Caspase-1-driven neutrophil pyroptosis and its role in host susceptibility to Pseudomonas aeruginosa.

Multiple regulated neutrophil cell death programs contribute to host defense against infections. However, despite expressing all necessary inflammasome components, neutrophils are thought to be generally defective in Caspase-1-dependent pyroptosis. By screening different bacterial species, we found that several Pseudomonas aeruginosa (P. aeruginosa) strains trigger Caspase-1-dependent pyroptosis in human and murine neutrophils. Notably, deletion of Exotoxins U or S in P. aeruginosa enhanced neutrophil death to Caspase-1-dependent pyroptosis, suggesting that these exotoxins interfere with this pathway. Mechanistically, P. aeruginosa Flagellin activates the NLRC4 inflammasome, which supports Caspase-1-driven interleukin (IL)-1ß secretion and Gasdermin D (GSDMD)-dependent neutrophil pyroptosis. Furthermore, P. aeruginosa-induced GSDMD activation triggers Calcium-dependent and Peptidyl Arginine Deaminase-4-driven histone citrullination and translocation of neutrophil DNA into the cell cytosol without inducing extracellular Neutrophil Extracellular Traps. Finally, we show that neutrophil Caspase-1 contributes to IL-1ß production and susceptibility to pyroptosis-inducing P. aeruginosa strains in vivo. Overall, we demonstrate that neutrophils are not universally resistant for Caspase-1-dependent pyroptosis.

Elucidating allergic reaction mechanisms in response to SARS-CoV-2 mRNA vaccination in adults.

BACKGROUND: During the COVID-19 pandemic, novel nanoparticle-based mRNA vaccines were developed. A small number of individuals developed allergic reactions to these vaccines although the mechanisms remain undefined. METHODS: To understand COVID-19 vaccine-mediated allergic reactions, we enrolled 19 participants who developed allergic events within 2 h of vaccination and 13 controls, nonreactors. Using standard hemolysis assays, we demonstrated that sera from allergic participants induced stronger complement activation compared to nonallergic subjects following ex vivo vaccine exposure. RESULTS: Vaccine-mediated complement activation correlated with anti-polyethelyne glycol (PEG) IgG (but not IgM) levels while anti-PEG IgE was undetectable in all subjects. Depletion of total IgG suppressed complement activation in select individuals. To investigate the effects of vaccine excipients on basophil function, we employed a validated indirect basophil activation test that stratified the allergic populations into high and low responders. Complement C3a and C5a receptor blockade in this system suppressed basophil response, providing strong evidence for complement involvement in vaccine-mediated basophil activation. Single-cell multiome analysis revealed differential expression of genes encoding the cytokine response and Toll-like receptor (TLR) pathways within the monocyte compartment. Differential chromatin accessibility for IL-13 and IL-1B genes was found in allergic and nonallergic participants, suggesting that in vivo, epigenetic modulation of mononuclear phagocyte immunophenotypes determines their subsequent functional responsiveness, contributing to the overall physiologic manifestation of vaccine reactions. CONCLUSION: These findings provide insights into the mechanisms underlying allergic reactions to COVID-19 mRNA vaccines, which may be used for future vaccine strategies in individuals with prior history of allergies or reactions and reduce vaccine hesitancy.

Adipose tissue is a critical regulator of osteoarthritis.

Osteoarthritis (OA), the leading cause of pain and disability worldwide, disproportionally affects individuals with obesity. The mechanisms by which obesity leads to the onset and progression of OA are unclear due to the complex interactions among the metabolic, biomechanical, and inflammatory factors that accompany increased adiposity. We used a murine preclinical model of lipodystrophy (LD) to examine the direct contribution of adipose tissue to OA. Knee joints of LD mice were protected from spontaneous or posttraumatic OA, on either a chow or high-fat diet, despite similar body weight and the presence of systemic inflammation. These findings indicate that adipose tissue itself plays a critical role in the pathophysiology of OA. Susceptibility to posttraumatic OA was reintroduced into LD mice using implantation of a small adipose tissue depot derived from wild-type animals or mouse embryonic fibroblasts that undergo spontaneous adipogenesis, implicating paracrine signaling from fat, rather than body weight, as a mediator of joint degeneration.

Rapamycin Perfluorocarbon Nanoparticle Mitigates Cisplatin-Induced Acute Kidney Injury.

