Surface functionalization of exosomes for chondrocyte-targeted siRNA delivery and cartilage regeneration.

Osteoarthritis (OA) is the most prevalent degenerative cartilage disease, but no effective treatment is currently available to ameliorate the dysregulation of cartilage catabolism. Cartilage degeneration is closely related to the change in the physiology of chondrocytes: for example, chondrocytes of the OA patients overexpress matrix metallopeptidase 13 (MMP13), a.k.a. collagenase 3, which damages the extracellular matrix (ECM) of the cartilage and deteriorate the disease progression. Inhibiting MMP13 has shown to be beneficial for OA treatments, but delivering therapeutics to the chondrocytes embedded in the dense cartilage is a challenge. Here, we engineered the exosome surface with the cartilage affinity peptide (CAP) through lipid insertion to give chondrocyte-targeting exosomes, CAP-Exo, which was then loaded with siRNA against MMP13 (siMMP13) in the interior to give CAP-Exo/siMMP13. Intra-articular administration of CAP-Exo/siMMP13 reduced the MMP13 level and increased collagen COL2A1 and proteoglycan in cartilage in a rat model of anterior cruciate ligament transection (ACLT)-induced OA. Proteomic analysis showed that CAP-Exo/siMMP13 treatment restored the altered protein levels in the IL-1ß-treated chondrocytes. Taken together, a facile exosome engineering method enabled targeted delivery of siRNA to chondrocytes and chondrocyte-specific silencing of MMP13 to attenuate cartilage degeneration.

Chondrocyte-Targeted Delivery System of Sortase A-Engineered Extracellular Vesicles Silencing MMP13 for Osteoarthritis Therapy.

Targeted drug delivery and the reduction of off-target effects are crucial for the promising clinical application of nucleic acid drugs. To address this challenge, a new approach for treating osteoarthritis (OA) that accurately delivers antisense oligonucleotides (ASO) targeting matrix metalloproteinase-13 (ASO-MMP13) to chondrocytes, is developed. Small extracellular vesicles (exos) are ligated with chondrocyte affinity peptide (CAP) using Sortase A and subsequently incubated with cholesterol-modified ASO-MMP13 to construct a chondrocyte-targeted drug delivery exo (CAP-exoASO). Compared with exos without CAP (ExoASO), CAP-exoASOs attenuate IL-1ß-induced chondrocyte damage and prolong the retention time of ASO-MMP13 in the joint without distribution in major organs following intra-articular injection. Notably, CAP-exoASOs decrease MMP13 expression (P < 0.001) and upregulate COL2A1 expression (P = 0.006), resulting in reorganization of the cartilage matrix and alleviation of progression in the OA model. Furthermore, the Osteoarthritis Research Society International (OARSI) score of articular cartilage tissues treated with CAP-exoASO is comparable with that of healthy rats (P = 0.148). A mechanistic study demonstrates that CAP-exoASO may reduce inflammation by suppressing the IL-17 and TNF signaling pathways. Based on the targeted delivery effect, CAP-exoASOs successfully accomplish cartilage repair and have considerable potential for development as a promising therapeutic modality for satisfactory OA therapy.

Volatile communication in plants relies on a KAI2-mediated signaling pathway.

Plants are constantly exposed to volatile organic compounds (VOCs) that are released during plant-plant communication, within-plant self-signaling, and plant-microbe interactions. Therefore, understanding VOC perception and downstream signaling is vital for unraveling the mechanisms behind information exchange in plants, which remain largely unexplored. Using the hormone-like function of volatile terpenoids in reproductive organ development as a system with a visual marker for communication, we demonstrate that a petunia karrikin-insensitive receptor, PhKAI2ia, stereospecifically perceives the (-)-germacrene D signal, triggering a KAI2-mediated signaling cascade and affecting plant fitness. This study uncovers the role(s) of the intermediate clade of KAI2 receptors, illuminates the involvement of a KAI2ia-dependent signaling pathway in volatile communication, and provides new insights into plant olfaction and the long-standing question about the nature of potential endogenous KAI2 ligand(s).

Monitoring drug metabolic pathways through extracellular vesicles in mouse plasma.

