Biology subjects

Wang, K.-C.

Publications and source records attributed to Wang, K.-C..

12 recordsLinked to original sources

bioRxiv2026

Monocyte-Mimetic Nanoparticle Delivery of Verteporfin Promotes Plaque-Stabilizing Remodeling in Atherosclerosis

Background: Despite advances in lipid-lowering and anti-inflammatory therapies, pharmacological strategies that act directly within established atherosclerotic lesions to suppress plaque progression remain limited. We recently developed a monocyte-mimetic nanoparticle (MoNP) platform that selectively targets inflamed endothelium and delivers Verteporfin (VP), termed MoNP-VP, to suppress YAP/TAZ-associated endothelial activation and plaque development. Here, we extend this biomimetic nanotherapy by defining the pharmacological effects of MoNP-VP within atherosclerotic vessels and evaluating its therapeutic efficacy in pre-existing plaques under distinct lipid burden. Methods: MoNP-VP were formulated by encapsulating VP in polymeric cores followed by cloaking with mouse monocyte membranes. Partial ligation was performed in ApoE-deficient mice, and single-cell RNA sequencing was used to define pharmacodynamic responses in carotid lesions. Proprotein convertase subtilisin/kexin type 9 (PCSK9) gain-of-function was induced in wild-type mice to generate pre-existing aortic lesions, followed by continued high-fat-diet or switching to chow to evaluate MoNP-VP efficacy under persistent hyperlipidemia and lipid-lowering conditions. Results: Single-cell analysis revealed that MoNP-VP remodeled the plaque microenvironment in carotid arteries, selectively reducing foamy macrophage populations while enriching vascular stromal populations associated with fibrotic remodeling. In mice with pre-existing aortic plaques, MoNP-VP suppressed lesion progression and reduced macrophage content under persistent hyperlipidemia. Following diet-induced lipid lowering, MoNP-VP further reduced macrophage accumulation despite no significant change in plaque size. Across both settings, MoNP-VP increased fibroblast-like cell populations and collagen deposition accompanied by activation of TGF{beta} signaling, features consistent with a more stable plaque. Importantly, repeated MoNP-VP administration was well tolerated and elicited no overt systemic toxicity. Conclusion: These findings bridge mechanistic insight and therapeutic efficacy in pre-established plaques by demonstrating that MoNP-VP remodels the inflammatory plaque microenvironment and promotes lesion stabilization, supporting its translational potential as a precision nanotherapy for atherosclerosis.

molecular biology↗

bioRxiv2026

Biomimetic MRI Nanoprobe for Mapping Cerebrovascular Inflammation After Traumatic Brain Injury

Cerebrovascular inflammation is a critical driver of secondary injury following traumatic brain injury (TBI), yet no noninvasive tool currently exists to map its spatial heterogeneity across the injured brain. Here, we report MoNP-SPION, a monocyte-mimetic nanoprobe that selectively targets activated cerebrovascular endothelium after TBI and enables quantitative MRI of spatiotemporally heterogeneous cerebrovascular inflammation. MoNP-SPION binding to inflamed brain endothelium scaled with activation state, enabling rapid, spatially heterogeneous targeting of injured cerebral vasculature. MoNP-SPION-enhanced MRI revealed dynamic reductions in T2* throughout the brain that extended beyond the injury penumbra and captured both subacute vascular injury and therapy-induced recovery. Importantly, spatiotemporal reductions in T2* strongly correlated with VCAM1 upregulation, supporting MoNP-SPION as a molecularly specific readout of endothelial activation. Together, these findings establish MoNP-SPION as a promising platform for longitudinal, noninvasive monitoring of cerebrovascular inflammation in TBI and other neurological disorders involving vascular dysfunction. TEASERCell-mimicking MRI nanoprobe maps cerebrovascular injury and recovery dynamics following brain trauma.

bioengineering↗

bioRxiv2026

Monocyte-Mimetic Nanoprobe Enables Longitudinal MRI of Atherosclerotic Inflammatory Dynamics

Noninvasive monitoring of plaque inflammatory dynamics remains an unmet need. We previously developed a monocyte-mimetic nanoprobe, termed MoNP-SPION, for MRI detection of atherosclerotic lesions. Here we demonstrate MoNP-SPION enables longitudinal tracking of plaque inflammatory status in a clinically relevant mouse model. Following 16 weeks of plaque induction, mice were maintained on high-fat diet or switched to chow for 6 weeks to model persistent versus resolving plaque inflammation. MoNP-SPION-enhanced MRI was performed at 3- and 6-weeks post-adjustment, and arterial tissue was collected for histological assessment. Mice maintained on high-fat diet exhibited persistent hypointense T2* signal at the carotid bifurcation and aortic root, whereas chow-transitioned mice showed progressive signal attenuation, consistent with histological evidence of reduced plaque burden and inflammation. These findings establish MoNP-SPION as an effective molecular MRI probe for longitudinal assessment of plaque inflammatory dynamics, supporting its potential for monitoring atherosclerosis progression and therapeutic response.

