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Day, Le Z.

Publications and source records attributed to Day, Le Z..

10 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2026

Astrocyte Reactivity by Alcohol Dependence in the Central Amygdala

Astrocytes play essential roles in the brain, but prolonged insult can result in the loss or diminishment of homeostatic functions and increased neuroinflammatory response. The central amygdala (CeA) is an important limbic region in reward and stress responsivity as well as in the development of alcohol dependence. Using a multi-omics approach with Aldh1l1-EGFP/Rpl10a mice and the chronic intermittent ethanol – two-bottle choice exposure model, we have characterized the translational response of CeA astrocytes, as well as the proteomic and phosphoproteomic changes in dependent, non-dependent, and naïve mice. We identified astrocyte-specific alterations in neuroimmune functions and antioxidant/oxidative stress pathways in dependent animals while non-dependent EtOH drinking altered cytoskeletal plasticity related pathways. Proteomic analysis showed down-regulation of astrocyte physiological functions in dependent animals while phosphoproteomic analysis identified cytoskeleton remodeling in both dependent and non-dependent animals suggesting the development of a reactive astrocyte phenotype. Astrocyte morphological reconstruction showed increased CeA astrocyte complexity in dependent and non-dependent groups compared to naïve mice. The astrocyte-specific increase in neuroimmune functions, down-regulation of astrocyte homeostatic functions, alteration in protein phosphorylation-mediated cytoskeleton remodeling, and increased astrocyte complexity demonstrate EtOH induced astrocyte activation in the CeA and suggest the presence of both adaptive and maladaptive reactive astrocytes. These findings highlight the roles CeA astrocytes play in the progression of alcohol intake to dependence and are the first step in the identification of novel astrocyte-specific therapeutic approaches for alcohol use disorder through the potentiation of adaptive changes and inhibition of maladaptive changes in astrocytes.

Hashimoto, Joel↗
DOE OSTI2026

Longitudinal Plasma Proteomic Profiling Reveals Divergent Immune Responses in Durably Cured and Relapsed Pulmonary Tuberculosis

Background: Predicting the risk of tuberculosis (TB) relapse is vital to improving treatment outcomes. Although clinical risk factors of relapse are well characterized, the biological mechanisms driving relapse, particularly host immune responses, remain poorly understood. Elucidating these mechanisms is necessary to better predict relapse risk. Methods: We conducted a longitudinal, global proteomic study on 60 participants with active pulmonary TB, half who were durably cured and half who relapsed. Plasma was collected at seven time-points: at treatment initiation (baseline), during therapy, and 52 weeks post-baseline. Samples were analyzed by high-resolution LC-MS/MS. Results: 2,418 proteins were identified across all samples, with 1,756 being differentially expressed relative to baseline (unadjusted p < 0.05). 956 proteins were differentially abundant between cured and relapsed participants. Relapsed participants showed heightened humoral immunity throughout treatment, as well as upregulated complement activation and HDL particles. Cured participants exhibited elevated recovery-related pathways by week 4, including downregulated epithelial invasion and upregulated oxygen transport. Conclusions: Heightened humoral and innate immune responses were associated with relapse, whereas recovery signatures were associated with durable cure. These findings advance our understanding of host responses to treatment and provide a basis for developing blood-based biomarkers to identify patients at increased risk of relapse.

LC-MS/MS↗
DOE OSTI2024

Lipid droplet-associated proteins in alcohol-associated fatty liver disease: A proteomic approach

The earliest manifestation of alcohol-associated liver disease (ALD) is steatosis characterized by deposition of fat in specialized organelles called lipid droplets (LDs). While alcohol administration causes a rise in LD numbers in the hepatocytes, little is known regarding their characteristics that allow their accumulation and size to increase. The aim of the present study is to gain insights into underlying pathophysiological mechanisms by investigating the ethanol-induced changes in hepatic LD proteome as a function of LD size. Adult male Wistar rats (180–200 g BW) were fed with ethanol liquid diet for 6 weeks. At sacrifice, large-, medium-, and small-sized hepatic LD subpopulations (LD1, LD2, and LD3, respectively) were isolated and subjected to morphological and proteomic analyses. Morphological analysis of LD1-LD3 fractions of ethanol-fed rats clearly demonstrated that LD1 contained larger LDs compared with LD2 and LD3 fractions. Our preliminary results from principal component analysis showed that the proteome of different-sized hepatic LD fractions was distinctly different. Proteomic data analysis identified over 2000 proteins in each LD fraction with significant alterations in protein abundance among the three LD fractions. Among the altered proteins, several were related to fat metabolism, including synthesis, incorporation of fatty acid, and lipolysis. Ingenuity pathway analysis revealed increased fatty acid synthesis, fatty acid incorporation, LD fusion, and reduced lipolysis in LD1 compared to LD3. Overall, the proteomic findings indicate that the increased level of protein that facilitates fusion of LDs combined with an increased association of negative regulators of lipolysis dictates the generation of large-sized LDs during the development of alcohol-associated hepatic steatosis. Several significantly altered proteins were identified in different-sized LDs isolated from livers of ethanol-fed rats. Ethanol-induced increases in specific proteins that hinder LD lipid metabolism led to the accumulation and persistence of large-sized LDs in the liver.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