The ability to monitor the response of metabolic enzymes to drug exposure in individuals is highly appealing and critical to personalized medicine. Although pharmacogenomics assesses genotypic differences, it does not report changes in metabolic enzyme activities due to environmental factors such as drug interactions. Here, we report a quantitative proteomics strategy to monitor drug metabolic pathways by profiling metabolic enzymes in circulating extracellular vesicles (EVs) upon drug exposure. Mass spectrometry (MS)-based measurement revealed that changes in metabolic enzyme abundance in EVs paralleled those in hepatic cells isolated from liver tissue. Coupling with multiplexed isotopic labeling, we temporally quantified 34 proteins involved in drug absorption, distribution, metabolism, and excretion (ADME) pathways. Out of 44 known ADME proteins in plasma EVs, previously annotated mouse cytochrome P450 3A11 (Cyp3a11), homolog to human CYP3A4, and uridine 5'-diphospho (UDP) glucuronosyltransferase 2A3 (Ugt2a3), increased upon daily rifampicin dosage. Dasatinib, a tyrosine kinase inhibitor to treat leukemia, also elevated Cyp3a11 levels in plasma EVs, but to a lesser extent. Altogether, this study demonstrates that measuring drug enzymes in circulating EVs as an effective surrogate is highly feasible and may transform today's drug discovery and development for personalized medicine.

Targeted Phosphoproteomics of Human Saliva Extracellular Vesicles via Multiple Reaction Monitoring Cubed (MRM3).

Oral squamous cell carcinoma (OSCC) has become a global health problem due to its increasing incidence and high mortality rate. Early intervention through monitoring of the diagnostic biomarker levels during OSCC treatment is critical. Extracellular vesicles (EVs) are emerging surrogates in intercellular communication through transporting biomolecule cargo and have recently been identified as a potential source of biomarkers such as phosphoproteins for many diseases. Here, we developed a multiple reaction monitoring cubed (MRM3) method coupled with a novel sample preparation strategy, extracellular vesicles to phosphoproteins (EVTOP), to quantify phosphoproteins using a minimal amount of saliva (50 µL) samples from OSCC patients with high specificity and sensitivity. Our results established differential patterns in the phosphopeptide content of healthy, presurgery, and postsurgery OSCC patient groups. Notably, we discovered significantly increased salivary phosphorylated alpha-amylase (AMY) in the postsurgery group compared to the presurgery group. We hereby present the first targeted MS method with extremely high sensitivity for measuring endogenous phosphoproteins in human saliva EVs.

Regenerative Signatures in Bronchioalveolar Lavage of Acute Respiratory Distress Syndrome.

Background: In patients with severe acute respiratory distress syndrome (ARDS) associated with sepsis, lung recovery is considerably delayed, and mortality is much high. More insight into the process of lung regeneration in ARDS patients is needed. Exosomes are important cargos for intercellular communication by serving as autocrine and/or paracrine. Cutting-edge exomics (exosomal proteomics) makes it possible to study the mechanisms of re-alveolarization in ARDS lungs. Aims: This study aimed to identify potential regenerative niches by characterizing differentially expressed proteins in the exosomes of bronchioalveolar lavage (BAL) in ARDS patients. Methods: We purified exosomes from BAL samples collected from ARDS patients by NIH-supported ALTA and SPIROMICS trials. The abundance of exosomal proteins/peptides was quantified using liquid chromatography-mass spectrometry (LC-MS). Differentially expressed exosomal proteins between healthy controls and ARDS patients were profiled for functional annotations, cell origins, signaling pathways, networks, and clinical correlations. Results: Our results show that more exosomal proteins were identified in the lungs of late-stage ARDS patients. Immune cells and lung epithelial stem cells were major contributors to BAL exosomes in addition to those from other organs. We enriched a wide range of functions, stem cell signals, growth factors, and immune niches in both mild and severe patients. The differentially expressed proteins that we identified were associated with key clinical variables. The severity-associated differences in protein-protein interaction, RNA crosstalk, and epigenetic network were observed between mild and severe groups. Moreover, alveolar type 2 epithelial cells could serve as both exosome donors and recipients via autocrine and paracrine mechanisms. Conclusions: This study identifies novel exosomal proteins associated with diverse functions, signaling pathways, and cell origins in ARDS lavage samples. These differentiated proteins may serve as regenerative niches for re-alveolarization in injured lungs.

Hands-Free Proteomic Profiling of Urinary Extracellular Vesicles with a High-Throughput Automated Workflow.