bioengineering↗

bioRxiv2026

Robust volumetric multiplex staining of centimeter-scale FFPE tissues guided by neural network-based optimization

Archived formalin-fixed, paraffin-embedded (FFPE) tissues, including brain-bank specimens, are valuable resources for disease research, but rapid cyclic immunolabeling of intact specimens remains challenging. Here, we present FIDELITY, an epoxy-free workflow that balances optical transparency, immunostaining performance, and tissue integrity for multi-cycle protein profiling in centimeter-thick FFPE specimens. Neural network-guided optimization identified an SDS-glycine formulation that integrates delipidation and epitope retrieval while preserving tissue stiffness, replacing prolonged epoxy reinforcement and subsequent high-temperature antigen retrieval with a single aqueous pretreatment. Application to human Alzheimer's disease specimens enabled volumetric imaging and quantitative analysis while retaining compatibility with conventional histopathology. Its compatibility with active electrophoretic staining further supported rapid labeling and five rounds of immunolabeling in a whole FFPE mouse brain. Two-round labeling of a human glioma specimen further demonstrated the applicability of the cyclic workflow to human pathological tissue. Together, FIDELITY provides a practical platform for large-volume, multiplexed protein profiling of intact FFPE specimens.

neuroscience↗

bioRxiv2026

Integrin Activation Enhances Lesion-Specific Targeting of Monocyte-Mimetic Nanoparticles in Atherosclerosis

PurposeEndothelial cell (EC) activation, characterized by upregulation of adhesion molecules that drive monocyte recruitment, contributes to plaque progression while also providing an opportunity for targeted therapeutic delivery. Leveraging the cell membrane cloaking strategy, we recently developed a monocyte-mimetic nanoparticle (MoNP) platform that exploits the natural inflammatory tropism of monocytes for site-specific delivery to atherosclerotic vessels. Recognizing that integrin activation is a key determinant of monocyte adhesion to ECs, this study investigates whether pre-activating integrins on MoNP enhances their binding affinity and accumulation at atherosclerotic lesions. MethodsMouse bone marrow-derived monocytes were pretreated with CCL2 or Mn2{square} to activate membrane integrins. Isolated monocyte plasma membranes were cloaked onto fluorescently labeled polymeric cores to generate integrin-activated MoNPs (IA@MoNPs). The targeting capability of IA@MoNPs toward endothelial ligands, inflamed ECs, and atherosclerotic lesions was evaluated using in vitro and in vivo models. ResultsIA@MoNPs exhibited markedly enhanced binding to VCAM1, the primary endothelial ligand mediating integrin-dependent monocyte adhesion, and significantly increased uptake by ECs under atheroprone conditions compared to standard MoNPs. In vivo, IA@MoNPs demonstrated enhanced accumulation in atherosclerotic arteries without increasing nonspecific binding, and blocking {beta}1-integrins on IA@MoNPs abolished this targeting effect. Importantly, integrin activation on IA@MoNPs did not compromise circulatory stability or induce immune or organ toxicity. ConclusionIntegrin activation represents a simple yet effective strategy to enhance MoNP targeting to inflamed ECs and atherosclerotic lesions. This mechanism-driven approach improves targeting performance while maintaining specificity and safety, advancing the translational potential of the biomimetic nanomedicine platform for atherosclerosis.

bioengineering↗

bioRxiv2025

Structural mechanisms for the recruitment of factor H by Streptococcus pyogenes

The major bacterial pathogen Streptococcus pyogenes (Group A Streptococcus, or Strep A) recruits the negative regulator of the alternative complement pathway factor H (FH) to its surface. Antigenically sequence variable regions of several Strep A M proteins, including M5 and M6, bind FH but have no obvious sequence homology. A second Strep A surface-localized protein, FbaA, binds FH through a purported coiled-coil region, suggesting mimicry of the well-known coiled coil of M proteins. We determined the structures of fragments of M5 protein, M6 protein, and FbaA complexed with FH domains 6 and 7 (FH(6-7)). M5 and M6 proteins formed dimeric -helical coiled coils, as expected, while FbaA instead consisted of a monomeric three-helix bundle preceded by a loop. FH(6-7) accommodated different binding modes in these three proteins, with very few common interacting amino acids. Based on contributions to binding, distinct FH-binding sequence patterns were constructed for M5 and M6 proteins, enabling identification of FH-binding sequences in M or M-like Enn proteins in 32 strains of differing M types. While FbaA was allelically sequence variable, its critical FH-binding amino acids were absolutely conserved in 95 strains of differing M types. Together, FH-binding sequences were identified in about half of the known 250 Strep A strains, with the majority due to FbaA. Our structural and functional elucidation of the mechanism of FH recruitment is applicable to precise investigation of its role in Strep A virulence.