For nearly five decades, cisplatin has played an important role as a standard chemotherapeutic agent and been prescribed to 10-20% of all cancer patients. Although nephrotoxicity associated with platinum-based agents is well recognized, treatment of cisplatin-induced acute kidney injury is mainly supportive and no specific mechanism-based prophylactic approach is available to date. Here, we postulated that systemically delivered rapamycin perfluorocarbon nanoparticles (PFC NP) could reach the injured kidneys at sufficient and sustained concentrations to mitigate cisplatin-induced acute kidney injury and preserve renal function. Using fluorescence microscopic imaging and fluorine magnetic resonance imaging/spectroscopy, we illustrated that rapamycin-loaded PFC NP permeated and were retained in injured kidneys. Histologic evaluation and blood urea nitrogen (BUN) confirmed that renal structure and function were preserved 48 h after cisplatin injury. Similarly, weight loss was slowed down. Using western blotting and immunofluorescence staining, mechanistic studies revealed that rapamycin PFC NP significantly enhanced autophagy in the kidney, reduced the expression of intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1), as well as decreased the expression of the apoptotic protein Bax, all of which contributed to the suppression of apoptosis that was confirmed with TUNEL staining. In summary, the delivery of an approved agent such as rapamycin in a PFC NP format enhances local delivery and offers a novel mechanism-based prophylactic therapy for cisplatin-induced acute kidney injury.

Evaluation of Serum Sodium Correction Rates for Management of Hyponatremia in Hospitalized Patients.

BACKGROUND: Appropriate correction of hyponatremia can reduce complications such as osmotic demyelination syndrome (ODS). OBJECTIVE: To evaluate rates of serum sodium correction in hyponatremic hospitalized patients and identify factors associated with higher rates of overcorrection. METHODS: This is an institutional review board-approved single-center, retrospective chart review of patients ≥18 years of age with at least 1 serum sodium <130 mEq/L during hospitalization. The primary end point was percentage of patients appropriately corrected for hyponatremia. Appropriate correction was defined as a sodium change ≤12 mEq/L over 24 hours and 18 mEq/L over 48 hours, and overcorrection was defined as an increase in serum sodium exceeding these cutoffs. Secondary end points included incidence of ODS, poor neurological outcome, intensive care unit (ICU) and hospital lengths of stay (LOSs), and in-hospital mortality. RESULTS: Of 234 patients evaluated, 100 were included. Mean age was 72 ± 16 years, and 47% were male. Overcorrection occurred in 14 patients. There was no incidence of ODS. Rates of poor neurological outcome (P = 0.77), ICU (P = 0.09) and hospital LOS (P = 0.13), and in-hospital mortality (P = 0.20) were similar between appropriately corrected and overcorrected patients. Using a logistic regression analysis, severe hyponatremia (serum sodium < 120 mEq/L; P = 0.0122) and history of alcohol use disorder (P < 0.001) were risk factors found to be associated with overcorrection. CONCLUSION AND RELEVANCE: Overcorrection of hyponatremia occurred in 14% of patients in this study. To minimize this risk, further caution should be taken when managing patients presenting with identified risk factors.

Side-chain modification of collagen-targeting peptide prevents dye aggregation for improved molecular imaging of arthritic joints.

Near-infrared (NIR) dye-peptide conjugates are widely used for tissue-targeted molecular fluorescence imaging of pathophysiologic conditions. However, the significant contribution of both dye and peptide to the net mass of these bioconjugates implies that small changes in either component could alter their photophysical and biological properties. Here, we synthesized and conjugated a type I collagen targeted peptide, RRANAALKAGELYKCILY, to either a hydrophobic (LS1000) or hydrophilic (LS1006) NIR fluorescent dye. Spectroscopic analysis revealed rapid self-assembly of both LS1000 and LS1006 in aqueous media to form stable dimeric/H aggregates, regardless of the free dye's solubility in water. We discovered that replacing the cysteine residue in LS1000 and LS1006 with acetamidomethyl cysteine to afford LS1001 and LS1107, respectively, disrupted the peptide's self-assembly and activated the previously quenched dye's fluorescence in aqueous conditions. These results highlight the dominant role of the octadecapeptide, but not the dye molecules, in controlling the photophysical properties of these conjugates by likely sequestering or extruding the hydrophobic or hydrophilic dyes, respectively. Application of the compounds for imaging collagen-rich tissue in an animal model of inflammatory arthritis showed enhanced uptake of all four conjugates, which retained high collagen-binding affinity, in inflamed joints. Moreover, LS1001 and LS1107 improved the arthritic joint-to-background contrast, suggesting that reduced aggregation enhanced the clearance of these compounds from non-target tissues. Our results highlight a peptide-driven strategy to alter the aggregation states of molecular probes in aqueous solutions, irrespective of the water-solubilizing properties of the dye molecules. The interplay between the monomeric and aggregated forms of the conjugates using simple thiol-modifiers lends the peptide-driven approach to diverse applications, including the effective imaging of inflammatory arthritis joints.

