Project description:The RNA binding protein IGF2BP2/IMP2 alters the cargo of cancer cell-derived extracellular vesicles supporting tumor-associated macrophages
Project description:The N-methyl-D-aspartate (NMDA) receptor is a glutamate-activated cation channel critical to many processes in the brain. Genome-wide association studies (GWAS) suggest that glutamatergic neurotransmission and NMDA receptor-mediated synaptic plasticity is important for body weight homeostasis1. Here, we report the engineering and preclinical development of a first-in-class bimodal molecule that integrates NMDA receptor antagonism with glucagon-like peptide-1 (GLP-1) receptor agonism to effectively reverse obesity, hyperglycemia, and dyslipidemia in rodent models of metabolic disease. We demonstrate that GLP-1-directed delivery of the NMDA receptor antagonist MK-801 affects NMDA receptor-mediated synaptic plasticity in the hypothalamus. Importantly, peptide-targeting of MK-801 specifically to GLP-1 receptor-expressing brain regions circumvent adverse physiological and behavioral effects associated with MK-801 monotherapy. In sum, our approach demonstrates the feasibility of cell specific ionotropic receptor-modulation via peptide targeting and highlights the therapeutic potential of unimolecular mixed GLP-1 receptor agonism and NMDA receptor antagonism for obesity treatment.
Project description:Myeloproliferative Neoplasms (MPNs) are a heterogenous group of hematologic cancers characterized by excessive JAK/STAT signaling. Mutations of JAK2 signaling components are among the most common drivers of MPN, but alterations in Suppressors of Cytokine Signaling (SOCS) proteins have been implicated in MPN pathogenesis and progression. Cullin 5 (Cul5) is an E3 ubiquitin ligase known to work with suppressors of cytokine signaling (SOCS) proteins which regulate the JAK/STAT pathway. Here we report that mice lacking Cul5 in hematopoietic stem and progenitor cells (HSPCs) develop an MPN-like disease with characteristic features including splenomegaly, extramedullary hematopoiesis, thrombocytosis, and anemia. Cul5-deficient HSPCs have higher phospho-STAT5 (pSTAT5) levels following stimulation with IL-3 and outcompete WT HSPCs in bone marrow transplants. Immunoprecipitation of Cul5 in cultured HSPCs showed interactions with STAT5 as well as several well-studied substrate receptors including SOCS2, SOCS6, ASB2, ASB3, ASB6 and CIS, as well as lesser-known WSB1 and LRRC41. Proteome analysis of Lin- Sca-1+ c-kit+ (LSK) cells from Cul5Vav-Cre bone marrow shared many upregulated genes and signatures with MPN patient cells. Finally, treatment with ruxolitinib, a JAK1/2 inhibitor, ameliorated MPN symptoms in Cul5-deficient mice. These studies demonstrate a novel function of Cul5 in hematopoiesis, delineating a contributing role in MPN.
Project description:Pancreatic cancer remains as one of the most challenging malignancies to diagnose and treat due to its silent progression and limited early diagnostic options. The intraductal papillary mucinous neoplasm (IPMN) provides an early-stage lesion for studying pancreatic cancer progression. However, proteomic analyses of IPMN and its comparison with pancreatic ductal adenocarcinoma (PDAC) and normal duct (ND) have been limited due to their low cellularity. In this study, we present an optimized spatial tissue proteomics workflow, termed SP-Max (Spatial Proteomics Optimized for Maximum Sensitivity and Reproducibility in Minimal Sample), designed to maximize protein recovery and quantification from limited laser micro dissected (LMD) samples. Our workflow enabled the identification of more than 6,000 proteins and the quantification of over 5,200 protein groups from FFPE tissue contours of actual pancreatic cancer patients. Comparative analyses across ND, IPMN, and PDAC revealed critical molecular differences in protein pathways and potential prognostic markers. SP-Max provides a systematic, reproducible approach that significantly enhances our ability to study precancerous lesions and cancer progression in pancreatic tissues at unprecedented resolution.