Proteomics identifies complement protein signatures in patients with alcohol-associated hepatitis

Diagnostic challenges continue to impede development of effective therapies for successful management of alcohol-associated hepatitis (AH), creating an unmet need to identify noninvasive biomarkers for AH. In murine models, complement contributes to ethanol-induced liver injury. Therefore, we hypothesized that complement proteins could be rational diagnostic/prognostic biomarkers in AH. Here, we performed a comparative analysis of data derived from human hepatic and serum proteome to identify and characterize complement protein signatures in severe AH (sAH). The quantity of multiple complement proteins was perturbed in liver and serum proteome of patients with sAH. Multiple complement proteins differentiated patients with sAH from those with alcohol cirrhosis (AC) or alcohol use disorder (AUD) and healthy controls (HCs). Serum collectin 11 and C1q binding protein were strongly associated with sAH and exhibited good discriminatory performance among patients with sAH, AC, or AUD and HCs. Furthermore, complement component receptor 1-like protein was negatively associated with pro-inflammatory cytokines. Additionally, lower serum MBL associated serine protease 1 and coagulation factor II independently predicted 90-day mortality. In summary, meta-analysis of proteomic profiles from liver and circulation revealed complement protein signatures of sAH, highlighting a complex perturbation of complement and identifying potential diagnostic and prognostic biomarkers for patients with sAH.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

Identification of integrated proteomics and transcriptomics signature of alcohol-associated liver disease using machine learning

Distinguishing between alcohol-associated hepatitis (AH) and alcohol-associated cirrhosis (AC) remains a diagnostic challenge. In this study, we used machine learning with transcriptomics and proteomics data from liver tissue and peripheral mononuclear blood cells (PBMCs) to classify patients with alcohol-associated liver disease. The conditions in the study were AH, AC, and healthy controls. We processed 98 PBMC RNAseq samples, 55 PBMC proteomic samples, 48 liver RNAseq samples, and 53 liver proteomic samples. First, we built separate classification and feature selection pipelines for transcriptomics and proteomics data. The liver tissue models were validated in independent liver tissue datasets. Next, we built integrated gene and protein expression models that allowed us to identify combined gene-protein biomarker panels. For liver tissue, we attained 90% nested-cross validation accuracy in our dataset and 82% accuracy in the independent validation dataset using transcriptomic data. We attained 100% nested-cross validation accuracy in our dataset and 61% accuracy in the independent validation dataset using proteomic data. For PBMCs, we attained 83% and 89% accuracy with transcriptomic and proteomic data, respectively. The integration of the two data types resulted in improved classification accuracy for PBMCs, but not liver tissue. We also identified the following gene-protein matches within the gene-protein biomarker panels: CLEC4M-CLC4M, GSTA1-GSTA2 for liver tissue and SELENBP1-SBP1 for PBMCs. In this study, machine learning models had high classification accuracy for both transcriptomics and proteomics data, across liver tissue and PBMCs. The integration of transcriptomics and proteomics into a multi-omics model yielded improvement in classification accuracy for the PBMC data. The set of integrated gene-protein biomarkers for PBMCs show promise toward developing a liquid biopsy for alcohol-associated liver disease.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

Macrophage-derived MLKL in alcohol-associated liver disease: Regulation of phagocytosis

