Search

A critical factor in thrombus formation during venoarterial extracorporeal membrane oxygenation (VA ECMO) is prothrombotic flow dynamics generated by the drainage cannula’s design. This study aimed to create and evaluate a novel drainage cannula design which optimized blood flow dynamics to reduce thrombus formation. Computational fluid dynamics (CFD) was used to iteratively vary drainage cannula design parameters such as inner wall shape and side hole shape. The final novel design was then placed in an ex vivoblood circulation loop, and compared against a Bio-Medicus cannula (n = 6, each). Clot volume, hemolysis, and other parameters were measured to assess thrombus formation markers. The novel design consisted of a parabolic inner wall profile with closely spaced side holes angled at 30º to align with flow. When tested in the ex vivoloop, the novel design resulted in lower instances (two vs. four) and volumes of clot in the cannula (360.5 ± 254.8 vs. 1258.0 ± 651.7 µl) when compared to the Bio-Medicus cannula. Results from tests assessing hemolysis, platelet activation, and other thrombotic markers revealed a noninferior relationship between the novel and Bio-Medicus designs. Future work will explore the clinical applicability of these findings.

•Reduced PI3K(p110α) in cardiomyocytes in mice led to atrial dysfunction, fibrosis, arrhythmia, and increased susceptibility to atrial enlargement and thrombi.•Circulating PI3K-related lipids were dysregulated in athletes with atrial fibrillation (AF).•Open-access resource of the mouse atrial proteome and comparison with the human atrial proteome for the discovery of new drug targets & biomarkers.•Assessment of PI3K-related markers in tissue and plasma may identify individuals at risk of AF.

Thrombolysis with tissue-type plasminogen activator (tPA) remains the main treatment for acute ischemic stroke. Nevertheless, tPA intervention is limited by a short therapeutic window, low recanalization rates, and a risk of intracranial hemorrhage (ICH), highlighting the clinical demand for improved thrombolytic drugs. We examined a novel thrombolytic agent termed “SCE5-scuPA,” comprising a single-chain urokinase plasminogen activator (scuPA) fused with a single-chain antibody (SCE5) that targets the activated glycoprotein IIb/IIIa platelet receptor, for its effects in experimental stroke. SCE5-scuPA was first tested in a whole blood clot degradation assay to show the benefit of platelet-targeted thrombolysis. The tail bleeding time, blood clearance, and biodistribution were then determined to inform the use of SCE5-scuPA in mouse models of photothrombotic stroke and middle cerebral artery occlusion against tenecteplase. The impacts of SCE5-scuPA on motor function, ICH, blood–brain barrier (BBB) integrity, and immunosuppression were evaluated. Infarct size was measured by computed tomography imaging and magnetic resonance imaging. SCE5-scuPA enhanced clot degradation ex vivo compared with its nonplatelet-targeting control. The maximal SCE5-scuPA dose that maintained hemostasis and a rapid blood clearance was determined. SCE5-scuPA administration both before and 2 hours after photothrombotic stroke reduced the infarct volume. SCE5-scuPA also improved neurologic deficit, decreased intracerebral blood deposits, preserved the BBB, and alleviated immunosuppression poststroke. In middle cerebral artery occlusion, SCE5-scuPA did not worsen stroke outcomes or cause ICH, and it protected the BBB. Our findings support the ongoing development of platelet-targeted thrombolysis with SCE5-scuPA as a novel emergency treatment for acute ischemic stroke with a promising safety profile.

The clinical success of monoclonal antibody therapy has inspired research in understanding the fundamental molecular basis of antibody–antigen interactions and the engineering of antibodies and antibody assemblies with enhanced or novel properties. In particular, colloidally stable antibody assemblies can enhance dosing strategies and enable combined therapy of a mixture of antibodies or biologics. Herein, nanoassemblies of therapeutic antibodies were fabricated with controlled physicochemical properties using a versatile template-mediated assembly method. The antibody nanoparticles (AbNPs) cross-linked with poly(ethylene glycol)-N-hydroxysuccinimide were monodispersed, with particle diameters consistent with the template size (250 nm). When assembled using Herceptin or Kadcyla as a model antibody and antibody–drug conjugate, respectively, the nanoparticles retained the selectivity of the monoclonal antibody and recognized >98% of cells expressing the target receptors on cell membranes. Unlike the free Herceptin antibody, which was predominantly localized at the surface, the AbNPs were internalized via receptor-mediated endocytosis, presenting opportunities for delivering monoclonal antibodies intracellularly at high concentrations and/or against intracellular targets. With the vast array of antibodies that could be applied and different cross-linking chemistries possible, the reported antibody assembly strategy provides a versatile platform for the development of antibody assemblies for therapeutic, diagnostic, and clinical applications.