Extracellular vesicles (EVs) have emerged as a promising source of disease biomarkers for noninvasive early stage diagnoses, but a bottleneck in EV sample processing restricts their immense potential in clinical applications. Existing methods are limited by a low EV yield and integrity, slow processing speeds, low sample capacity, and poor recovery efficiency. We aimed to address these issues with a high-throughput automated workflow for EV isolation, EV lysis, protein extraction, and protein denaturation. The automation can process clinical urine samples in parallel, resulting in protein-covered beads ready for various analytical methods, including immunoassays, protein quantitation assays, and mass spectrometry. Compared to the standard manual lysis method for contamination levels, efficiency, and consistency of EV isolation, the automated protocol shows reproducible and robust proteomic quantitation with less than a 10% median coefficient of variation. When we applied the method to clinical samples, we identified a total 3,793 unique proteins and 40,380 unique peptides, with 992 significantly upregulated proteins in kidney cancer patients versus healthy controls. These upregulated proteins were found to be involved in several important kidney cancer metabolic pathways also identified with a manual control. This hands-free workflow represents a practical EV extraction and profiling approach that can benefit both clinical and research applications, streamlining biomarker discovery, tumor monitoring, and early cancer diagnoses.

FAM210A is essential for cold-induced mitochondrial remodeling in brown adipocytes.

Cold stimulation dynamically remodels mitochondria in brown adipose tissue (BAT) to facilitate non-shivering thermogenesis in mammals, but what regulates mitochondrial plasticity is poorly understood. Comparing mitochondrial proteomes in response to cold revealed FAM210A as a cold-inducible mitochondrial inner membrane protein. An adipocyte-specific constitutive knockout of Fam210a (Fam210aAKO) disrupts mitochondrial cristae structure and diminishes the thermogenic activity of BAT, rendering the Fam210aAKO mice vulnerable to lethal hypothermia under acute cold exposure. Induced knockout of Fam210a in adult adipocytes (Fam210aiAKO) does not affect steady-state mitochondrial structure under thermoneutrality, but impairs cold-induced mitochondrial remodeling, leading to progressive loss of cristae and reduction of mitochondrial density. Proteomics reveals an association between FAM210A and OPA1, whose cleavage governs cristae dynamics and mitochondrial remodeling. Mechanistically, FAM210A interacts with mitochondrial protease YME1L and modulates its activity toward OMA1 and OPA1 cleavage. These data establish FAM210A as a key regulator of mitochondrial cristae remodeling in BAT and shed light on the mechanism underlying mitochondrial plasticity in response to cold.

One-Pot Analytical Pipeline for Efficient and Sensitive Proteomic Analysis of Extracellular Vesicles.

Extracellular vesicle (EV) proteomics emerges as an effective tool for discovering potential biomarkers for disease diagnosis, monitoring, and therapeutics. However, the current workflow of mass spectrometry-based EV proteome analysis is not fully compatible in a clinical setting due to inefficient EV isolation methods and a tedious sample preparation process. To streamline and improve the efficiency of EV proteome analysis, here we introduce a one-pot analytical pipeline integrating a robust EV isolation approach, EV total recovery and purification (EVtrap), with in situ protein sample preparation, to detect urinary EV proteome. By incorporating solvent-driven protein capture and fast on-bead digestion, the one-pot pipeline enabled the whole EV proteome analysis to be completed within one day. In comparison with the existing workflow, the one-pot pipeline was able to obtain better peptide yield and identify the equivalent number of unique EV proteins from 1 mL of urine. Finally, we applied the one-pot pipeline to profile proteomes in urinary EVs of bladder cancer patients. A total of 2774 unique proteins were identified in 53 urine samples using a 15 min gradient library-free data-independent acquisition method. Taken altogether, our novel one-pot analytical pipeline demonstrated its potential for routine and robust EV proteomics in biomedical applications.

FAM210A mediates an inter-organelle crosstalk essential for protein synthesis and muscle growth in mouse.

Mitochondria are not only essential for energy production in eukaryocytes but also a key regulator of intracellular signaling. Here, we report an unappreciated role of mitochondria in regulating cytosolic protein translation in skeletal muscle cells (myofibers). We show that the expression of mitochondrial protein FAM210A (Family With Sequence Similarity 210 Member A) is positively associated with muscle mass in mice and humans. Muscle-specific Myl1Cre-driven Fam210a knockout (Fam210aMKO) in mice reduces mitochondrial density and function, leading to progressive muscle atrophy and premature death. Metabolomic and biochemical analyses reveal that Fam210aMKO reverses the oxidative TCA cycle towards the reductive direction, resulting in acetyl-CoA accumulation and hyperacetylation of cytosolic proteins. Specifically, hyperacetylation of several ribosomal proteins leads to disassembly of ribosomes and translational defects. Transplantation of Fam210aMKO mitochondria into wildtype myoblasts is sufficient to elevate protein acetylation in recipient cells. These findings reveal a novel crosstalk between the mitochondrion and ribosome mediated by FAM210A.

