Biology subjects

Jimenez, J. A.

Publications and source records attributed to Jimenez, J. A..

5 recordsLinked to original sources

bioRxiv2022

The open field assay is influenced by room temperature and by drugs that affect core body temperature

The open field assay is used to study anxiety-related traits and anxiolytic drugs in rodents. This assay entails measuring locomotor activity and time spent in the center of a chamber that is maintained at ambient room temperature. However, the ambient temperature in most laboratories varies daily and seasonally and can differ substantially between buildings. Here, we sought to evaluate how varying ambient temperature and core body temperature (CBT) affected open field locomotor activity and center time of wild-type (WT) and Trpm8 knock-out (Trpm8-/-) mice. TRPM8 is an ion channel that detects cool temperatures and is activated by cooling agents, including icilin and menthol. We found that the cooling agent icilin increased CBT and profoundly reduced distance traveled and center time of WT mice relative to vehicle controls. Likewise, cooling the ambient temperature to 4{degrees}C reduced distance traveled and center time of WT mice relative to Trpm8-/- mice. Conversely, the TRPM8 antagonist (M8-B) reduced CBT and increased distance traveled and center time of WT mice when tested at 4{degrees}C. Predictably, the TRPM8 antagonist (M8-B) had no effect on CBT or open field behavior of Trpm8-/- mice. The anxiolytic diazepam reduced CBT in both WT and Trpm8-/- mice. When tested at 4{degrees}C, diazepam increased distance traveled and center time in WT mice but did not alter open field behavior of Trpm8-/- mice. Our study shows that environmental temperature and drugs that affect CBT can influence locomotor behavior and center time in the open field assay, highlighting temperature (ambient and core) as sources of environmental and physiologic variability in this commonly used behavioral assay.

neuroscience↗

bioRxiv2022

Development and validation of PainFace, a software platform that simplifies and standardizes mouse grimace analyses

Facial grimaces are now commonly used to quantify spontaneous pain in mice and other mammalian species, but scoring remains subjective and relies on humans with very different levels of proficiency. Here, we developed a Mouse Grimace Scale (MGS) for black-coated (C57BL/6) mice consisting of four facial action units (orbitals, nose, ears, whiskers). We used this scale to generate ground truth data from over 70,000 images of black mice in different settings. With this large data set, we developed a deep neural network and cloud-based software platform called PainFace (http://painface.net) that accurately scores facial grimaces of black mice on a 0-8 scale. PainFace generates over two orders of magnitude more MGS data than humans can realistically achieve, and at superhuman speed. By analyzing the frequency distribution of grimace scores, we found that mice spent >7x more time in a high grimace state following laparotomy surgery relative to sham surgery controls. The analgesic carprofen reduced the amount of time animals spent in this high grimace state after surgery. Specific facial action unit score combinations were overrepresented following laparotomy surgery, suggesting that characteristic facial expressions are associated with a high grimace state. We performed validation experiments in two labs located in different countries to demonstrate reproducibility of the PainFace platform. To further enhance rigor and reproducibility, we will invite pain researchers to beta test PainFace and then incorporate their feedback into the software and manuscript prior to peer review. While this study is focused on mice, PainFace was designed to simplify, standardize, and scale up grimace analyses with many other mammalian species, including humans.

neuroscience↗

bioRxiv2022

HOXD13 is a direct EWS-FLI1 target and moderates fusion-dependent transcriptional states

Oncogenic fusion proteins display exquisite tissue specificity, revealing that malignant transformation requires cooperation with cell-autonomous factors. Recent studies have also demonstrated that tumorigenicity of Ewing sarcoma requires precise regulation of the transcriptional activity of the EWS-FLI1 oncogenic driver. Here we show that the developmentally and anatomically restricted transcription factor HOXD13 is a direct target of EWS-FLI1. Transcriptomic and CUT&RUN studies revealed that HOXD13 binds active, fusion-bound enhancers, resulting in altered expression of EWS-FLI1-induced targets. More strikingly, HOXD13 was found to bind and activate cis-regulatory regions of genes that are normally repressed by EWS-FLI1. Single-cell sequencing demonstrated marked intra-tumoral heterogeneity of HOXD13 transcriptional activity and revealed that antagonism between HOXD13-mediated gene activation and EWS-FLI1-dependent gene repression confers a spectrum of transcriptional cell states along a mesenchymal axis. Thus, HOXD13 serves as an internal rheostat for EWS-FLI1 activity, providing a paradigm for tissue-specific transcription factors as critical partners in fusion-driven cancers.