Thrombolytic (clot-busting) therapies with plasminogen activators (PAs) are first-line treatments against acute thrombosis and ischemic stroke. However, limitations such as narrow therapeutic windows, low success rates, and bleeding complications hinder their clinical use. Drug-loaded polyphenol-based nanoparticles (NPs) could address these shortfalls by delivering a more targeted and safer thrombolysis, coupled with advantages such as improved biocompatibility and higher stability in vivo. Herein, a template-mediated polyphenol-based supramolecular assembly strategy is used to prepare nanocarriers of thrombolytic drugs. A thrombin-dependent drug release mechanism is integrated using tannic acid (TA) to cross-link urokinase-type PA (uPA) and a thrombin-cleavable peptide on a sacrificial mesoporous silica template via noncovalent interactions. Following drug loading and template removal, the resulting NPs retain active uPA and demonstrate enhanced plasminogen activation in the presence of thrombin (1.14-fold; p< 0.05). Additionally, they display lower association with macrophage (RAW 264.7) and monocytic (THP-1) cell lines (43 and 7% reduction, respectively), reduced hepatic accumulation, and delayed blood clearance in vivo (90% clearance at 60 min vs 5 min) compared with the template-containing NPs. Our thrombin-responsive, polyphenol-based NPs represent a promising platform for advanced drug delivery applications, with potential to improve thrombolytic therapies.

Extensive reports of pulmonary embolisms, ischaemic stroke and myocardial infarctions caused by coronavirus disease 2019 (COVID-19), as well as a significantly increased long-term risk of cardiovascular diseases in COVID-19 survivors, have highlighted severe deficiencies in our understanding of thromboinflammation and the need for new therapeutic options. Due to the complexity of the immunothrombosis pathophysiology, the efficacy of treatment with conventional anti-thrombotic medication is questioned. Thrombolytics do appear efficacious, but are hindered by severe bleeding risks, limiting their use. Nanomedicine can have profound impact in this context, protecting delicate (bio)pharmaceuticals from degradation en route and enabling delivery in a targeted and on demand manner. We provide an overview of the most promising nanocarrier systems and design strategies that may be adapted to develop nanomedicine for COVID-19-induced thromboinflammation, including dual-therapeutic approaches with antiviral and immunosuppressants. Resultant targeted and side-effect-free treatment may aid greatly in the fight against the ongoing COVID-19 pandemic.

We report the development of a theranostic collagen targeted cell penetrating drug delivery system toward treatment of cardiovascular disease. Caused by the action of matrix metalloproteinases (MMP), degraded collagen is a hallmark of unstable atherosclerotic plaques that are highly susceptible toward rupture. Targeting unstable plaques and delivering MMP blockers directly to plaque to inhibit MMP activity is a promising new strategy that requires the benefits and possibilities of nanodelivery approaches. The presented delivery system is designed to (a) target and bind to a cryptic epitope on collagen IV exposed through the degradative action of matrix metalloproteinase 2 (MMP2), (b) to image the targeting and cell uptake, and (c) to deliver the MMP14 inhibitor naphthofluorescein. In detail, the novel targeting unit is composed of a collagen-homing T-peptide and bound to a MMP2-cleavable activatable cell penetrating peptide (ACPP) which upon cleavage by MMP2 deposits the MMP14 blocker drug into cells, directly into contact with MMP 14 activating enzymes. To selectively attach both the targeting peptide and a reporting imaging dye, a nanosponge nanoparticle network is modified to present orthogonal aldehyde and thiol functional groups as surface units. The MMP14 inhibitor naphthofluorescein is loaded into the nanoparticle delivery system after postconjugation chemistries and finalizes the synthesis of this novel theranostic delivery system. The ability to evade phagocytosis is confirmed in vitroby using murine RAW cell line and effective cell uptake in vitroby using the MMP2 producing the HT1080 cell line is demonstrated. In this work, we have combined a highly specific targeting peptide directed against degraded collagen and a tailorable nanosystem that is deemed to deliver its potent drug load directly into cells, to inhibit the cascade for MMP activation which breaks down collagen structures leading to plaque rupture, the underlying cause of myocardial infarction and strokes.