biochemistry↗

bioRxiv2025

ChipNMR: Hyperpolarized NMR for non-invasive metabolic flux analysis in perfused microfluidic chips

Dissolution dynamic nuclear polarization NMR Spectroscopy (dDNP-NMR) has become a transformative tool for metabolic studies by significantly enhancing signal sensitivity more than three orders of magnitude compared to traditional NMR. However, NMR detection probes are optimized for round narrow glass tubes typically 5 mm in diameter, which impose constraints on their utility for metabolic studies of adhernt cells. Here, we present a novel NMR probe head integrated with a custom microfluidic chip that facilitates real-time monitoring of hyperpolarized substrate conversion from adhernt cells. This system enables metabolic flux analysis in a controlled, in vitro environment, as demonstrated by tracking the conversion of [1-13C] pyruvate to [1-13C] lactate in HeLa cells over 48 hours. To the best of our knowledge, this is the first demonstration of cell metabolism from an adhering monolayer of mammalian cells in combination with hyperpolarized NMR. The custom microfluidic chip design is modular and adaptable allowing expansion to dual-chamber chips, demonstrating its potential in applications for more complex cellular environments, such as Organ-on-a-Chip systems.

biochemistry↗

bioRxiv2025

Endothelial AGO1 Drives Vascular Inflammation and Atherosclerosis via a Non-Canonical Nuclear Mechanism

BACKGROUNDEndothelial cell (EC) dysfunction is a cause and consequence of vascular inflammation and lipid dysregulation in atherosclerosis, yet the molecular drivers linking EC dysfunction to systemic metabolic derangements remain incompletely understood. Moreover, whether inhibiting an endogenous gene in ECs can impact liver function, lipid profile, and the vascular inflammation in the context of atherosclerosis has not been demonstrated. We previously identified Argonaute 1 (AGO1), a component of the RNA-induced silencing complex, as a regulator of EC function in angiogenesis and obesity. However, the role of endothelial AGO1 in vascular inflammation and liver function in the context of hyperlipidemia and atherosclerosis is unknown. METHODSEC-conditional AGO1 knockout (EC-AGO1-KO) and wildtype mice were subjected to pro-atherosclerotic models induced by AAV9-PCSK9 coupled with a Western diet or partial carotid ligation. Metabolic and vascular phenotype and gene expression were analyzed. In human liver sinusoidal and aortic ECs, AGO1 was knocked down using antisense oligonucleotides (ASO), followed by assessment of inflammatory responses (qPCR, RNA-seq, ELISA, and monocyte adhesion assays). To identify the molecular mechanisms linking AGO1 and EC inflammation, Cut&Tag sequencing, chromatin immunoprecipitation, immunofluorescence, proximal ligation assay, and co-immunoprecipitation were performed. The therapeutic effect of AGO1 inhibition was assessed using ASO-delivered via lipid nanoparticle (LNP) for systemic distribution and monocyte membrane-coated nanoparticles (MoNP) to target the inflamed endothelium. RESULTSEC-AGO1-KO mice exhibited improved plasma lipid profiles, reduced hepatic steatosis, inflammation, and fibrosis, and decreased aortic atherosclerotic burden. AGO1 knockdown in ECs attenuated inflammatory responses. Mechanistically, AGO1 interacted with NF-{kappa}B p65 and promoted p65 nuclear translocation and the transcriptional activation of pro-inflammatory genes, including ICAM1 and THBS1. AGO1-ASO delivered through LNP or MoNP achieved the anti-inflammatory, anti-hyperlipidemic, and anti-atherosclerotic effects, recapitulating the phenotypes observed with EC-AGO1-KO. CONCLUSIONSEndothelial AGO1 promotes vascular inflammation and liver dysfunction in the context of hyperlipidemia and atherosclerosis, in part through a non-canonical nuclear action of AGO1 as an NF-{kappa}B coactivator. Inhibition of endothelial AGO1 provides the dual benefits of ameliorating lipid dysregulation and suppressing vascular inflammation. These results highlight EC-AGO1 as a possible therapeutic target for atherosclerosis and cardiometabolic diseases.

pathology↗

bioRxiv2023

Lesion-specific suppression of YAP/TAZ by biomimetic nanodrug ameliorates atherosclerosis development

