Biology subjects

Valencia, M.

Publications and source records attributed to Valencia, M..

6 recordsLinked to original sources

bioRxiv2025

Biomarker Quantification in Breast Cancer using Xenium In Situ

Advances in spatial transcriptomics enable high-throughput quantitation of both established and novel biomarkers at single cell resolution, offering the potential to transform diagnostics. Using Xenium in situ technology in FFPE human breast samples, we address two challenges in the cancer field: 1. achieving reliable normalization of gene expression across heterogeneous sample populations; 2. identifying biomarkers that predict invasion or metastasis. We describe a scalable approach to identify low-variation housekeeping (HK) genes within any given sample set, then use those HK genes for cross- and intra-sample normalization of biomarkers. Analyzing 12 FFPE human breast samples-primarily ductal carcinoma in situ (DCIS)-with a custom 280-gene panel, we identified four HK genes (EEF1G, EEF2, MALAT1, and RPLP0) that exhibited minimal variability in tumor cells, four tumor cell biomarkers (LDHA, SDC1, PIGR, SFRP1) that increased or decreased with tumor grade, and one tumor-associated myoepithelial biomarker (LAMC2). Normalizing biomarkers to the four HK genes preserved the dynamic range of expression necessary for distinguishing tumor grades, outperforming HKs from legacy RT-PCR diagnostic panels. Lastly, we employed a cell-agnostic approach in the tumor periphery to quantify MMP11, a biomarker correlated with proliferative and potentially pre-invasive ducts. Our results establish a single cell normalization method for spatial in situ transcriptomics and reveal and quantitate biomarkers relevant to DCIS risk and progression.

cancer biology↗

bioRxiv2023

Cultivating efficiency: High-throughput growth analysis of anaerobic bacteria in compact microplate readers.

Anaerobic microbes play crucial roles in environmental processes, industry, and human health. Traditional methods for monitoring the growth of anaerobes, including plate counts or subsampling broth cultures for optical density measurements, are time and resource intensive. The advent of microplate readers revolutionized bacterial growth studies by enabling high-throughput and real-time monitoring of microbial growth kinetics but their use in anaerobic microbiology has remained limited. Here, we present a workflow for using small-footprint microplate readers and the Growthcurver R package to analyze the kinetic growth metrics of anaerobic bacteria. We benchmarked the small-footprint Cerillo Stratus microplate reader against a BioTek Synergy HTX microplate reader in aerobic conditions using Escherichia coli DSM 28618 cultures. The growth rates and carrying capacities obtained from the two readers were statistically indistinguishable. However, the area under the logistic curve was significantly higher in cultures monitored by the Stratus reader. We used the Stratus to quantify the growth responses of anaerobically grown E. coli and Clostridium bolteae DSM 29485 to different doses of the toxin sodium arsenite. The growth of E. coli and C. bolteae was sensitive to arsenite doses of 1.3 M and 0.4 M, respectively. Complete inhibition of growth was achieved at 38 M arsenite for C. bolteae, and 338 M in E. coli. These results show that the Stratus performs similarly to a leading brand of microplate reader and can be reliably used in anaerobic conditions. We discuss the advantages of the small format microplate readers and our experiences with the Stratus. Importance statementWe present a workflow that facilitates the production and analysis of growth curves for anaerobic microbes using small-footprint microplate readers and an R script. This workflow is a cost and space-effective solution to most high-throughput solutions for collecting growth data from anaerobic microbes. This technology can be used for applications in which high-throughput would advance discovery, including microbial isolation, bioprospecting, co-culturing, host-microbe interactions, and drug/toxin-microbial interactions.

microbiology↗

bioRxiv2023

Staphyloccocus aureus biofilm, in the absence of planktonic bacteria, produces factors that activate counterbalancing inflammatory and immune-suppressive genes in human monocytes