The orientation-specific immobilization of antibodies onto nanoparticles, to preserve antibody–antigen recognition, is a key challenge in developing targeted nanomedicines. Herein, we report the targeting ability of metal–phenolic network (MPN)-coated gold nanoparticles with surface-physisorbed antibodies against respective antigens. The MPN coatings were self-assembled from metal ions (FeIII, CoII, CuII, NiII, or ZnII) cross-linked with tannic acid. Upon physisorption of antibodies, all particle systems exhibited enhanced association with target antigens, with CoIIsystems demonstrating more than 2-fold greater association. These systems contained more metal atoms distributed in a way to specifically interact with antibodies, which were investigated by molecular dynamics simulations. A model antibody fragment crystallizable (Fc) region in solution with CoII-tannic acid complexes revealed that the solvent-exposed CoIIcan directly coordinate to the histidine-rich portion of the Fc region. This one-pot interaction suggests anchoring of the antibody Fc region to the MPN on nanoparticles, allowing for enhanced targeting.

Drug carriers typically require both stealth and targeting properties to minimize nonspecific interactions with healthy cells and increase specific interaction with diseased cells. Herein, the assembly of targeted poly(ethylene glycol) (PEG) particles functionalized with cyclic peptides containing Arg-Gly-Asp (RGD) (ligand) using a mesoporous silica templating method is reported. The influence of PEG molecular weight, ligand-to-PEG molecule ratio, and particle size on cancer cell targeting to balance stealth and targeting of the engineered PEG particles is investigated. RGD-functionalized PEG particles (PEG-RGD particles) efficiently target U-87 MG cancer cells under static and flow conditions in vitro, whereas PEG and cyclic peptides containing Arg-Asp-Gly (RDG)-functionalized PEG (PEG-RDG) particles display negligible interaction with the same cells. Increasing the ligand-to-PEG molecule ratio improves cell targeting. In addition, the targeted PEG-RGD particles improve cell uptake via receptor-mediated endocytosis, which is desirable for intracellular drug delivery. The PEG-RGD particles show improved tumor targeting (14% ID g–1) when compared with the PEG (3% ID g–1) and PEG-RDG (7% ID g–1) particles in vivo, although the PEG-RGD particles show comparatively higher spleen and liver accumulation. The targeted PEG particles represent a platform for developing particles aimed at balancing nonspecific and specific interactions in biological systems.

The treatment and management of kidney disease present a significant global challenge, affecting over 800 million individuals and necessitating innovative therapeutic strategies that transcend symptomatic relief. The application of nanotechnology to renal therapies, while still in its early stages, holds transformative potential for improving treatment outcomes. Recent advancements in nanoparticle-based drug delivery leverage the unique physicochemical properties of nanoparticles for targeted and controlled therapeutic delivery to the kidneys. Current research is focused on understanding the functional and phenotypic changes in kidney cells during both acute and chronic conditions, allowing for the identification of optimal target cells. Additionally, the development of tailored nanomedicines enhances their retention and binding to key renal membranes and cell populations, ultimately improving localization, tolerability, and efficacy. However, significant barriers remain, including inconsistent nanoparticle synthesis and the complexity of kidney-specific targeting. To overcome these challenges, the field requires advanced synthesis techniques, refined targeting strategies, and the establishment of animal models that accurately reflect human kidney disease. These efforts are critical for the clinical application of nanotherapeutics, which promise novel solutions for kidney disease management. This review evaluates a substantial body of in vivoresearch, highlighting the prospects, challenges, and opportunities presented by nanotechnology-mediated therapies and their potential to transform kidney disease treatment.

MOFs have demonstrated outstanding properties for the protection and controlled release of different bio-entities, from proteins to living cells. Carbohydrates, as pure molecules or as a component of proteins and cells, perform essential biological functions. Thus, an understanding of the role of carbohydrates in the formation of MOF-based bio-composites will facilitate their application to biotechnology and medicine. Here, we investigate the role of carbohydrate molecular weight and chemical functionalization in the formation of carbohydrate@MOF composites. We find that chemical functionalization, such as carboxylation, that leads to an enhancement of metal cation concentration at the surface of the molecule triggers the rapid self-assembly of the MOF material, zeolitic-imidazolate framework 8 (ZIF-8). Furthermore, we determine the encapsulation efficiency and measure the release properties of the carbohydrate under controlled conditions. Our findings show that MOFs can be used to prepare a new class of biocomposites for the delivery of carbohydrate-based therapeutics.