Atherosclerosis, characterized by the buildup of lipid-rich plaque on the vessel wall, is the primary cause of myocardial infarction and ischemic stroke. Recent studies have demonstrated that dysregulation of yes-associated protein 1 (YAP) and transcriptional coactivator with PDZ-binding domain (TAZ) contributes to plaque development, making YAP/TAZ potential therapeutic targets. However, systemic modulation of YAP/TAZ expression or activities risks serious off-target effects, limiting clinical applicability. To address the challenge, this study develops monocyte membrane-coated nanoparticles (MoNP) as a drug delivery vehicle targeting activated endothelium lining the plaque surface and utilizes MoNP to deliver verteporfin (VP), a potent YAP/TAZ inhibitor, for lesion-specific treatment of atherosclerosis. The results reveal that MoNP significantly enhance payload delivery to inflamed endothelial cells (EC) while avoiding phagocytic cells, and preferentially accumulate in atherosclerotic regions. MoNP-mediated delivery of VP substantially reduces YAP/TAZ expression, suppressing inflammatory gene expression and macrophage infiltration in cultured EC and mouse arteries exposed to atherogenic stimuli. Importantly, this lesion-targeted VP nanodrug effectively decreases plaque development in mice without causing noticeable histopathological changes in major organs. Collectively, these findings demonstrate a plaque-targeted and pathway-specific biomimetic nanodrug, potentially leading to safer and more effective treatments for atherosclerosis.

bioengineering↗

bioRxiv2022

A versatile method to archive and delipidate brain tissues for ensuing multiplexed immunolabeling and organ-level imaging

Tissue clearing and labeling techniques have revolutionized brain-wide imaging and analysis, yet their application to clinical formalin-fixed paraffin-embedded (FFPE) blocks remains challenging. We introduce HIF-Clear, a novel method for efficiently clearing and labeling centimeter-thick FFPE specimens using elevated temperature and concentrated detergents. HIF-Clear with multi-round immunolabeling reveals neuron circuitry regulating multiple neurotransmitter systems in a whole FFPE mouse brain, and is able to be used as the evaluation of disease treatment efficiency. HIF-Clear also supports expansion microscopy and can be performed on a non-sectioned 15-year-old FFPE specimen, as well as a 3-month formalin-fixed mouse brain. Thus, HIF-Clear represents a feasible approach for researching archived FFPE specimens for future neuroscientific and 3D neuropathological analyses.

bioengineering↗

bioRxiv2022

An M protein coiled coil unfurls and exposes its hydrophobic core to capture LL-37

Surface-associated, coiled-coil M proteins of Streptococcus pyogenes (Strep A) disable human immunity through interaction with select proteins. However, coiled coils lack features typical of protein-protein interaction sites, and it is therefore challenging to understand how M proteins achieve specific binding, for example, with the human antimicrobial peptide LL-37, leading to its neutralization. The crystal structure of a complex of LL-37 with M87 protein, an antigenic M protein variant from a strain that is an emerging threat, revealed a novel interaction mode. The M87 coiled coil unfurled and asymmetrically exposed its hydrophobic core to capture LL-37. A single LL-37 molecule bound M87 in the crystal, but in solution recruited additional LL-37 molecules, consistent with a protein trap neutralization mechanism. The interaction mode visualized crystallographically was verified to contribute significantly to LL-37 resistance in an M87 Strep A strain, and was identified to be conserved in a number of other M protein types that are prevalent in human populations. Our results provide specific detail for therapeutic inhibition of LL-37 neutralization by M proteins.

microbiology↗

bioRxiv2021

Design of M protein immunogens to elicit broadly reactive antibodies against Streptococcus pyogenes

Coiled coil-forming M proteins of the widespread and potentially deadly bacterial pathogen Streptococcus pyogenes (Strep A) are immunodominant targets of opsonizing antibodies. However, antigenic sequence variability into >220 M types, as defined by the M protein hypervariable region (HVR), has been considered to limit its utility as a vaccine immunogen due to type-specificity in the antibody response. Surprisingly, a multi-HVR immunogen in clinical vaccine trials elicited M type cross-reactivity. The basis for this cross-reactivity is unknown but may be due in part to antibody recognition of a three-dimensional (3D) pattern conserved in many M protein HVRs that confers binding to human C4b-binding protein (C4BP). To test this hypothesis, we asked whether a single M protein immunogen carrying the 3D pattern would elicit cross-reactivity against other M types carrying the 3D pattern. We found that a 34-amino acid sequence of M2 protein bearing the 3D pattern retained full C4BP-binding capacity when fused to a coiled coil-stabilizing sequence from GCN4. This immunogen, called M2G, elicited cross-reactive antibodies against a number of M types that carry the 3D pattern but not against those that lack the 3D pattern. The M2G antiserum recognized M proteins as displayed natively on the Strep A surface, and promoted the opsonophagocytic killing of Strep A strains expressing these M proteins. As C4BP-binding is a conserved virulence trait of Strep A, targeting the 3D pattern may prove advantageous in vaccine design.

immunology↗