Phosphoproteome analysis of cerebrospinal fluid extracellular vesicles in primary central nervous system lymphoma.

Primary central nervous system lymphoma (PCNSL) is a rare but highly aggressive extra-nodal non-Hodgkin's lymphoma, mostly of the diffuse large B-cell lymphoma (DLBCL) type. The present invasive diagnosis and poor prognosis of PCNSL propose an urgent need to develop molecular markers for early detection, real-time monitoring and treatment evaluation. Cerebrospinal fluid (CSF)-derived extracellular vesicles (EVs) are promising biomarker carriers for liquid biopsy of CNS diseases and brain tumors; however, research remains challenging due to the low concentration of EVs in the limited available volume of CSF from each individual patient and the low efficiency of existing methods for EV enrichment. Here, we introduce functionalized magnetic beads called EVTRAP (extracellular vesicles total recovery and purification) for rapid and efficient EV isolation from CSF. By coupling with high-performance mass spectrometry, over 19 000 peptides representing 1841 proteins were identified from just 30 µL of CSF. Furthermore, up to 3000 phosphopeptides representing over 1000 phosphoproteins were identified from about 2 mL of CSF. Finally, we analyzed the EV phosphoproteomics of CSF samples from PCNSL patients and non-PCNSL controls. Among them, multiple phosphoproteins related to PCNSL, including SPP1, MARCKS, NPM1 and VIM, were shown to be up-regulated in the PCNSL group. These results demonstrated the feasibility of the EVTRAP-based analytical strategy in CSF EV phosphoproteomic analysis of PCNSL molecular markers.

The EXO70 inhibitor Endosidin2 alters plasma membrane protein composition in Arabidopsis roots.

To sustain normal growth and allow rapid responses to environmental cues, plants alter the plasma membrane protein composition under different conditions presumably by regulation of delivery, stability, and internalization. Exocytosis is a conserved cellular process that delivers proteins and lipids to the plasma membrane or extracellular space in eukaryotes. The octameric exocyst complex contributes to exocytosis by tethering secretory vesicles to the correct site for membrane fusion; however, whether the exocyst complex acts universally for all secretory vesicle cargo or just for specialized subsets used during polarized growth and trafficking is currently unknown. In addition to its role in exocytosis, the exocyst complex is also known to participate in membrane recycling and autophagy. Using a previously identified small molecule inhibitor of the plant exocyst complex subunit EXO70A1, Endosidin2 (ES2), combined with a plasma membrane enrichment method and quantitative proteomic analysis, we examined the composition of plasma membrane proteins in the root of Arabidopsis seedlings, after inhibition of the ES2-targetted exocyst complex, and verified our findings by live imaging of GFP-tagged plasma membrane proteins in root epidermal cells. The abundance of 145 plasma membrane proteins was significantly reduced following short-term ES2 treatments and these likely represent candidate cargo proteins of exocyst-mediated trafficking. Gene Ontology analysis showed that these proteins play diverse functions in cell growth, cell wall biosynthesis, hormone signaling, stress response, membrane transport, and nutrient uptake. Additionally, we quantified the effect of ES2 on the spatial distribution of EXO70A1 with live-cell imaging. Our results indicate that the plant exocyst complex mediates constitutive dynamic transport of subsets of plasma membrane proteins during normal root growth.

FAT-switch-based quantitative S-nitrosoproteomics reveals a key role of GSNOR1 in regulating ER functions.

Reversible protein S-nitrosylation regulates a wide range of biological functions and physiological activities in plants. However, it is challenging to quantitively determine the S-nitrosylation targets and dynamics in vivo. In this study, we develop a highly sensitive and efficient fluorous affinity tag-switch (FAT-switch) chemical proteomics approach for S-nitrosylation peptide enrichment and detection. We quantitatively compare the global S-nitrosylation profiles in wild-type Arabidopsis and gsnor1/hot5/par2 mutant using this approach, and identify 2,121 S-nitrosylation peptides in 1,595 protein groups, including many previously unrevealed S-nitrosylated proteins. These are 408 S-nitrosylated sites in 360 protein groups showing an accumulation in hot5-4 mutant when compared to wild type. Biochemical and genetic validation reveal that S-nitrosylation at Cys337 in ER OXIDOREDUCTASE 1 (ERO1) causes the rearrangement of disulfide, resulting in enhanced ERO1 activity. This study offers a powerful and applicable tool for S-nitrosylation research, which provides valuable resources for studies on S-nitrosylation-regulated ER functions in plants.