Project description:In vitro maturation (IVM) of oocytes retrieved from ovum pick-up (OPU) or ovarian tissue (OT) is a standard approach for patients with specific conditions where prior hormonal stimulation is contraindicated. However, the developmental competence of oocytes matured in vitro is still inferior to that of oocytes matured in vivo. Capacitation-IVM (CAPA-IVM) includes an extra step of pre-maturation culture (PMC) with c-type natriuretic peptide (CNP) as a meiotic arrestor to better synchronize cytoplasmic and nuclear maturity in oocytes. This study aims to evaluate the effect of CAPA-IVM on equine oocyte quality and developmental competence. Immature cumulus-oocyte complexes (COCs) were retrieved from slaughterhouse ovaries and matured in vitro either in CAPA-IVM (short or long) or standard IVM. Matured oocytes from each group were analyzed for calcium-releasing potential and single-oocyte proteomics, and embryo development was assessed after fertilization with Piezo-drilled intracytoplasmic sperm injection (ICSI). Genetic analysis of developed blastocysts was performed to detect chromosomal aberrations. Our findings demonstrate that CAPA-IVM of equine COCs yields significantly higher maturation rates than controls. Moreover, short CAPA-IVM with six hours pre-maturation culture showed substantially higher embryo development potential than the control group. Genetic analysis revealed a high euploidy rate in equine blastocysts regardless of the maturation conditions. Live calcium imaging of the fertilized oocytes demonstrated the majority of oocytes with non-continuous calcium oscillation patterns, irrespective of maturation conditions. Single oocyte proteomics reveals a comparable proteomic landscape between matured oocytes from short CAPA-IVM and standard IVM. However, a trend of differential expression was observed in specific proteins related to cytoskeleton, cell cycle, and hemostasis in the short CAPA-IVM group. Our findings indicate that CAPA-IVM holds the potential to improve oocyte quality and competence in horses. However, further fine-tuning of culture conditions based on omics analysis would benefit the effective use of these IVM systems. Moreover, given that the mare serves as an excellent model for human reproduction, the molecular trends identified in this study could provide valuable insights for advancing human artificial reproductive technologies.
Project description:Background: Aggressiveness guides treatment in IDH-mutant gliomas. Objective grading of oligodendrogliomas is therefore urgently needed. Material and Methods: 211 primary and recurrent resections from 111 oligodendroglioma patients were collected, complemented with 91 samples for validation. Samples were subjected to Ki-67 staining, proteomics and DNA-methylation profiling. Data were analyzed using various multivariate differential models and algorithms. Results & conclusion: We developed CGCψ, a continuous grading coefficient independent of tumor typing, and demonstrate its prognostic value in oligodendrogliomas. Its prognostic value outperformed WHO grade at tumor recurrence. CGCψ is linked to large scale DNA de-methylation, but increased DNA methylation of polycomb transcription factors, aging, Ki-67, and losses on chr4 and chr9p. DNA de-methylation at tumor recurrence/higher grade is sequence context specific and associated with TET recognition sites. Both oligodendrogliomas and astrocytomas progress along this shared epi-genetic axis. Oligosarcomas are characterized by a high CGCψ and low tumor purity.
Project description:This work was designed to profile major proteins of extracellular vesicles (EVs) from mouse tanycytes with target gene knockdown (KD) versus control. The assay was based on using in vitro culture models of mouse hypothalamic tanycytes (predominantly alpha-tanycytes) which were engineered through lentivirus-mediated KD of a target gene, including CRISPR/Cas9-based Rab27a KD or Rictor KD or shRNA-based KD of pre-pro-insulin (ppIns). Scramble gRNA-based control was used as the control group for both Rab27a KD and Rictor KD, and scramble shRNA-based control was used as the control group for ppIns KD. EVs were released from these tanycytes in culture, purified and subjected to proteomics. EV samples of KD groups included Rab27a KD EVs (251015-1, 251015-2, 251015-5), ppIns KD EVs (251015-12, 251015-13, 251015-15), and Rictor KD EVs (251015-3, 251015-6, 251015-14). EV samples of control groups included scramble gRNA control EVs (251015-4, 251015-10, 251015-11) and scramble shRNA control EVs (251015-7, 251015-8, 251015-9). Proteomic profiling suggested that the overall protein composition of tanycyte-derived EVs was not substantially changed by either of these KD approaches.
Project description:Advances in mass-spectrometry (MS)-based technologies have leveraged our understanding of protein-wide adaptations in human skeletal muscle in response to exercise. However, there is a lack of such data in females, particularly pertaining to already trained females and menstrual cycle phase-based sprint interval training (SIT) despite its efficacy and popularity. Here, we present a comprehensive global proteome analysis of skeletal muscle adaptations to high-frequency SIT during different menstrual cycle phases in endurance-trained females. We randomized 49 eumenorrheic females to either high-frequency SIT in the follicular (FB) or luteal phase (LB) over one menstrual cycle comprising eight sessions of 6×30-s all-out efforts. MS-proteomics, covering 4155 proteins after filtering, revealed notable differences in muscle adaptations to phase-based SIT. LB suppressed mitochondrial pathways of the tricarboxylic acid cycle and electron transport chain while enriching ribosomal complexes. Conversely, FB enriched filament organization and skeletal system development. Mitochondrial repression during LB was linked to reduced O2max, whereas exercise capacity improved in FB only. Our findings show that menstrual cycle phase-based high-frequency SIT induces distinct protein-wide muscle adaptations and affects phenotype in endurance-trained eumenorrheic females.
Project description:This part of the data submission of PXD046505. LD3 knock out cells were generated in HEK293T and HMC3 cells. Proteomics was done to test if they have any significant changes in the abundance of proteins that metabolize sugar lipids like gangliosides.