Biology subjects

Campos, S.

Publications and source records attributed to Campos, S..

5 recordsLinked to original sources

bioRxiv2025

Cryopreservation of Platynereis dumerilii larvae

The marine annelid Platynereis dumerilii is a functional molecular model organism for developmental, evolutionary and chronobiological studies. Research on Platynereis is rapidly growing, and with it, the number of genetic variants that laboratories isolate or generate and must subsequently maintain and propagate. Therefore, there is an urgent need to alleviate the burden of live culture maintenance by developing cryopreservation techniques for this species. We report the first cryopreservation protocol for P. dumerilii larvae, which combined with a careful post-thawing culturing regime, allowed us to obtain animals that survived to adulthood and successfully reproduced. Our experiments show highest survival rate in 6 - 8 day-old larvae. Equilibration with cryoprotecting agents takes 1h in 5% (v/v) Me2SO + 0.1%(v/w) sucrose, followed by transfer to 0.25ml straws. The protocol cools larvae at 2,5{degrees}C/min from 20{o}C to -35{degrees}C using a programmable freezer, followed by a rapid transfer to liquid N2. Larvae are thawed in a water bath at 18{degrees}C. The post-thaw larvae feeding regime consisted of 50% Tetraselmis + 50 % diatom strains mixture (Grammatophora marina and Nitzschia laevis). The maximum survival obtained with this protocol so far produced 34% survival after [~]5 months.

zoology↗

bioRxiv2024

Brain-body circuit by which midbrain dopamine neuronal activity modulates splenic immunity

Midbrain dopamine neurons are well-known to shape central nervous system function, yet there is growing evidence for their influence on the peripheral immune system. Here we demonstrate that midbrain dopamine neurons form a circuit to the spleen via a multi-synaptic pathway from the dorsal vagal complex (DVC) through the celiac ganglion. Midbrain dopamine neurons modulate the activity of D1- and D2-like dopamine receptor-expressing DVC neurons. In vivo activation of midbrain dopamine neurons induced dopamine release in the DVC and increased immediate early gene expression in both the DVC and celiac ganglion. Activation of this midbrain-to-spleen circuit reduced spleen weight and decreased naive CD4+ T-cell populations without affecting total T-cell numbers. These findings unveil a functional midbrain- DVC-celiac ganglion-spleen pathway, through which midbrain dopamine neurons modulate splenic immunity. These novel insights into the neural regulation of the immune system have important implications for diseases involving altered dopamine neurotransmission and highlight potential targets for immunotherapeutic interventions.

neuroscience↗

bioRxiv2024

Comprehensive Proteomic, Ultrastructural and Functional Analysis of Cryopreservation Effects on Marine Mussel Oocytes

Cryopreservation is a valuable tool for preserving marine genetic diversity and supporting selective breeding in aquaculture. The blue mussel (Mytilus galloprovincialis) is an important species for the aquaculture industry and a useful model for studying cryopreservation in marine invertebrates. Declining mussel seed availability over the past decade, combined with the growing demand for year-round hatchery production, has increased interest in developing effective cryopreservation methods for gametes and embryos. However, cryopreservation remains challenging, particularly for marine invertebrate oocytes, as it requires balancing cryoprotectant cytotoxicity with sufficient cell protection during freezing and thawing. Moreover, the molecular, structural, and functional changes associated with oocyte cryopreservation in invertebrates are still poorly understood. This study examines the effects of two commonly used cryoprotectants, dimethyl sulfoxide (DMSO) and ethylene glycol (EG), on M. galloprovincialis oocytes under a monitored slow freezing (MSF) protocol. Using an integrative analytical approach combining proteomics, functional assessments, and ultrastructural analyses, we provide a comprehensive understanding of cryoprotectant-induced changes. Our findings reveal that cryoprotectants cause significant proteomic alterations, with DMSO having more pronounced effects. These alterations are associated with increased oxidative stress, an insufficient antioxidant response, and disruptions in meiosis restart mechanisms, ultimately delaying larval development. Cryoprotectant-induced oxidative stress may also weaken the oocyte membrane, leading to rupture during the MSF protocol and subsequent fertilisation failure post-thawing. These results provide evidence that EG is less cytotoxic than DMSO and suggest that supplementing cryoprotectants with antioxidants and membrane stabilizers could enhance success rates in the cryopreservation of oocytes from mussels and other marine invertebrate species.

developmental biology↗

bioRxiv2021

Development of novel MOG analogues with increased stability to explore MCT2 and α-ketoglutarate biology in vivo

-ketoglutarate (KG) is a central metabolic node with a broad influence on cellular physiology. The KG analogue N-oxalylglycine (NOG) and its membrane-permeable pro-drug derivative dimethyl-oxalylglycine (DMOG) have been extensively used as tools to study prolyl hydroxylases (PHDs) and other KG-dependent processes. In cell culture media, DMOG is rapidly converted to MOG, which enters cells through monocarboxylate transporter MCT2, leading to intracellular NOG concentrations that are sufficiently high to inhibit glutaminolysis enzymes and cause cytotoxicity. Therefore, the degree of (D)MOG instability together with MCT2 expression levels determine the intracellular targets NOG engages with and, ultimately, its effects on cell viability. Here we designed and characterised a series of MOG analogues with the aims of improving stability and exploring the functional requirements for interaction with MCT2, a relatively understudied member of the SLC16 family. We report MOG analogues that maintain ability to enter cells via MCT2, and identify compounds that do not inhibit glutaminolysis or cause cytotoxicity but can still inhibit PHDs. We use these analogues to show that glutaminolysis-induced activation of mTORC1 can be uncoupled from PHD activity. Therefore, these new compounds can help deconvolute cellular effects that result from the polypharmacological action of NOG.

biochemistry↗

bioRxiv2020

Isoform-level transcriptome Atlas of Macrophage Activation

RNA-seq is routinely used to measure gene expression changes in response to cell perturbation. Genes that are up or down-regulated following perturbation in RNA-seq studies are designated as target genes for follow-up. However, RNA-seq is limited in its ability to capture the complexity of gene isoforms, defined by the exact composition of exons and transcription start sites (TSS) and poly(A) sites they contain, as well as the expression of these isoforms. Without knowing the composition of the most dominant isoform(s) of a target gene, a minority or non-existent isoform could be selected for follow-up solely based on available annotations for that target gene from databases that are incomplete, or by their nature not tissue specific, or do not provide key information on expression levels. In all, this can lead to loss in valuable resources and time. As the vast majority of genes in the human genome express more than one isoform, there is a great need to identify the complete range of isoforms present for each gene along with their corresponding levels of expression. Here, using the long-read nanopore-based R2C2 method, we generated an Isoform-level transcriptome Atlas of Macrophage Activation (IAMA) that identifies full-length isoforms in primary human monocyte-derived macrophages (MDMs). Macrophages are critical innate immune cells important for recognizing pathogens through use of Toll-like receptors (TLRs), culminating in the initiation of host defense pathways. We characterized isoforms for most moderate to highly expressed genes in resting and TLR-activated MDMs and generated a user-friendly portal built into the UCSC Genome Browser to explore the data (https://genome.ucsc.edu/s/vollmers/IAMA). Our atlas represents a valuable resource for innate immune research as it provides unprecedented isoform information for primary human macrophages.

immunology↗