Safety Profile of Thread Lifts on the Face and Neck: An Evidence-Based Systematic Review.

BACKGROUND: "Thread lifting" has quickly gained popularity as a minimally invasive treatment for facial rejuvenation. However, the effectiveness is questionable, and the safety and adverse effects are often not discussed. OBJECTIVE: To identify and discuss the adverse effects associated with various types of threads. MATERIALS AND METHODS: Studies describing the use of thread lifts were identified using a PubMed search. Inclusion criteria included studies in which barbed and nonbarbed threads were used for the face and neck. RESULTS: Fifty-nine articles consisting of 14,222 patients (14,134 barbed, 81 nonbarbed, and 7 combined cases) were included. The most common side effects overall were facial asymmetry (n = 6,143), edema/tumefaction (n = 453), and ecchymosis (n = 407). Serious adverse effects were rare and consisted of paresthesias, alopecia, and injuries to vessels/glands. Most adverse effects were transient and self-resolving, with the exception of contour irregularities, injuries to vessels/glands, infections, and inflammatory reactions. CONCLUSION: Most side effects associated with threads were self-resolving, whereas more serious cases subsided with treatment. Future studies are critical to further determine whether thread lifting provides long-lasting, safe, and satisfying results.

Monitoring Response to Platelet-Rich Plasma in Patients with Alopecia Areata with Optical Coherence Tomography: A Case Series.

Alopecia areata (AA) is an autoimmune hair loss condition that is difficult to treat and frequently disruptive to the psychosocial well-being of patients. Platelet-rich plasma (PRP) is an innovative therapy that provides concentrated GFs that impart anti-inflammatory effects. Optical coherence tomography (OCT) is a noninvasive imaging modality with the potential for providing quantitative monitoring of AA response to PRP. Our objective is to share our experience using OCT to monitor the therapeutic progress of patients with AA treated with PRP. Two patients with patchy AA and one with alopecia universalis were treated with PRP three times at 6-week intervals as part of a larger clinical trial. Patients were followed from baseline to week 24 with OCT imaging. OCT demonstrates an increase in hair density associated with improvement in inflammation at week 24. Conversely, the patient with alopecia universalis did not experience any significant change in follicular activity. This case series exemplifies the potential of PRP in inflammatory regulation as well as hair regrowth in patchy AA, whereas there is no notable advantage in alopecia universalis. Our findings add evidence on the possible value of OCT in quantitatively assessing hair growth progress throughout a treatment course.

The human papillomavirus vaccine as a treatment for human papillomavirus-related dysplastic and neoplastic conditions: A literature review.

BACKGROUND: Human papillomavirus (HPV) infections are associated with common dermatologic and nondermatologic diseases. Although HPV vaccines are well established as preventive measures for genital warts and cervical neoplasia, their use as therapeutic agents deserves greater attention. OBJECTIVE: To evaluate the use of HPV vaccine(s) as a treatment modality for cutaneous and/or mucosal disease. METHODS: A primary literature search using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines was conducted in January 2019 by using the PubMed and Cochrane databases. RESULTS: A total of 63 articles with 4439 patients were included. The majority of patients with cutaneous warts, recurrent respiratory papillomatosis, and squamous and basal cell carcinomas were successfully treated with HPV vaccination. Preliminary data on patients with pre-existing anogenital warts, cervical intraepithelial neoplasia, anal intraepithelial neoplasia, and vulvar intraepithelial neoplasia is promising. LIMITATIONS: This review was limited by the lack of controls, patients' previous HPV vaccination status, and publication bias. CONCLUSION: The commercially available three-dose, quadrivalent HPV vaccine is a potential therapeutic option for the treatment of cutaneous warts, recurrent respiratory papillomatosis, and squamous and basal cell carcinomas. Noncommercially available HPV vaccines demonstrate therapeutic response for treating anogenital warts, cervical intraepithelial neoplasia, anal intraepithelial neoplasia, and vulvar intraepithelial neoplasia. The vaccine's efficacy as an adjunct therapy for HPV-associated cutaneous and/or mucosal disease warrants further exploration.