Staphyloccocus aureus (S. aureus) is a major bacterial pathogen in orthopedic periprosthetic joint infection (PJI). S. aureus forms biofilms that promote persistent infection by shielding bacteria from immune cells and inducing an antibiotic-resistant metabolic state. We developed an in vitro system to study S. aureus biofilm interactions with primary human monocytes in the absence of planktonic bacteria. In line with previous in vivo data, S. aureus biofilm induced expression of inflammatory genes such as TNF and IL1B, and their anti-inflammatory counter-regulator IL-10. S. aureus biofilm also activated expression of PD-1 ligands that suppress T cell function, and of IL-1RA that suppresses differentiation of protective Th17 cells. Gene induction did not require monocyte:biofilm contact and was mediated by a soluble factor(s) produced by biofilm-encased bacteria that was heat resistant and > 3 kD in size. Activation of suppressive genes by biofilm was sensitive to suppression by Jak inhibition. These results support an evolving paradigm that biofilm plays an active role in modulating immune responses, and suggest this occurs via production of a soluble vita-PAMP. Induction of T cell suppressive genes by S. aureus biofilm provides insights into mechanisms that suppress T cell immunity in PJI, and suggest that anti-PD-1 therapy that is modeled on immune checkpoint blockade for tumors may be beneficial in PJI.

immunology↗

bioRxiv2023

JAMS - A framework for the taxonomic and functional exploration of microbiological genomic data

Shotgun microbiome sequencing analysis presents several challenges to accurately and consistently depict sample composition and functional potential. Here we present a two-part framework - JAMS (Just a Microbiology System) - whereby with raw fastq files and metadata as input, meaningful analysis within a sample and between a sample can be performed with ease for either shotgun or 16S sequences. JAMS is the first package to provide seamless deconvolution of functions into their taxonomic contributors. We validated our JAMS framework on two human gut shotgun metagenome test datasets against the popular tool MetaPhlAn 4. We further demonstrate the application of the JAMS package, particularly the plotting functions, on a mouse shotgun dataset.

bioinformatics↗

bioRxiv2018

Cholinergic Midbrain Afferents Modulate Striatal Circuits and Shape Encoding of Action Control

Assimilation of novel strategies into a consolidated action repertoire is a crucial function for behavioral adaptation and cognitive flexibility. Acetylcholine in the striatum plays a pivotal role in such adaptation and its release has been causally associated with the activity of cholinergic interneurons. Here we show that the midbrain, a previously unknown source of acetylcholine in the striatum, is a major contributor to cholinergic transmission in the striatal complex. Neurons of the pedunculopontine and laterodorsal tegmental nuclei synapse with striatal cholinergic interneurons and give rise to excitatory responses that, in turn, mediate inhibition of spiny projection neurons. Inhibition of acetylcholine release from midbrain terminals in the striatum impairs action shifting and mimics the effects observed following inhibition of acetylcholine release from striatal cholinergic interneurons. These results suggest the existence of two hierarchically-organized modes of cholinergic transmission in the striatum where cholinergic interneurons are modulated by cholinergic neurons of the midbrain.

neuroscience↗

bioRxiv2018

Oscillatory cortical forces promote three dimensional cell intercalations that shape the mandibular arch

Multiple vertebrate embryonic structures such as organ primordia are composed of a volume of confluent cells. Although mechanisms that shape tissue sheets are increasingly understood, those which shape a volume of cells remain obscure. Here we show 3D mesenchymal cell intercalations, rather than cell divisions and biophysical tissue properties, are essential to shape the mandibular arch of the mouse embryo. Using a genetically encoded vinculin tension sensor, we show that cortical force oscillations promote these intercalations. Genetic loss and gain of function approaches show that Wnt5a functions as a spatial cue to coordinate cell polarity with cytoskeletal oscillation. YAP/TAZ and PIEZO1 serve as downstream effectors of Wnt5a-mediated actomyosin bias and cytosolic calcium transients, respectively, to ensure appropriate tissue form during growth. Our data support oriented 3D cell neighbour exchange as a conserved mechanism driving volumetric morphogenesis.

developmental biology↗