Nanomedicine holds great promise for vascular disease diagnosis and specific therapy, yet rapid sequestration by the mononuclear phagocytic system limits the efficacy of particle-based agents. The use of low-fouling polymers, such as poly(ethylene glycol), efficiently reduces this immune recognition, but these nondegradable polymers can accumulate in the human body and may cause adverse effects after prolonged use. Thus, new particle formulations combining stealth, low immunogenicity and biocompatible features are required to enable clinical use. Here, a low-fouling particle platform is described using exclusively protein material. A recombinant protein with superior hydrophilic characteristics provided by the amino acid repeat proline, alanine, and serine (PAS) is designed and cross-linked into particles with lysine (K) and polyglutamic acid (E) using mesoporous silica templating. The obtained PASKE particles have low-fouling behavior, have a prolonged circulation time compared to albumin-based particles, and are rapidly degraded in the cell’s lysosomal compartment. When labeled with near-infrared fluorescent molecules and functionalized with an anti-glycoprotein IIb/IIIa single-chain antibody targeting activated platelets, the particles show potential as a noninvasive molecular imaging tool in a mouse model of carotid artery thrombosis. The PASKE particles constitute a promising biodegradable and versatile platform for molecular imaging of vascular diseases.

Liver accumulation of nanoparticles is a major challenge in nanoparticle-mediated delivery as it can reduce the delivery of the nanoparticles to their intended site and lead to liver damage and toxicity. Recent studies have shown that particle engineering, e.g., nanoparticle composition, can influence liver uptake and allow homing of nanoparticles to specific organs or tissues. Herein, we investigated the role of nanoparticle cross-linking on liver uptake. We developed a series of antibody nanoparticles (AbNPs) using various poly(ethylene glycol) (PEG) molecule (e.g., different arm numbers and arm lengths) cross-linkers. Specifically, AbNPs based on Herceptin were engineered with PEG cross-linker architectures ranging from 2-arm (at molecular weights of 600 Da, 2.5 kDa, and 5 kDa) to 4-arm and 8-arm via a mesoporous silica templating method. The molecular architecture of PEG modulated not only the targeting ability of the AbNPs in model cell lines but also their interaction with phagocytes in human blood. Increasing the PEG arm length from 600 Da to 5 kDa also reduced the uptake of the nanoparticles in the liver by 85%. Tumor accumulation of Herceptin AbNPs cross-linked with a 5 kDa 2-arm-PEG was 50% higher compared with control AbNPs and displayed similar liver uptake as free Herceptin. This study highlights the role of PEG cross-linking in receptor targeting and liver uptake, which influence tumor targeting, and combined with the versatility and multifunctionality of the antibody nanoparticle platform could lead to the development of organ-selective targeted antibody nanoparticle assemblies.

Thrombolysis with tissue-type plasminogen activator (tPA) remains the main treatment for acute ischemic stroke. Nevertheless, tPA intervention is limited by a short therapeutic window, low recanalization rates, and a risk of intracranial hemorrhage (ICH), highlighting the clinical demand for improved thrombolytic drugs. We examined a novel thrombolytic agent termed “SCE5-scuPA,” comprising a single-chain urokinase plasminogen activator (scuPA) fused with a single-chain antibody (SCE5) that targets the activated glycoprotein IIb/IIIa platelet receptor, for its effects in experimental stroke. SCE5-scuPA was first tested in a whole blood clot degradation assay to show the benefit of platelet-targeted thrombolysis. The tail bleeding time, blood clearance, and biodistribution were then determined to inform the use of SCE5-scuPA in mouse models of photothrombotic stroke and middle cerebral artery occlusion against tenecteplase. The impacts of SCE5-scuPA on motor function, ICH, blood–brain barrier (BBB) integrity, and immunosuppression were evaluated. Infarct size was measured by computed tomography imaging and magnetic resonance imaging. SCE5-scuPA enhanced clot degradation ex vivo compared with its nonplatelet-targeting control. The maximal SCE5-scuPA dose that maintained hemostasis and a rapid blood clearance was determined. SCE5-scuPA administration both before and 2 hours after photothrombotic stroke reduced the infarct volume. SCE5-scuPA also improved neurologic deficit, decreased intracerebral blood deposits, preserved the BBB, and alleviated immunosuppression poststroke. In middle cerebral artery occlusion, SCE5-scuPA did not worsen stroke outcomes or cause ICH, and it protected the BBB. Our findings support the ongoing development of platelet-targeted thrombolysis with SCE5-scuPA as a novel emergency treatment for acute ischemic stroke with a promising safety profile.