Contribution of Adipose-Derived Factor D/Adipsin to Complement Alternative Pathway Activation: Lessons from Lipodystrophy.

Factor D (FD) is an essential component of the complement alternative pathway (AP). It is an attractive pharmaceutical target because it is an AP-specific protease circulating in blood. Most components of the complement activation pathways are produced by the liver, but FD is highly expressed by adipose tissue. Two critical questions are: 1) to what degree does adipose tissue contribute to circulating FD levels and 2) what quantity of FD is sufficient to maintain a functional AP? To address these issues, we studied a novel mouse strain with complete lipodystrophy (LD), the fld mouse with partial LD, an FD-deficient mouse, and samples from lipodystrophic patients. FD was undetectable in the serum of LD mice, which also showed minimal AP function. Reconstitution with purified FD, serum mixing experiments, and studies of partial LD mice all demonstrated that a low level of serum FD is sufficient for normal AP activity in the mouse system. This conclusion was further supported by experiments in which wild-type adipose precursors were transplanted into LD mice. Our results indicate that almost all FD in mouse serum is derived from adipose tissue. In contrast, FD levels were reduced ∼50% in the sera of patients with congenital generalized LD. Our studies further demonstrate that a relatively small amount of serum FD is sufficient to facilitate significant time-dependent AP activity in humans and in mice. Furthermore, this observation highlights the potential importance of obtaining nearly complete inhibition of FD in treating alternative complement activation in various autoimmune and inflammatory human diseases.

Adverse Events of Injectable Deoxycholic Acid.

BACKGROUND: Injectable deoxycholic acid (DCA) may be used to remove excess submental fat and off-label for local adipose reduction. Despite DCA's widespread use, rare incidences of severe, systemic, long-term adverse events (AEs) have been reported. OBJECTIVE: To evaluate the potential side effects associated with injectable DCA. METHODS AND MATERIALS: A systematic review was conducted using PubMed, Cochrane, CINAHL, and Web of Science using PRISMA guidelines to gather the literature relating to DCA or deoxycholate-associated AEs and their management. RESULTS: Twenty-eight manuscripts were included after full article review. Most commonly, patients experienced mild localized AEs, whereas a small number of patients experienced severe pain, alopecia, nasopharyngitis, dysphagia, dizziness/lightheadedness, and gastrointestinal upset. Severe, long-term AEs were reported as rare in the evaluated literature. Deoxycholic acid injections in large volumes were more likely to cause severe adverse effects. CONCLUSION: Self-resolving, mild side effects and severe but rare adverse effects have been reported with DCA use making it a safe treatment for local adipose reduction. Further studies are necessary to determine its safety profile, especially when using DCA in off-label areas.

Combined Experimental Approach and Finite Element Modeling of Small Molecule Transport Through Joint Synovium to Measure Effective Diffusivity.

Trans-synovial solute transport plays a critical role in the clearance of intra-articularly (IA) delivered drugs. In this study, we present a computational finite element model (FEM) of solute transport through the synovium validated by experiments on synovial explants. Unsteady diffusion of urea, a small uncharged molecule, was measured through devitalized porcine and human synovium using custom-built diffusion chambers. A multiphasic computational model was constructed and optimized with the experimental data to extract effective diffusivity for urea within the synovium. A monotonic decrease in urea concentration was observed in the donor bath over time, with an effective diffusivity found to be an order of magnitude lower in synovium versus that measured in free solution. Parametric studies incorporating an intimal cell layer with varying thickness and varying effective diffusivities were performed, revealing a dependence of drug clearance kinetics on both parameters. The findings of this study indicate that the synovial matrix impedes urea solute transport out of the joint with little retention of the solute in the matrix.

Suppression of NF-κB activity via nanoparticle-based siRNA delivery alters early cartilage responses to injury.

