens from liver resections at a tertiary academic center were procured and precisely cut with a Vibratome to create 250 μm × 6 mm slices. These fixed-dimension TSCs were grown on polytetrafluoroethylene inserts, and their metabolic activities were determined by a colorimetric assay. Correlation between baseline activities and clinicopathologic parameters was assessed. Tissue CEA mRNA expression was determined by RNAseq. Results: By standardizing the dimensions of a slice, we found that adjacent tumor slices have equivalent metabolic activities, while those derived from different tumors exhibit \u3e30-fold range in baseline MTS absorbances, which correlated significantly with the percentage of tumor necrosis based on histologic assessment. Extending this to individual cancers, we were able to detect intra-tumoral heterogeneity over a span of a few millimeters, which reflects differences in tumor cell density and Ki-67 positivity. For colorectal cancers, tissue CEA expression based on RNAseq of tumor slices was found to correlate with clinical response to chemotherapies. Conclusions: We report a standardized method to assess and compare human cancer growth ex vivo across a wide spectrum of tumor samples. TSC reflects the state of tumor behavior and heterogeneity, thus providing a simple approach to study of human cancers with an intact TME

Synthesis of Molecularly Imprinted Polymers for Protein Detection and Sample Depletion

Molecular Imprint Polymers (MIPs) constitute a unique class of chemically-synthesized antigen recognition materials. As microparticles and nanoparticles, they can act as synthetic antibodies with numerous potential applications. We will develop protocols for the rapid generation of albumin-binding MIPs, characterize the affinity and specificity of the resulting particles, and investigate their application towards serum depletion for proteomics

Novel metrics for quantifying bacterial genome composition skews

Abstract Background Bacterial genomes have characteristic compositional skews, which are differences in nucleotide frequency between the leading and lagging DNA strands across a segment of a genome. It is thought that these strand asymmetries arise as a result of mutational biases and selective constraints, particularly for energy efficiency. Analysis of compositional skews in a diverse set of bacteria provides a comparative context in which mutational and selective environmental constraints can be studied. These analyses typically require finished and well-annotated genomic sequences. Results We present three novel metrics for examining genome composition skews; all three metrics can be computed for unfinished or partially-annotated genomes. The first two metrics, (dot-skew and cross-skew) depend on sequence and gene annotation of a single genome, while the third metric (residual skew) highlights unusual genomes by subtracting a GC content-based model of a library of genome sequences. We applied these metrics to 7738 available bacterial genomes, including partial drafts, and identified outlier species. A phylogenetically diverse set of these outliers (i.e., Borrelia, Ehrlichia, Kinetoplastibacterium, and Phytoplasma) display similar skew patterns but share lifestyle characteristics, such as intracellularity and biosynthetic dependence on their hosts. Conclusions Our novel metrics appear to reflect the effects of biosynthetic constraints and adaptations to life within one or more hosts on genome composition. We provide results for each analyzed genome, software and interactive visualizations at http://db.systemsbiology.net/gestalt/ skew_metrics

Single-cell analysis of erythropoiesis in Rpl11 haploinsufficient mice reveals insight into the pathogenesis of Diamond Blackfan anemia.

Rpl11 haploinsufficient mice develop a macrocytic anemia similar to patients with DBA. Here, we fully characterize this model from clinical and pathophysiological perspectives. Early erythroid precursors have increased heme content and high cytoplasmic ROS, impairing erythroid differentiation at the CFU-E/proerythroblast stage and subsequently. Using single-cell analyses that link a cell\u27s surface protein expression to its total transcriptome and unbiased analyses, we show GATA1, GATA1 target gene and mitotic spindle pathway gene transcription were the pathways most decreased. Expression of ribosome protein and globin genes were amplified. These changes, as well as the other transcriptional changes that were identified, closely resemble findings in mice that lack the heme export protein FLVCR, and thus suggest that heme excess and toxicity are the primary drivers of the macrocytic anemia. Consistent with this, treating Rpl11 haploinsufficient mice with corticosteroids increased the numbers of earliest erythroblasts but failed to overcome heme toxicities and improve the anemia. Rpl11 haploinsufficient mice uniquely upregulated mitochondrial genes, p53 and CDKN1A pathway genes, and DNA damage checkpoint genes, which should contribute further to erythroid marrow failure. Together our data establish Rpl11 haploinsufficient mice as an excellent model of DBA that can be used to study DBA pathogenesis and test novel therapies