Proteomic Discovery and Array-Based Validation of Biomarkers from Urinary Exosome by Supramolecular Probe.

Exosomes are nanoscale, membrane-enclosed vesicles with contents similar to their parent cells, which are rich in potential biomarkers. Urine, as a noninvasive sampling body fluid, has the advantages of being simple to collect, stable in protein, diverse and not regulated by homeostatic mechanisms of the body, making it a favorable target for studying tumor biomarkers. In this report, the urinary exosomal proteome was analyzed and high-throughput downstream validation was performed using a supramolecular probe-based capture and in situ detection. The technology demonstrated the efficient enrichment of exosomes with a high concentration (5.5 × 1010 particles/mL) and a high purity (2.607 × 1010 particles/mg) of exosomes from urine samples. Proteomic analysis of urine samples from patients with hepatocellular carcinoma and healthy individuals combined with proteomic screening techniques revealed that 68 proteins were up-regulated in patients with hepatocellular carcinoma. As a proof-of-principle study, three of these differentially expressed proteins, including OLFM4, HDGF and GDF15, were validated using the supramolecular probe-based array (48 samples per batch). These findings demonstrate the great potential of this approach toward a liquid biopsy for the discovery and validation of biomarkers from urinary exosomes, and it can be extended to various biological samples with lower content of exosomes.

Quantitative proteomics and phosphoproteomics of urinary extracellular vesicles define putative diagnostic biosignatures for Parkinson's disease.

BACKGROUND: Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene have been recognized as genetic risk factors for Parkinson's disease (PD). However, compared to cancer, fewer genetic mutations contribute to the cause of PD, propelling the search for protein biomarkers for early detection of the disease. METHODS: Utilizing 138 urine samples from four groups, healthy individuals (control), healthy individuals with G2019S mutation in the LRRK2 gene (non-manifesting carrier/NMC), PD individuals without G2019S mutation (idiopathic PD/iPD), and PD individuals with G2019S mutation (LRRK2 PD), we applied a proteomics strategy to determine potential diagnostic biomarkers for PD from urinary extracellular vesicles (EVs). RESULTS: After efficient isolation of urinary EVs through chemical affinity followed by mass spectrometric analyses of EV peptides and enriched phosphopeptides, we identify and quantify 4476 unique proteins and 2680 unique phosphoproteins. We detect multiple proteins and phosphoproteins elevated in PD EVs that are known to be involved in important PD pathways, in particular the autophagy pathway, as well as neuronal cell death, neuroinflammation, and formation of amyloid fibrils. We establish a panel of proteins and phosphoproteins as novel candidates for disease biomarkers and substantiate the biomarkers using machine learning, ROC, clinical correlation, and in-depth network analysis. Several putative disease biomarkers are further partially validated in patients with PD using parallel reaction monitoring (PRM) and immunoassay for targeted quantitation. CONCLUSIONS: These findings demonstrate a general strategy of utilizing biofluid EV proteome/phosphoproteome as an outstanding and non-invasive source for a wide range of disease exploration.

Parkinson's disease (PD) is a progressive neurological disorder that affects body movement because some brain cells stop producing the chemical dopamine. PD is often not diagnosed until it has advanced, making early detection crucial. To enable early detection, we investigated tiny packages called extracellular vesicles released from a variety of cells, including the brain cells, that can be found in urine as a potential source for diagnosing PD. These tiny packages contain different kinds of molecules from inside the cells. We analyzed urine samples from 138 individuals and found several proteins involved in PD development that could be biological indicators for early detection of the disease. We used various techniques to make sure that our findings were accurate. Our study suggests that looking at these proteins in urine could be a good way to detect PD in a non-invasive manner.

Mass spectrometry-based phosphoproteomics in clinical applications.

Protein phosphorylation is an essential post-translational modification that regulates many aspects of cellular physiology, and dysregulation of pivotal phosphorylation events is often responsible for disease onset and progression. Clinical analysis on disease-relevant phosphoproteins, while quite challenging, provides unique information for precision medicine and targeted therapy. Among various approaches, mass spectrometry (MS)-centered characterization features discovery-driven, high-throughput and in-depth identification of phosphorylation events. This review highlights advances in sample preparation and instrument in MS-based phosphoproteomics and recent clinical applications. We emphasize the preeminent data-independent acquisition method in MS as one of the most promising future directions and biofluid-derived extracellular vesicles as an intriguing source of the phosphoproteome for liquid biopsy.