Osteoarthritis (OA) is a major cause of disability and morbidity in the aging population. Joint injury leads to cartilage damage, a known determinant for subsequent development of posttraumatic OA, which accounts for 12% of all OA. Understanding the early molecular and cellular responses postinjury may provide targets for therapeutic interventions that limit articular degeneration. Using a murine model of controlled knee joint impact injury that allows the examination of cartilage responses to injury at specific time points, we show that intraarticular delivery of a peptidic nanoparticle complexed to NF-κB siRNA significantly reduces early chondrocyte apoptosis and reactive synovitis. Our data suggest that NF-κB siRNA nanotherapy maintains cartilage homeostasis by enhancing AMPK signaling while suppressing mTORC1 and Wnt/ß-catenin activity. These findings delineate an extensive crosstalk between NF-κB and signaling pathways that govern cartilage responses postinjury and suggest that delivery of NF-κB siRNA nanotherapy to attenuate early inflammation may limit the chronic consequences of joint injury. Therapeutic benefits of siRNA nanotherapy may also apply to primary OA in which NF-κB activation mediates chondrocyte catabolic responses. Additionally, a critical barrier to the successful development of OA treatment includes ineffective delivery of therapeutic agents to the resident chondrocytes in the avascular cartilage. Here, we show that the peptide-siRNA nanocomplexes are nonimmunogenic, are freely and deeply penetrant to human OA cartilage, and persist in chondrocyte lacunae for at least 2 wk. The peptide-siRNA platform thus provides a clinically relevant and promising approach to overcoming the obstacles of drug delivery to the highly inaccessible chondrocytes.

Cathepsin G Controls Arterial But Not Venular Myeloid Cell Recruitment.

BACKGROUND: Therapeutic targeting of arterial leukocyte recruitment in the context of atherosclerosis has been disappointing in clinical studies. Reasons for such failures include the lack of knowledge of arterial-specific recruitment patterns. Here we establish the importance of the cathepsin G (CatG) in the context of arterial myeloid cell recruitment. METHODS: Intravital microscopy of the carotid artery, the jugular vein, and cremasteric arterioles and venules in Apoe-/-and CatG-deficient mice (Apoe-/-Ctsg-/-) was used to study site-specific myeloid cell behavior after high-fat diet feeding or tumor necrosis factor stimulation. Atherosclerosis development was assessed in aortic root sections after 4 weeks of high-fat diet, whereas lung inflammation was assessed after inhalation of lipopolysaccharide. Endothelial deposition of CatG and CCL5 was quantified in whole-mount preparations using 2-photon and confocal microscopy. RESULTS: Our observations elucidated a crucial role for CatG during arterial leukocyte adhesion, an effect not found during venular adhesion. Consequently, CatG deficiency attenuates atherosclerosis but not acute lung inflammation. Mechanistically, CatG is immobilized on arterial endothelium where it activates leukocytes to firmly adhere engaging integrin clustering, a process of crucial importance to achieve effective adherence under high-shear flow. Therapeutic neutralization of CatG specifically abrogated arterial leukocyte adhesion without affecting myeloid cell adhesion in the microcirculation. Repetitive application of CatG-neutralizing antibodies permitted inhibition of atherogenesis in mice. CONCLUSIONS: Taken together, these findings present evidence of an arterial-specific recruitment pattern centered on CatG-instructed adhesion strengthening. The inhibition of this process could provide a novel strategy for treatment of arterial inflammation with limited side effects.

Neutrophil Proteases Promote Experimental Abdominal Aortic Aneurysm via Extracellular Trap Release and Plasmacytoid Dendritic Cell Activation.

OBJECTIVE: We previously established that neutrophil-derived dipeptidyl peptidase I (DPPI) is essential for experimental abdominal aortic aneurysm (AAA) development. Because DPPI activates several neutrophil serine proteases, it remains to be determined whether the AAA-promoting effect of DPPI is mediated by neutrophil serine proteases. APPROACH AND RESULTS: Using an elastase-induced AAA model, we demonstrate that the absence of 2 neutrophil serine proteases, neutrophil elastase and proteinase-3, recapitulates the AAA-resistant phenotype of DPPI-deficient mice. DPPI and neutrophil serine proteases direct the in vitro and in vivo release of extracellular structures termed neutrophil extracellular traps (NETs). Administration of DNase1, which dismantles NETs, suppresses elastase-induced AAA in wild-type animals and in DPPI-deficient mice reconstituted with wild-type neutrophils. NETs also contain the cathelicidin-related antimicrobial peptide that complexes with self-DNA in recruiting plasmacytoid dendritic cells (pDCs), inducing type I interferons (IFNs) and promoting AAA in DPPI-deficient mice. Conversely, depletion of pDCs or blockade of type I IFNs suppresses experimental AAA. Moreover, we find an abundance of human cathelicidin peptide, a 37 amino acid sequence starting with 2 leucines and the human orthologue of cathelicidin-related antimicrobial peptide, in the vicinity of pDCs in human AAA tissues. Increased type I IFN mRNA expression is observed in human AAA tissues and circulating IFN-α is detected in ≈50% of the AAA sera examined. CONCLUSIONS: These results suggest that neutrophil protease-mediated NET release contributes to elastase-induced AAA through pDC activation and type I IFN production. These findings increase our understanding of the pathways underlying AAA inflammatory responses and suggest that limiting NET, pDC, and type I IFN activities may suppress aneurysm progression.