cancer biology↗

bioRxiv2021

Systemic delivery of a CXCR4-CXCL12 signaling inhibitor encapsulated in synthetic protein nanoparticles for glioma immunotherapy

Glioblastoma multiforme (GBM) is an aggressive primary brain tumor, with poor prognosis. Major obstacles hampering effective therapeutic response in GBM are tumor heterogeneity, high infiltration of immunosuppressive myeloid cells, and the presence of the blood-brain barrier. The C-X-C Motif Chemokine Ligand 12/ C-X-C Motif Chemokine Receptor 4 (CXCL12/ CXCR4) signaling pathway is implicated in GBM invasion and cell cycle progression. While the CXCR4 antagonists (AMD3100) has a potential anti-GBM effects, its poor pharmacokinetic and systemic toxicity had precluded its clinical application. Moreover, the role of CXCL12/ CXCR4 signaling pathway in anti-GBM immunity, particularly in GBM-mediated immunosuppression has not been elucidated. Here, we developed a synthetic protein nanoparticle (SPNPs) coated with the cell-penetrating peptide iRGD (AMD3100 SPNPs) to target the CXCR4/CXCL12 signaling axis in GBM. We showed that AMD3100 SPNPs effectively blocked CXCR4 signaling in mouse and human GBM cells in vitro as well as in GBM model in vivo. This results in inhibition of GBM proliferation and induction of immunogenic tumor cell death (ICD) leading to inhibition of GBM progression. Our data also demonstrate that blocking CXCR4 sensitizes GBM cells to radiation, eliciting enhanced release of ICD ligands. Combining AMD3100 SPNPs with radiotherapy inhibited GBM progression and led to long-term survival; with 60% of mice remaining tumor-free. This was accompanied by an anti-GBM immune response and sustained immunological memory that prevented tumor recurrence without further treatment. Finally, we showed that systemic delivery of AMD3100 SPNPs decreased the infiltration of CXCR4+ monocytic myeloid-derived suppressor cells to the tumor microenvironment. With the potent ICD induction and reprogrammed immune microenvironment, this strategy has significant potential for future clinical translation. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/457953v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@17c9377org.highwire.dtl.DTLVardef@1b5a4dcorg.highwire.dtl.DTLVardef@5aa4eaorg.highwire.dtl.DTLVardef@1011a86_HPS_FORMAT_FIGEXP M_FIG Immunological mechanism targeting Glioblastoma (GBM) upon blocking CXCR4 signaling pathway with AMD3100-conjugated nanoparticles (SPNPs). (1) Radiotherapy induces glioma cell death, followed by Damage-associated molecular patterns (DAMPs) release. Dendritic cells (DC) are activated by DAMPs and migrate to the regional lymph node where they prime cytotoxic T lymphocyte immune response. Tumor-specific cytotoxic T cells infiltrate the tumor and attack glioma cells. (2) Glioma cells express CXCR4, as well its ligand CXCL12. CXCL12 induces glioma cell proliferation and, (3) as well as mobilization in the bone marrow of CXCR4 expressing myeloid MDSC, which will infiltrate the tumor, and inhibit tumor-specific cytotoxic T cells activity. GEMM of glioma when treated systemically with SPNPs AMD3100 SPNPs plus radiation, nanoparticles block the interaction between CXCR4 and CXCL12, thus (4) inhibiting glioma cell proliferation and (5) reducing mobilization in the bone marrow of CXCR4 expressing myeloid MDSC, (6) generating a reduced MDSC tumor infiltration, as well as releasing MDSC inhibition over tumor specific cytotoxic T cell response. C_FIG

cell biology↗

bioRxiv2020

Drainage of inflammatory macromolecules from brain to periphery targets the liver for macrophage infiltration

Many brain pathologies are associated with liver damage, but a direct link has long remained elusive. Here, we establish a new paradigm for interrogating brain-periphery interactions by leveraging zebrafish for its unparalleled access to the intact whole animal for in vivo analysis in real time after triggering focal brain inflammation. We reveal that drainage of inflammatory macromolecules from the brain led to a strikingly robust peripheral infiltration of macrophages into the liver independent of Kupffer cells. We further demonstrate that this macrophage recruitment requires signaling from the cytokine IL-34, Toll-like receptor adaptor protein MyD88, and neutrophils. These results highlight the ability for circulation of brain-derived substances to serve as a rapid mode of communication from brain to the liver. Understanding how the brain engages the periphery at times of danger may offer new perspectives for detecting and treating brain pathologies.

immunology↗