The safe administration of thrombolytic agents is a challenge for the treatment of acute thrombosis. Lipid‐based nanoparticle drug delivery technologies present opportunities to overcome the existing clinical limitations and deliver thrombolytic therapy with enhanced therapeutic outcomes and safety. Herein, lipid cubosomes are examined as nanocarriers for the encapsulation of thrombolytic drugs. The lipid cubosomes are loaded with the thrombolytic drug urokinase‐type plasminogen activator (uPA) and coated with a low‐fouling peptide that is incorporated within a metal–phenolic network (MPN). The peptide‐containing MPN (pep‐MPN) coating inhibits the direct contact of uPA with the surrounding environment, as assessed by an in vitro plasminogen activation assay and an ex vivo whole blood clot degradation assay. The pep‐MPN‐coated cubosomes prepared with 22 wt% peptide demonstrate a cell membrane‐dependent thrombolytic activity, which is attributed to their fusogenic lipid behavior. Moreover, compared with the uncoated lipid cubosomes, the uPA‐loaded pep‐MPN‐coated cubosomes demonstrate significantly reduced nonspecific cell association (<10% of the uncoated cubosomes) in the whole blood assay, a prolonged circulating half‐life, and reduced splenic uPA accumulation in mice. These studies confirm the preserved bioactivity and cell membrane‐dependent release of uPA within pep‐MPN‐coated lipid cubosomes, highlighting their potential as a delivery vehicle for thrombolytic drugs. The synthesis of thrombolytic drug (uPA)‐loaded lipid cubosomes coated with a low‐fouling peptide and incorporated within a metal–phenolic network and their application as nanocarriers of thrombolytic drugs are demonstrated. The metal–peptide–phenolic network‐coated lipid cubosomes display low nonspecific uPA activity, reduced nonspecific cell association, a prolonged circulating half‐life, and reduced splenic uPA accumulation in mice.

We report the engineering of poly(ethylene glycol) (PEG) hydrogel particles using a mesoporous silica (MS) templating method viatuning the PEG molecular weight, particle size, and the presence or absence of the template and investigate the cell association and biodistribution of these particles. An ex vivoassay based on human whole blood that is more sensitive and relevant than traditional cell-line based assays for predicting in vivocirculation behavior is introduced. The association of MS@PEG particles (template present) with granulocytes and monocytes is higher compared with PEG particles (template absent). Increasing the PEG molecular weight (from 10 to 40 kDa) or decreasing the PEG particle size (from 1400 to 150 nm) reduces phagocytic blood cell association of the PEG particles. Mice biodistribution studies show that the PEG particles exhibit extended circulation times (>12 h) compared with the MS@PEG particles and that the retention of smaller PEG particles (150 nm) in blood, when compared with larger PEG particles (>400 nm), is increased at least 4-fold at 12 h after injection. Our findings highlight the influence of unique aspects of polymer hydrogel particles on biological interactions. The reported PEG hydrogel particles represent a new class of polymer carriers with potential biomedical applications.

This protocol describes sortase A-mediated site-specific bioconjugation of single-chain antibodies to iron oxide particles for magnetic resonance imaging and to layer-by-layer capsules for drug delivery.