Neutrophil serine proteases: specific regulators of inflammation.

Neutrophils are essential for host defence against invading pathogens. They engulf and degrade microorganisms using an array of weapons that include reactive oxygen species, antimicrobial peptides, and proteases such as cathepsin G, neutrophil elastase and proteinase 3. As discussed in this Review, the generation of mice deficient in these proteases has established a role for these enzymes as intracellular microbicidal agents. However, I focus mainly on emerging data indicating that, after release, these proteases also contribute to the extracellular killing of microorganisms, and regulate non-infectious inflammatory processes by activating specific receptors and modulating the levels of cytokines.

Activation of proteinase 3 contributes to Non-alcoholic Fatty Liver Disease (NAFLD) and insulin resistance.

Activation of inflammatory pathways is known to accompany development of obesity-induced non-alcoholic fatty liver disease (NAFLD), insulin resistance and type 2 diabetes. In addition to caspase-1, the neutrophil serine proteases proteinase 3, neutrophil elastase and cathepsin G are able to process the inactive pro-inflammatory mediators IL-1ß and IL-18 to their bioactive forms, thereby regulating inflammatory responses. In the present study, we investigated whether proteinase 3 is involved in obesity-induced development of insulin resistance and NAFLD. We investigated the development of NAFLD and insulin resistance in mice deficient for neutrophil elastase/proteinase 3 and neutrophil elastase/cathepsin G and in wild-type mice treated with the neutrophil serine proteinase inhibitor human alpha-1 antitrypsin. Expression profiling of metabolically relevant tissues obtained from insulin resistant mice showed that expression of proteinase 3 was specifically upregulated in the liver, whereas neutrophil elastase, cathepsin G and caspase-1 were not. Neutrophil elastase/proteinase 3 deficient mice showed strongly reduced levels of lipids in the liver after fed a high fat diet. Moreover, these mice were resistant to high fat diet-induced weight gain, inflammation and insulin resistance. Injection of proteinase 3 exacerbated insulin resistance in caspase-1(-/-) mice, indicating that proteinase 3 acts independently of caspase-1. Treatment with alpha-1 antitrypsin during the last 10 days of a 16 week high fat diet reduced hepatic lipid content and decreased fasting glucose levels. We conclude that proteinase 3 is involved in NAFLD and insulin resistance and that inhibition of proteinase 3 may have therapeutic potential.

Quantifying progression and regression of thrombotic risk in experimental atherosclerosis.

Currently, there are no generally applicable noninvasive methods for defining the relationship between atherosclerotic vascular damage and risk of focal thrombosis. Herein, we demonstrate methods to delineate the progression and regression of vascular damage in response to an atherogenic diet by quantifying the in vivo accumulation of semipermeable 200-300 nm perfluorocarbon core nanoparticles (PFC-NP) in ApoE null mouse plaques with [(19)F] magnetic resonance spectroscopy (MRS). Permeability to PFC-NP remained minimal until 12 weeks on diet, then increased rapidly following 12 weeks, but regressed to baseline within 8 weeks after diet normalization. Markedly accelerated clotting (53.3% decrease in clotting time) was observed in carotid artery preparations of fat-fed mice subjected to photochemical injury as defined by the time to flow cessation. For all mice on and off diet, an inverse linear relationship was observed between the permeability to PFC-NP and accelerated thrombosis (P = 0.02). Translational feasibility for quantifying plaque permeability and vascular damage in vivo was demonstrated with clinical 3 T MRI of PFC-NP accumulating in plaques of atherosclerotic rabbits. These observations suggest that excessive permeability to PFC-NP may indicate prothrombotic risk in damaged atherosclerotic vasculature, which resolves within weeks after dietary therapy.