Profiling Phosphoproteome Landscape in Circulating Extracellular Vesicles from Microliters of Biofluids through Functionally Tunable Paramagnetic Separation.

Many biological processes are regulated through dynamic protein phosphorylation. Monitoring disease-relevant phosphorylation events in circulating biofluids is highly appealing but also technically challenging. We introduce here a functionally tunable material and a strategy, extracellular vesicles to phosphoproteins (EVTOP), which achieves one-pot extracellular vesicles (EVs) isolation, extraction, and digestion of EV proteins, and enrichment of phosphopeptides, with only a trace amount of starting biofluids. EVs are efficiently isolated by magnetic beads functionalized with TiIV ions and a membrane-penetrating peptide, octa-arginine R8 + , which also provides the hydrophilic surface to retain EV proteins during lysis. Subsequent on-bead digestion concurrently converts EVTOP to TiIV ion-only surface for efficient enrichment of phosphopeptides for phosphoproteomic analyses. The streamlined, ultra-sensitive platform enabled us to quantify 500 unique EV phosphopeptides with only a few µL of plasma and over 1200 phosphopeptides with 100 µL of cerebrospinal fluid (CSF). We explored its clinical application of monitoring the outcome of chemotherapy of primary central nervous system lymphoma (PCNSL) patients with a small volume of CSF, presenting a powerful tool for broad clinical applications.

Isolation and Identification of Plasma Extracellular Vesicles Protein Biomarkers.

Extracellular vesicles (EVs) have emerged as a valuable source for disease biomarkers and an alternative drug delivery system due to their ability to carry cargo and target specific cells. Proper isolation, identification, and analytical strategy are required for evaluating their potential in diagnostics and therapeutics. Here, a method is detailed to isolate plasma EVs and analyze their proteomic profiling, combining EVtrap-based high-recovery EV isolation, phase-transfer surfactant method for protein extraction, and mass spectrometry qualitative and quantitative strategies for EV proteome characterization. The pipeline provides a highly effective EV-based proteome analysis technique that can be applied for EV characterization and evaluation of EV-based diagnosis and therapy.

Meeting Report on the 2nd Chinese American Society for Mass Spectrometry Conference: Advancing Biological and Pharmaceutical Mass Spectrometry.

The 2nd CASMS conference was held virtually through Gather. Town platform from October 17 to 21, 2022, with a total of 363 registrants including an outstanding and diverse group of scientists at the forefront of their research fields from both academia and industry worldwide, especially in the United States and China. The conference offered a 5-day agenda with an exciting scientific program consisting of two plenary lectures, 14 parallel symposia, and 4 special sessions in which a total of 97 invited speakers presented technological innovations and their applications in proteomics & biological mass spectrometry and metabo-lipidomics & pharmaceutical mass spectrometry. In addition, 18 invited speakers/panelists presented at 3 research-focused and 2 career development workshops. Moreover, 144 posters, 54 lightning talks, 5 sponsored workshops, and 14 exhibitions were presented, from which 20 posters and 8 lightning talks received presentation awards. Furthermore, the conference featured 1 MCP lectureship and 5 young investigator awardees for the first time to highlight outstanding mid-career and early-career rising stars in mass spectrometry from our society. The conference provided a unique scientific platform for young scientists (i.e., graduate students, postdocs and junior faculty/investigators) to present their research, meet with prominent scientists, and learn about career development and job opportunities (http://casms.org).

Epitope Imprinting of Phospholipids by Oriented Assembly at the Oil/Water Interface for the Selective Recognition of Plasma Membranes.

Phospholipids, as fundamental building blocks of the cell membrane, play important roles for molecule transportation, cell recognition, etc. However, due to the structural diversity and amphipathic nature, there are few methods for the specific recognition of lipids as compared to other biomolecules such as proteins and glycans. Herein, we developed a molecular imprinting strategy for controllable imprinting toward the polar head of phospholipid exposed on the surface of cellular membranes for recognition. Phosphatidylserine, as unique lipid on the outer membrane leaflet of exosome and also hallmark for cell apoptosis, was imprinted with the developed method. The phosphatidylserine imprinted materials showed high efficiency and specific targeting capability not only for apoptotic cell imaging but also for the isolation of exosomes. Collectively, the synthesized molecularly imprinted materials have great potential for selective plasma membrane recognition for targeted drug delivery and biomarker discovery.