Imaging of activated platelets using an activation specific anti-GPIIb/IIIa integrin single-chain antibody (scFvanti-LIBS) conjugated to a positron emitting copper-64 complex of a cage amine sarcophagine chelator (MeCOSar) is reported. This tracer was compared in vitroto a 64CuIIcomplex of the scFv conjugated to another commonly used macrocycle, DOTA. The scFvanti-LIBS-MeCOSar conjugate was radiolabeled with 64CuIIrapidly under mild conditions and with higher specific activity than scFvanti-LIBS-DOTA. The utility of scFvanti-LIBS-MeCOSar as a diagnostic agent was assessed in vivoin a mouse model of acute thrombosis. The uptake of scFvanti-LIBS-64CuMeCOSar in the injured vessel was significantly higher than the noninjured vessel. Positron emission tomography (PET) was used to show accumulation of scFvanti-LIBS-64CuMeCOSar with high and specific uptake in the injured vessel. ScFvanti-LIBS-64CuMeCOSar is an excellent tool for highly sensitive in vivodetection of activated platelets in PET and has the potential to be used for early diagnosis of acute thrombotic events.

The ecto-nucleoside triphosphate diphosphohydrolase CD39 represents a promising antithrombotic therapeutic. It degrades adenosine 5′-diphosphate (ADP), a main platelet activating/recruiting agent. We hypothesized that delayed enrichment of CD39 on developing thrombi will allow for a low and safe systemic concentration and thus avoid bleeding. We use a single-chain antibody (scFv, specific for activated GPIIb/IIIa) for targeting CD39. This should allow delayed enrichment on growing thrombi but not on the initial sealing layer of platelets, which do not yet express activated GPIIb/IIIa. CD39 was recombinantly fused to an activated GPIIb/IIIa-specific scFv (targ-CD39) and a nonfunctional scFv (non–targ-CD39). Targ-CD39 was more effective at preventing ADP-induced platelet activation than non–targ-CD39. In a mouse carotid artery thrombosis model, non–targ-CD39, although protective against vessel occlusion, was associated with significant bleeding on tail transection. In contrast, targ-CD39 concentrated at the thrombus site; hence, a dose ∼10 times less of CD39 prevented vessel occlusion to a similar extent as high-dose non–targ-CD39, without prolonged bleeding time. An equimolar dose of non–targ-CD39 at this low concentration was ineffective at preventing vessel occlusion. Thus, delayed targeting of CD39 via scFv to activated platelets provides strong antithrombotic potency and yet prevents bleeding and thereby promotes CD39 toward clinical use.

The ecto-nucleoside triphosphate diphosphohydrolase CD39 represents a promising antithrombotic therapeutic. It degrades adenosine 5′-diphosphate (ADP), a main platelet activating/recruiting agent. We hypothesized that delayed enrichment of CD39 on developing thrombi will allow for a low and safe systemic concentration and thus avoid bleeding. We use a single-chain antibody (scFv, specific for activated GPIIb/IIIa) for targeting CD39. This should allow delayed enrichment on growing thrombi but not on the initial sealing layer of platelets, which do not yet express activated GPIIb/IIIa. CD39 was recombinantly fused to an activated GPIIb/IIIa-specific scFv (targ-CD39) and a nonfunctional scFv (non–targ-CD39). Targ-CD39 was more effective at preventing ADP-induced platelet activation than non–targ-CD39. In a mouse carotid artery thrombosis model, non–targ-CD39, although protective against vessel occlusion, was associated with significant bleeding on tail transection. In contrast, targ-CD39 concentrated at the thrombus site; hence, a dose ∼10 times less of CD39 prevented vessel occlusion to a similar extent as high-dose non–targ-CD39, without prolonged bleeding time. An equimolar dose of non–targ-CD39 at this low concentration was ineffective at preventing vessel occlusion. Thus, delayed targeting of CD39 via scFv to activated platelets provides strong antithrombotic potency and yet prevents bleeding and thereby promotes CD39 toward clinical use.

Molecular imaging is a fast emerging technology allowing noninvasive detection of vascular pathologies. However, imaging modalities offering high resolution currently do not allow real-time imaging. We hypothesized that contrast-enhanced ultrasound with microbubbles (MBs) selectively targeted to activated platelets would offer high-resolution, real-time molecular imaging of evolving and dissolving arterial thrombi.