Mixed lineage kinase domain-like pseudokinase (MLKL), a key terminal effector of necroptosis, also plays a role in intracellular vesicle trafficking that is critical for regulating liver inflammation and injury in alcohol-associated liver disease (ALD). Although receptor interacting protein kinase 3 (Rip3) -/- mice are completely protected from ethanol-induced liver injury, Mlkl -/- mice are only partially protected. Therefore, we hypothesized that cell-specific functions of MLKL may contribute to ethanol-induced injury. Bone marrow transplants between Mlkl -/- mice and littermates were conducted to distinguish the role of myeloid versus nonmyeloid Mlkl in the Gao-binge model of ALD. Ethanol-induced hepatic injury, steatosis, and inflammation were exacerbated in Mlkl -/- →wild-type (WT) mice, whereas Mlkl deficiency in nonmyeloid cells (WT→Mlkl -/- ) had no effect on Gao-binge ethanol-induced injury. Importantly, Mlkl deficiency in myeloid cells exacerbated ethanol-mediated bacterial burden and accumulation of immune cells in livers. Mechanistically, challenging macrophages with lipopolysaccharide (LPS) induced signal transducer and activator of transcription 1–mediated expression and phosphorylation of MLKL, as well as translocation and oligomerization of MLKL to intracellular compartments, including phagosomes and lysosomes but not plasma membrane. Importantly, pharmacological or genetic inhibition of MLKL suppressed the phagocytic capability of primary mouse Kupffer cells (KCs) at baseline and in response to LPS with/without ethanol as well as peripheral monocytes isolated from both healthy controls and patients with alcohol-associated hepatitis. Further, in vivo studies revealed that KCs of Mlkl -/- mice phagocytosed fewer bioparticles than KCs of WT mice. Together, these data indicate that myeloid MLKL restricts ethanol-induced liver inflammation and injury by regulating hepatic immune cell homeostasis and macrophage phagocytosis.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

The Mycobacterium tuberculosis protein O -phosphorylation landscape

Bacterial phosphosignaling has long been synonymous with the histidine kinases of the two component systems (TCSs), but many bacteria, including Mycobacterium tuberculosis (Mtb), also code for Ser/Thr protein kinases (STPKs). STPKs are the main phosphosignaling enzymes in eukaryotes, but the full extent of phosphorylation on protein Ser/Thr and Tyr (O-phosphorylation) in bacteria is unclear. Here, we explored the global signaling capacity of the STPKs in Mtb. We generated a panel of STPK loss- and gain-of-function strains and measured the resulting O-phosphorylation and transcriptional changes. This deep phosphoproteome shows that O-phosphorylation in Mtb is a vastly underexplored protein modification that affects >70% of the proteome. The substrate-kinase interactions of STPKs show an extensive interface with the transcriptional machinery, resulting in STPK regulation of gene expression of over 30% of Mtb genes. Finally, Mtb O-phosphorylation gives rise to an expansive, distributed, and cooperative network of a complexity that has previously only been associated with eukaryotic phosphosignaling networks.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Proteomic Profiling of Intra-Islet Features Reveals Substructure-Specific Protein Signatures

Despite their diminutive size, islets of Langerhans play a large role in maintaining systemic energy balance in the body. New technologies have enabled us to go from studying the whole pancreas, to isolated whole islets, to partial islet sections, and now to islet substructures isolated from within the islet. Using a microfluidic nanodroplet-based proteomics platform coupled with laser capture microdissection (LCM), we present an in-depth investigation of protein profiles specific to regions within the islet. These regions studied include the vascular tissue containing interface boundary, micro-vascular tissue internal to the islet, isolated endocrine cells, islet sections with vasculature intact, and finally acinar tissue from around the islet. Unique protein signatures observed in the inner vasculature potentially indicate increased innervation and intra-islet neuron-like crosstalk compared to external vasculature. We also demonstrate the utility of these data for isolating localized structure-specific drug-target interactions using existing protein/drug binding databases.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Integrated Transcriptomic and Proteomic Analysis Identifies Plasma Biomarkers of Hepatocellular Failure in Alcohol-Associated Hepatitis

Alcohol-associated hepatitis (AH) is a form of liver failure with high short-term mortality. Recent results have shown that HNF4a defective function and systemic inflammation are major disease drivers of AH. Plasma biomarkers of hepatocyte function could be useful for diagnostic and prognostic purposes. Herein an integrative analysis of hepatic RNAseq and liquid chromatography-tandem mass spectrometry (LC-MS/MS) was performed to identify plasma protein signatures for mild and severe AH patients. Alcohol-related liver disease cirrhosis (ALD)(AC), non-alcoholic fatty liver disease (NALFD), and healthy subjects (HC) were used as comparator groups. Identified proteins primarily involved in hepatocellular function were decreased in AH patients which included hepatokines, clotting factors, complement cascade components, and hepatocyte growth activators. A protein signature of AH disease severity was identified including thrombin (THRB), hepatocyte growth factor alpha (HGFA), clusterin (CLUS), human serum factor H-related protein (FHR1) and kallistatin (KAIN), which exhibited large abundance shifts between severe and non-severe AH. The combination of THRB and HGFA discriminated between severe and non-severe AH with high sensitivity and specificity. These findings were correlated with the liver expression of genes encoding secreted proteins in a similar cohort, finding a highly consistent plasma protein signature reflecting HNF4A and HNF1A functions. This unbiased proteomic-transcriptome analysis identified plasma protein signatures and pathways associated with disease severity, reflecting HNF4A/1A activity useful for diagnostic assessment in AH.

60 APPLIED LIFE SCIENCES↗