In the hippocampus, the synaptic vesicle protein synaptoporin (SPO) has been reported to be exclusively enriched in the granule cell axons, the mossy fibers. In this study, we show that in adult rats and mice SPO immunoreactivity (IR) is also detectable in strata oriens, radiatum, and lacunosum‐moleculare of CA1–CA3, as well as perisomatically in the hippocampus proper and fascia dentata. In situ hybridization confirmed that SPO mRNA was present in granule cells and CA3 pyramidal cells but not in CA1 pyramidal cells. Importantly, cells scattered throughout the hippocampal layers resembling the distribution of interneurons were found to synthesize high amounts of SPO mRNA, too. Thus, these findings indicate that SPO expression in the hippocampus was underestimated until now. Moreover, double‐labeling immunohistochemistry and confocal microscopy revealed selective colocalization of SPO and glutamate decarboxylase (GAD 65), a marker for γ‐aminobutyric acid (GABA)ergic terminals. To identify SPO expressing interneurons, in situ hybridization was combined with immunocytochemistry against parvalbumin (PV), calbindin (CB), calretinin (CR), cholecystokinin (CCK), and vasoactive intestinal polypeptide (VIP). We found that SPO transcripts were differentially expressed by various interneuron subpopulations in the hippocampus of C57Bl/6 mice (PV 44.2%, CB 46.3%, CR 19.3%, CCK 38.6%, VIP 59.9%). Immunoelectron microscopy for SPO labeled synaptic vesicle profiles in distinct symmetric and asymmetric synapses. In conclusion, our data demonstrate that hippocampal principal cells and interneurons display a variety of synaptic vesicles that are likely to contribute to the functional characteristics of their output synapses. J. Comp. Neurol. 452:139–153, 2002. © 2002 Wiley‐Liss, Inc.

Several cytochrome P450 subfamilies are inducible by specific exogenous compounds like the antiepileptic drug phenytoin. Some of these P450 enzymes are involved in the metabolism of gonadal hormones also contributing to neuronal differentiation. CYP3A enzymes have the capacity to catalyze the hydroxylation of testosterone and a wide variety of therapeutic agents, but little is known about the expression and potential function of this subfamily in mouse brain. Here, we report the identification of mouse CYP3A isoforms, their induction and localization in mouse brain. Western blot analysis with anti-CYP3A1 antibodies revealed the phenytoin-inducible expression of CYP3A in brain microsomes, and also a constitutive expression of members of this subfamily in brain mitochondria. Using RT-PCR with a consensus primer pair for known mouse liver CYP3A-isoforms we could demonstrate the expression of CYP3A11 and 3A13 mRNA in mouse brain. Finally, using double immunofluorescence labeling we analyzed the histoanatomical distribution of CYP3A throughout the brain with confocal laser scanning microscopy. We found strong immunoreactivity in neurons of the hippocampus and hypothalamic areas which are sensitive to steroid hormones. CYP3A immunoreactivity was apparent also in neurons of the cerebellum, the thalamus and the olfactory bulb. Non-neuronal expression of CYP3A could be found in some astrocyte populations and in vascular as well as ventricular border lines. The presence of CYP3A predominantly in neurons but also in cells contributing to the blood-brain and blood-liquor barrier suggests important roles of this subfamily in mediation of steroid hormone action in mouse brain as well as in preventing the brain from potentially cytotoxic compounds.

One dose of rHDL after myocardial ischemia increases both cardiac glucose uptake and heart function in healthy and insulin-resistant mice.

Therapeutic nanoparticles hold clinical promise for cancer treatment by avoiding limitations of conventional pharmaceuticals. Herein, a facile and rapid method is introduced to assemble poly(ethylene glycol) (PEG)‐modified Pt prodrug nanocomplexes through metal‐polyphenol complexation and combined with emulsification, which results in ≈100 nm diameter nanoparticles (PtP NPs) that exhibit high drug loading (0.15 fg Pt per nanoparticle) and low fouling properties. The PtP NPs are characterized for potential use as cancer therapeutics. Mass cytometry is used to quantify uptake of the nanoparticles and the drug concentration in individual cells in vitro. The PtP NPs have long circulation times, with an elimination half‐life of ≈18 h in healthy mice. The in vivo antitumor activity of the PtP NPs is systematically investigated in a human prostate cancer xenograft mouse model. Mice treated with the PtP NPs demonstrate four times better inhibition of tumor growth than either free prodrug or cisplatin. This study presents a promising strategy to prepare therapeutic nanoparticles for biomedical applications. A polyphenol‐based therapeutic nanoparticle system for cancer therapyis prepared by self‐assembly and emulsion templating. The nanoparticles have a long blood half‐life, accumulate in tumors in vivo, and exhibit better inhibition of tumor growth than either free prodrug or cisplatin.